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Documente Profesional
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A,k no\1i lco.l~emenIS •
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Preface
InuothK110D 7
r an 1 Jl
The red cells and their precursors
,\'0 , ,"<11fo rms and ahnomwl \'arjaTlfJ
Pari 2 «r
., Granu lUl;)tes. monocyte» and
,ne!lilk ,t TytX-Y' CS
... N urnllll and nhnormal [n rm'
" Parl3 f'JI
...yrnphocy tcs , plasma cells a nd
their de rivatives and prccu r""rs
in t>1<ltlo.l a nd bone marro w
Nomla' and otmormat for....,
Part 4
:\h lol:c ll.:incous cell~ trom hone marro..
or Mood S OU:"IS. relicul()oenJot he lial
cc 11s. oaeoclasts and O!ItlNh!as,s.
foret,," cells and parasites
PanS 32<
Im prin ts a nd sections 0 1lymph nodd>
aed spleen
APf'C odi x: staining techniq ues .i T·
A hbrc\ iat ion s
Index
Part 1
The nome nclat ure of red-cell prec ursors is co nfusing. Reti culocytes spe nd up to two days in the bo ne
The earliest recognizable member of the red-cell series. marrow before being released into the peripheral blood.
the procryrhro btast , has the cytoplasmic basophilia. the T here they make up nor mally less than I % of the red-
nuclco latcd and moderately lcptochrornatic nucleus cell populatio n and within a no ther o ne to two days lose
and the large cell size gene rally characteristic of the remn ant s of cytop lasmic basophilia which give them
primitive ce lls. It gives rise to a seq ue nce of nucleat ed their cha racte ristic staining properties. and become
cells. the erythroblasts , which progressively develop orthochromati c mature red ce lls.
increasing ly pac hychromatic nucl ei . lose their nucleoli Mature red cells survive some 120 da ys before de-
and their cyto plasmic basophili a and acq uire a rising struction. They ar e normally circula r and fairly uniform
haemoglobi n con te nt, Th is seq uence is subject 10 an in diam eter. but arc readil y distorted by exte rna l
arbi trary divisio n into stages. the co mmonest division pressure . as fro m neig hbo uring cells in a smear. Their
being into three : stru ctu re as bico ncave discs leads to wea ker eosinophil
staini ng at the centre than at the pe riphery. a fea ture
l. Th e baso philic o r early ery throblast . or no rmoblast which is least prominent at the tail of a blood smea r
A whe re the ce lls are most spread o ut and flatten ed . In
2. Th e polychro matic o r inte rmed iate e rythro blast . o r the body of the smea r it becom es more conspi cuou s and
normob last B this normal phenomenon must be appreciated and
3. Th e o rthoc hromatic or late ery thro blast. o r norm o- distinguished from hypochromia .
blast C.
Abnor ma l var iants: nucleated prec ursors
T he re are objectio ns to the use of many of these ter ms.
but the y are all so firmly entrenched in co mmon usage Th e chief cytological variants involving altera tion in
that they must he accepted . When autho rs use diffe rent morph ology rathe r than numbers or relative pro portions
or more elaborate staging and nomenclature they usually o f c ryth rob lasts are as follows:
de fine their terminology. but those who use any of the
syno nyms abov e expect them to he under stood without Macronormoblasts. Cells havin g the general nuclear
furt he r explanatio n. and cyto plasmic morpho logy of normoblasts but an
T he proeryth robla st is not itself the function al ste m increased average cell diamet er. Th ey occur especially
cell se rving as a self-maintai ning progenito r of the in sta tes of active ery throhlast ic hyperpl asia without
nor mo blast series. but is de rived from an earlier haem atinic defects. such as haemolytic anae mias , but
func tiona l myeloid stem cell of unide ntified ' mo rph- may be seen also in the early stages of d isorders that
ology. having pluri pote nt ial capacity for giving rise to subseque ntly become megalob lastic,
cells of eryt hro id. gra nulocytic . monocytic and
megakaryocyte -plate let lines. Mcgaloblasss. The esse ntial morphological cha nge in
Kine tic studies with radi o-isotopically labelled ce lls mcgnlobla sts as co mpared with normobl asts lies in the
suggest that four cell cycles culminating in mitoses more open chromatin pattern of the nucleus at all
occur duri ng developme nt from proerythroblast to late stages of developm ent. T he re is often a de gree of
no rmoblast. three at the procr yth robl ast and ea rly haemoglobini zation of the cytoplasm which appea rs
basophilic no rmoblast stages. and the last at the po ly- excessive for the sta te of nucle ar maturation . and later
chro matic intermediate nor moblast . stage. Nests of megaloblasts may appea r fully or thoc hromic. Mcgalo-
erythroblasts of diffe re nt stages of maturity co mmo nly blasts are lar ger than no rmo blasts of comparable
occur in appositio n around a ce ntrally situated reticulo- maturi ty_ Th ey may sho w mitoti c irregularities . with
endothelial cell . T ra nsfer of iro n may he effect ed in o nc multipolar mitoses.' sometimes asymmetrical. producing
or ot her d irectio n. and the ce ntra l macrophage is ofte n un equal daughter ce lls.'Cyto kinesis may not occu r and
rich in stai nable free iron . Late normoblasts do not giant cells with two or mor e nucle i. so metimes uneq ual
unde rgo a furt he r cell cycle but lose their nucl ei by in size . result . Chromosomes or fragments of chromatin
ext rusion and give rise to marrow re ticulocytes . may become separated from the spindle and constitut
accessory small nuclear masses : Th ese ' Howcll-Jolly or co nta in abnormal inclusio ns. such as the r
bodies' may rem ain in the red cells afte r the mai n bulk chro mat in materia l of the Howell -Jolly bodies.
of nuclear materia l has bee n extrude d. Rosette for- All these ab nor malities may be seen in vaf)i n ~
deg ree and frequ ency and severa l of the m may occur
mation. wit h several chrom atin masses linked by bridges.
togeth er in the same smea r.
may follow a halt of the mito tic process in metaph ase .
Megalo bla ts are present in the marrow in- state s of
B 1:, and folic acid deficie ncy. the com mo nest of which Disorders tnvotving chiefly the red- cell series. and thri r
are Add isonian pernicio us anae mia and cond itions of associated main cytological features
gastro-i ntcsti nal malabso rption d ue eithe r to disease or -
to ga_trie o r intestinal resectio ns. Th e folic acid
Megalo blastic anaemias. These disord ers are charac-
deficiency of pregnancy may lead to megalob lastic
terized by striking erythro blastic abnormalities arising
anae mia. Megalo blasts arc see n also in so me cases of
from deficiencies of e ither vitamin B l 2 or folic acid . The
refracto ry siderob lastic anaemia and in £!)'1bmclJliL
und erl ying bioch emical defect involves a slowing of
m~s . The y may be seen in acute leukaemi as.
DNA synthes is in (he S phase of the cell cycle . due
especially followin g treatme nt with antimet abolitcs .
principally to defective formation of thymidylate . which
is normally depend ent on bot h BJ2 a nd folate as eo-
Micronor moblasts. Sma ll normob lasts with a tcnd ency
facto rs in its synthesis. Nuclear division is thus retarded .
10 ragged cyto plasmic ou tline a nd irre gular sta ining a re
and the patt ern of nucle ar chro matin disposition and
found in slates of iro n deficien cy. Th e cha nges a re most
nuclear- cytopl asmic synchro niza tio n in develop ment is
evident in interme diate , and late normob lasts. which
affected . with the result s illustrat ed in 39-69. In fact,
also show defective haemog lohini zation .
the changes of megalohl asto sis. with increas e in ce ll
Side roblasts. Normobl asts containing free . non -haemo- size and opening of the nuclear chroma tin network . arc
globin, iron are best detected by the use of Prussian detect ahie in most prol iferating cells. such as those of
blue staining , but occasionally coarse accumulat ions of the gut mucosa. and includi ng granulocyte and platelet
free iron may be visible in Romanowsky preparations. precur sors in the bo ne marrow.
The findings in bo ne marrow aspirates and biopsies
include high ce llularity . the hyperplasia involving mainly
Bizarre cytological variants. G ross mitoti c and nuclear
the red -cell series. with mycloid-erythro id (M :E) ratio
abno rmalities. of the same kind as describ ed und er
comm onl y I :1 or even 1:2 - co mpared with the nor mal
megaloblas ts but much more ex tre me in degree . occ ur
in e rythraem ic myelosis. range of 5-20: 1. The re is a shift to the 1eft in red -eell
pre cu rso rs. with a n increased proport ion of large pro-
Reticule cytes e rythro blasts and ea rly basophil ic e rythrohl asts,
num e rous mito tic figures - often sho wing Howell-J olly
bodies - and the typ ical prem ature haemoglobinizat ion
Blood normally contai ns less than I % of ret iculccyt cs
best seen in interme di ate c rythrob lasts. G iant myclo-
and an increase in numbers indicates a heighten ed
cytes and mct am yelocyte s ca n be fo und. as can multi-
o utput of young red cells from the bon e marr ow. Th is
nuclea ted po lymorphs ( 7~) . and par allel changes
may occur durin g the co rrectio n by increase d marrow
involving asynchrony of nuclear- cytoplasmic maturation .
activity of any anaemic sta te . whet her spo ntaneo usly
with gra nular cyto plasm and relatively primitive nuclear
(as afte r an acute haem orrhage ) or following trea tment
structure. so metime s in mitosis. occur in promegak ar-
(as in pern iciou s anae mia aft er BJ2 or iro n deficiency
yocytes , while nuclear hypcrse gment at ion may be
anae mia afte r iron) . Reticul ocyto sis is especia lly
co nspicuo us in mature mega karyocy tes. Macrop hagcs
promine nt in haem olytic ana e mias , whe re great
usu ally show an increase d load of haem osiderin . with
e rythropoie tic activity att empts to co mpe nsate for the
the fast turnover patt ern of small particle s. resulting
shorte ned life spa n of periphe ral erythroc ytes .
from the con sider able co mpo nent of ineffective
In Ro manowsky prepara tion s reticulocyrcs may
erythro poiesis and co nseq uent intra med ullary break-
sometim es be detected as po lychro matic cells. with a
down of non- viable red-ee ll precurs ors.
blue or pu rple tinge. -or much less often may show a
The periphe ral blood find ings include a more or less
scatte ring of fine blue ' baso philic stipples'. Th ey arc
severe ana emi a with normoc hromic but ma rkedly
best recognized . ho wever . by the use of supravit al
macrocytic erythrocytes. the mean cell volume (MCV )
staini ng with brilliant cresyl blue . which reveals a
usually exceedi ng 115 fe mtolitres (ft) and reaching as
netwo rk of fine filaments and dots. Re ticulocytes a rc
high as 150ft. Th e mean ce ll haemoglob in (MCII ) is
ofte n a little lar ger in diam ete r than mature red cells.
raised abo ve 32 pico grams (pg) . but the mean cell
Matu re red cells haemoglobin concent ration (MC HC) is normal . not
over 36% . A nisocyto sis and poi kilocytos is appear ea rly
in the co urse of the anae mia and become increasingly
Red ce lls may diffe r from norm al in their appea rance in
seve re . large ovalocytcs bei ng particularly charac-
stained films by varia tio ns in size (a nisocytos is. micro-
teri stic. but other shapes . including tear-drop poiki lo-
cytosi s. macrocytosis). in shape (po ikilocyt osis. ellipto-
cyte s, may also be see n (711-73) . Th e red cells may
cytosis. sickle cell formatio n. cre scent formati o n.
occasion ally show basophilic stippling ( 126) and I lnwcll-
crenatio n) and in dept h of sta ining (hypoc hro mia .
Jolly nucle ar fragmen ts ( 146). but reticulocytes arc not
ani sochrnmi a . spheroc ytos is. target cell formatio n) .
usually increase d and may be depresse d. As the anaemi a
Th ey may also und ergo fragme ntatio n (schisto cytosis)
pro gresses. occasional late eryt nrob lasts with residual priate immunologica l studies. e nzymology. or haemo-
meg alobl ast ic features appea r in the circ ulation . globin electrop horesis. as the case may be . Neverthe-
Leuc openia aro und 2-t x 10"'/1 is usual in mega lo- less. red-cell morph ology can offe r some useful
blast ic an aemias. pe rha ps especia lly in perniciou s pointers .
anaemia . with neutrop e nia and a relative Iymp ho- Spherocy tes ( 135-137). apparen t in stained blood
cytosis . The neutrophils show a shift to the right with smears as small. deeply stained red cells. are always
increase d nuclear lobulation. most cells having fOUT present in HS and commo nly de tectab le in A IHA .
lobes a nd some hypcrse gmentc d cells having five or six. Elliptocytcs (138) formin g more than 5% of the red-
l Neutrophils may also manifest cellular gigantism - these cell populat ion are virtually confined to her editary
exce ptio nally large cells are known as macrop olycytes . e lliptocytosis. but a few marked ly oval or cigar-sh aped
Platelets may be present in normal number s. but red cells may be found wheneve r gross anisocytosis is
thrombocytope nia is more commo n and is sometimes presen t, as in seve re megalobla stic or iro n deficien cy
severe . while giant platelet s and occas iona l megak ar yo- anae mia and in thalassa emia. and a localized elllpro-
cyte fragme nts provi de furthe r evide nce of megalo- cytotic appe arance may be prod uced in some areas of a
blastic dyst hro mbopoi esis. smear as a spread ing artefact (139).
Echinocytes or 'burr cells' , erythrocytes with numerou s
Haemol ytic anaem ias. Excessive destruction of red cells
surface undul at ions or spicules ( 141), may be pro-
arises from two ver y broad gro ups of causes. intri nsic
duced by exposu re of nor mal 'discocytes' to an alka line
and ext rinsic. In the first group are defec ts of the cell
pH or to vario us cchinocytoge nic factors such as bile
constitution. ranging from heredit ary spherocy tosis acids, lysolecithins and fatty acids. Although the change
( HS). enzyme defects such as pyruvate kinase (PK) and is rever sible, it is associa ted with a tend ency to sphe ring
glucose -6-phos pha tc de hydroge nase (G6 PD) deficien -
a nd subseq ue nt haemolysis. In path ological states.
cies, and paroxys mal noctu rnal haemoglobinuria
echinocytes occu r especia lly in uraem ia. chronic liver
(PNH) . to haemoglobin opathi es such as the th al- disease , and acutely in severe bums and toxic infections.
assae mias and sickle cell disease. The secon d grou p
Acanth ocytes or 's pur cells". erythrocy tes with sha rp
includes acqui red auto- immune hae molytic anaemias spiny irreg ular projections ( 1.13). arc also associated
(A IHA) due to warm lytic antibod ies active at the red- with a haemolytic tende ncy. and occur in heredita ry
cell surface: cold hae magglut inin disease (C HAD) due acanthocytosis or congenital abeta ltpopro teinacm ia.
to high titre agglutinating antibod ies. usually with ant i-I and may develop in liver disease or after splenec tomy.
specificity: and hae molysis seconda ry to toxins. foreign Stomatocytes , cup-shaped erythrocytes which in
an tibodies as in mismatched transfusions, and circu- stained smears sho w an elo ngated slit of central pallo r
latory impedime nts such as occur in microangic pathy. instead of the norm al circular area ( 128 and 158).
thromh otic thrombocytopen ic purpura (TIP) . ren al are found conspicuously in heredita ry stomatocyrosis.
glomerular disease . dissemina ted intravascu lar coagu-
and not infrequently in haemolytic anaemias with altered
lation ( DIC) and artificial vascular prost heses . membra ne pe rmeabil ity. A few may usually be found in
All these conditions share features in common. arising normal blood smea rs. and thei r numbers may be
from the increased breakdown of red cells. with resulting increase d in hep atic cirr hosis and in many generalized
raised levels of seru m bilirubin and disappe ara nce of neoplastic sta tes .
haptoglobins, and attempt s at compensatory marrow
D repanocytes or sickle cells ( 155) are characteristic
erythro poietic hyperpl asia. with norm oblastic prolifer-
of haemoglobin S disease , being produ ced by the
ation. a redu ced M:E rat io. and an outpo uri ng of
crystallization of reduced HbS within the cell.
rcticulo cytes, sometimes accompa nied by late normc-
Codocytes or target cells . bell-sha ped eryt hrocytes
blasts. into the pe ripheral blood . The intense normo-
with a striking target appeara nce in stai ned smea rs
blastic hyperplasia. with oblite ration of fat spaces and
(150. 151 and 153). are especia lly promin ent in thal-
100% cellularity in trephin e sections . from a seve re
assaemia, but are found also in othe r haemoglob ino-
hae molytic anaemia due to PK deficiency . is shown in
pat hies , such as HbC and HbSC disease .
19 and 20. and a less marked but substantial erythro id
Leptocytes, or thin flat ery throcytes (150) . occur in
component in sections from a case of AIHA seco nda ry
the same conditions as target cells. and also in liver or
to non-Hodgkin's lymphom a (N HL) is shown in 21 and
gall bladd er disease with biliary obstruc tion. and after
22.
splenecto my.
Tbe anae mia of hae molytic disease is usually normo-
Schistocytes , red cells with irreg ularly fragme nted
cbromic and neither macrocytic nor microcytic. although
sha pes due mostly to mechan ical da mage (134. 152. 163
~1~ and poikilocytosis are common. Neutrop hil and 164). a rc found especia lly in micro angiopath ic
~1osb may be seen. especially in acute haemolytic haem olytic anae mia and the related states o f DIC . TIP
reaction . but the count is ofte n normal. Apa rt from
and microva scular nep hropath y.
(he acqu ired hae molytic processes associated with
Heinz bodies (160) may be dem onstrable in the red
microan giopa thy, TIP and DIC. where there is seve re
cells in supravitally stained preparation s of blood from
thrombo cytope nia . platelet levels show no consiste nt
patient s with haemol ytic anae mias due to G6PD
trend . The chief morpho logical findings of differential
deficie ncy. unstable hacmoglobins, chemical dam age or
diagnostic value among this wide range of diseases are
thalassa emia. They arc someti mes remov ed by the
therefore to be found in the circu lating ery throcytes
spleen a nd therefor e are parti cularl y conspicuous after
themse lves. alth ough they are rare ly pathognomonic
splenec to my.
and need confirm ation or supplementation by appro-
· leucocyte alkaline phosph atase (LA P) sco re.
Haemoglobin H inclusio ns. which stain with supra-
vital cresyl blue a fte r prolo nged exposure ( 162), are A late transition to a myelo fi brotic. state occ urs in
found in the red cells of alpha-thalassaemia. around 15-20% of PRY cases . and when this change
ta kes place the marrow sections show . of course.
A part fro m these vario us pointers. the differential increasing reticulin and fibrosis (as in 30). and the
diagnosis bet ween haemolytic disorders is not greatly peripheral red cells manifest anisocy tosis and teardrop
assisted by morphological featu res present in the pe ri- poikilocytosis. Terminal tr ansformation to a pre-
pheral blood. and the only contributory differential leu kaemi c myelodysplastic state or to fra nk acute
findi ng in the bone marro w. where normo blastic or myelo id leukae mia (A ML) may also occur.
macrono rmoblastic hyperplasia is the co mmon ru le. is Seco ndary polycythaemia or erythrae mia resembles
the existence of strong periodic acid-Schiff ( PAS) PRY in having raised haemoglobin. haematocrit .
posiuvity in the erythroblasts of beta-thalassaemia erythrocyte count and red -cell mass. bu t differs in the
major and to a lesser exte nt in th ose of be ta- tha l- absence of leucocytosis and th rom bocytosis. and . when
assaemia minor. Ot her haemo lytic sta tes o nly ra rely d ue to chro nic cardiovascul ar or pulmonary disease , in
sho w any eryt hro blastic PAS pc sitivity. altho ugh this showing decrea sed arterial oxygen saturation. Morpho-
was a fea ture of foetal haemo lytic d isease . now fortun- logically. erythroid hypcrpla-..ia predominates in the
ately preventable and therefore uncommon where there bone marrow . withou t paralle l increase in o ther
are adeq uate medical services. lineages. and the overall cellularity of sections is
ge nerally less marked than in PR Y. as exemplified in
Aplastic and hypoplastic anaemias. Despite the titular 23.
e mphasis o n anae mia. these diseases arc usuall y
pancyto penias, with acco mpa nying neut ropen ia and Congenital dyserythrop oietic anaemias (CDA s). This
thrombocytopenia. all beco ming progressively mo re group of relatively uncommon disorders is of parti cular
severe as the disease adva nces . and associated with interest to the cytologist because the dyseryt hro poie nc
failure of myelopoiesis. The bone marrow is poorly abnormalities manifest in the erythroolasts are so striking
ce llular. with a predominance of fatty spaces and a and bizarre . There are thr ee main varieties of CDA .
reactive o r inflammatory inliltrat e ranging from a few Type I affects the later basophilic and intermediate
residual islands of Iymphocytes and plasma cells to erythroblasts predominan tly. with megaloblastoid
widespread d iffuse Iympho plasmacytosis. Marrow features and spo ngy chromatin patterns sometimes giving
aspirate s are generally acellular. and firm diagnosis an appearance of irre gularity in the nuclear membranes.
req uires trephine biopsy. Typical appearances at and with occasion al internuclear chromatin hridges.
diffe rent stages of disease arc shown in 24--27. Type 11 . the com mone st form . affects later ervthroblasts
The anae mia itse lf is usually no rmocytic and normo - especia lly. with conspicuous muitinuclea n ty and with
chromic with litt le anisocytosis o r poikilocytosis . and double nud ear membranes seen b)' electro n microscopy.
re ticulocyres arc scanty. In this type of CDA the erythrocytes are susceptible to
Pure red-cell aplasia . witho ut concomitant decrease haemolysis in an acid medium. hence the alte rnative
of granulocyt es and plate lets. and wi th a cellular marrow na me . HEMPA S - hereditary erythroblastic multi -
de void of red -cell precurso rs. occ urs o nly rarely. nuclearity with positive acidified serum test. Type III is
characterized by giant eryth roblasts. up to 70 micro ns
Primary and seconda ry po tycytnaemios. Polycythaemia in d iamet er. with either num ero u separa ted nuclei or a
rubra vera ( PRV) o r primary proliferative poly- single rnultiplo id lobul ated nucleu .
cytbae mia is characte rized by high haemoglobin . Exa mples of all these vari an ts of CDA and their
haematocrit and red-cell count . a raised red -cell mass. mai n cytological and cytochemical features are illu-
oonnal arterial oxygen saturation. rommonly significant strated in 78-88 .
plenomegaly. and usually elevat ed to tal leucocyte and
platelet counts. It is thus a pancytosis. and the high Erythruemic myelosis and erythroleukaemia. Alt hough
peripheral blood figures reflect a chron ic neopl astic an acute neoplastic state with vast preponderance of
myeloprolife ra tive state involving all cell series in the erythroblasts has lon g been recognized as erythrae mic
bone marrow (28 and 29). whe re fat spaces arc much myelo sis or Di Gu glielmo's disease. it is probable that
reduced in trephine sectio ns and trilineage hyperplasia this occ urs rarely if ever as a pu rely monolineage
ma) approach 100% ceJl ularity. Cytogenetic st udies in disorder. but that it is almost always. if not always. a
the stable ph ase of PR V re vea l non -random chro mo- bilineage o r trilineage disease from the sta rt, with
somal abnormalities in many patients. trisomy H. trisomy involve ment to a greater or less exte nt of the gra nu -
9. 5q- and. especially. 2Oq- being the most com mon . locytidmonocytic line and the megakar yocytic line .
thus co nfinning the clo nal neoplastic nature of the The name 'eryth roleukaemia' is the refore strictly more
disea se . Morpho logica lly. there arc few changes from co rrect in all cases. although erythrae rnic myelosis is
normal. with e rythro poiesis normoblastic and eryt hro- still co mmonly us-ed for cases where the erythroid
cytes no rmoc hro mic and normocytic. and with bot h co mpon ent is clearly dominant.
gra nulopoiesis and th rom bopoiesis regular and cyto- Cytoge netic abno rma lities are found in the maj ority
lo gically typical. apart from an occa sional increase in of cases . including (he co mmones t AM L alte ra tion.
basophils and a few gian t plat elets. at least during the trisomy 8 (8 + ), and defects of 5 and 7. but multipl e
main course of the disease . The one abnorm al cyto- nume rical and structural abe rra tions are usual. Th e
chemical fea ture almost invariably fo und is a raised cyto logy of the neoplastic erythroblasts in the bo ne
marrow and in the blood . where they may be numerous. cvt osi s and poikilocytosis ( 0 1 and 133 •
vane... fro m relauvelv normal to zrosslv bizarre (89-95) . indices, ~t C \' . ~ICH
and MCHC. aD O«_I<cla::1_ .....
C~ tochemica llv. the~ usually show conspicuous PAS The leucocyte co unt tends to be 10.... or DOl t
IX Iti\ ity (98- 102). and ofte n strong localized acid platelet count is often raised . U ntil ,rOIl U·. .......
pho-phatasc reactions ( 103 and I~ ) . started the rericulocvte count is 10.... and
pe ripheral polyc hromasia . Ervthropoese, iD
Refractory sideroblastic illJaemias (RSA 'i ). These dis- ma rrow is hype rp last ic. but with a bnonnal ~106~
o rde rs are characterized by a n overload of iro n in the e ryt hroblasts being small. poorly hac TTlOgIobiDizrd.
bo ne marrow ( 112) . and by hypercellular micronormo- ofte n with irregular and ragged outlines ( I aDd I
bla stic or occasiona lly rne galo basti c e rythropo iesis. The y may contai n PAS-po siti\·e mat e rial (l J
Coa rse sidero tic gra nules . demon strabl e by the Prussian marrow is not abl y de void of free iron whether in ~
blue stain. a re found a rra nge d in a rin g around th e phages o r e xtra-ce llula rly in part icles a nd flee s ( I l l
nucleus of man y c rythro btas rs. T he a ppea ra nce is mo st and there ar c virtually no sideroblasts or siderocy
st riking a nd involves the highest proportion o f These cyto logical features a re really diagnostic.
c ryth rob lasts in cases of primary acquired RSA . a nee- co nfirmat ion is give n by the findin g of lo w serum lTOO
pla...tic mye lodys plastic sta te which may tran sform to a nd rai sed total iro n-binding ca paci ty (TIBe) , .... ltb
• A ~ lL. Exa m ples o f the cyto logy a nd of ringed sidcro- decreased sa tura tio n and a low se ru m ferritin level.
blu ... ts are shown in 113-116. In hereditary RSA and in Other microcytic hypochromic a na e mia- generally have
acq uired cases seco nd a ry to age nts such as iso niazid, clear distinguishing features : the anaemia of chronic
cyclose rine . chloram phe nicol. a lco ho l a nd lead. th ere disorders. for e xa mp le. shows a n increase o f large-
a rc ue ne ra l!v fewe r sidc roblasts. a nd ontv interme dia te particle free iron in th e mar ro w. decreases in both serum
a nd ~I ate ervth roblas ts a rc affected. . iro n a nd TIB e . and norm al or ra ised ferritin . while
PAS posit ivity is found in a p ropo rtio n of the crythro- thalassaemia a nd HbC a nd H bS C diseases show con-
blas ts in so me cas es o f RSA . alt ho ug h the reaction is spicuo us target ce lls a nd increased marrow iron .
usua lly m uch we ak er than in c rythracrnic crvthroblasts
1118). Leucoervshrobtastic anaemia (myelophthisic anaemia.
myeloid me/aplasia). T his co mp lex form of a naemia -
A1W('mia ofblood loss und iron deficteucv. Acute blood with th e a ppea ra nce of bot h e ryth roblas ts a nd granulo-
lov, insuffici e nt to produce ca rd iovascular collapse cytc prec urso rs in the periphe ral blood . and with
re... ults in haemodilution and th e de velop me nt. o ver a moderat e ly seve re a nisocytosis a nd po ikilocytos is . the
pe riod of 2-1. hours or so. o f an anaemi a which is normo- latt er ofte n including dacr yocytcs - a rises when the
ch ro mic and normocyti c. hut usu all y associa te d with marrow is infiltra ted with met ast ati c tumour cells or
come increase in rcti culocytc s and a pa ralle l rise in othe r forei gn tissue . Megakaryocyt e fra gmen ts may he
le ucoc yte and platelet co unts as marrow act ivity is see n in the blood , and platele t morphology is frequ e ntly
-tim ulare d . a bnormal with distorted o r giant forms .
Ch ro nic blood loss leads to the developme nt of iro n Bone marrow aspira te s o r trephine biopsies re vea l the
dcficic ncv a naem ia. a nd is th e commonest cause by fa r nature of the infiltrate . Exampl es of ca usative invasion
of this d;s0rde r. cha rac te rize d by fallin g hae moglobin a rc illu strat ed and de scr ibed in Part -1.. but the resulting
.... 1Ih initia lly microcy tic a nd lat er hypoch ro mic red cells red-cell morphology ge nera lly resem bles that show n in
.... ith increasing ce ntral pall or a nd progressive aniso- 1.' 1.
•
- (
•
•
I 2
I
t
i6
,I , s
I
.. .. . h w 5ayJl
7
I'
10 11
-
I~
13
•
•
• •••
•
•
22. A higher-power view of part of the field sho wn in 21
'A herevirtually all of the ce lls can be c1carl y identified as
o f the erythroid series. Th e largc proeryth ro blasts with
leptochro matic nuclei and conspicuo us nucleo li arc •
pa rticularly well shown .
•
• •
•
• •
/
33
26
35
•
• •
•
to:
•
•
41
pa n icularly co m m o n .
-15 -16
4S and -U.. T wo fields fro m a per iphe ral blo od h uffy coat
smear from a pat ient with chronic mye loid leukaemia
(C t L) treated with the antimetabolite hyd roxyurea .
In -15 two basophil's and two neut roph il stab ce lls
accompany two lar ge intermedia te mc galo blasts . a nd in-
..u, arc sho wn a sta b ce ll. two neutrophil polymorphs. a
met am yeloc yte and two late mcg!Uohl asts. widely
sep arated but with inco mple te nuclea r' reconstitu tio n
after a p rece ding mitosis. . /
30
so
3
55
53
I
• I
56
53 and 54. Late megalo blast!'. showin g typical nuclear
rosette for mation . probably arising from an incomplete
mitos is with hold- up at metaphase .
.. ...".,.
~"" •.-'"L
~ ~
-
.
• ~:"$"
, 1"-
65
6.'. A range o f cryr hrob lasts with minimal mc galo -
blastic cha nge . T he confus ing terms ' intermed iate' and
' tra nsitio nal" rncgalobl asts arc so metimes applied to
such c rythroblasts, whet her ea rly or late in the rnuturu-
tiu n sequ en ce . A late megaloblast with densel y pycnou c
pai red nuclei shows hasop hilic stippling of th e ortho-
chro matic cyto plasm . At the top o f the field is a
ne utro phil metamyelocyte .
•
3
~ .
83
81. CD A, type II. A furthe r example showing two
erythrob lasts with doubl e nuclei and on e with four
uneq ual nuclei.
..
•
91- 94. Hi ghly ab no rmal e rythro hlas ts fro m c ryt h-
rac micrnvclosis.
Cc llula"r gigantism. mu ltiple a nd irr egu la r nucl ear
ma..-.c:s and ab norma l mitotic figures occ ur in c rythro -
bla cts at d ifferent stuucs of nuclear ma tur at ion a nd
~ to plasrnic haem oglobin izat io n .
Ab norma lities as st riking and bizarre as these are
virt ua llv restricted 10 c rvth racrnic mvc losis and e rvt hro -
Icul acmias. hut they ' are probab ly not cssc~( ia lly
diffe re nt in kind fro m the simi lar but much less frequent
nd promine nt nuclea r ab norma lities illust rat ed earlier
a, occ u rring in the co mmon megalo blastic anae rnias , or
the freq ue nt hut more reg ular rnulu nuc lca rity of C DA .
'"
- .'
95 96
-.
and 99. Two examp les of PAS positivity in eryt hro- .---- .. ilK)
IS • from erythraernic myelosis. Coa rse granul ar
positiviry is conspicuous in earl y erythroblasts. with •
diffuse positivity. sometimes very intense . in later
eryt hroblasts. Both these fields contain gran ulocytcs ,
four mctarnyclocytes at the top and a possible myelo-
blast ne xt to the diffusely PAS-positive late eryt hroh last
. . and three myelobla sts or early prom yelccyres and
a tab cell in 99. The se cells show the expect ed pattern of
PAS positivity for their respective stages of maturity.
and thei r presence again demonstrates the existence of
multilineage involve ment in eryth raemic myelosis.
-5
101 102
10-'
-16
106 ~1 07. Erythroblasts fro m th e bo ne marro w in IIIK
~de~i~ ncy anae mia. Erythro poiesis is normoblastic.
1 the no rmo hlasts tend to he sma ll. with ragge d
lliI1t.~ arid defective hacmoglon mlzation . The
Ill. joio-crvt broid ratio appea rs no tab ly low in the se
fiellh. with few gra nuloc ytcs prese nt . bu t although this is
ot en th e case in iro n deficiency a nae mia. ery thro id
b~pe rp l as i a is not always cons p icuo us .
--=_..~
109 III
110
..
• :
~.
,-
.' I
... ..
; "..
~
I'
.- . ..
~
.. .
:.~. r.
~ .
"
• ,
•
. ...
•
116
/
- •
•
117 11 . Free iron in erythroblasls (siderohlasts) with a
on-ringed distribution. Th is exa mple is of the 'normal'
pattern of free iron dist ribution in ery throblasts , hut the
positive cells (sideroblasts) arc mo re nu merous than
usual.
118
50
I 11
"
.-
:
,
: •
12 1
" .
51
122 123
•
l
., I
•
•
I~ 1~6
129 ~
.-•
54
•
56
136
• •
••
•
.-•• •
•
- ..
IJS a nd 136. Respective ly low - and high-power views ol~
sDherocytcs. Th ese are red cells wiffi more sphe rica l
1.17
•
•
•
• •
1 ~2
1 ~2 .
Ery throcytes showing a tend ency to cre nat ion and
~ eVidence o t ph ySIcal da mage o r distortIOn--
-ing the ce ll mem br ane. o ne red cell in the up per
pan of the field showing a co nstrictive defect so that it
rubles a co ttag e loaf in sectio n and a lower cell
ari ng to have lost a bite from the pe riphe ry.
Prooablv bo th these defects result from mechanical
ma (ro m fibrin thr eads.
59
1+1 145
00
,~
J
150 151
.\",
i~l
~"
o .\!-> .,..
,d' , .
r'<'
c
I!'-I
•
C" J,. !' M
JuJ/1/ t/10
JjT n" t~~
lJJ
156 158
157
•
-
•
159. Prus ian blue stain for free iron on the same
blood speci me n as in 158. Siderocytes (eryt hrocy tes with
free iron) are seen to be numerou s.
•
161
••
, •
•
6,
163 IM
165
163 and 164. Targe t ce lls. sp ur ce lls o r acant hocyte s
with mult iple sharp pro ject ion s. schistocytcs of irregular
frag mented shape and 'sput nik' cells. aca nthocytes with
two or three elongated spurs. Such changes are see n in
micro angiopathy especially and may he conspicuo us in
sple necto mized pati ent s. Th ese preparat ions are in fact
fro m a patient with congenita l PK deficiency and severe
haemolytic a nae mia. who had nee" sple nectomized 20
years previously, and had recen tly developed obstruc-
tive jaund ice as a result o f the accum ulat io n of pigm ent
sto nes in the gall bladd er .
66
Part 2
67
Fo r the A"ILs. the origi na ting cell m3Y be the ge ne ral into clinical use, different factors rnav we ll be found to
mye lo id precursor. the CF - G E ~ I ~ 1. capable of giving have more appropriate use in mo nolincage and multi -
rise 10 colonies ill vitro containing grunulocytcs . ".10no- lineage cases , res pectively.
C)tc_ macrophugcs . erythrob last- and mcgakaryocy tes : The second broad de fect of the FAH class ification is
o r perhaps the mo re narrowly co m mi tted granuloeyte ~_u!1.9ue. rel~nc~ o.n Rqm anowsk~' ~)10!ogy as a n ind~x 7"
monocyte macrophage pre cur sor , the C FU-G ~ I : or the of ce ll d lftCrentlatJOn kSudanophlha , often shown by
corre pend ing e ryt hroblas t or mega ka ryocy te pre- mat uring rnyclo blasts before azurophil granula tion
CUN.l~, HF U-E a nd C FU- Meg , res pective ly. Pur e becomes e vide nt in Rom an owsk y sta ins . is pr o babl y the )
ervt hrob lastic or mcgak aryo blast!c -lcukacmias' mus t be hest sing le prognost ic factor in AML. a part fro m age .
\eT) rare , if they occur at <I ll, since a predomi na nce of Surface antigcnic markers hav e not proved of gre at
these ce lls see ms al ways to be acco mp anied by at least discri mi natory value in A I\IL. a lt ho ugh po sitivitv to the
so me ncoplas t!c co m pone nt of gra nulocyrcs a nd/or major G M marke rs (CR..t.~ and/or C D33) hel ps to
mo nocvtcs . d isti nguish poorly dif fer enti at ed AML from Al. T'(ucut e
The causes of A M L a rc sti ll not de termin ed , a ltho ugh lym phoblastic leukaemi a ). a nd reactions with anti-
activa tio n o r ot he r functiona l change of proto - bod ies to glycophorin A or to platelet glycoproteins m,ly
o nco gcncs. which norm a lly pla y a controll ing role in furt her aid the recognition of c rvt hrob lastic or rnc aa-
myelopoiesis , ma y be importa nt in this re gard , and ma y ku ryo blast ic co mpo n"'e nts. · "-
res ult from the ch ro moso mal rcarra ngc rnents tha t arc Cy toge netic cha nges ma y we ll ha ve pa t hog e net ic
"readi ly demonstrab le in about SO 'Yo of A ML cases, and significance a nd arc of some pr ognostic va lue , t( :21).
probably occur at a less easi ly recognized level in t he t( 15:17) , im'( 16 ), und trisomy X as a n isol ated defect. all
rem ammg cases . havi ng rel a tivel y good prognos is , ttv : 11), monosorny 7,
It is not sur p rising, give n th e co mplex ity of hon e del e tion s of 7 o r 5, a nd most m ultiple ab norma lities,
marrow cytology. that ma ny cyto logical varia nts of ha vin g .1poor one, a nd ot her str uct ura l ah nor ma lities
A ML occ ur . T he)' have bee n cla ssified traditionally for having inte rmed iate prognostic significance. How eve r .
the last 6U yea rs or more according to the nature of the diffe rences o f this kind tend to d iminis h as treatme nts
pre domina ting ce ll in the ho ne mar row , a nd this becom e mo re effec tive . In a nv case . since so me SO% of
a pp roa ch fo rmed t he basis of the FAH subdivisio ns AML cas es do nut e xhibit an o bvio us chro mos oma l
proposed by a group of Fren ch , American and British abno rmality. cytogenet ic da ta do nu t a t pr esent provide
( FA R) haem a tol ogists in 1976, subse q ue ntly widely a satisfactory bas is for a gene ra l cla ssifica tion .
ado pted. For A M L the divisio ns ar c : ~1 I . jnyeloblastic The a uthors hav e deve loped a simple cla ssificat io n of
(>90'X) myc fob tasts}: r\'12, mye lo blastic wit h AML whi ch tak es th ese vari ou s co nsiderat ions int o
gr an ul.ocy.!iE.-matura tio n (> 10°(0 pnj myclocytcs or Inter account . Mon olin cagccascsu rc gro uped .tS T ).:r.£J and
granulocytcs] : ~t3 . p-.IQ!TIyeloc}tic (either coarsely mult ilineuge a~ Ty~_U . Each type is furthe r sP.!!t into A
gra nular or rnicrogranular promyclocyrcs prcdominat- and B uccordinu to whet her > or < 50% oft he lcukucrnic
ing): ~.I-t , myelornonocy tic (>20% gran ulocytcs , or G M cc lls a re ;uda no philic. T ype lA thus incl udes .all
pr ecursors a nd > 20'Yo mo nocytcs o r precurso rs) ; :\15, t(X;21) . inv (16) a nd 1(15 ;17) cascs . a nd may be o f an y
monocyt ic (M5a predomina nt ly monoblastic. M5b wit h FAB gro up fro m ~1l _ 1o--,\ I5 . Type IIJ. with little
diffe re ntia tio n): ~1 6 , erythro leukaemi a (>50% erythro- diffe rent iation , includes most t(\' ;1 I) , and again crosses
blasts}: a nd ~1 7 , mega kuryo blastic (>SO% mega kuryo- the FA ll groups widel y from Ml to MS, excluding ~ 13 .
blasts}. A M L with mini ma l differentiation hut po sitive T ype 11 cas es include most wit h poo r progn osis cyto-
antige nic markers ma y be gro uped as MO. ge net ics, as we ll as FAB groups !\.16 a nd M7 a nd <I
Th is classification is based on a rb itrary borde rlines proportion of all the remaining groups except M3.
bet ween the gro ups, a nd alt hou gh broadly -in co nformity Type I A~1 Ls , including cases wit h the assoc ia te d
with thc ge ne ra l usage of pred omin a ting cel l classifica - chromosoma l defects listed above . a re illustrated in
(io ns in the e a rly IlJ70s it has two import ant defect s . One pa rallel with ot he r disor der s a ffec ting gran ulocy rcs
conce rn !'! its fa ilure to discr iminate bet wee n mon o - a nd/or monocytes early in thi s sectio n. T ype 11 A MLs ,
lineage (co unting th e pro geny of the CF U-G~ I as of one with multiple line age invol ve ment. a rc grouped at the
lineage ) and m ulti lincage cases (with granukx... yte / end of the section . wit h t he myclodysplasttc sta te!'! wh ich
mo noc yte (G ~ 1l involve men t hut a lso erythroid an d/o r a rc mostl y also clona l neo plastic diso rd e rs with multiple
mc ga ka rvoc vn c ). Thi s ma y he of so me im po rtance in linea ge exp ression .
pr ogno sis . y, ith m ulrilineagc ca se- po ssibly surviving less The c ha racte ristic feat ures o f all these var iants of
well overall , although with adva ncing cfficacv of treat - A ML arc de scribed in the respective captions to the
ment this diffe renl:e may well dimLini sh . Of greater re leva nt cyto log ical and cy tochem ical illustrations, as
sig nifican ce , how ever . ma y be the different pat ho- arc t hc fea t ures found in bo ne marrow as pira tes a nd
ge neti c mec han ism , ac tive in pr oducing. resp ectively. tr ephi ne biopsies d urin g emerging remi ssion or de velop -
monoline ae c or multilincaee disease , with e it he r a ing relapse .
d ifferent 1~\·e1 o f stem cell 'affected or a d ifferen ce in
ste rn ce ll e xpression dict at e d by biologica l co ntr o lling
mech ani sms. A s bioloeical ac tiva to rs such , IS colo nv- Chro nic mvctoid leukucmiu (CM I.) , This d isease is
stimulati ng factor!'! (G~ I -CSF etc.) co me mo re widel\'.
- a not he r clo na l neoplasttc state , probably a rising in t he
mined plu ripote ntial haemopoietic ste m cell. but and/or the presence of A uer n S - hence presumed to
lh differentiatio n exp ression initi ally co nfined to th e be close r to transformation to A~1L ( RAE Bt) - and.
doid ce ll lines . Som e X5 ft!c. of cases mani fest th e Ph fina lly. chronic rnyelomonocytic leuk aem ia (C ~ I ~ 1L ) .
cmosome. a sho rt ch ro moso me 22 resu lting fro m a Th is nom en clature is tha t propo sed hy th e FA R group
reci procal tra nslocation betw een parts o f its I~Jn g arm for con d itions descr ibed over man y yea rs under var ious
- ' the long arm of chromosom e 9.1(9 ;22) (q 34;ql I) . nam es. which pro bably mak e up a bro ad spec tru m uf
lh the transfe r o f th e c-ahl on cogenc from 9q to a site indolent or smouldering c1on al1cukaernic states, alread y
~ q .nown as th e breakpoint cluster region (b.£r ). associated with non -random cytoge netic abnormali ties.
Tbe combi ned chime ric gene at thi s site produces an Th e way in which th ese d iso rder s evo lve is errat ic:
.iboormal protei n kinase . characte ristic for C ~ 1 L Most transformation to a floridl y lcukae mic p ictur e o f multi-
the rem aining 15% of -cases with the cyto log ical lineage (Type 11 ) AM L m;;I Y occ ur in a ny of them ,
ure of C ~ I L - with a high leu cocyte co unt and perhaps more commonly in RA EB and RA EB t. but
nee o f granutocvtc p rec ursors in the blood , many patients with MDS die from chronic anaemi a.
~ ulocy t i c hyperplasia of th e bo ne marrow . freq ue nt infectio n or haemorrhage without any suc h over-
philia and basophilia. and a low leu cocyte whelm ingly blast ic meta morphosis .
ine phospha tase (LA P) sco re in the neu tro phils c- T he histo logy. cytology and cytoche mistry o f RAEB
evide nce that a morph ologicall y inapparen t trans- and CMML <Ire illustrated exte nsively in 560-565 and
tio n has occ ur red. with production of the bcr-ab l 569-594. T he distin ction betw een RA EB and RA EBt is
cbimeric gene . 10 the ext ent that th e same abnormal esse ntially·a qu antliah vC Otit' , with up to l Ot?., hlasts I
pro tein kinas e ge ne product as in Ph +vc Cfvl l, can be the marrow III the lormer an Irom 2(}:j 0 10 thc.lauer
de tec ted and c-ab l probes locat e this on cogcn c at the her : alt ho ug h the presen ce of re adil y detectab le A uer rod s
le o n chro moso me 21. allows less b lastic cases to be c asse as B t.
C ML is well co ntrolled by single agent che mo the ra py. -./ E\'Idence of mu ltilincagc IIwol veme nt in e r\'t hrobla"sts
ally busulphan o r hyd roxyurea . for var iable pe riods includ es-nuclear fluddm"g o r ro...ct te formation . multiple
from a few mon ths to many years. unt il a change in th e nuCl eI. sldero6htstoslS. megalotilastos ls. and PAS
pa tern o f disease happen s. with mal ignan t progression pos . ega .arvocytcs atypical mononuclear
hypop las ia . myelofi brosi s. or an ac ute lcuk acm ic or multi-nuclea r forms and ei the r giant or micro-
pecture , usually th at o f Al\1L but occasionally ALL in megakar yocyte.... Defective gra nu la rity in myelocytes
. -pe. In tra nsfo rma tio ns to an AML·like sta te there is and acquired Pclger-Hu ct nuclear anomaly in
lten an eryt h rae mic and/o r a megak .eyocyric co m- ...cg mc ntc d cells provide evide nce of dysplasia in the
pone nt. with a multiline age T ype 11 cyto logy - althou gh , gra nuloc ytc se ries. while mon ocytcs may shO\..· biza rre
som etimes. a more purel y monolm eage GM form [Type nuclear morph olo gy espe ciall y in C MML.
It develo ps. A t thi s stag e furth er chro moso me ub nor -
IDalilies arc freque ntly found . not abl y one or mor e ex tra Reactive chunges ill granu tocytes and monocytes.
.22 trans locatio ns, struct ural ano ma lies o f 17q. and Leu cocytosis with incre ase in the WBC above the
trisomies o f S or 19. normal up pe r limit o f I I x 10"'/1 occurs chiettv in
Th e hae matolo aical findi ngs in bo th chro nic and infe ctive and mftammato ry states with jI reacii\'e
transfo rmed phases of C ~ ll . a~l" illustrated in 320-37.$. inc rease In neu trophils accompanied by a ' shift to the
be rc cha racteristic features of histology, cytology and left ":""'wtth a IJ ised pro po rt ion Of stab cells and po ly·
. tochc mistrv arc fur ther descr ibed in the ca ptions. morphs with fcw nuclear lobes . Occasiona lly. mor e
immature pre cursor s. mctamyclocytes - o r even a n odd
JIII enile C \I L. Thi s is a form o f subac ute ncopl astic myelocyte - may be found in the peripheral bloo d. Th e
~oIi fe ra t i o n with Ic.uurcs inte rmedi ate be twee n th e neutrophils may sho w coa rse darkly _stain ing 'to xic'
It type of Ph +ve C~lL (which itself occurs in granules. and ther e may he bluish Dohlcbodics visible
dliIdIiKx)(J. hut o nly rarely) a nd a myelom on oc ytic acut e ift ~Romanowsky p!.cpa ra ti(H1S -in- th cir cyto plas m,
ae mia . This d isease is not associated with tl 9:22). r~pre~entin g ureasof. hj-peractive aggregated ro ugh or
about 50% of cases sho w chro mosomal ub no rmali- smoo th end oplasmic.reticulum .
much the co mmo nes t being mOllosom y 7. Th e Le ucopemas with gra nulocyte co unts < 1.5 x IU"/l arc
is illustrated in 375-383. where ad dit ional seen in states o f pri mary or secondary marrow
PtI\C material is included in the captions. hypoplasia or aplasia . with generally defective myclo-
poicsis and assoc iated ana e mia and thro mbocvtopcrna :
vsplassic states (.\ 1DS ) ami other preleukaemic ho wever. selective neut ro pe nia may occur in o ver -
rs. A mong the conditions gro uped toge the r whelmingly seve re infec tions. whether bac terial. viral.
me title o f ~1 D S arc refr acto ry a nae mia ( RA) and rickett sial o r pro tozoal, an d may proceed to ag ra nulo-
-dn.:to~ a nae mia with side ro hlas ts (RA S), bo th o f cytos is. with circula ting neut rophil s < O.S x 10"/1 . A
......1 . D,,~ e bee n discussed and illustrated alr ead y in Part similar selective neutropenia may result from se nsitivity
prese nt sectio n includes the remaining MDS to certa in dru gs. includi ng sulpho namides , ant i-thy roid
. re fracto rv anaemia with e xcess o f blast.. agents and some anti -con vulsan ts.
.-..E8 . the- ~m~ with a hi-g:hc"'r- prorXlTli on of bElsts Cytoplas mic inclusio ns o f degeneratin g nuclear
material resulting from the action of an anti-nuclear T he Ched lak-H jgashi-Steinh rlnck syndro me is a ra re
facto r (ANF) form a special feature of systemic lupus inherited d isease . usually fatal in early life . with severe
c rvt hc rnatosi-, (SL E) . and are of cyto logical inte rest as cytopc nias and a lyrnphorna-Iike clinical picture. associ-
we ll as pro vidi ng a rapid diagnostic test. although their atcd with rem ark abl y lar ge cyto plas mic gra nules in
pract ical impo rtance has been largel y replaced hy va rious tissue ce lls - incl ud ing man y lcu coc ytcs o f blood
immunological testing for ANF. Examples arc illus- a nd bo ne marrow . especially gra nulocy tcs (398). T hese
trated in .'S-a and 385. coa rse gran ules arc probably lysoso mal in nature .
The reverse of the infective ' left shift" in neutrophils is Pseu do-C hed iak inclusions may develop as an acq uired
seen in megaloblastic anaemia». where deficiency of B l.:! phen o menon in so me acute lcukaemias. notably cases of
o r folat e leads to nuclear changes involving a ' right shif t' A ML with the 8;2 1 tra nslocation (211 and 213 ).
with an increase in the ave rage number of nuclear lobes
in neutrophil polymorph, and the presence of some cells Platelet disorders , \Vhile qualitative defects of platelets
with six or more nuclear lobes. macropolycytes or arc best det ected by platelet funct ion tes ts and arc not
hvpcrscgmcntcd ne utrophi ls. reliably recogn ized by changes in pla telet morphology.
Eosinophilia . with blood levels over O.5 x 10"/1. may quanti tative cha nges thrombocytopcnias and
he seen a, a familial ano maly. but occurs as a reactive thrombocvtosis or th rombocvt haernia - arc associated
phenome non - especia lly in skin disorders. allergic with char~cteristic cyto logica l findings . not on ly in ter ms
states, and pu rasitic infestat ion - and has add itio na lly of numbers of circulating platelets hut a lso with regard
Peen fou nd wit h associated mya lgia in pa tie nts la king to the cytology of mcgakarvocyrcs and the process of
dietary supple me nts of Ltrypro ph nn. Buso ph ilia. with thrombopoiesis as observed in marrow smears and
counts ovcr u.zx 10<)/1 . may also be seen in so me allergic sections.
or inflammatory states, altho ugh it is relat ively T hro mbocytopenia. with reduction in circ ulat ing
uncom mo n. p latelets be low 150x 10"/1. arises in most instances fro m
Mo nocytosis. above the nor mal upper level of o ne or two broad causes: defective formation of plate lets
I x 10"/ 1. may occur in more chro nic infective states or in (as in prim ary or secondary aplastic or hypopl astic
those d ue to non -pyogenic bacteria . viral or rickettsia l sta tes . inclu din g th e effects of myelotoxic che mo the rapy
infections and so me p rotozo al or oth er pa rasitic infes ta- and so me less co mmo n d rug sensitivities) . and increased
tion s. A reacti ve mon ocytosis may a lso he seen in so me de struction o f plat elets in the periphera l circulat ion and
auto- immune disorde rs and co llage n diseases, and in splee n (a rising ge ne ra lly from auto-i mmune sc nsitizu-
so me cases o f disseminated malignancy. tion of platel ets. render ing them liable to phag ocy tos is
by macrop hagcs ). Th e fo rm er type of thru mbocy to pcnia
Congr nitul anom alies and acquired parallels. Alth ou gh is associated with a reduction or abse nce of mcgakaryo-
ma ny an o ma lies o f leu coc ytc s have bee n described . cytcs in the hon e marr ow - an amcgakaryocy tic
most a rc ve ry uncommo n; on ly the four most important thrombocytop eni a - wher eas, in the latt er . mcgak ar yo -
arc described and illustr ated her e . cytcs arc gene ra lly plentifu l and ofte n incre ase d in
The l)l'IJ,:l·r·llut't nuclear ano ma ly occu rs as a ra re nu mb er s, alth ou gh they may a ppear cyto logica lly
benign familial inh erited diso rder P S7) . hut it is see n unu sual in hav ing immature nuclea r feat ures and
very co mmon ly as an acqui red o r ' pse udo' fo rm in man y defective gra nularity. with often promi nent glycoge n
cases of myelo dys plasia an d bo th ac ute (e .g . 215) and inclusion bod ies and few peripheral p latele ts (467- *73) .
ch ro nic (3AA) myeloid leuk aem ia . Thrombocytos is. with counts over 500 x 1tI<J/ I occurs
Th e Ma~' - l h1:.g l i n ano maly. like the Pe tge r-Hu ct us a co mpo ne nt fea ture of many states invol ving marrow
anomaly. is a ge nera lly be nign co ndition inherited as an hyperplasia . fro m iron-deficiency and haem olytic
autoso mal domi nant . It is associated with the for ma tio n anaemias to acute infections. It mav also be a feat ure of
of large oval or crcsccnuc basophilic inclusions (394 and disse mina ted malignancy . This tyre of platelet increase
395) in th e cytoplasm of gra nulccyrcs and occasionally is a secondary or reactive phenomenon of no great
also o f monocytcs. These represe nt a special type of clinica l significance.
inclusio n material rather than an aggregation of no rmal A mo re severe and ~rsistent increase in plat elets,
cytoplasmi c structures. and arc thus essentially different often to over 1000x 10 11. occurs in csse ntiul thrombo-
horn Doh le bodies. Giant platelets may also be found in cythernia or megakaryocytic myelosis. a chronic nco-
this ano ma ly. and there is sometimes thrombocytopenia plastic myeloproliferative disorder whic h appears to he
and leucopenia . largely co nfined to the megakaryocyte-plate let line .
The Aldrr-Rrilh' a no ma lv. which mav occur in associ- Th ro rnbocythcmia of similar degree m;'IY also he seen in
anon with various mucopolysaccharidcscs. involves the mullilineage proliferations. especially polycythemia
development of coarse azurophific granules in the cyto- ve ra. but also occasionally in C ML. In all these co n-
plasm of gr anulocyrcs (389) . The material of the ditions ther e is a conspicuous increase in megak aryo-
gra nules is derived from the mucopolysaccharide and cytes in the hone marrow. often with imma ture or
resem bles basophil granules somewhat. in reacting ot herwise atypical cyto logy (477--U«2). and clumps of
met ach rom atically with toluidine blue. aggregated pla telets are freq ue ntly found in the pc ripb-
711
C'f'JII blood smears . where megakaryocyte nuclei and rubru vera (PRV) or CML (339-348). More mature
-=lear fragments may also sometimes be present. megakaryocyte proliferation . arising in a s.ir:nilar fashion
Icga .arvoblastic proliferation in the bone rnarrov as either a primary or a secondary condition. ap~a rs
. be promine nt in some trilineage acute leukaemia- often to stimulate the development of myelofibroti c or
may occasionally predominate. This occurs in some osteomyelosclerotic changes. prohahly. thr,!ugh t~e
. apparently arising de nm 'o (48R--l95) . hut also production of growth factors and other biological a~tI\"·
rg as a form of blastic transfo rmation or malignant atcrs of fihroblast proliferation and collagen formation .
eeceression in the ter minal phases of polycythemia
11>6
•
•
• •
. #, I
~
,
•• • • •
•
\
167
•
169. A r1!1~ h ) ~ l as t w i t h _1 W O-C( mspi c:.uo us....!!.~ c1co l i . a 171
\acuo lcy nd a few uzuro philic gran ules. a larger-pro-
myelocyte . a myelocyte and a sta b cell o f th e neutro ph il
series. toge the r with a lat e no rmoblast. Bot h th e cells at
the pro mye locyte and early myelocyte stages show
fad ing nucleo li and azurophil gra nules. th e smaller and
mo re ma ture myelocyt e also having less sharply de fined
and less stro ngly coloured neutrophil spec ific gra nules.
li .e tho se in the acco mpa nying stab cell.
\
j
172 173
17~
172. A myelocyte and four metamyelocytcs of in Teas-
ing maturit y (with increa sin gly ind ented nucl ei) . Th e
myelocyte is the sma llest cell in thi s gro up and has the
most basophilic cytop lasm . th ou gh without any det ect-
able azu ro phil granu les. and its nucl eu s has sev era l sma ll
but fad ing nucleoli . Spe cific granules are inconspicuous
in th is ce ll but numerous in all the metamyelocytes.
The re is aga in hypoch romia of the red cells and a
stomatocyte with a nei ghbouring dacryocyrc towards the
upper left.
!
I ~
5
17 179
76
••
'7
I~ 185
7S
; .
..' -,
"
,
..
.., ...
".'~ . ~,
,. , .
I . Acid pho sphatase in grunulocytcs in normal IH9
aarr m" ; myelobl ast . promyelocyte , st ab and two
~ e n t c d neutrophils. All these gra nulocytcs sho w a
- ueri ng of fine granules . probably represent ing the
prima ry lysoso mal azuro phil granules which first appea r
• the late mye loblast to ea rly promyelocyte stage and
in in the cyto plasm of later gra nulccyres . altho ugh
led in co nce ntratio n by division at the myelocyte
. A cid phosphatase positivitv in the norm al cell
is the rclore generally most conspic uo us an
) cloc vtes.
192
201
S2
•
'.
2f1K 2119
210
20K-215. Humun n wsky stains of how! mllrrow and
blood SUI i' ll rS f rom cases of A M L wilh tilt' X;21 l rans -
locution - 1(8;21) (q22;q22).
In thi... A MI. variant the b last cells ar e gen era lly la rge
hut ... how vunab ilit v in size and in the amount of
/
cvtoplu ...m. which is basop hilic , oft e n con tai n... lo ng a nd
...lender A ucr rod s, and may also co ntain vacuole ... and
ulcbular indu... ion s. T he nucleu s may he eccent ric. with
commonly an inde nta tion o n the side next to the mai n
mass 01' cvtopla... m . v. here t he re may he a pale -... raining
area representi ng th e Gnlgi body and where inclu... ions
o ften tend to he localized. Th ere arc so met ime ... uro.....er
disto ru o n-, of the nucleus and sepa ra ted nuclear
trua mc m , mac he foun d . N ucle o li are ac ne ra llv laru e
and. pale. and- tw 0 or three are co mmo nly de tectable ,
u... u .J1I~ ",idd~ spaced. Th e- pro mye locy te ... o fte n po.........·......
many coa r-e azurophil (p rima ry ) gra nules and may
...how large pveudo-Chcdiak inclusions as well as slende r
Auer rod .... u...uallv ...inulc . Later staces o f maturati on .
fn~m myelocyte... ~mv.;rds . show defect ive pro d uction
(lCs~cifiC' gr -notc ~ o plas-m--- t h a l -a p pc a r ... -
pJ!lkish:)dlo", aitht=myeTocyfe sfagC ...omcti mc... with
a residual pcnpherartlmofl1eejJoa...opfii~ he re arc
par allel defects in nuclear maturation. with occasio na l
ring nuclei and. at the end of the maturation ...cquc ncc .
some neu tro phil... with pscudo- Pclgc r-Huct nucle i in
the ir agra nula r cytoplasm . Occasional abnor ma l
cosinophib may he see n. with atypical greeoic h-grcv or
blue-staining gra nules (St'e also 220).
21-l
liS
216 2 17
22-1
225
•
226 227
1.. A PL with coa rse azurop hil (prima ry) gra nu les 232
_e1 ~' filling the cyto plasm of all the le ukaem ic pro·
jocytcs. Nucleo li arc co nspicuo us and va riab le in
bcr hut the ove rlapping o r dumb-bell type of
ar shape often see n in A Pt is no t shown he re , a nd
Iller rods arc not visible in this l.cis hma n-staincd
ration.
90
I )
242
92
• 0
, .•..0; '
4
'.
_...a..I Dua l este rase reaction in APL: th er e is a negative 2 ~7
. throhlast pr esent hut all live promyclocytc s e xhihit
g gra nula r Cfi po sitivity a nd two of them contain
In ple Aue r rod s which ar e her e int en sel y CE-
tivc. T he strong reactio n for CE, with or without
_ativc or Bfi-posuivc ce ntra l cores, is not usuall y see n
Auer rods except in the 15: 17 a nd 8:2 1 tru nstocati on s.
U9 . Anot her sect ion from the same biops y sta ined with
Gie msa. hut not providin g better gra nule definiti on o r
clea re r cell differentia tion . although the lcptochrom utic
nucle i with pro minent nucleoli arc well visua lized and
the prese nce of occas io nal nuclear twinni ng can be
... •
detected .
94
~-
95
•
257 258
257. Exce ptio nally lar ge pro mycloc ytcs. myclo cytcs 259
and rnctu myelocytes may he enco untere d in the marrow
of pernicio us anae mia ( PA) and someti mes of other
megaloblastic (and rar ely nor mob lastic) anacmias. Her e
arc e xamples of gia nt pro myelocyte and myelocyte from
rA.
258. A neutrophil sta b cell and a segmented cell from
nq rmal periphera l blood .
96
16 1
265
_.al.-\In>ma
smear fro m the buffy coat of a blood sample
pat ient who developed se vere deficien cy o f
.. and folic ac id while o n parenteral nutrition in
."''''·C the rapy uni t. The field shows an ea rly
';::::;me
~ galoblast and a mult i-lobed pol ymo rph or
• le. Two red cells co ntain Howell-Jo lly
99
271. LClU..ocytc a lkaline pho sphatase ( LA P) reaction . 27J
w eak and stro nger povitivity in neu troph ils. with a
nega tive lymphocyte . Positive reaction s in hae mic cells
arc virtually confined to neutrophil stabs and segme nted
cells. although macropbagcs may also show positivity.
276
101
279 280
281 2M2
I _
•
•
286
::;-
: :ic no rmal eos inophil po lymorph. showing the
'spectacle' arrangement of the nuclear
. which are usua lly two in nu mber. The sur-
red ce lls show slight hypochromia and there is
relet o verlying one of them .
I •
,
105
295 296
29H
,
.
.. . \ ... ......: '-.... ..
. . s : t- ..
/~
~
106
- I
107
30(,
.111
•
Ira
JI ~ JI5
110
318
111
32 1
310
113
J 16 1ioI'!7'~ · 327
"
'. '
"
:: . '~ ' ''.
• ,
I A se ries of views at low, intermed iate and -'-' I
rs o f sectio ns made from a biopsy o f skin and
_ ' . . . .""'''' tissue tak en from a pati ent with C ML.
a k ukaemic infiltration. The first two figur es
a section stained with H& E and th e third is
"~~~~::,-stJ i ned section. Th e first field shows the
- . with the infiltrate ext ensively involving
.....rt.!iolg_ de rmis. producing vascular erosion and
- . . orrhagc . In the second field the nature of
:
:::~.~ceIlS can be d istinguished mor e clea rly,
ntly nuclcolatcd myeloblasts and fewer
ytes spreading bet ween th e subcutaneous
_ _oeti,..e tissue . Th e third field shows th e edge
e as it reaches the epide rmis. where the
ma tu re granulocyte precursors inter-
co llagen fibrils and macrophagcs lad en
.... .. .L eithe r free iron or possibly ca rbon particles.
__ .... «11 is visible towards th e lower left co rne r.
no w skin deposits arc no t unco mmo n in
_ times nume ro us and wide spread so a,s
-,......, a lru aemic skin rash . es pecially. as in the
~ at the time o f gene ral metamorphosis o f
fro m the chro nic to a more acute
appearance may therefo re have a poor
. nee. More bulk y a nd restri cted
"",,,,,,,... may also occu r. less co mmon ly. at any
. and are sometimes referred to as
• •• •
115
JJ2 333
116
3J6
11
3-10
339
11 8
,
•
:
:~:~.; \\ith the intramedullary blasts. despite the
. of their cytop lasm. having more unifonnly
- --.... fea tures o f Iept och rom at ic nuclear staining,
ible nucleoli. especia lly in the G iemsa
".,- - - • • hereas the intravascular cells have a
r nuclea r pattern and also show the
• topIasmic o utlines cha rac te ristic of mcga -
aod early mcgaka ryoc ytcs when see n in
......ontions. Alt ho ugh th e PAS reac tion whe n
CMl plastic sectio ns usually gives much less
.[\ in haemic cells than is found in PA S
"(d oJ.Ul). some peri phe ra l cytop lasmic
disce rned in th e primitive cells in 344 .
....,_.... in the intravascular than in th e intr a-
tio ns. The differences in cyto logy
ites may be partly a consequence of
_ ......... from contiguo us cells in the marrow
the vascula r lume n. but may also
ease in maturity associated with
vessel wall.
119
345-348 . Cyto logical and cytoche mical details of m ega-
karvoblastic and m egakaryocytic transformation in CM L
as seen in smear preparations of bone marrow aspirates.
345. Rom anow sky stain shows two hlast cells. to left
and right. several megak aryocyte nuclei and masses of
variably sized platelets.
,. ..
346. PAS reaction sho ws the coa rse pos itivity in
plat elet and megakar yocyte precur sors.
120
1. C \ f L progression to myeloblast-typ e crisis.
of plastic-embedded bone marrow trephine
sectio ns (3-19- 35 7) and cYIOI01-:Y and cy lO-
ry of smears fro m bone marrow aspinnrs 351
1 ).
•
I ~l
...~ --- --.." ...- - - -... .\53
351 ,
•
Ano ther exa mple of myelob lastic transfo rmatio n
~'5 2 .
develo ping in CML. In this thin sectio n of plastic-
embedd ed trephine biopsy mater ial fro m the iliac crest
bone marrow . the edge of a blastic nod ule is on the right .
betw een bon y trabeculae above and be low. with
resid ual chro nic phase cells havin g gene rally da rker
nuclei o n the left.
122
:
•
•
• I . #' •
• ,,
• of
•
..... • • 0'
. :.. . • •
0
.. , , t• 0
•
F
•
,
;
0 • o.
• ,
.. . • ,
0
•
't
-
-S-J57. T hree furt he r thin sectio ns from the same 357
ine bio psy as in 354. Th e first (355) shows at higher
"ficatio n an area similar to that reproduced at low
in 3.5-4 . with a clea rly visib le co ntrast bet wee n the
_ _dve nucleo latcd and lep toch rom atic myclobJasts
... the irregularly eroded trabecular margin and the
e of more mature cells space d furt her away . The
reil wit h large darkly stai ned nucleus. direc tly
........ a sem i-circular bo ny erosion at the middle of
I ma rgin of the field. is probably a poly ploid
_:udlast . The second field. in 356. from a Ponceau S·
pre paratio n. reveals an absence of collagenous
aDd fibro b lasts fro m an area of myeloblastic
t
....n1>e<" lar proliferation very similar to that shown
preceding two figures . T he third similar field (357)
3 prepa ratio n sta ined for argyro phil retic ulin
~ n . the re is a notable absence of such fibrils in
_ urhood of the tr abecula e , wher e the myelo-
proliferatio n is co ncentra te d.
123
359
36U
358 , A smea r pre parat ion fro m a bon e ma rrow aspi rate
take n fro m a pa tien t in the blastic phase o f CM L. A
.,
Ro ma nows ky sta in shows a grou p o f blas t lTUS with little
evide nce of differ enti ati on .
-"
•
, ..
I•
•
• 4 f
•
• •
t
•
• •
t,
•
• •
\...370. C \! L progression to a lymphobla st-type crisis.
C·A L L untigrnic findin gs. Histology of trephine •
(363- 365 ) ami cytology and cytochemistry of
_ ....Don , m ears (J 66-J70) .
366. Roma nowsky stai n shows agran ular blast cells 369
with high nuclear-cytopl asmic ratio . in this smear of
bone marrow aspirate from blastic crisis of CML.
368. The PAS sta in shows occa sional block o r coa rse
gra nula r positivity of lymphoblasti c type .
126
J 1
127
-
•
376
•
37K
•
·
...·, •
"
--
•
385
1.-
•
Dl
3 391
392 393
:
;;;iII~diam
~.~ eter, weakly basophilic and apparently
-bound, in granulocyte precursors and in
from a child with a transient pancytopenia
. syndrome. The material seems likely to
..._ _,i) protein.
133
~O l
0$02
1: .
Various mOlOcytes f rom norm al periph eral ~07
,. •
""JO(~.- t es .
one with vacuoles. and a lyrnpho-
..c1_" are probably all phagocytic. one ' till
~ _ in :- ted platele t and the other two having
d( este d their contents.
_ _lC).lCS and. a stab cell. The m onocyte-
cytoplasmic granularity and typical
135
•
408 and 409. Co nsecutive Rom anow sky and d uplicate 411
Prussian btue sta ins on the same field , fro m a buffy coat
smea r made from the peripheral blood of a pat ient with
chronic myelomon ocytic leuk aem ia (C MML). la show
more bizarr e forms of nucl ear convo lutio n than are
usua lly see n in normal monocytes. The furth er tr ans-
for mation of mon ocytes to macrophages normally
occu rs in tissues. but may occasionally. as her e , take
place in the blood. Th e transforming mon ocyte co ntai ns
phagocytosed nuclear fragments and also a heavy load of
free iron . as demon str ated in 409.
...
ca_.ooly observed in acute leukaemias of any kind and
seen when o the r malignant processes invade the
.".
•
~1 6
-
• .
,
.-.
J
41
419
' .
•
.
-lIK. A gro up of mo nocyte pre cur sors from an A~lo llL. 421
.l)9. The same licit! . co nsec utively stained with the PAS
react ion. Mon ocyte precur sor s show co nsiderable •
variability in thei r positivity to PAS . fro m co mp letely
negative reaction s to coarsely gran ular. heavy pos itive
ones. Her e the ce lls show a mixt ure of d iffuse cyto-
plasmic stai ning and fine granules.
139
423
.'
.......
.: , . , ',
•
.. ' ..
,
•• ' 4j.
•
141
~2 9
•
•
t
• • •
•
•
t
."
..
.•....
~
0
•
•
,
t·
0
'. .. • • •
• • •
•
•• •
• • .. .• .,
•
..• •• • ., •
." •• ... . • •• •
• •
·U H- B O. Var iou s histological fields fro m thin sections
of bone marro w tre phine biopsi es fro m cases of acute
mo nocytic le uk ne mlas. In 428 and 429 . respectively low-
an d higher -power views of th e same sect ion ( H&E
stai n ). co nside rab le resid ual fat spaces re main . and
islands o f erythropoietic tissue can be d istingui shed : but
the cel lular a reas arc predo mina ntly occu pied by lar ge .
poo rly di ffer ent iated blast cells wit h nucle i sufficie ntly
widely spaced 10 ind icate a mp le cytoplasm . and wit h
several nucleo li o fte n visible under the high er po wer . A
suggestion of nuclear twisting ca n be d iscerned in ·*29.
This is a more rcadil v recog nized feat ure of ·B O. a
thin ne r section sta ined 'with Gie msa fro m anot her case.
whe re there is denser ce llula rity wit h a sca ttering of
cryt hroblasts having da rk nuclei a nd eosi no philic cyto-
plas m - but . aga in. a preponder an ce of nuclco lat cd
pr imitive ce lls wit h nuclear twist ing of the mo nocyto id
type . T hes e sectio ns prov ide valuable supple mentary
info rmatio n o n o vera ll ce llula rity and relat ive pro -
po rtions of differen t ce ll types. but for cytologi cal deta il
and co mpre hensive cytoc he mistry it is nece ssar y to
stud y smea r prepar at ion s in parallel.
•
~31
1 ~3
•
435
1-44
~39
J5
i
~.
'"
~I
36
• -
37 439. PAS reaction o n the same speci men as in 434.
T he re rsa negati ve o r weak reaction in the least mature
blast cells. increasing to moderately coa rse gra nular
positivity against a diffusely positive hackgro und in the
more mature pro mon ocytes.
138 442 and 443. Dual esterase reaction s in I( 11; J7) and
1(9; 11). respect ively . The low-po wer view in 442 sho ws
the variable positivity for BE in the leukaem ic cells. with
gene rally negative blasts and mor e marked posi tivity in
the most mature-looki ng promonocytes. The higher-
po wer view in 443 illustrates the same pattern . with four
nega tive monoblasts and a BE -pos itive promo nocyte .
•
.' ...
A zeneral view of a bone marrow smea r from the
case as shown in the previo us figures. sta ined by
PAS reaction. T he diffuse and finely gra nular
- -ity of granulocyte precurso rs. increasing in in-
: with increasing maturity. is shown ; the stro ng
. reaction in some ery throblasts is very character-
of erythraernic involvem ent in a mixed leukac mic
147
~51
149
2
~S 7
•
.,
•
~59
.,
.$56 . A normall y granular mature mega ka ryocyte . with
minimal plat elet formation at the periphery.
458 . A poo rly gra nula r meg akaryoc yte with minim al
platelet formation .
151
46 S
• •
IS ~
i
153
e
~ I , ~ Z
, • t
•
.. • •
e
, ••
.,•
• e
•
, .~.e
• • ,• e , &
.. •
e
" . • .:.; . 0
o•
•
• •• •
, ...". • • 0
, • ••
, e. •
•"
0
••
,
•• ,
;
•
t. • ".; •
•
•
•
•,
, e ,. • .• •
•
••
0
•
..,..
471. Fo ur mcgakaryocytcs from a ho ne marrow smea r
.
-. " ,.
't·,
~ 73
_,-=... . .
....•. -'
gra nulocytcs within th eir cytoplasm . mor e probably
, . ~ • .; ~
.,". _, 4.',.
resultin g from a passive incor poration or emperipo lcsis
-: . ,.e
than an active phagoc ytic process (ef. ~ 77-479 ) .
S
- . .......'.,.
.... --...
sti mulate erythro id hyperplasia - but without prod ucing •• • .: . /# ",.e.... ~ , . . .
any marked increase in ove rall mar row cellularity. since I'. .... ,.. ," ·A e_a.
-
fat spaces are plenti ful. Megakar yocytosis is clea rly
evident with generally somewhat prim itive . defectively ' ",
~~ .
.• :e!. ,.. .., e/.tf ' _.
lobu lated nuclei and poo rly gra nular cyto plasm in most
rnegakaryocytes . Th e absence of per iphe ral plate lets in :" ".
'. ...
. _" ,..1
megakaryocytcs in ITP reflects their ra pid release into
the circulation and destru ction there. rath er than am:
r·:.....,t."..",. ,.1, .
~.
.-., .. .
defect in productio n. for the o utput of functionally ~ .. ~ . ,
active platelets in IT P is usually several times nor ma l. \ -.-. "' .. '."> ..
,"" . ,~-", , .
.,~ ,. , ..
_.7... . "
L
-~
•
•
155
478
479
477 and 478. Gross phagocytosis of an erythrobl ast
clump by one megakaryocyte and phagocytosis of other
cellular debri s and red cells by othe r megakaryocytes in
a myelop rolifer at ive disorder with megakaryocytic
hyperplasia .
479. PAS stai n in the same case . showing coa rse PAS
pos itivity in two megak aryoc ytes, eac h of which co ntai ns
ingested leucocytes.
•
•
.."'...
..
- -
.,(
•
•
• • -.. •
•
•"
• - •
•
•
'.
••
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•
158
486. S8 reaction in the same case . A megakaryocyte
wi th thre e phagocytosed lymphocytes or erythroblasts is
negative. as is the neighbour ing megakaryoblast.
s,
~87
159
-189
493
•
•
/0 /
-196-502 . Examples uf the cytology and cytochemis try of
AM L with trisom y 8.
I h'
.-
'-'
•
•
163
503-5 20. Examples of the cyto logy and cytochemistry of
AM L associated with an interstitial deletion of 'he 10nK
•
arm of chromosome 9 between bands ql 3 to q22. 9q-
(q/3 ;q22)(Type /lA ). 505
I/H
•
510
512
51 ~
le
/65
516
515
•
518
166
S11
1"/
5U 526
525
168
•
~
...-.
~'l; ... .'
531
I 'll
5~'6-S47 . Earlv cytological signs of emerging myeloid 53ll
haemopoietic ;lctivity during remission development in
aClde leuk aemia. The fea tures illustrated are usually
fo und in ma rrows which otherwise remain hypoplastic
fo llo wing cytotoxic therapy,
171
539
- •
172
1
• -,
•
•
542. A group of typical rem ission -associated cells:
plasma ce lls, RE cells. megakaryocyte-plate lets . a single
blast cell. granulocytc precursors and a giant megalo-
blast following an timetabolite chemot herapy.
543. A myelo blast . two coa rsely gran ular prom yelo-
cvtes (as typically found in ea rly remissio n) and a
meta myelocyte.
17J
545-547 . Examp les of giant o r mult ilobed neutrophil 547
mcta myclocyt cs or pol ymor ph » in ho ne mnTTOW aspi r-
at cs taken du ring ea rly stages of re mission e me rge nce in
ac ute leukae mia. An ac tivated Iymph ocyte or imrn uno-
eyre approaching pla sma -cell morphology is also shown
in 547 , Patho logical though the mu ltilohed neut rophils
a ppe a r, they do see m freque ntly to precede remission
de ve lopme nt .
174
p
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551-553. A ve ry low -po we r field a nd two high e r-power
views of hon e marro w tr e phin e bio psy sect io ns from a
pat ien t wi th AML a fte r remi ssion induction th e rap y. In
551 the ma rrow id ve ry po o rly cell ula r wit h mult iple fa t
spaces . scatte red haem orrh age a nd m acrophages lad e n
with haernoside rin . and a residual nodu le of what ar c
seen, in 552 (a high e r magnification of pa rt of the sa me
Held), to he lymph o id ce lls. Th is picture is one co m-
monly fo und immed iatel y after agg ressive co mbi natio n
ch emot he rapy and re pr esen ts the hypo plastic phase
-
preceding e mergence of remiss ion . T he th ird Iield in this
gro up is fro m another biopsy ta ken a week later fro m the
same patient . T here is no w a mixed pict ure p resent , with
islands of crythroblast ic activi ty e vident a mong the fat
•
spaces. especially in the uppe r part of the field. hut also
recu rr e nt myelornonocytic blast cell proliferation
e lsewhe re . Pa ra lle l aspiration smears confirmed thi s
lcukaemic recurrence and furt her induction chcmo-
therapy was indicated . •
•
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55~ and 555, Low- and higher -power views. res pect- 556
iv·:ly. o'f a bo ne mar row trephine biopsy sectio n ta ken
from a pat ie nt with seco ndary AML. afte r repeated
courses of re mission induction therapy were followed by
a pro lo nged period of marrow hypoplasia and pe riphe ral
pancytope nia, A t this stage the marrow under low
power (55-4) shows a reticular stroma with recoven ng
<;,Ci.l tlered ce llularity and fresh cell uli.lr proli ferati on .
e!'oopccially along cap illaries . Th e higher magnifici.ltion of
•
555 revea ls that the cytology is pleom orphic. with
plasma cells along the cap illaries and cells of vi.lrio us
lineages sci.lttered through the stro ma . a mong them.
•
primitive immi.lture cells. possibly represent ing leu -
kac mic recurre nce . hut possibly nor mal e merge nt
prec urso rs.
556. A fur ther biopsy ta ken a week lat er from the same
patient as in 554 and 555 resolves the qu estion of re lapse
or early remi ssion , Thi s thin sectio n shows a few resid ua l
plasma ce lls but also goo d mat ur ation in the gra nuloeyte
~ri c s. with all stages fro m pro myelocyte o nwards
recognizable. toge ther with scatte red eryt hrob lastic
pro liferation. Clearly. remission is now eme rging.
"
,
•
177
•
• •
• •
• •
•
,
•
•
•
•
557. A thin section o f ho ne marrow tr ep hine bio psy
fro m a patient with A ML afte r inducti on o f a temporary
remissio n . now showing evide nce o f re lapse . This high -
po wer field conta ins <lOOU! 50% lightly sta ining large
blas t ce lls. with conspicuo us nucl eoli a nd va ria ble
myclomon ocytic nuclear sha pes a nd nuclear-
cyto plasmic ratio . Th er e arc res id ua l no rmal er yth ro-
blas ts a nd late r gra nulocy tcs . b ut full rela pse is now
inevita ble unle ss fresh inten sive treatmen t is sta rted .
17S
I
I!IO
;
181
569 5 0
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:;:69-571. Thin sections of bon e marr ow treph ine 571
bio psy fro m a pa tient with a n MDS involvi ng com-
po nents o f ref rac tory sidc roblustic anaemia a nd
excessive blastic pr oliferat ion toge ther with a n increase •
in mcgak a ryocytcs . In 569 th e low-powe r view sho ws
genera lly high ce llula rity with a very pleo morphic •
picture. including numerous scatte red macro phagcs
laden with haem osiderin a nd seve ral multin ucleated
rnegakaryocytes. T he higher -pow er view in 570 is of a
neig hbo uring area with more residu al fat spaces . whe re •
the megakaryocytic cytology is well shown . a nd where
there a rc several areas of haemo rrh age of the kind from
which the haemosidcr in in the macroph agcs in 569 was
do ub tless de rived . T he rem aining ce lls in this field
include grn nulocyrcs chie fly fro m myeloc yte o nwa rds
and ervt hroblasts with densclv sta ined nuclei. In the
reti cultn-staincd prepa ration ' at still high er power
illustrated in 571 . the abse nce o f a ny increase in re ticuli n
a nd the pre se nce o f lade n macroph ages. a nd also o f
nuc lco la tcd blast cells with lightl y sta ined nucl ei . a rc
clea rly evide nt.
•
182
572. Prussian blue stain for free iron on cells in a hone 57.t
marro w smear made from the same patient whose
trephine sections arc shown in 569-571. Th e cluste r of
ervthrc biasts in this field shows a high conte nt of free
iro n with occasional ringed distributi on .
573. In this Rom anow sky stain. a buff y coat prepa ra-
tion from pe ripheral blood - agai n. from the same
patient - shows a myelobl ast , a prom yelocyte . later
neutrophils, two lat e erythrohlasts on eith er side of a
megakaryocyte nuclear fragment . a nd a mass of giant
platelets.
183
5 5 576
-•.- .'........
. "
.al
••
•
•
•
:•.....•
'.
•
57:-577. Sections of trephine biopsies from furthe r 577
pa tients with MDS . In 575 and 576. an exampl e is shown
of the mor e hypocellu lar variant of MDS . in this case
associa ted with mark ed dysmega karyocytosis (micro-
forms and multinu cleat ed ce lls be ing especially con-
spicuo us in 575) ; the presence of erythrob lastic island s
and the abse nce of collagenous fibrosis are mani fest in
the Van G ieson stai n of 576. Th e section appearing in
577 is fro m anot he r pat ient with the RAEB form of
MDS , associa ted also with the chro moso me anomaly
5q-. This defec t occurs most common ly in elde rly
females . as in this case . and is usually found in associa-
tion with ref racto ry a naemia. dysgranulo poiesis with left
shirt. and a cha racte ristic megakaryocyte picture with
mononuclear or bilobed ce lls. Typical dysplastic mega-
karyoc yres showing this feature are seen in the upper
part of the field. A mo ng the scattered islands of e rythro-
blastic hyperplasia and later granulocytes occupying
much of the lower part of the field are several small
groups of blast cells. well away from the endosteal
surfaces of bony trabeculae whe re early myeloid pre -
cursors are norma lly concent rated . Th is 'abno rmal
local ization of immature precursors' (All ?) may
pe rhaps predicate immi nen t leukaemic transfo rmat ion.
184
578-580. Two low-powe r views and a higher-power 5110
field. respectiv ely . from a thin sec tio n o f bon e mar row
tre phine biopsy from a pat ient with MD S. showing
RAEB with trilin eage dysp lasia . In 578 there is a
generally pleo mo rphic picture with mon onuclear mega-
.aryocytcs and ot hers with multip le perip heral nuclei.
an evident increase in early granulocytes - though with
poorly gra nular cytoplasm - a few later neutrophi ls and
inophils. and a sca ttered c ryth roblastic component
' th deeply stai ned co mpact nuclei. A broadly similar
field is shown in 579 . but w ith several abnormal rnega-
ryocytcs having lar ge sing le o r do uble nuclei. In both
fields occas iona l cells with elo ngated nuclei
~ est the prese nce of fibroblasts . The high-po we r view
of 5SO clearly illustrat es the nucleo lat cd myelo blasts
ich predo mina te . toge the r with eryt hroblastic islands
• top a nd bottom of th e field showing some rncga lo-
tic feat ures. and var ious ce lls of th e megakar yocyte
. includ ing a hi nucleat ed cell at mid -centre left and a
-Kro megakaryocyte at the right edge . a thi rd up from
bo tto m.
•
.-.,
•
5KI-583. Two furt her examples of typica l bo ne marrow 5
trephine b iopsy appea ran ces in va ria nts o f MDS with
incre ase in blasts (RA Ell) . Both show hyperplastic
ccllularity with few rat spaces a nti marked pleo-
morphism . the first case - with la\\'- and highe r-power
field s shown in 581 and 582 - havin g all myelo id cell
series well represent ed. but with an evide nt increase in
immatu re cells. especially of the gra nulocyte se ries.
bo th neutrophil and eosinophil. as revealed in more
det ail in 582. The thicker section from anothe r case
shown in 583 is agai n highly cellu lar. with all cell series
.
pre se nt . hut with a co nspicuou s shift to the left in the
grun ulocy res. with se ve ra l clumps of nuclcol a te d myc lo-
blasts. Both these cases illustrate A LII' , possibly sug-
gestive of early progression to florid trilin cagc A~t L.
•
,
•
187
5
.'
188
592
19 1
CD3 and CD7 , and usually with CD4. Th ese cases are
ntial diffe re nce in cell type o r o rigin, since cases
prese nting as II ofte n show a change to U in relapse . classifiabl e as T -Cl L and are to be disting uished from
The disaimi nating morpho logical fea tures are never- T-proly mphocytic leuk aemia and also fro m CD +
illustrated in subsequ ent figures and furth er granula r T -ce ll lymp hocytosis. wheth er in an ea rly no n-
descri bed in the ir caption s. malignant sta ge . o r frankl y neoplas tic . Th ey co mmonl y
I unologically, A LL ca n be subd ivided into man y show skin infiltrati on and a mo re aggressive clinical
diffe rent types. but ther e arc five main gro ups: T-ALL co urse th an B-CL L.
derived from 'f-cell precursors (TdT+ , CD3+ , CD7+);
' null" ALL and C· ALL fro m ea rly B-<:ell precurs ors Prolymp hocytic leukaem ia (PL L) . This subacu te and
(TdT+ , CD 19+ and respecli vely CD I()- and CD I() + ); clinicall y intracta ble varia nt may eme rge by a form of
pre- B ALL from B precurs ors at a somewhat later malignan t progres sion in Cl. L. or may arise de novo .
sta ge (TdT + /-, cl gM + ); and B-ALL fro m a still usually in association with gross spleno megaly and
late r stage of Bccell develop menl (TdT-, srnlg -i- ). C- ofte n also he pat ome galy. The periphe ral cell count is
ALL makes up abo ut 70% of childho od cases, T-ALL usually very high , with prolymp bocyr ic feat ures of
abo ut 15% , null ALL about 12% , and B-ALL < 2%, In nucleolat ed but moderately pachych rom atic nuclear
ad ults. where only abo ut 15% of acute leukaem ias are chro matin and variable amount s of gran ular cyto plasm.
lympho blastic , C-A LL is relatively infreque nt , co m- Mosl cases of PLL sho w a Bcce ll phenot ype , with
prising perhaps 40% of all cases. o ther early B and pre - stro nger expressio n of sml g than in B-CLl, and with
B cases making up the differen ce . A small proport ion CD l9 positlvit y. but up to 20% of cases have a T-cell
of cases «2%) with dubiou s or mixed reactions rem ain phenotype , most often CD4+ .
d ifficult to classify immun ologi cally . Ap art from the
part ial association of B-AlL immuno logy with L3 Hairy cell leukaem ia (HC L) or leuk uem ic reticulo -
mor phology. there are few distin guishing cyto logical endosheliosis (LR E). Thi s cytolo gically fascinatin g
feat ures in parallel with the rem ainin g sub type s, except leukaem ia of B-eell lineage occurs in middl e years or
for the pres ence of localized par anuclea r dot positi vity lat er . four times as commo nly in men as in women. and
to acid phosphat ase and less markedly to o ther acid is associated with ge neral malaise , spleno megaly.
hyd rolases in T-ALL. occasio na l abdo minal lymph adenop ath y, mod er ate
Cytoge netically , seve ral non -random chro moso mal anae mia , th rom boc ytopeni a and neu tropeni a , and
abno rma lities have been identified in ALL. The se con spicuou s monocytop eni a . A few e ryth roblasts are
includ e t(9:22 ). occurring in nearly 20% of adult ALL usuall y to he found in the periphe ral blood . Th e bo ne
case s, h ut in only abo ut 5% of child hood cases; t(8;14) marrow is d iffuse ly infiltrate d with hairy ce lls (HCs)
in most cases of B-ALL ; t(I ;19), t( v; 12) and 6q- in a a nd also shows d iffuse fibrosis. so that atte mpts at
small proport ion of earl y B-cell cases; t(4; 1I ) in aspiration usually result in a dr y tap. T rephine biopsies
uncomm on cases with a tenden cy to develop mixed sho w a very characte ristic picture . as illustrat ed in
lymphob lastic and mon ob lastic fea tures ; and t(11;14) in 767-772 . The HCs ca n be recog nized and studied in
T-A lL. More import ant than these individual structural blood smea rs or in buffy coa t prepar at ions (7~766);
ano malies in terms of overall prognosis for the ALL the y have a striking and diagnostic cyto logica l appear -
ance in Rom anowsk y stains and have tartrat e-resistant
gro up as a whole arc chromo some num bers. hyper -
diploidy with 50--60c hromos omes be ing associated with acid phosph at ase (T RA P) positivit y and typica l PA S
a relatively good response to treatme nt , dipl oidy and cytoc hemistry . Th e lA P score is not abl y high .
near haploid y with an inte rmed iate one , and minor lmmun ologically, He . , like B-CLL cells, have receptors
hypodi ploid y o r pse udodi plo idy with most structura l for C3h (CD l1+) hUI , unl ike CLL ce lls, not for C3d
ab no rma lities a worse o ne . (C D21- ). They express activatio n ma rke rs such as IU
recepto rs (CD25 +) and have certa in relatively specific
Chronic lympho cytic leukaemia (C LL) . Thi s disease is mark ers such as CD22 antigen , strongly pres ent o n
rare before middle years and has an increasing incidence HCs but o nly weak ly present o n a minori ty of normal
with rising age . There is a periphe ral lymphocytosis B ce lls. Biochem ically, the ce lls produce mon oclon al
usually over lSx 10' /1, with variable lymph adenopathy , Ig, and DNA analysis indicate s the pre sen ce of clon al
sple no megaly, hepatom egaly a nd defe cts of marrow rearr an gements of both light and hea vy chain genes .
function manifest by anae mia and thromboc ytopenia. All these fea tures sugges t that HCs are activated
The extent of thes e clinical features forms the basis of B ce lls. at a stage before maturat ion to plasma cells.
various stagi ng systems, now in internat io nal use . Some HeL is of furth er unique int erest in bein g the onl y
10% of pat ients with ClL also show a more o r less haem atological malignancy to show almost inva ria bly a
seve re auto-im mune haem olyt ic anaemi a with sphero- dr amatic therape utic response to the biological age nt
cytos is and a positive antiglo bulin test. Hypoga mma- alpha-in terferon .
globulinaemia is co mmo n, especia lly in mor e advance d
stages of disease, and a monocl onal par aprot ein may be Sezary syndrom e (SS) . Thi s chronic T-eell neoplasm
produce d . proba bly always in case s with a Iymph o- represe nts the leukaemi c ph ase of mycosis fungoid es .
plasmac yto id co mpo nent to the cytology of both peri- combin ing the skin infiltr ation . nodular and exfoli ative
pher al blood and lymph nod es. CLL is usually a Bccell dermatitis and epide rmal Pautrier 's microabscesses,
disease with CDl9 positivity and at least wea k expression with the presence of typical Sezary cells in the circulating
of smlg, but a small proport ion «5%) of cases with blood . Th e cha racteris tics of Sezary cells includ e , most
CLL morphology and sma ll ag ranular lymph ocyte s importantl y. a deeply convoluted o r cerebrifo rm
with non -convo luted nuclei show par anuclear focal acid nucleus . not always clearly manifest . altho ugh it may be
phospha tase posi tivity. and react with the pa n-T MA bs suspecte d . in smea rs viewed under the light micro-
tQl
sco pe . but striking in electro n microsco pic preparati on s diagnostic. as illustrated in 798-830. although indivi
(cf. 77&-788). Th ere is genera lly a high nuclear-cyto- cytological features such as mult inuclearity. fragme u .
plas mic rati o . surface mem br ane reactivity with the ing cells, flaming cells. the saurocytes. Mott cells. grape
pa n-T MAb CD3 and usua lly with the helper cell cells. cells with intranuclear or cyto plasmic inclusion
marker CD4. although rarely with the suppressor cell and so forth are not in themselves indicative of neo-
marke r COS instead. T he cells are usually PAS·positive plastic change . since any of them may be found in
but rarely show localized acid phosp hatase reaction . It reactive plasmacytosis. An incr ease in plasma cells
seems likely that Sezary cells are the same as the con- abov e 20% in the bo ne marrow . or a smaller increase
volu ted T cells of mycosis fungoides. alt ho ugh the latt er with demon str ab le light chain restricti on . provides
sho w some immunological differences. notably CD? accept ab le evidence of myelo ma . confirmed when
and C 0 2S positivit y. suggesting that a possible activa- taken in conjunction with Xcray evidence of oste olytic
tion effect may follow infiltration into the dermis. lesions and the dem on strat ion of a circulatin g mono-
clonal paraprotein .
Large granular lymp hocytosis (L C L) and granular Clinically. the bony destru ction ofte n leads to hyper-
lymphocytic leukaemia (CLL) . This chro nic diso rder calcae mia. and myelo matou s infiltratio n of the kidn ey
with over lu x lU'J/l large gra nular Iymph ocytes in the freq uentl y produces renal failure and urae mia. Blood
peripheral blood, associated with splenomegaly but findings include a raised ES R and variable anaemia.
little other organ invo lvement. may have a ben ign but plasma cells are not often seen. except occasion ally
co urse when found as a respo nse to variou s chro nic in buffy coat preparations, and leucocytes and platelets
infective or auto-immune co nditions. but may also are generally present in no rma l nu mbers. altho ugh
occur in an appa rently pri mary neoplastic form when it ther e may be a mild leucoer yth roblastic picture with
is best regarded as a form ofT·CLL. T he large granular infrequent e rythroblasts and myelocytes de tect able .
Iymphocytes, with typically lymphocytic nuclei but with Levels of seru m beta 2 microglo bulin become in-
a mode rate ly low nuclear-cytop lasmic ratio and fine creasingly elevated with advancing disease and probably
scattered azurophilic granules in the cytopla sm. as pro vide thc best single prognostic index . although the
illustrated in 643. express the pan-T cell marke r CD? degree of ana emia and of ren al failure are also of
and are usuall y COR+ve. They may have cytotoxic progno stic importance .
functions, includin g natural killer (N K) cell activity. w aldenstro m's macroglobulinaemia is a form of
The y do not usua lly show focal paranuclear acid Iymphoplasmacytoid immunocytoma with bone marrow
phosphatase positivit y. but scattered granul ar react ion involvement and prod uction of a circulating monoclonal
(644 ). IgM paraprotein . As in myeloma, the re is usually a
high ESR and mod erat e-to -severe ana emia. but
A dult T-cell leuka emia/lymphoma lA TL ). This neo - leucocytes and platelets are genera lly normal. Th e
plastic disease has cyto logical feat ures somewhat bo ne marrow shows infiltration with Iympho plasma-
resemb ling. thou gh mo re exaggera ted than . those of cytoid cells and com monly an increase in macrophage
SS. with circulating T ce lls having con spicuously co n- iron and in tissue mast cells. manifest in both aspirat es
voluted nuclei . scattered acid phosp hat ase positivity. (834-842 and 844) and histological sections (843 and
and an imm unophenotype includ ing CD3 and C0 4 845-851). where the large intranuclear inclusions known
positlvity. but unlike SS. the disease is du e to infection as Dut che r bod ies and characteristic of this dise ase may
with a retroviru s. human Teccll Iymphotrophic virus 1 also be conspicuo us. Clinically. the course is ge ner ally
( HT LV l) , and is largely restricted to ce rtai n areas in more indolent than that of myeloma . and there is mor e
Japan and to Caribbeans of African descent. Clinically. resemb lance to a low-grade no n-Hodgkin's lymphoma.
the co ndition is associated with lymphadeno path y. with lymphade nopathy and some degree of hepa to-
hepatosple no megaly and co mmo nly hype rcalcaemia , splenomegaly mor e often fou nd .
and generally pu rsues an acute aggressive co urse .
alt ho ugh chronic fo rms exist. Lym phoma. Th is malignant disease arising in lymph
node s and involving ce lls of more or less matur e de gree
Mvetoma and waldenstrom:s macroglob ulinaemia . in the lympho cytic line . commo nly extends at some
T hese disorde rs involve a neoplastic gro wth of the stage to involve the bo ne marr ow and to liberate
plasma ce lls or Iymphoplasmacytoid cells at the end of abno rmal lymph ocytes in the pe riphe ral blood . If the
the Bvccl l maturation chain . with the form ati on and cell type is mature the disease is cytologically ind istin-
secretio n of a mo noclon al immunoglobulin or its guishable from chronic lympho cytic leukaem ia (C LL),
co mpo nent parts. In myelom a the plasma cells in an y and, if very primitive. as it may be in children especially.
individ ual case are usually of a single clone (rarely it is equally indistinguishable from acute T- or B-cell
biclonal). pro liferating in and displacing the hon e lymphoblastic leukaemia. In some cases. however.
marrow and producing a specific Ig. biochemically and abnormal 'lymphoma cells'. unlike either mature Iympho-
immunologically uniqu e. This is usuall y IgG or IgA . cytes o r the Iymphoblasts of acute le ukaemia bu t with
rarely IgM and even mor e rarely IgD or IgE ; but in characte ristics inte rmediate between the two . mav be
abo ut a third of cases . sometimes called Bence-Jo ncs see n. T hese cells arc usuall y of the B-cell line bu t 'J1l3.~
myeloma. o nly the light chain. kap pa or lambda. is be of T-ce ll lineage and include neoplastic vari ants ot
produced . There is associated bo ne destruction . d ue to the centrocytcs . ccntrobl asts and immunoblasts preseee
increased osteocla stic act ivity. secondary to production in lymph nodes and illustrat ed mor e fully in Pan S.
by the neop lastic cells of an oste oclast activatin g facto r The cytology and cytochemistry of all these
(O AF). Th e histological picture in treph ine biopsies is and abnor ma l variants as seen in blood and man
characte ristic and the cytology in aspirates usually illustrated in the following pages.
S9S
l 194
~ --
.;.
195
601 602
•
.
J
•
/
601. Another example of PAS staining in ALL. In this
case so me ce lls co ntain blo cks in additio n to coa rse
granules .
196
•
••
197
607
198
610 11
610
./
~O l
621 622
202
615
627
203
631
204
633 6.!
,~ ,. ,
, I
,•
.
/}
.. 4
,. •
•••••• • • • •
.
635
•• • • •.:,- •• • •
.-.. ....
0
•• • •• •
•• 0.
" • ,."
, ••
'. • ..- ..... ..
• .....
..
. •••• • e. • • e - •
•
•
• •·t •
•
633--635. A sequence of bone marrow tre phi ne biopsies 636
fro m a pati ent with C-A LL. und er going remission
ind uctio n. In 633. ta ken at the sta rt of therapy . the
specime n shows an overwhelming prep ond er ance of
primitive cells with large and leptochr o matic nuclei
often co ntaini ng visible nucleol i. There are only a few
cells of the ery thro id or later granulocyte series present.
wit h more pach ych ro matic nucl ei. A fte r two weeks of
01010xic combination chemot herapy. marrow aspirates
gave o nly a poorly ce llula r speci me n with too few cells
for differential assess ment , but the tr ep hine biopsy
sho wn in 634 revealed a severely hypopl astic picture
still containing a substa ntial pr opor tion of blast ce lls.
vith the co ntinuation of treatmen t . a furthe r trephin e
biopsy two weeks later reveal ed the emergence of a
remi ssio n picture with active pro lifera tion of normal
mopoietic cells of all myelo id lines, alt hough with
ervthroblasts predomina ting , as shown in 635.
Th ese fields illustra te again the impo rtant additional
-.formatio n to be o btained from mar ro w sect ions in
e le ukaemias, especially with regard to ove rall •
rity and the differe ntial compos ition of hypo-
. marrow du ring phases of att empte d re mission
. n.
205
6..'7
•
I
• •• ••
•
•
. - ••
637-639. Sections of bone marr ow tre phine biopsy
fro m the iliac cres t of an ad ult pa tient with sm Ig+ B-
•
ALL. moving from a period of chemo therapy-induced
remission to relapse. T he H&E-stained paraffin sectio n
show n in 637 illustrates the very mixed cell pop ulation
at this stage . with a megakaryocyte at the left and
numerous scatte red e rythroblasts and later gra nule-
cytes, but with blast cells having large leptochromatic
and freque nt ly nucleolated nuclei making up some
50% of the whole. The PAS stai n in 638 reveals
positively reacting neu trophi l polymorphs and earlier
granulocyte precu rsors . but the blast cells appear
negative. A furt her H& E stain , this time of a plasti c-
embedded thin section. again shows the mixed cytology.
with cells of different lineage readily distinguishable .
The B lymphoblasts. with their pale nuclear chromatin
and conspicuous nucleoli . co nstitute about 50% of the
pop ulation.
206
640
207
&B . Vari at ion s in lyrnphocyte cyto logy in a norm al
buffy coat smear . On e of the five Iymphocytes has litt le
cyto plasm and is prob ably either a B or a T I-l cell; o ne
has some marginal hair-li ke processes . not infrequentl y
seen in nor mal lymphocytes and not to be confused •
with the appeara nce of ha iry cells (see 748-750 ); and
the remaini ng thr ee Iymphocytes have relatively large
amo unts of cytopl asm with a few scatte red granules.
and are probabl y T y cells. A persiste nt increase in ce lls
of this cyto logical cha racte r. with positivity to the pan-
T MAb CD? and the suppresso r subset MAh CD S. is
found in large gra nula r lymphoc ytosis and granular
lymp hocytic leuk aem ia.
208
•
209
649-660. Reactive 'immunoblusts' and 'virocytes' arising
in various infective states. These activated lymphocy tes
are generally PAS, acid and alkalin e ph osphatase und
esterase negative.
210
652
211
655. Two neutrophil s. two lymp hocytc s. a mo nocyte
and four immun oblasts (activa ted Iymphocytes) - from
the buffy coat of a leuk aemic pat ient in remissio n but
with a mycop lasma infect ion. T hese cells must be
distingu ished fro m ea rly mo nocytes o r leukaemic blast
ce lls.
212
,
213
661-663. Sections ofbone marrow trephine biopsy from ",1_ ,- ... 66..'
a patient with diffu se marro w involvement with angio- (1 '7
im munoblustic lymphadenopathy (A IL) . . ,
A IL is generally regarded as a reactive process. although
with a high risk of transformation to a floridly neoplastic
non-Hodgkin :S lympho ma (NH L) . Mar row involve-
ment may be either focal or diff use . The low- and
higher-power views sho wn in 661 and 662 reveal the
cha racte ristic morphological picture , with a swirling
patte rn of branching endothel ium-lined blood vessels,
an interspers ed proli feratio n of activated lymp hocytes -
plasmacytoid immunobl asts and plasma cells. and a n
interstitial deposit of weakly staining amorphous eosino-
philic materi al. Th ere is often a compo ne nt of haem o-
lytic anaemia pr esen t in the se cases, and the marrow
may show areas of nor moblastic hyperplasia . as in 662.
wher e there is an island of normoblastic proliferation at .- ~
-
~
~ ,-
,
~
,
--. -.
;;- ,
the upper right corner. In the low-power view of a ,
reticulin stain in 663. the characteristic increase in , ",0
..... -...
reticulin fibres and their distribution in a swirling patt ern
around the pseudo-vessels is well sho wn.
-./,,.... .
1./-;--. ,
'. I
Y
:,.1/';
- '-- ,
,(
,.
,
~ I.. ..
214
66+-675. Protymphocytic leukaemia (PL L) . 666
:!15
667
•\.
i'.
2 16
670 "I
\72 673
.' -,
67~
670. P [ ~L: acid phosphatase reactio n. In this T-cell case
all the pro lymphocytes show strong localized reaction .
217
676
2 1S
678
..
220
68~ -
". (
,
.'
, f r> ,
) /:)
-, --
!
!
687 ~ . .:~.....
.. "'\ -r: -,
: . ' . '.
........ ",
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••
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.. .
;~ ~ :.~
.'-
. -"0- .. ~
.( ; -~":.
,;
,
·1 '
:.,..;~ ~
- .',..
• .'
.,.
,
." , -,.'. :
• •
687-692. Trephine biopsy sections from various cases
of e L L to illustrate the common morphological patterns
of infiltration.
-
222
690 r---,• o •.
• ·. I
•• ••
.' ..
. •.
......-,1..,. .• .
~
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e • • ".
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o
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to
.
• •_ e,
• • .'
••
'-
o • •
,
o •
690. A high-power view of bo ne marrow in late e L L 692
showing the diffu se nature of the infiltrate and its uni-
formity in bot h interstitia l and parat rabe cular areas,
and also the overwhelming predom inance of Iympho-
cytes and the pau city of resid ual no rma l haemopoietic
tissue . T his type of picture is usu ally found at advanced
sta ges of disease , commonly in association with marked
anaemia and thrombocytopenia.
~
:~~rp~h iC character of the interstitial cells, with
..res. various granulocyte stages, and a pre-
of erythroblasts, including numerous nudeo-
peoe ryt hro blasts. There is a suggestion of megalo-
change among the early erythroblasts.
223
693 69-1
69::
224
,
• , 0
•,., ,
• '.J •., , •
· '.. o" •
• •
v-, .·,. . "
• • ••• •
t.
,..(. ,
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...i- . ..........-.,-#-
• ~.I ....e' •
..., ,.
w
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•
•
0
'. " .11 '.
~
~
..- . -.
, •,
' ' •
11
;,
";1, . . ... .. . "
,et" .. ••••
-.- ,¥••
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•
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o
•
....0,.ro '';
,
,, •
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...•,. ... : • • • 0
•
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;
;::~_ of the large cells with vesicular nucl ei and
co nspicuo us nucleoli amo ng a mixed po pu-
c idual small Cl.L lymphocytcs. occasional
_lIP:" and erythroblasts, capillary endot helial
scantv fibrob lasts. Several of the large ce lls
ed nuclei and bear a gene ral resemblance
"_C!'1O and it is not surprising that Richter' s
at one time thought to represent a
tic tra nsforma tio n of Cl.l.. but both
aDd cytochemistry of these cells in smears
:EqII<n1 de mo nstra tion that they exp ress mono-
un m uno globulin co nfirm that they are
_ _obIasts.
701 a:
702. Anot her exam ple of lymp homa cells in the blood :
this represents a leukaemic phase of a secondary
ce ntroblastic lympho ma. There are multiple small
nucle oli visible in the irregular nuclei and cytop lasmic
basophilia is moderate.
.. .'
"
r.
•
713
1
•
--
•
••
716 117
231
7 19 ....- ------------- 7
."
7_
233
726 i
"
'.
731
235
73-1
...
738
· ' ...
I . .
.l..
• •
=',
•• •
• ••
• '.
•
,. -. ••
..~
. \.
• •• • •
."
•
--
~is hab le from those found infiltrat ing the marrow In
eLL.
~I l
238
744
747
239
•
751 754
n
759 ;
--- - - - -- -
762
763. SB sta in. sho wing thr ee negative HCs. two posi-
tive neutrophil s and two monocytes with var iable
scatte red positiv e granu les.
-.....••....._.......- ..
# ~
•
•~
• •
- , -,
s. ."
#.
I
t •
' .,.
..- .'.•
• iI •
• a ""
.
..,
•
••
•
•
•
•
770 and 771. T hick and thin sectio ns. respective ly,
from par affin- a nd plastic-em bedded bone ma rro w
tre phin e biopsy prep ar at ion s taken from a pati en t with
Be l d uring th e course o f remi ssion-induction treat-
ment with alpha-in terferon . In bot h sect ions, cha rac-
te ristic HCs are now in a minority, thoug h readily
recog nizable; no rma l haemopoietic mar row elements
predomina te, with erythroblasts most numerous bu t
with megakar yocytes and eve n occas ional gran ulocytcs
present. In 770 a tissue mast cell is see n at the lower
right and a plasma ce ll at the upper left ncar a mitotic
figure . In th is pai r of slides the thin section offers little
additional info rma tio n .
246
773
, ,
•
•• 774
•
...
• •
• ~
• •
•
•
•
• , •
• •
77~7 75 . Sections ofbon e marrow trephine biopsy fro m 775
patients with p eriph eral Tscell lymp ho m a and marrow
infiltration . Th is type of lymphoma is m orphologically
variable, composed of T cells ranging f rom sm all m ature
/ymphocy tes to large im munoblasts, and classifiable as
of either low or high-grade severity. They show a high
incidence of bone marrow involveme nt, mostly diffuse
but occasionally with a randomly scattered foca l distri-
bution. The histological structure of infiltrates mimics
that of the parent lymphoma , with an inflammatory
element and prominent vascularity, difficult to distin-
guish f rom pleomo rphic Hodgkin 's disease or various
reactive stales such as A I L , connective tissue diseases.
or im m une deficiency syndrom es. including A IDS.
2 ~8
77 9
249
782
250
786
251
7H9 7
/
792
• •
794 795
•
. .,.. ~. 0 ,
. ..• ..
'-~
.., .,
.. .,.,.. \
.. . '
79H
254
SOl
-
803
255
H05
256
•
8 11
813
257
814
..- ,-
815
" -
258
H20 11
•
•
822
823 and 824. PAS reac tions in plasma cells fro m case s
of mu ltiple myelom a. Reaction s ra nge fro m nega tive to
we akly positive . with occa siona l gran ules against a H2~
we a ke r diffu se ba ckground of faint positive tinge . T he
sphe rical bluish incl usions illustrated in 814 a nd 815
a re PAS-negat ive .
259
825
."
..
I
2Nl
829
.
•
.:,
••• • -
.........._ resid ual nor mal haem opo ietic mar row. with ,:.. ~.. ...,
stic. gra nulocytic an d megak aryocytic co rn-
all visible . at the lower left . Ov er all ccllulariry
ar ea is relative ly low . with num erous fat spaces.
. ::iI
.2
.
..
.... . ~ :oX ...
'\.. .. ,. .. ,.
~-J.
.
•
•
"
..--.
••
...
,
..-... '\.
..
•
• •
..
• ~
2hl
K3 \
262
,
IU6
...... ';: .
.I.... ~~ ~':
. ;"':'.; ~ ' .
' •• JII/jt'••' ,:••~'
.....' . .
263
83H. PAS stain in macroglobulinaemia: there is scattered
granular PAS positivity in the cytoplasm of two R E
,
cells. hut the lymphocytes and plasmacytoid cells in this
•
disease show little or no reaction .
•
839. Dual esterase stain in macroglobulin aemia . show-
ing several tissue mast cells with coa rse CE positivity in
a marrow fleck. The neoplastic lymphoi d cells a nd RE
cells in the fleck show only weak scatte red gra nular
pos itivity to both BE and CE .
... '.::.
'
~~~.
..
.....P7 . " w ~
"
264
R43. A high- powe r view of a sectio n of hone marr ow
trep hine hiopsy from a patient with Iymphoplasmacytoid
immunocyto ma and macroglo bulin aemia. providi ng a
clea r demonstration of the mixed lymphocytic. plasma-
eyrie. and inte rmediate cytology of the predom inant
infiltrating cells. Several binucleated plasma ce lls can
he see n.
265
lWi •
•
#
..
~
.. '•
,. .
• •.. •
"
•
." •
•••
•
, , ••
4
•
. .•, .
'.: •
'
•
•n. • ••
•
residual marrow with lymp hoplasmacytoid cells and
furthe r 'eosinophilic' mast cells. and also a conspicuo us
scatte red background deposi t of amo rphous eos ino phil
deposit. probably represe nting amyloid mat erial. T he
higher magnificatio n of 848 reveals qu ite clea rly the
Iympho plasmacytoid nat ure of the infiltrate and the
presen ce of various normal myelo id precursors, and
a lso confirms the prese nce of num erous tissue mast
cells.
266
849 xsn
•• •
•
•
•
849-851. Sections from a hone marro w trephine biopsy H51
tak e n from anot her patient with macro globulinaemia .
1149 sta ined by H&E . and 850 and 851 by the PAS
reaction to show various examples of Dut chcr body
formatio n in the neapl astic Iyrnphcplasmacyto id cells.
Th ese are PAS-positive nuclear inclusion bod ies pro-
hab ly derived from cyto plasmic im ·aginations . Similar.
but ge nera lly much smaller. int ra nuclear inclusions
occur in myeloma and less often in reactive plasma
cells. but the lar ge inclusio ns know n as Dutcher bod ies
arc see n especially in macroglobulin aemi a. where they
may be so lar ge . as in 849 , as to ca use fragment ati on of
the nucleus. A PAS-positive body stretching the nuclear
membrane and occupying most of the cell is shown in
850. while in 851 a who le se ries of these bodies , at
vario us stages of deve lopm ent . appears in the lower
pa rt of the field . Th ere is a single PAS · positive poly-
morph near the top.
Part 4
Reticulo-endoth elial ( RE) cells (re ticulum ce lls. arising in the bon e marrow. and wit h mixed cytol ogical
histiocytes. macrophagcs) are co mmon in the bo ne features of macr op hagcs and dend ritic RE cells.
mar row and have bee n illustr ated several times pre- The Leishman-D ono van bodies . the protozoal
viously in this boo k. Th ey take up foreign particl es. free par asites of Leishmani asis. appea r most prominently in
iro n. fat globules, specific gra nules from disrupted RE macrophagcs.
granulocytes, and o the r cell fragments. and therefore Malignant histioc ytosis is a ra re neopla stic state of the
ofte n co ntai n pha gocytosed mat er ial. Their cyto plasm RE cell and even rarer is histiocytic leuk aemia. Examples
appea rs fragile and is readily broken up in the smea ring of each ar e illustrated in this sectio n.
process o r str etched out between neighbourin g cells - so Apart fro m RE cells . most of the other ce lls illustrated
that the cytoplasmic outlines may be difficult to recog- here ar c less commonly encountered. hut they have
nize. T hey may co ntain various inclusions - sea-blue highly cha racteristic cyto logical features. and o nce these
histiocyte material, pscudo-Gaucher cell bire fringent features are appreciated the cells arc unlikely to be
lipid or blue crystals and gre y-green crystals. In ce rta in co nfused with normal or abn ormal varian ts of more
lipid sto rage disea ses the y appear grossly swollen with common cell lines in the bone marrow or blood.
ahno rmal fibrillary or globular deposits of lipids. A non-malignant form of histioc ytic proliferation with
The nature of this lipid inclusion materi al in the two enhanced phagoc ytosis. arising in var iou s oppo rt unistic
main forms of lipidosis. Ga ucher's disease and Niemann- virus infection s. especially in immun ocomp rom ised
Pick's disease . is illustr at ed in this section and discussed subjects . is virus -associated haem ophagocytic synd rome
in the acco mpanying captions. (VAHS); examples of this co ndi tion in trephine and
Glycogen stor age disease is also associa ted with the aspira tio n marrow biopsies are illust rated here.
accumulatio n of inclusion materi al. parti cularl y in Several different examples of tum o ur cells invadin g
rnacrophages. but 10 a lesser exte nt in man y other the marrow a re shown. Alth ou gh an isolated tumour cell
hae mic ce lls. may rarel y be iden tified as such in the abse nce of more
In chro nic infective or reactive states involving the typical cell clumps, the feature which allows identifica-
bone marrow. rangin g from tuberculosis to sarco idosis. tion in most instances is the occ urrence of ce ll nest s.
macrophage proliferation may lead to forei gn-bod y frequently partially syncytial. of cells not belon ging to
giant-ce ll (or Langhans' ce ll) forma tion . with appea r- any hae mic serie s. The identifi cation cannot often go
ances similar to those encountered in the lymph nod e beyond ' metastasizing tumour ce lls', but comparison
imprints illustrated in 1098 and 1099. with the histology. cytology and cytochemistry of the range
Anothe r group of gra nuloma to us states frequ en tly of illustrative metastascs shown in %(~I024 may provide
involving the bon e marrow a rc those of the ' histiocytosis a suggestive indicat ion of likely origin . Apart from
X: family. including Hand-Sch uller-Chri snnu syn- neuroblastoma. medullobl astoma and chemodectoma -
drome. eosi nop hilic gran uloma and l.cttcrc r-Siwc all relatively rare tum ours with characteristic cytology
disease. A ll arc thou ght to involve proliferation of found in yo ung people the meta sta ses most often found
-c
Langerhans' cells. pro minent in the skin but probably in marrow aspirat es o r trephin e biop sies from o lder
269
patient s. usually with lcucoery throblastic anaemia. are tropical zones, but their appe arance should be recog-
from primary malignant tumou rs of lung. breast, kidney nized by all haematologists: although what may actually
or thyroid. and examples of all these are shown here. be seen unde r the microscope (as in the photo graphs
Certain of the more common parasites which may be her e) does not always provide detail comparahle with
seen in smears of blood or bone marrow are also the diagrams and painti ngs in parasitology texts.
illustrated. They are found mostly in tropical or sub-
152
._ • ..27J
S55
... .
't
r-
272
•
,,
.• ,.
• '"
""
274
"
•
.. •
,
,
. ,,... '
." .
"
,
,
K64 . Fain tly visible blue crysta ls toge ther with coarser 1167
iron gra nules in a distend ed RE ccl l from the marro w in
a case of congenital dyscrythropoietic anaem ia (CDA)
type I. R E cells co ntaining such blue crystals. and
pseudo-Ga ucher cells. RE cells with crumpled swollen
cytoplasm. are found not uncommonly in C DA .
275
1169
. ~
r' •
.'
•
.-
. #-.,.. ' • •.
I
.
.. .. -r . . . ~~
•
-,
.~
~.
'\'"
.
.
.-
~
•
•
. " ,,
-.
276
874
•
•
876
277
879 -
.
•
•
• • •
•
880. Anot her exa mple of the non -birefrin gent type of
'grey-green crystal in a macroph age from the bone
marrow of a pat ient with A ML. Th e flat nucleu s and
spread ing cytoplasm of the macro phage are typical. and
the ce ll conta ins othe r phagocytosed mater ial , probably
chiefly iron. in addition to the grey-green crysta l.
278
882
885
886-889 . Marrow trephine sectio ns and aspirate smear
fro m ano ther patient with G auc her's disease . In the low-
po wer field in 886, the nodul ar ap peara nce results from
the pe rsistence of normal hae mopoieti c marrow , with
little infiltrat ion in the vicinity of the bony trabeculum at
the upper left. but elsewhere the Gaucher cell infiltrate
was very extensive in this case , Th e characte ristic
histo logical appeara nce of these cells in marrow sections
stained with H&E is well sho wn in the higher-power
view of 887, with their ample and swollen cyto plasm.
diste nded with fibrill ar mate rial. In the reticulin silver
sta in of 888 a scatt erin g of interl acing ret iculin fibrils,
somewhat increased ove r normal. is revea led. while the
accumulated cytoplasmic materi al has a mo re gran ular
appe arance in this stain a nd there is eve n a suggestion of
weak argyrophilia in the cytoplasm of some hlstioc ytcs.
Th e smear prepa ratio n in 889, with the usu al
Romanowsky stain , shows the same Gaucher cells as in
the preced ing sections. here with a mixed fibrillary and
granular or foa my appearance.
T he striking and highly charac teristic appea ra nce of
Gaucher cells is d ue to the accumulation within ex-
panded Iysosomes o f the lipid glucosy l ccramide. result-
ing from deficiency of the enzyme glucocc rebrosidasc .
890 .. 891
28 1
H9J
282
896 897
•
,. •
•
•
./
Co •
•
J •
••
1
896-898. Furt her fields from the same tr eph ine biops y 89H
of hon e marrow as shown in 895 , with two high-power •
ileitis from an II& E sta in and o ne from a G iemsa-stai ned
prepa ration. Th e histiocyte cyto plasm her e shows so me
• •
•• • • ••
granularity and there is evide nce of phagocytic activity.
especially in 897. where there arc also sever al mult i-
nucleated hisriocytes. In the Gicmsa stain a few of the
histiocy tes show a suggest ion of sea-blue inclusio n
materi al. • • •• •
As scc n previo usly. this appearance is not co nfined to
a single spec ific lysosom al enzyme deficiency as in the • .. •
•
.-
ma in her editar y lipid sto rage diseases. but appears to
occur most often as a manifestation of myelodysplasia
and perhaps accompanying increased ce ll dest ruct ion • •
•
invo lving especially the gra nulocyte and th rom bocyte
••
lines .
e. • •
•
• •
e. •
•
283
899
,
..
90 1
.~
284
905 906
907
912
'~
I'
,. . '....,
, :::-
'~ ,
'
:/ y.-
286
916 917
91K 919
'0
,
•
/
" •
; I
,
•
.. •
·c
922. Malignant histiocytosis (histiocytic medullar y
ret iculosis) showing erythro phagocytosis by bon e
marrow RE ce ll (histiocyte).
288
125 926
• ••
• •
•
•
•• •
289
928
•
• •
,' ,
.
, ., •
•
1',
t.
i
•
•
•
••
. •' • f
•
".
•
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•
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.'",..,...
.' ~. •
1
, • •
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.,- ,
,, -.., . " -
•
-, . •• • • , ••
, •• J
'I
-" . •
•
•
.'" ". • .. , . J:.
r:
' .-~,
•
• 1•
• •
"
• •
•
290
32 933
935
291
93ll
.. • ••
. .. • , .•
•
,.
"
. • ..;
• • •
. .1#. , "
•
, •"
'. ..
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I
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!
'. • •• • • I
• • • • ••
•• ••
• •
• • • •
,• .,
939
• \
.'. •• •
•
• • • •
• • •
• •
• •••
•
938 and 939, Two furth er eytoehemieally sta ined
field s from the same marrow aspira te as in 936 and 937.
The PAS stain in 938 sho ws a central ph agocyti c
macrophage , aga in with p redo mina ntly polymorph
debris baving the expected PA S po sitivit y. T he field also
co nta ins several early. PAS -negative . gra nulocyte pre -
cursors. The dual esterase reaction shown in 939 is
unu sual , in that the mass of ingested po lymo rp hs
appea rs predominan tly BE-positive rather tha n having
the normal C E positivity sho wn by gra nulocytes . as
man ifest for example in the ne ighbouring myelocytes in
this field . Some residu al CE reac tion ca n still be
made o ut. however. and it see ms likely that this e nzyme
is bei ng ove rridden by high activity o f the endoge no us
BE of the engulfing macrophage,
292
942 • 943
...
'
293
9~5
•
t"
•
I
.'
•
! , ...... ' .
.
. .. ., .~.
294
9511 951
•
.
• •
296
957
959
-.
•
297
962 • j•
• .
962-966. Examples of vascular endothelial cell clump s
in the peripheral blood . The very regular nuclear
structure and size and the tendency for these cells to
occur in sheets or streaks along the dire ction of spread-
ing of the film assist in recognition. Th ese ce lls arc
for eign to blood, and are lifted from the intima of the
vein during insertion o r withdrawa l of the needle used
for collecting the blood sample. Th e nuclei in 965 both
sho w conspicuous Ban bodies. indicating the presence
of the inactive X-chromosom al materi al of no rmal •
female cells . In 966 can be seen alkaline phosph atase
pos itivity in a strand of vascular endothelium. cross ing a
field of negative immature marrow cells.
298
7 969
•
•
•
299
971
-
•
fJ73 and 974. Stro mal fat cells in bone marrow smears.
300
5 976
,
• ,
,- I
"
• "
-.
,,
.
, e• • •~
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•
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7 97H
r
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. .
•
•
,
•
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'... ,
megaka ryocy tcs and residua l hairy cells. while the • "
detailed cyto logy o f all th ese ce ll types is seen in 976. The .' ••• • , •••
foamy ce lls a rc d iste nded with lipid . no dou bt derived
fro m dis integrat ing hairy cells. Th ey hea r a supe rficial
resemblance to the sphingo mye lin-lade n rnacro pbagcs
•
,• •
•• , .....
of Niema nn-Pick disea se. ."
, -, •• •
• ....
977 and 1)78. SB stain and consecutive iron stain on the
same field from a marrow aspirate ta ken from a patient
• .' . C-
-..'••
with A ML illustrating two cytoplasmic frag ments with
spurious loc aliza tio n of S8 positivi ty on coarse particles
• •••
of free: iron . probably derived from a disrupted macro-
phage. The two blast cells present show the expected • .• ,
...., , •
sudano philia of early grunulocytc precu rsors . - •
979. A thin sectio n fro m a plast ic-em bed ded bo ne
•, •
marro w tr ephine bio psy, showi ng an artefac tual scat-
tering of spherical globules due to the presence of bubbles •
in the embedding medium . T he bubb les ar c con spicuo us •
and rea d ily recognizable o ver e mpty fat spaces. h ut give
rise to co nfusion when over cellular area s.
•
•
•
•
."
•
.'DJ
980 981
302
9HS
••
Mixed cell clumps with ce ntral neuroblastom a cells
and peri pheral haem ic cells some times occur . whe n they
may be confused with norm al mar row Decks, especially
when dense . Examples are illustr ated sta ined with the 9H7
du al esterase rea ction in 985 and the Prussian blue sta in
for free iron in 986. In 985 there is the usual CE positivit y ,
in granulocytes and so me BE positivity in later norrno-
blasts and a megakaryocyte at the edge s of the cell
clump , while the neuroblastoma cells show moderat ely
..'
strong, finely granular, BE posiiivity . Th ere is no
det ect able free iro n in the tum our cell nest see n in 986.
• ' .
nor in the pe ripheral normobla sts . Th is contrasts with
the finding of norm al qu ant ities of free iron in the >
marro w Deck of haemopoietic tissue from the same , ."'. ,
slide . illustrated in 987, where neuro blastoma cells, if
prese nt. arc no t easily distinguished . , , . •
~
."
J()J
•• •
,
• • •• • ...
.\
• •
:.
•
•
988-992. Thi n sect ions fro m bo ne marrow tre phine
bio psies showing the histological appearance of neuro-
blasto ma infiltrat ion. In the low-power view of 988. the -,
pale-staining tumo ur cells form swirling colu mns
c, •
.• . - .
between the residual islands of normal hae mic tissue.
and in the higher- power field seen in 989 the typical '
features of these cells arc well shown: their large size .
their flimsy and indefinite cytoplasm, and their pale-
staini ng nuclei without obvious nucleoli but with
..
,
..
304
• . ;
~
•
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,
• •
~ ..•'. .
~
, ., .. '"
,
•
,"
,
.,
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./
,
,l
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,
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:;,
, , \ •
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••• •
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•
993-997 . Thin sectio ns from bo ne marrow tre phine
biopsies from pat ients with medull obl astom a invading
the marrow . Thes e tumour cells bea r some resemb lance
to ne uroblasto ma cells , but they are ge ne rally mo re
stro ngly stai ning , and form sma ller ce ll clumps or chai n-
like co lumns o f single cells in the bo ne marrow. and do
not usually ma n ifest th e swirled or who rled arra ngeme nt
co mmon in ne uroblastoma. The low-power fields of 993
and 994 show th ese cha racteristic gro wth patterns well,
with resid ual no rmal marrow ele men ts in the bac k-
gro und and a sectioned capi llary at the left in 994 . Th e
higher magnification of 995 and 996 reveals the
moderately dense nuclear chromatin and tende ncy to
fo rm ce ll chai ns and small syncytia l clumps . In a still
higher-pow er field (997), th e nuclei o f the medu llo-
blasto ma cells can be see n to have the sa me type of
hyperch rom atic spo ts as previou sly sho wn in neuro- 997
blasto ma ce lls.
305
•
Ii
999
• 11
1000. SB stain with a norm ally react ing polymo rph and
negative chemod ectoma cells.
.-
306
1006
307
oos
308
I 14
•
f
- •
.-.-• •
•., •
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; ~ -ii-,
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310
1019
,
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00 0
,
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0
or
1020. A high-power view tumour metasta sis in a bo ne ,
mar row sectio n. in this case fro m a pati ent with a thyroid
carcino ma. Th e cytology of the three co mpo ne nts ,
tumou r ce lls. fibrotic reaction , and resid ual marro w .
cells. can be see n in some de tai l.
311
•
22 ,..----------~,..------ 1 13
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312
.
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1027
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1028
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313
,
1030 ,
• , • : 100
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1030. A very low-power view of a section from a bon e
marrow tr ephine biopsy, showing a rou ghly circul ar
nodule of HD infiltrate in the lower half of the field . T he
'.
,.... ----. •
nature of the infiltra te cannot be det ermined at this
magnification , of co urse, but the field illustr at es how
.
infiltr ativ e nodul es ca n be det ected by sectio n scanning
at very low power .
••
••
1031. A high-power view of a neighbouring nodule to
that in 10~O , from the same section and sho wing the
mixed cyto logy with num erou s Hodgkin 's cells and
reactive eosinophilia. together with lymphocytcs and a •
co nside rab le fibrobl astic reaction ; the latt er is almost
always present in area s of marrow infiltra tion and is not
co nfine d to th e nodular scle ros ing vari an t of HD (which
does not ofte n involve the marrow),
•
314
1034
315
1036
1036. Ano the r exa mple of Leis hma nia, with o rganisms
within a mon ocyte and ot he rs liberated amo ng th e
surrounding cells. Below the monocyte is a clump of
platelets for comparison with the more sharp ly and •
positive ly staining Leishmania bodies.
316
0 ICJ.I
r
•
•
• , 'I
3 18
1 ~7
• •
•
-, •
•
•
:
,.
•
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319
10•
...
320
;3 105-1
...
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•
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.
1056
• •
•• • " • • • •
•
•
• •
". •
3:!1
1057
322
,\ \ 062
.'
. .
1061. Tryp anosoma gumbiense, the most import ant 1063
species causing African trypan osomiasis. Th e kinet o-
plast is much smaller th an the centra l nucleus, the
undul atin g membrane is broad and the contor tions of
shape variable.
323
•
Part 5
Cyto logical study of imprin t or needl e aspirate samples immunocy toc he mica l sta ins which fre que ntly re nder
of lymph nodes an d splee n co mp leme nts histo logical them particul arl y co nspicuous . Such stains may hel p in
st udy o f biopsy specimens and is especially va luable in ide ntifying the vari an ts listed above - fibro blastic R E
the diagn osis of infecti ve and reactive lymp hade n- ce lls are describe d as stro ngly positive for alkaline
opat hy, lympho mas and so me ot her metastatic tumo urs . phosph atase bu t o nly weakly so for es terases an d acid
and . in th e case of splenic mat erial. also in lipid oses, phos p hatase; den dr itic ce lls (C D 14 + , C D23 +) , mostly
mali gn an t histiocytosis and so me pri marily haem aro- fro m germina l ce ntres . are negative for phos phatases
logical d isorders with ex tra me du llary haemopoiesis. and weak in es terase ; interdigitating ce lls (C D40+ ).
T he no me ncla ture of lym ph nod e ce lls is confusing and chiefly from extrafollicular T vcel l zones , are nega tive for
several systems exist. In the illustrati on s here , certa in of alka line phos pha tase and react wea kly for acid
the more widely used synonyms are given initially, afte r phos pha tase and es te rase . The aut hors ' impressio n is
which the simple Kicl classification is used chiefly. that the se crite ria are no mor e th an ro ugh guidel ines;
readers bei ng expec ted to tra nslate as necessar y. T he ce rtai nly. the cytoc he mical po sitivity o f clea rly ph ago-
cytological differences between lymphomas, for example, cytic RE cells varies wide ly and is probab ly mu ch
are ge ne ra lly clear in the slides de picted, eve n if influe nced by the nature of the ingested ce llular
se ma ntic d ispute re ma ins . material. T he degree of variability of reaction in the cell
Phagocytic reticu lo-endoth eli al cells (RE cells) in gro up as see n in imprints is illustrated in this sect ion .
bo ne mar row and blood have been illustrate d pre viously We have includ ed illustrations of a wide ran ge of
in thi s volume; simila r cells arc conspicuo us in lymph malign an t disor ders affecti ng lymph no des. fro m
nodes whe re their development fro m mo nocytes Hod gk in's d isease (H D) and non -H od gkin ' s lymp homa
through an inte rme d iate mon ocyte/m acrophage or (N H L) . to seconda ry met astases fro m various tumours
epit he lioid cell may be envisaged . Ce lls o f th e same of non -haem ic cell origi n. Primar y lymph nod e tumou rs
family - perhap s the sa me ce lls in differ ent to pog raphi- are ofte n immedia tely diagn osab le with reason able
cal sites or at differ ent functiona l stages o f activity - ce rtai nty from impri nts mad e at biopsy, and typi cal
have be en separated by histo logists and electron micro- findings for variants of H D and N HL are pro vided .
scopists int o a se ries of type s. A pa rt fro m th e actively Histol ogical appeara nces in lymp h no de sections arc
phagocytic R E ce ll (histiocy te , macrophage , starry sky also incl ude d , par ticul arl y when th e struct ural infor-
cell, histiocytic reticulum cell) (CD 11b/c+) and th e ma tio n th ey pro vide is necessary for , or especially
ep ithe lioid cells, fibrob lastic, dendriti c and inte rdi gi- relevan t to , di agnostic classificati on . Fo r HD , the con-
tating R E or reti cu lum cells have been describ ed . Th ese ventiona lly accepted div ision s into Iymphocyte pre-
latter are no t ofte n clearly distin guishable in imp rint s, do mina nt, nod ular sclerosi ng , mixed cellularity and
perha ps beca use they te nd to rem ain in the suppo rting Iymp hocyte dep leted are followed , while for NH L the
stru cture of the node an d do not easily come free in Kiel classificat ion is gene ra lly used . The table here
tou ch or smear prepara tions . Th e R E cells th at do sho ws the co nd itio ns included in th at class ificatio n.
appear in imp rints are show n in Ro man owsky prep- together with their equivalence to an int ernatio nally
arations and also with va rious cytoche mica l and agree d 'working for mu lati on ' .
324
Classification of NUL Descriptions o f the histology and cytol ogy of
lymphoid tissu es and cells and of th e reactive and ne o-
\\'o rkin~ Form ulation Kiel Eq uivalent plastic conditions illustrated are given in further de tail in
Low grade Low grade the captions. Chromosome abnormalities frequen tly
A. Sm all lymphocytic. SL A . Lymphocytic: C LL occur in NHL. These include especially t(14 ;18) and
cons istent with CLL. Lymphoplasmacytic t(II ;14) in follicular lymphomas. involving the bel genes
Plasmacytoid Lymphoplasmacytoid on ISq and Ilq with the H-chain locus on 14q : the t(8 ;14)
Lymphocpithelioid of Burkitt's lymp homa and lymphoblastic lymphomas o f
T-zone lymphoma sim ilar cyto logy. involvi ng c-rnyc o n 8q with the sam e H-
B. Fo llicular, sma ll F-SC B .lC. Cc ntrocyticJ chai n locus ; and a ran ge of transloca tions in T-c ell
cleaved ccntroblastic fo llicular lymp homas invo lving th e various tcl ge nes o n lOq. lip
and diffuse and 14q. or the c-myc gene on Sq. with the alpha T-cell
C. Follicular, mixed F-M receptor locus on 14q.
small cleaved and Several examples of sections or imprints from nodes
lar ge ce ll with seco ndary metastases of solid tumours are given ,
interm ediate grade mostl y fro m nod e biopsies made before the primary
D . Ccntroblasridce ntro-
D . Follicular . larg e cell F-L tumo ur had been di agn osed and when lymphoma had
eyrie follirular o r diffuse been suspected . Since haematologists a re frequ ently
E. Diffuse. small cleaved D-SC E . Cenlrocytic . diffu se involved in the interpretation of such node biopsies. an
F. Diffu se. mixed sm all D-M F. Ccntrocytidcentro- appro priate range is illustrated he re .
and large cell +1- blastic diffu se Needle biopsy of the splee n may occasionally be
sclerosis/epit hc lioid diagnostica lly va luab le in co nd itions such as lipidoses
cells High grade and certain tropical d iseases like Leishm ani asis; a few
G . Diffuse lar ge cc ll D-L G . Ccntroblastic. diffuse illustrat ive examp les of the appearances of sp leen
High grade as pirates or imprints are provided .
H . Immunoblastic .largclB H . Immunobl astic Th e occur rence o f Icukaemic or lymphomato us infil-
ce ll (plasmacyto id . (B or T ) tra tio n in pleura , peritoneum or meninges, may lead to
po lymo rpho us. +1- C D 30 (ki - I)+ la rge the presence. o f neoplastic ce lls in pleural. ascitic o r
epithelioid cells) ce ll ana plastic. T o r B ce rebrospina l fluids. The neoplastic cells which may be
I. Lymphoblastic . LB I. Lymphoblastic see n in ce ntrifuge d deposits from these fluids look mu ch
co nvo luted convoluted T -cell as th ey do in buffy coat preparations from blood o r in
non -convoluted unclassified marro w. as illustr ated alrea dy in earlier sections of th is
J . Lymphoblast ic. small SNC J . Lymphoblastic . atlas. They are acco rd ingly not shown again here . bu t
non-cleaved Burkitt Bur kill ce ll type some illustrations are given of pleural lining cells . which
cell -type (B -ce ll) must be differentiated fro m neoplastic ones.
Miscellaneous: Mycosis fungoides and Sezary synd rome
(T< ell). extramedullary plasmacytoma (Bccell), histiocytic
and non -classifiable lymphomas.
325
1064
326
7
3'"27
1070 10
32 '
1074
329
, -
1076. Acid pho sphat ase: lymph nod e impr int : very
strong positivity is seen in a phagocytic RE cell contain-
ing much ingested material, while at the opposite corner
of the field a smaller monocyte-m acrophage shows less
intense butstill striking po sitiviry. Most of the remaining
cells are of the lyrnph ocyte , centrocyte, centroblast
familyand show no more than occasional weak granular
'. .
;
-,
9 1080
331
IOS3
" . •
•
..
or
.-
,
r
•
••
•
333
1091 1091
1093
335
11
1100
•
•
11
1102
•
,
11
1100- 1105. Cytoc he mical features of typical Langha ns
giant cells as see n in imprint s of a lymp h node biop sy
fro m a fema le patient with chronic gra nulo matous
disease . Th ese cells a re probably de rived fro m multiple
nuclear division s witho ut cytokinesis occurring in macro-
pha ges . The seque nce o f stains illustr ated her e includes .
first . a Ro manowsky prepar ation . with flat nuclei of
moder ate chro ma tin co nce ntra tion. witho ut clear
nucleoli bu t with single are as o f localized chroma tin
co nde nsation. usually at the pe riphe ry . eac h probab ly
represe nti ng the inac tive X chromo some or Bar r body.
The bluc agra nular cytoplas m is eviden tly fra gile and
eas ily disru pted . In ll Ol the SR stain is esse ntially
negative except for a few scatte red granules probably
derived fro m an ingested neut roph il polymo rp h. T he
haem ato xylin co unte rsta in used in the PAS reaction of 11
1102 again shows the Barr bod ies well. while the cyto-
plasm has weak diffuse PAS-positivity. 'The dual esterase
react ion of 1103 has a modera tely stro ng gran ular
positivity fo r BE in the giant cell. with aCE-pos itive
neutrop hil top left. In 1104 ther e is the strong positive
acid phospha tase reactivity to be expecte d in a cell of
macrop hag ic o rigin. but the alkaline ph osph atase stai n
in 1105 is negative .
336
Uh
1107
337
1109 • 11
1111 Ill.
"'i' . ,. . \-
..
1117
III
.•• .. . ....
I .
:;. ,.
,4 / ). .... ,
1119 and 1120. Acid and alkaline phosphat ase stai ns,
respectively. o n the lymph nod e biop sy materia l, bo th
field s showing positi vity in ph agocyti c macroph ages.
Lan gerh an s cells a re difficult to identify in these pre-
parations but appea r generall y negative for both
pho sph at ases.
339
1121
340
1125
., .' .
11 26
-,
, -;
, (
- .....
_ • 4 4
- : , .;
'.. _ 4 .
• ~ · ;-I " .-
r:
-: . "-."
34 1
128
343
•
113.3
9 11-10
1137. A field showing various bin uclca ted R-S cells and
mon onu clear Hod gkin's gian t ce lls. A ce ntroblast at th e
lower right corner and a centra l immunoblast show
morphological similarities to the smaller of the Hodgkin's
ce lls and suggest a possible derivation of th e R-S ce lls
from activated B cells.
3-15
1I ~3 11
"
I
•
• ;'
3~ 7
1J ~9 11:
~9
1155 1156
."
350
L58 ..,......,.... ..,.......- - - . . - - - - - - - . . , 1159
<
•
."
•
.... •
'.
z- ... ~
..
"
If
o
o. •
1160
.'
.' , .:
:. r:
.' . "
351
1163 11
11 •
1163-1168. Lymph node impr ints: lymphoblastic
lymphoma, B-cell type. I
352
11 -
33
1169 11
354
2 1173
-.
355
11 -
356
79 1180
357
182 11 ,
"
35
85 11
359
11
•
•
11
•
•
360
92 1193
361
11
1196 and 1197. Cha racte ristic tumou r cell clumps: the
cytological structure of these teratoma cells is quite
distinct from any normal or lymphomatous component
to be found in lymph nodes .
362
198 1199
1200
1202
363
103 12O-a
205 I~
-
•
j ...
, .
'
365
1111 120
367
•
219 1220
....
369
•
12'
370
•
28 1229
. ,
..
1231
371
1232 11
312
1235
• ,
•
.,....
••
I
\
r •
•
•
1238 123
374
IUI 12-1
• •
• •
:'
376
Appendix: staining techniques
ROM ANOW SK Y STA INS
R~agents: Reagents:
(a) Azur e Bvthiocyanate (He yl): 1.5 g. in 200 ml. dimeth yl (a) May-Griinwald sta in: Prepare 0.3% so lution of powder in
sulphoxide (DMSO) . methanol by grinding with pestle and mortar . Filter after
(b) Eos in Y (G) (Heyl): 0 .5 g. in 300 ml. methanol. 2-3 days. Befor e use dilute 1:1 with buffer solution
(c) Stock. stai n : Mix: (a) and (b) togeth er. Store in da rk. (phos pha te buffer. pH 7.2) . Diluted so lution should be
(d) Buffer : 2.38 g. HEPES in 1 I. distilled water (O.OI M); pH discarded afte r o ne da y.
adjus ted 10 6.8 with IN NaGH . Store at + 4°C. (b)Gie msa : Add 0.6 g. Gie msa powder to 50 ml. methanol.
(e) Wor king stain: Add 3 ml. stock stain to 41.5 mI. buffer plus Sha ke to dissolve. Add 25 mt. glycerine. Filter after 2-3
2.5 ml. DMSO. Fresh working slain should be made up days . Befor e use dilute 10 ml. stoc k solution with 90 ml . of
dail y. phosph at e buffer (pH 7.2).
Techn ique:
1. Fix.air-dried smears in und iluted Leishman for 3 minutes.
2. Di lutel:2 with phospha te buffer for 8-10 minutes.
3. Wash in distilled wate r or phosphate buffer .
4. BIoI dry .
N.B. 8-/0 minutes staining fo r periph eral blood sm ears is quite
sufficie nt. but bone marrow specimens require 10-20 minutes
dep ending on the cellutarity .
-
FREE IRO :-lSTA IN SUDAN BLACK B
(after MacFadu an and Da vis, 1947) (aftn Sheehan and Storey. 1947)
1. Fix. air-d ried smea rs in formalin vapour for 30 minut es. Reag,nls:
2. Wash in distilled water for 2 minutes. (a) Suda n black B (SB) (Gurr) : 0.3 g. in 100 ml. abso lute
3. Im merse in Co plin jar co ntaining equal parts 2% potas - etha nol.
sium fer rocy anide (Prussian blue) a nd 2% dil ution of pure (b) Buffer : Disso lve 16 g. crys talline phe no l in 30 ml. absolute
(37%) hydr ochloric ac id for I hou r . et ha nol. Add to 100. ml. distilled wat er in which 0.3 g.
4. Rinse with distilled water . hydrated disodium hyd rogen phosph ate (Na2HPO..+ 12H 20 )
5. Counterstain with 0 . 1% nucle ar fast red made up in 5% has been dissolved .
aluminium sulpha te solutio n for 30 minu tes. (c) Work ing stai n: add 40 ml. buffe r to 60 ml. S8 so lution a nd
filte r by suctio n. Kee ps 2- 3 months. Store in refriger ator .
Staining jars must be iro n-free .
Technique:
- I. Fix a ir-d ried smea rs in formalin vapour for ~IO minutes.
2. Wash briefly in distilled water a nd blot dry .
L fmmerse in work ing sta in for 1 hour.
4. Wash off with 70% e thanol.
5. Co unte rstain with Leishman o r MGG .
-40% wlv fo rm alde hy de saturated filter pap " in bottom of
Copiin jar.
377
PAS REACTION PE ROXIDASE
imodified f rom McMonus. /946) Mod ified Graha m-Knetl technique
Rtagt lflS: I. Fix air-d ried smears for 30 seconds (use stopwatch ) in
(a) Peri odi c acid sol ution: Dissolve 5 g. pe riod ic acid crystals 10 ml. -to% form alin a nd 90 ml. ethanol sol ution at room
in 500 ml. dist illed water . Sto re in dark bot tle. Keep s for 3 temperature .
months . 2. Wash with tap water for 10 seco nds and blot dry .
(b) Basic fuchsin: Dissolve 5 g. basic fuchsin in 500 ml. hot 3. Dissolve app rox . 250 mg. of hen zidine or o-t otidine " in
distilled wate r. Filt er when coo l. Saturate with SOl gas by 6 ml. eth anol a nd dilute with 4 ml. distilled wate r- Add
bubbling for 1 hour. Shak e with 2 g. activated cha rcoa l in 0 .02 ml. hydrogen peroxide (20 vol. ). When solution i
conical flask for few minut es unt il j ust clear a nd filter co mplet e. pour o nto slide without filtration a nd a llow to
imme diately through Wha tm an No . I filter into a dark react for 7 minutes .
bottle. Charcoal extraction should be done in fume cup- 4. Wash wi th tap water for 10 seco nds a nd allow to dry in ai r .
boa rd . Solution kee ps for 3--6 mo nths de pending o n how 5. Counters tai n with Lcishm an , diluted immediate lv with
often used. buffer, or use stand ard Ma y-Grunwald-Gi em sa tech niq ue
for co unte rsta ining .
Technique: • N .B. Both these compounds are carcinogenic and should be
1. Fix air-dried sme ars for 10 minutes in IQ ml . 40'Yo forma lin
handled with care.
and 90 ml. e tha nol so lutio n.
2. Wash bri efly in tap wat er .
Diaminohenzidin e ((lAB ) method for peroxid ase
3. Treat with periodic ac id so lutio n for 10 minut es .
4. Wash in distilled water a nd blot dry .
R~agtn ts:
5. Imm er se in Schiff's basic fuchsin in Co plin ja r fo r 30
(a) Fixative : Buffer ed fonnol ace to ne (B FA) - Na 2HPO..
minutes. ( Return fuchsin solu tio n to stoc k bo ttle imm ed i-
40 mg. , KH 2PO.. 200 mg. , ace to ne 90 ml. . co nce ntrat ed
atel y after use. )
form alin 50 ml., distilled water 60 ml. Stor e a t 4°C a nd use
6. Wash in tap wate r for 5-10 minut es .
co ld for fixatio n.
7. Counterstain with aqueous haematoxylin for 10- 15
(b) Stock phosphat e buffer : Dulbecco A , pH 7.3 - NaCl-tO g..
minutes.
KCl 1 g. . anhydrous Na2H PO .. 5.75 g. , KH 2PO .. I g. .
Co ntro l smears ar e exposed to saliva ry digestion for 30 distilled water I 1.
minutes between stages 2 a nd 3. (c) Working buffer: Dilut e I I. stoc k buffer with 4 1. distilled
water.
(d) Subs tra te : 3.3 'd ia mino benzi dine (D A B) .
(c ) H yd ro gen pe rox ide ( 1(x) vol.).
ALKAI.ISE PH OSI'll,\TASE (f) Working incubatio n solutio n: Dissolve 30 mg. DAB in
60 mt. wo rking bu ffer and add 120 1. of lOO va t. HlO l ; use
Rt agtnLf: immedi at ely.
(a) Stock propanediol buffer . 0.2M: Dissolve 10.5 g. 2- (g) Car azzi's aqueo us haematoxylin : Disso lve 75 g. potassium
amino-I-methyl propane-I I :3)-diol in 5(X) ml. distilled water . aluminium sulphate in 121) ml. warm dist illed water ; add
Store at 4°C; discard after 3 months. 1.5 g. haematoxylin powder dissolved in 3(X) ml. glycerol
(b) Working buffer . 0.05M: Mix 25 ml. stock buffer with by grinding with pestle and mortar ; dissolve 0.3 g. sod ium
5 ml. O. IN HQ and make up to 100 mt. with distilled iod ate in a littl e wat er a nd add grad ually.
water .
(e) Subst rate. to be made up freshl y immediately before use : TtC'hniqut:
Sod ium alpha-na phthyl phosp hate: 35 mg. I. Fix air-dried smea rs in BFA fo r 45 seco nds .
Fa..t Garnet GB C salt: 35 mg. 2. Rinse with distilled water and drain dry .
Working buffer : 35 mt. 3. Incu bat e in working substrat e so lution for 10 minutes.
(d ) Methyl green : 2% in distilled water . freed from co ntamina- 4. Rinse with distilled water.
tion with met hyl violet by extraction with chloroform and 5. Counterstain with Ca razzi' s (or ot her wat er -soluble )
kept free by storage at room temperature in co ntinuo us haematoxylin for I minute .
cont act with chloroform . 6. Rinse with distilled wat er and a ir-d ry.
Tuh niqut:
1. Fix air-dried smears in 10% formalin in absolute methanol
for 30 seco nds (use sto pwatch) at Cl-5°C. AC ID PHOSPHATASE
2. Pour fres hly pr epared substrate directl y on to slides and
incubate for 5- 10 minut es at room temperature. Substrate Reagenu:
must be used with in 5 minutes of preparation . (a) Naphthol AS·B I phosphoric acid : 10 mg .
3. Rinse slides in tap water for 10-15 seco nds . (b ) Fast garn et GB C salt (G urr) : 10 mg .
4 . Counterst ain with meth yl green for 1ll-15 seco nds . (c) Walpolc's acetate buffer , O.IM, pH 5.0: 50 ml.
(d ) Filter freshly prepared substrate into Coplin jar .
Good posi tive co ntrols are provid ed by slides from pol y-
cythae mia. infection. hairy ce ll leukaemia o r Hodgkin's
TeC'h niq u~:
disease .
1. Fix ai r-dried smea rs in forma lin vapour for 4 minut es .
2. Wash briefly in ta p wat er and blot d ry .
3. Incubate in subs trate sol ution a t 3TC for 1-1 '12 ho urs.
4. Wash briefl y in tap water .
5. Co unterstain with aqueous haematoxylin for 11l- 15 minut es.
For assessment of tartrate resistance add HXJ mg. U + )
tartaric acid to the substrare.
•
DUAL ESTERASE
379
•
Ch loramphe nicol. effect in eryrhro blas ts 75 Hisriocyt es, sea blue 869-873. 898 (see also
381
....
- - PAS reacnon in 600 . 60 1. 603. 6l).S . 615 .
3 2
- histiocytosis-X 892 M velo blast 67. 169. 170. 173-175. 178 (set' ... cytopla...mic disruption in 805
- H od gki n's d isease 283. 893. 894 .10,:40- leukaemia. A ML. myeloblastic) - dua l est er ase in 803. 827
IO~U Mye locy te 67. 169-IU. 178- 180. 185. 188. -- flaming a ppearance of cytopla... m in 807-
- kala-azar 1l}.lJ. IO-U 199 (see leukaemia . C M L) 1lO'I.820
- leucocvtosis . infective a nd reactive 279- - basophi l 305 - inclusions in vacuoles in 820
283 ' - eosinophil .284 - iron . free stai na ble in 792-797
- macroglobulinaemia SoU...S51 - giant 257 - locular degenerati on in 818
- malignant hi...ti0C)1OSis 92$-927 Myelodys plastic synd ro me (M D S) 69 . 7-9. ... multiple nuclei in 806
- M DS 122-124. 167. 168.569-571.575- 119-121.560-565.569-59-1 (seealso - in mveloma 798-83~'
58..' • Leukaemia . C M M L. Refract ory anaem ia - nucl e ar bridging in 804
- - hi...uocvtosi ... in 895-898 (RA). with sideroblasts ( RA S o r RSA ). - nucl ear inclusions in 812. H13. 818
- - pse udo-Gaucher ce lls in 874 with excess blas ts ( RA E B)) - PAS re action in 813. 817. 822-S24
- medullobla...toma 993-997 Mvelofibrosis ... in plasma cell leukaemia 801-803
- megalob lastic a nae mia 39~ 40 - ~arrow histology in se, 339 . J.W. 559. - Russell bodies in 814-817
- myelofibrosis 30. 339. 340. 559. 940. 941 9-10.941 ... the ..a urocv te varian ts of 81U. 811
- mvelo ma SOO. 828-830 - red cellsi n 131 Pla s modi um
- n~uroblastoma 9AA-992 M yelogranular d ysp lasia 560-562 - falcipa rum 1046-1048
-N HL Mvelo ma 192.798-833 (.fet' also Plasm a - malari a e 1049- 1052
- - Bccell. Burkitt ce ll type 742. 743 Cells) -- ovale 1054'
- - cen troblastic 712 - ac ute leu kaemic tran sformation o f 83 1- - viva x 1053. 105.5-1lI60
- - cen troc ytic 7 13-715 lU3 Plat elets 70. 456-459. " 62-1 66
- - cemrocyuccentrob lastic 734-736 - lymp h nod e imprints in 1192-11 94 ... gianl 463
- - immu nobtastic j tu, 711 a-Naphth yl acetate es te rase in ... phagocytosis of 464
- - lymp hocytic-ce ntrocytic 737-739 megakaryccytes 476 Pleural effusion
- - T -cetl . convol uted lymphoblastic 719 a-Naphthyl butyrate es terase (see Dual ... cytology of pleura l linin g ce lls 1248
- - T-cell. periphera l 773-775 este rase) - - PA S reaction in 1249
- normal 166 a -Naphthyl chloroacetat e esterase (see Dual Poikilocytosis (see Red cell s)
-- polycythaemia 23. 28-.\0 esterase ) Pol ychromasia (see Red cell s )
- thrombocvtbaem ia 482 Neurobla..tom a cell.. 9~1J92. 1211 Polycythaemia . primary and secondary. I~ .
-- thrombocytopenia . autoimmune .lS. 472. Normal blood leu cocytes lS-JO
473 -- acid phosphatase in 318 . 6.U . 645 Polym orpbs 67
-- VAHS 928-931 . 940. 941 - dual esterase in 189. 319 . ~ - ag ra nula r 212. 214. 21~. 268. 269. 274. 32.."
Mast cells 953-961 - PAS react io n in 186.314 - basopbil Ixt , 183. 184. IK6. 269. 30.5-308.
- d ual esterase in 960 - peroxidase rea ction in 184.316 31J-316.321.337.3.'8 .371...373 .~24-532
- in macroglobulinaemia 8J.&-.837. 839 - Sudan black Sla in in 181-183.315 ... in emerging rem issio n of ac ute leuk aemi a
- PAS reaction in 961 Normal bone marrow cell s 166 (see also !-46,547
- Suda n black sta in in 9$9 indi vidual ce ll type s ) - eosi nophil 181. IS6. 181- .'ta-l. 31J-JI6.
May-Grun wald Giemsa sta in 377 - acid phosph at ase in 187 1131-1133
May-Hegglin anomaly 70. 394. 395 - d ua l este rase in 188 - multi-lobed in pern icious a naemia 73. 7·1-
Med ullobl asto ma 993-997 - PA S re action in 185 265
Megak ary o bla sts 67 . •\41-348. 460. 484. 4X5 - peroxidase re action in 179 - neutrophil 73. 74. 176.181...184 , 186. 187.
Megakaryoc yt es 67. 456-462. 465--195 - Pru ssian blu e sta in in 110 189.201. 204. 207. 258-2Mt . 262-265 .
-- in e me rg ing remi ssion o f ac ute leukaemia
536-53_
- fragments in periphe ral blood 345-~\4X .
- Su dan black sta in in 180
No rmoblasts It . 1- 15
- in iro n deficiency a nae mia 106-108
....
26H. 271-273. 284. 306 . 31J-316. ~'H(~399 .
383
- in RE cetb 14. 15. 109. 110. 87 1. 873 - acid phosphatase in 862. 1076-1 078. 11$6. Star ch gran ules. in smears a nd imprints
- in ringed side roblasts 11.1-116 1167 122..~12 Z7. 1230. 1231
- in sea-blue histiocvte, 871 , 873 - a lk.aline pbosphatase in 13. 861. 1074. - birefringence of 1226. 1231
- in sideroblasnc anaemia Ill. 11.1-116, 118 1075 - PAS re act io n in 12~
- technique 3n - in blastomvcosis 103-1 Stem cell s 1R hae mopoiesis 8. 67
Pse udo-elliptocy tosis (see Red ce lls) - blue crys tals in 864-866 Suda n blac k stain (see indi vidual cells and
Pseuco-Ga ucner ce lls 864-868. 87.1-87f, - ce llular de bris in 852-854. 858 d isorders)
Pse uoo-P e tger - Huet phen o men on 215. JX8 - dual esterase in 863. I07 9-WS.2. 1148. - techniqu e 377
Pyruvate-kinase (P K) def iciency 1157 T ce lls
- renculocvt osis in 129 - in emergi ng remission of acu te leukaemia - helper 6-U-6.a8
Re d ce lls' 540-542 - sup presso r n.&2-64-l. 646. 647
- aca m hocyt es or spur ce us 143. 163. 164 - fat -laden ?7J/!, .v : Tart ce ll JM
- anisocytosis 62. 72. 73. 130, I.B. 13-1. 137, - free -iron stain in 1:1. IS. IU'J. 110.871. Terato ma . testi cul a r , in lymph nod e 1195-
I~ 873.923 1197
- basophi l stippli ng 72. 126. 148 - in Gauc he r's d isease 881-889.1222-1227 Th a lassaemi a . re d cells in 150. 151. 153
- bu rr ce lls 13. 141 - - bir efringen ce of 884. K85. 1226 Th e sa uroc ytes 810 , 811
- co-ocy ses (set' tar get ce lls) - grey -gree n crys tals in 879 Th romboc ytopenia 70
- cot tage loaf shape defect 142 - in Ha nd- Scb uller -Chn stian d ise ase 891 - megakaryocytes in 70. 467-a73
- crenation }·n. 142 - interdigitating 1069. 1072 Thrombocvtosi.. a nd th rombocvthaerma 70.
- c re sce nt formation 140 - in kala-azar 1035-100U '477-482 ' .
- dacryoc ytes 131.165 - in lym ph nod e impri nts a nd sections 6 16. Toxoplasma HMS. 1068. 1083-1 0%
- d repanocytes 13. 155 1066-1070.1072-1082.1133,1139.1148, Try pa nosoma
- echi noc yte s 13. 141 1156, 1167 - bruce i l{){i~
-
-
elliptocytes 13. 138
helme t ce lls 152. 153
- multinuclea ted 858
- in Nie rna nn- Pick disease 890 .: , ....
" ",
- cruzi 1063
- gambiense 1061
- hypoc hromia 132-13-1. 150 - PA S rea ction in 860 , 92-1. 1072 Tubercu losis . lymp h nod e imp ri nt in 1099
- leptocytes JSO' - as pseudo-Gaucber ce lls sti7. '168. 87~ Tumour ce lls in marrow 269
- M3~TOCVIO'> i s 62. 72-74. 146, 153 1I78 - bre as t carcinoma 1010- 1018
- rmcrcc yrosis 132 - - birefringe nce of 868 - bronchial carcinoma 1007-1009. JOl9
- ·Jrma I 16. 17 - see -blue ma terial in 86473 - chemodectoma (paraganglioma) 99~
- ,>olkllocyt osis 12. 13. 62. 72. 73. 131. 133. - in sp lee n imprint s 1220. 1222-1227 HMI3
13J. ISO. 152. 154 - Sud an bla ck sta in in 859. 1070 - Ewing 's sarcoma of bo ne 10 24
- · '·ar· ,lro p 131. 165 - tox oplasma in 1045 .- fibrosa~rcoma 1023
- pu .. chro-nasia 70. 125. 137 Ried er ce lls 596 - gas tric ca rcino ma 1004-1006
- pseudo-clhptecytosis 13. 139 Romanow sk v stai ns 377 - hyperne phro ma 1022
- -ouleau- I·U . 145 - purified re agent method (Heyl} 377 - med ulloblastoma 993-997
- schis tocyt osis 143. 153. 163. 164 Ro ule au x {see Red ce lls ) ,. - neuroblastoma ~992
- vc; ling Is.! . 155 Rus..o:.ell bodies 814-817. 1169-11 ".pros icca rcinom a 1021
- - rder tic 158. 159 Sa liva ry glan d impri nt 1238-12-13 - thy 1020
- sp her oc ytcs 13. 135-137. 153 Sa rcoidosis. Iymph nod e imp rints.1097. Uraemia. red cell cha nges in 143
- SpUT' cells 13 . 143, 163. 164 1098 Vacu ol ation
"pu, nik ce lls 163. 164 Schistocv tosis (see Red ce lls) ": ~ '-' - in bla st ce lls
- stomatocvt , , 13, 128. 158 Sea- blue histioc ytes 869-873 ... - of AL L 602.603, 613
- target ct.li<; j3 . 1$0. IS'. 153. 154. 163. 164 Sebaceous skin cell . in mar row sme ..r 972 - - of A M L 226-228. 30'1-313. 489. 509 .
Reed- Sterub -rg cells Serosal cells (see sple nic imprints) 516
- ac«t ph-spbatase in 1147 Sezary's syndrome 191. 776-788 - in Bur kin lymp homa cells 744-747
- d ual e- ' erase in 1148 Sick le ce lls (see Red cells) - in e . yt hro blasts . as ch loramphenicol or
- Ieu kae mic invol vement 1029 Sideroblasts 12. 114-118 alcohol effect 75
- :n lymp h nod e im pri nt s and ..ec tio ns 1121- Siderocyt es (see Red ce lls) - in hairy cells 750. 755
114H Smear ce lls. in e LL 676-678 - in irnmunoblasts 65H
- ill marrow 1025-1029 Splenic aspirates . imprint s a nd sectio ns - in mon cc ytes aee . 649
- PA S re action in 1146 - cyto logy an d cytoc he mis try of 1220- - in mon ocyt e-m acro phage s 855-H57
Re fract o rv anae mia 1237 Vascul ar endothe lial cells . in blood smear
- with excess blasts ( RAE BI SfiO-565. 574. - in G aucher's disease 1222-1229 962-966
577- 587 - in HeL 1216-1221 - lkelinephos phatase in 966
- with sideroblasts ( RAS or RSA ) Il2- U4 - in malignant histiocytosis 1233-1 237 ,infect io n (set' Imm uno bla sts)
Re n culocyt e s 11.12.70. 71. 1~129, 161 - serosal ce lls in 1231 rocytes (see Imm unoblasts )
Reticulo-en dot he lial ( RE) ce lls 269 . 852- Spu r ce lls (set' Re d ce lls) Vi ru s-associ at e d haemo ph agocyti c
924 (seealso Leu kae mia . histiocytic, Sputnik ce lls (see Red ce lls) svnd ro me 269. 92H.-939
M acro ph ages . M alignant histiocy tos is. Stab ce lls 169-171 , 175. 178. 187.258. Wald en strtlm 's ma cro globulina em ia (see
Mo nocyt e-macroph ages) 266 M acro glo bu linaemia )
.{
6 16- 0 7
9S
/ 6:..6: 95"
384