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 LEUKOCYTE ALKALINE
PHOSPHATASE (LAP)
LAP is useful for differentiating CML from leukemoid
reaction.

PRINCIPLE
LAP is enzyme found in secondary granules of neutrophils.
The substrate naphthol AS-BI phosphate is hydrolyzed by
the enzyme at an alkaline pH.
This hydrolyzed substrate in combination with a dye such as
fast red violet LB or fast blue BB produces a colored
precipitate at site of the enzyme activity.
SCORING

LAP activity is scored in the mature

polymorphonuclear cells and bands only.
The activity scores range from 0 to 4+.
The scores of 100 mature polymorphonuclear cells
and bands are added for the LAP score.
 Score No. of Cells Score x No. of Cells

0 20 0
1 45 45
2 25 50
3 5 15
4 5 20
Total 100 130 = LAP score
 SUMMARY OF LAP INTERPRETATION

RESULTS OF LEUKOCYTE ALKALINE PHOSPHATASE

STAIN
Finding Score
Normal 20 – 100
Chronic myelogenous leukemia < 13
Leukemoid reaction > 100
Polycythemia vera 100 – 200
Secondary polycythemia 10 – 100
Negative neutrophil alkaline
phosphatase (NAP) reaction. A low NAP
score is seen in 95% of cases of
chronic granulocytic leukaemia. This
test is redundant if there is ready
availability of karyotypic or molecular A normal neutrophil alkaline
genetic analysis to detect t(9;22) and phosphatase reaction
BCR-ABL fusion respectively.
The Leukocyte Alkaline Phosphatase (LAP) test was used years ago to
determine if a patient had CGL or an infection.
Normal cells (PMNs) contain the enzyme leukocyte alkaline phosphatase (LAP).
In CML the PMNs lack or contain a decreased amount of LAP (Right).
(LAP Stain x Oil)
Both peripheral blood samples shown above were stained for LAP. The sample
on the left is a normal blood sample, and shows positive LAS staining. The
sample on the right is from a CML patient. LAP staining is characteristically low
or absent in CML.
      ALKALINE  PHOSPHATASE  REACTION

Reaction product: red 
granulation

Neutrophils: 
The more mature the cell, the 
more positive the reaction; 
maximum intensity occurs in 
segmented forms of neutrophilic 
leucocyte. The reaction intensity 
is evaluated according to 
Kaplow's scoring system: 
0  points - no reaction  
1 point - weak diffuse positive 
reaction or single granules 
2 points - diffuse positive 
reaction and a few granules  
3 points - strong positive reaction 
with numerous granules  
4 points - very strong reaction 
with a lot of granules, covering 
the nucleus. 
The score from evaluation of the 
100 neutrophils shall be between 
20 - 100 points. Other blood cells 
do not contain the enzyme.

Staining: azo-dye method

Comment: Very weak, diffuse only reaction of alkaline 
phosphatase in granulocytes, which is seen in a form of pink  Magnification: x 1000
coloured cytoplasm (score 1 according to Kaplow’ scale).
Comment: Diffuse and granular phosphatase reaction of little intensity in a
segmented neutrophil leucocyte (score 2 according to Kaplow’ scale).
Comment: Intense positive phosphatase reaction shown as red granules in a
segmented neutrophil leucocyte (score 3 according to Kaplow’ scale).
Comment: Very strong activity of alkaline phosphatase reaction is seen in a
segmented neutrophil leucocytes (score 4 according to Kaplow’ scale) and a weak
diffuse reaction (score 1) is seen in the segmented neutrophil leucocyte above.