BLOOD GROUPING

(Forward and Reverse)

Dr Darshita Sharma
JR – II
Pathology
RDGMC, Ujjain
OBJECTIVES

– Importance of blood grouping

– Landsteiner’s law
– Typing techniques
– Slide method
– Tube method
NEED FOR BLOOD GROUPING

– BLOOD TRANSFUSION
– HEMOLYTIC DISEASE OF NEWBORN
– PATERNITY DISPUTES
– MEDICOLEGAL USE
– SUSCEPTIBILITY TO VARIOUS DISEASES
– ROUTINE HEALTH CHECKUP
LANDSTEINER’S LAW

– If an antigen is present on a patient’s red

blood cells, the corresponding antibody will
not be present in the patient’s plasma under
normal conditions.
UNIVERSAL DONOR AND RECIPIENT

UNIVERSAL DONOR
GROUP O
– Neither A or B antigens

UNIVERSAL RECEIPIENT
GROUP AB
– Patient has no Anti A/Anti B present.
– Cannot lyse any transfused cell
ABO TYPING TECHNIQUES

– Slide test
– Tube technique
– Microplate
– Gel system
Slide Test
Procedure:
ANTISERA
FIRST ADD ANTISERA TO SLIDE
THEN SAMPLES ARE ADDED TO SLIDES
OBSERVE FOR AGGLUTINATION
Sample 1
Sample 2
ADVANTAGES:
– Preliminary typing tests
– Use during camps

DISADVANTAGES:
– Less sensitive
– Drying of reaction giving to false positive
results
SLIDE GROUPING PRECAUTIONS:

– Test should be done at room temperature or

lower
– Slides should be dry and labelled properly
– Antisera should always be added before
adding cells
– Results should be recorded immediately
after observation
– Agglutination is interpreted as positive
result
Test Tube Method
Recommended method (Gold standard)
– Allows longer incubation of antigen and antibody
mixture without drying
– Tubes can be centrifuged to enhance reaction
– Can detect weaker antigen / antibody

Two steps in ABO grouping

Cell grouping (Forward grouping)
– Tests the patients red cells with known Anti-A & Anti-
B to determine the antigen expressed
Serum grouping (Reverse grouping)
– Test the patients serum with known A & B cells to
determine the presence of antibody
CELL GROUPING ( Forward grouping)
– Prepare 5% suspension of test sample in normal saline
– Set three tubes and label them as A,B, D
– Add two drops of anti A , anti-B, anti D in three different
tubes
– Add one drop of 5% cell suspension (Ratio of 2:1)
– Mix contents well and centrifuge at 1500 rpm for 1
minute
– Observe for cell button
– Gently disperse cell button and check for agglutination
– Confirm negative results under microscope
TUBE METHOD
SAMPLE ADDED
MIX TUBE CONTENTS
CENTRIFUGE
VIEW FOR CLUMPING
GRADING AGGLUTINATION
SERUM GROUPING ( REVERSE
GROUPING)
– Prepare 2-5% suspension of pooled cells A,B,O
– Label three tubes A cells, B cells and O cells
– Place two drops of serum in each tube
– Add one drop of cell suspension ( A cell to A
tube, B cell to B tube and one drop of O cell to
O tube.
– Centrifuge tubes at 1500 rpm for 1 minute
– Gently disperse for agglutination
– Negative results check by microscope
GRADING OF AGGLUTINATION
Thank You