Introduction

General technique = agglutination

Antigen + antibody  red cell agglutination  visualisation

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 Introduction

Important factors  agglutination

1. Nature of antibody : IgM or IgG

2. Number of antigenic sites per cell

3. Localisation of the antigens in the membrane

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 Introduction

Important factors  agglutination

4. Temperature of the reaction : 37°C, RT,4°C

5. pH : no very important between 6 and 8

6. Zeta potential : natural repulsion of the RBC

To decrease the Zeta potential (IgG):

 Coombs antiglobulin
 Albumin
 Enzyme (papain, bromelin, trypsin, ficin)

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 Introduction

Important to note the importance of agglutination !

4+ 3+ 2+ 1+ (+) --

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 Blood grouping

Blood group = ABO + Rh(D) !!!! (At least)

ABO system : 3 alleles

L-Fucose
D-Galactose
H A B N-Acetylgalactosamine
N-Acetylgluosamine

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 ABO blood group

To define an ABO blood group :

1. Presence of antigen on the RBC membrane
2. Presence of the antibody in the serum (when the antigen is
absent) : natural regular antibody

3% 45% 8% 44% 6
 ABO blood group

6 tubes mandatory

Test repeated twice by two different technicians, different reagents

Globular test Seric test Autotest
(Beth-Vincent) (Simonin)

RBC  antigen
patient patient patient A B patient

Antibody
Anti-A Anti-B Anti-AB patient patient patient
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 ABO blood group

Interpretation

RBC
patient patient patient A B patient

Antibody
Anti-A Anti-B Anti-AB patient patient patient

+++ --- +++ --- +++ --- A

--- +++ +++ +++ --- --- B
+++ +++ +++ --- --- --- AB
--- --- --- +++ +++ --- O

Seric test = very important  Difference between an O blood group

and a misidentification due to outdated reagents

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 ABO blood group

Transmission : following Mendel’s laws

 A and B = codominants
 O = recessive
Parents

Phenotype

Genotype

Genotype

Phenotype

Children
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 ABO compatibility

Transfusion of whole blood

O AB

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 ABO compatibility

Transfusion of whole blood

==> double risk of incompatibility

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 ABO compatibility

Transfusion of red cell concentrates (packed cells) ans platelets concentrates

O AB

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 ABO compatibility

Transfusion of plasma : the rules are inverted !!!!!!!!!!!!!!!!!!!

O AB

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 Rh blood group

Old name : Rhesus

More than 50 antigenic specificities
5 more important antigens : D, C, c, E, e

Ag D : if present ==> Rh positive (85% of caucasians)

if absent ==> Rh negative (15% of caucasians) « dd »

Homozygotes D/D
When D+ (familial studies)
Heterozygotes D/-

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 Rh blood group

D variants : discordant results with different reagents !

Normal D Weak D (Du) Partial D

épitope antigène

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 Rh blood grouping

Reference technique : albumin

Patient’s RBC + anti-Rh(D) + albumin

 45 min, 37°C
Centrifuge

Note the importance of agglutination

Agglutination +  Rh positive
No agglutination  Rh probably negative

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 Rh compatibility

Rh(-) patients must receive Rh(-) red cells, except when :

 there are no Rh(-) RBC available
 in case of emergency
 and there is no anti-Rh(D) antibody in the recipient’s serum

Rh negative Rh positive

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 Pre-transfusion cross-match

Objectives :
 To verify ABO compatibility
 To detect irregular antibodies in the recipient’s serum
against RBC to be transfused (previous transfusion or
pregnancy)

Principle :
 Donor’s RBC + recipient’s serum, at least
 In saline medium  IgM antibodies
 With antiglobulin  IgG antibodies

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 Pre-transfusion cross-match

All antigens don’t have the same immunogenicity

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 Pre-transfusion cross-match

Precautions :
Tf°
1. One antibody ------------ several antibodies

2. The antibodies disappear with time (but be careful with

memory lymphocytes)

3. !!! « latent » immunizations  important to verify the

efficacy of a transfusion

4. When a cross-match is positive  the specificity of the

antibody(ies) has to be determined and compatible blood
has to be selected.

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 Consequences of an incompatible transfusion

Clinical consequences : very variable !!!

Ineffective transfusion

haemodynamic schock
with intravascular haemolysis

Biological diagnosis :
Haemolysis : bilirubin, LDH,…
Identify the responsible antibody

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