Anti – D Active & Passive

Titration in Pregnant Women

Speaker :
Ghulam Ullah
MSc Immunohematology
Senior Laboratory Technologist
QRI BB, HMC
Discovery of Anti D:
 First time in 1939 by Levine & Stetson.

 In women whose fetus had HDFN & who had hemolytic Tx

reaction to her husband’s blood.

 In 1940 Landsteiner and Wiener described an antibody obtained

by immunizing Guinea Pigs & Rabbits with the red cells of
Rhesus Monkeys.

 It agglutinated the red cells of approximately 85% of human

tested, and they called it the Rh factor.
Anti D Interaction:
Anti D production is a result of D antigen exposure after
transfusion or pregnancy.

D antigen integral (traverse 12 times) part of the RBC

membrane.

B Cells produce antibody molecules that are specific for a

target antigen. These B cells become part of the memory
against the targeted antigens and playing stronger and more
dangerous role (secondary response) at re exposure.
Significance of D antigen & Active Anti D:
After A & B antigens, D is the most important
red cells antigen in transfusion practices.
In Rh antigens D is the most potent immunogen
followed by c & E.
Formation of anti – D (active) almost always
results from exposure to D antigen through
transfusion or pregnancy .
Immunization to D can occur with a volume of
less than 100 ul of D POS fetal blood.
Active Anti D Cont….
Active Anti D persists for many years.
If serum antibody level fall below detectable
thresholds, exposure to D antigen will produce
a rapid secondary immune response and
causes significant hemolytic transfusion
reaction.
With rare exception anti D do not bind
complement, as a result it primarily causes
extravascular instead of intravascular
hemolysis.
Effect of ABO Incompatibility;
 ABO Incomp between mother and fetus has a substantial effect
against maternal immunization.
 The incompatible red blood cells of the fetus are destroyed by
maternal anti A or anti B.
Passive anti D (Rh Immune Globulin):
Cont…
 Rh immune globulin (RhIg) is a human plasma-derived
product first licensed in 1968.
 RhIg consisting of IgG antibodies to the D antigen.
 Full dose of RhIg (300 ug/1500 IU) is sufficient to counter the
immunizing effect of 15ml RBCs/30ml D-POS WB.
 The protective effect of RhIG on D NEG individual exposed to
D POS cells result from interference with antigen recognition
at the induction phase of primary immunization.
Mechanism of action & half life of RhIG :

 Possible mechanisms include rapid clearance of D+ red cells

from the maternal D NEG circulation.
 Interference in the processing of D antigen in the presence of
passively administered anti-D.
 Influence on the feedback mechanism between B and T cells.
 Coating of placental Fc receptors & inhibiting the transfer of
maternal IgG antibodies.
 Half life, 21 days.
 RhIg remained detectable for as long as 6 months.
Administration of RhIg:
 Antepartum:
• At 28 weeks of gestation.
• At Obstetric event that might allow fetal cells to enter mother
circulation.
• 50 ug dose of RhIg is adequate if D NEG preg is terminated before 13
wks of gestation. Above 13 wks full dose needed.
_________________________________________________
 Postpartum:
• With in 72 hrs of delivery.
• Partial protection could be provided as late as 13, possibly 28 days
after exposure.
• AABB st requires examination of a postpartum sample (1 hr) of all D
NEG women at risk of immunization, to detect the presence of FMH
that require more than one dose of RhIg.
Continued…
 Who are NOT candidates for RhIg:

a) D NEG women whose infant is D NEG.

b) Weak D POS women.
c) D NEG women known to be immunized to D.
Active vs Passive Anti D
 Passive ;
• Passive anti D is entirely IgG.
• AHG Titer rarely go above 4.
 Active;
• Immune anti D is a mixture of IgM + IgG.
• Reacting strongly at AB screening , resulting ↑titer.
• IgM part can be completely or partially deactivated
with 2mercaptoethanol or dithiothreitol.
Anti D Titration;
 Principle:
• Antibody titration is a semi quantitative method.
• Serial twofold dilutions of serum are prepared and tested for
antibody activity.
• The reciprocal of the highest dilution of plasma or serum that
gives a 1+ reaction is referred to as the titer.
• Critical titer for anti D is 16 at AHG phase.
 Titer should be performed with same method and same red
cells. Preferably with R2R2 cells. D antigen on R2R2 cells are
free of the position effect created by C.
 Testing frozen sample in parallel minimizes the possibility of
changes in results due to deference in technique.
 END

THANKS