1. CLS 2215 Principles of Immunohematology
Hemolytic Disease of
the Newborn
HDN

2. Hemolytic Disease of the Newborn
HDN occurs when the Mother has an antibody capable of crossing the placental barrier that is specific to an antigen present on the red blood cells of the fetus.
Fetal red cells become coated with the IgG alloantibody and undergo accelerated destruction both before and after birth.
Where does the baby get an antigen that is foreign to the Mom?
It’s the Dads fault!

3. Immunization and Production of Antibody
Mechanism of HDN
Immunization and Production of Antibody
Fetomaternal Bleed:
During subsequent pregnancies:

4. Categories of HDN 1. Rh System Antibodies 2.3.
Rh System Antibodies
Other Blood Group Antibodies
ABO Antibodies

5. Pathophysiology of HDN
Accelerated red cell destruction leads fetus to increase production of RBCs therefore there are increased numbers of nucleated RBCs.
Severe cases of HDN can result in:
Generalized edema of the fetus:
Severe anemia:

6. Pathophysiology of HDN
Bilirubinemia
Results from increased RBC destruction
Fetus in utero:
Newborn:

7. Prenatal Testing Patient History: Which pregnancy is this?
Has she ever been transfused?
Is she Rh negative? Has she had antenatal RhIg?
Does she have a previously identified unexpected antibody?

8. Test Mom for ABO, Rh (Weak D), and Antibody Screen
Prenatal Testing
Test Mom for ABO, Rh (Weak D), and Antibody Screen
Group O Mom :
Rh Negative Mom:

9. Positive Antibody Screen?
Prenatal Testing
Positive Antibody Screen?
Identify antibody and perform titration if antibody is clinically significant (anti-D, K, etc.). FREEZE the serum sample. If a subsequent titer is requested you need to compare the first titer results with the second titer. Run both titers in parallel and compare endpoints.
Has the titer increased? Two tube increase is clinically significant. May lead to more sensitive testing to determine severity of disease.

10. Rh Immune Globulin
What is it?
How does it work?

11. Rh Immune Globulin
Full Dose:
Mini dose:

12. Antenatal administration
When to give RhIg
Antenatal administration
Amniocentesis

13. Postpartum Administration
When to Give RhIg
Postpartum Administration
When Mother is Rh negative (and is negative for anti-D) and Baby is Rh positive. It is that simple.
How much?

14. Rosette Test
Purpose:
Qualitative:

15. Rosette Test - Principle
Add chemically modified anti-D to Mothers washed Post Partum (EDTA) red cells and incubate at 37oC. Anti-D will attach to Rh Positive cells present.
Wash cells and add R2R2 indicator cells. Indicator cells will “rosette” around anti-D that has attached to the Rh positive cells.
Centrifuge and resuspend the suspension and read microscopically looking for Rosettes.
Rosettes present?

16. Kliehauer Betke (Acid Elution)
Purpose:
Quantitative:

17. Kliehauer Betke (Acid Elution)
Principle: Draw a Post Partum EDTA sample from the mother and make and fix a blood smear on a glass slide. Flood the smear with an acid solution. The Hgb of adult red cells is washed out by the acid solution while red cells with Hgb F are not. Counter stain (Safranin) the smear. Cells with Hgb F stain red while the adult red cells remain transparent.
Count number of stained Hgb F red cells within 2000 adult (Hgb A) red cells.

18. Kliehauer Betke Stain Calculations
# Fetal cells / 2000 adult cells x 100 = % of Fetal cells present in the maternal circulation.
% of Fetal cells X 50 = number of mls of Fetal bleed
# of mls of fetal bleed / 30 = # vials of RhIg required

19. Cord Blood Studies
Required testing on the Cord Blood of Newborn’s with Rh Negative Moms (suggested on Group O Moms)
ABO group:
Rh typing:
Direct Antiglobulin Test:

20. Exchange Transfusion
Exchange transfusion may be definitive therapy for newborns with severe HDN.
A process where you exchange baby red cells with transfused red cells. Accomplishes the following:
Remove coated RBCs
Remove antibody
Lower bilirubin:

21. Compatibility testing for Exchange Transfusion
Crossmatch blood for exchange transfusion with Mothers serum. Why?
Can crossmatch with baby if mom is not available, but best indication of red cell survival is to crossmatch with the Mothers serum. Remember the source of the antibody is…

22. Which mothers are candidates for RhIg?
Mother Rh positive with anti-K
Baby Rh positive with negative DAT
Mother O negative with anti-C
Baby A negative with positive DAT
Mother A negative with negative IAT
Baby O positive with negative DAT

23. Which mothers are candidates for RhIg?
Mother: A negative with anti-D, C, K
Baby: B positive, +DAT eluate showed D, C
Mother: A negative with negative IAT
Baby: O positive with positive DAT, eluate neg
Mother: AB negative with anti-D
Baby: A negative with negative DAT