CLS 2215 Principles of Immunohematology Hemolytic Disease of the Newborn HDN

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!

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

Categories of HDN 1. Rh System Antibodies 2. 3. Rh System Antibodies Other Blood Group Antibodies ABO Antibodies

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:

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

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?

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:

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.

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

Rh Immune Globulin Full Dose: Mini dose:

Antenatal administration When to give RhIg Antenatal administration Amniocentesis

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?

Rosette Test Purpose: Qualitative:

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?

Kliehauer Betke (Acid Elution) Purpose: Quantitative:

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.

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

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:

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:

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…

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

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