Prevention

This is truly one of the great success stories in transfusion medicine! Preventing Hemolytic Disease of the Fetus and Newborn (HDFN), particularly the severe form caused by anti-D, relies almost entirely on preventing the mother from becoming alloimmunized (sensitized) to the specific fetal red blood cell antigen in the first place

The Cornerstone: Rh Immune Globulin (RhIG) Prophylaxis for Anti-D

This strategy focuses specifically on preventing RhD-negative individuals from developing anti-D antibodies after exposure to RhD-positive red blood cells

What is RhIG?: A sterile solution containing concentrated IgG anti-D antibodies, derived from the plasma of screened human donors who have high levels of anti-D. It’s a form of passive immunization
Mechanism of Action (How it Prevents Sensitization): The exact mechanism is complex, but the leading theory involves immune clearance and suppression:
1. When RhD-positive fetal red blood cells enter the RhD-negative mother’s circulation (Fetal-Maternal Hemorrhage - FMH), the injected RhIG antibodies (passive anti-D) quickly bind to the D antigens on these fetal cells
2. These antibody-coated fetal cells are rapidly cleared from the maternal circulation, primarily by macrophages in the spleen, likely via Fc receptor interactions
3. This rapid clearance occurs before the mother’s own adaptive immune system (B cells and T cells) has a chance to fully recognize the foreign D antigen and mount its own active, long-lasting immune response (i.e., produce her own anti-D and form memory cells)
4. Essentially, the passive antibody “masks” the antigen and facilitates its removal, preventing primary immunization
Crucial Point: RhIG is prophylactic, meaning it prevents sensitization. It is NOT effective if the mother has already developed her own anti-D antibodies (i.e., is already alloimmunized)

Standard Administration Protocols for RhIG

To be effective, RhIG must be given strategically around times when FMH is likely to occur

Antenatal Prophylaxis (Routine)
- When: A standard dose (typically 300 µg in the US) is recommended for ALL RhD-negative, unsensitized (anti-D negative) pregnant women around 28 weeks of gestation
- Why: Recognizes that small, often silent, FMHs can occur during the third trimester, potentially sensitizing the mother before delivery. This dose provides passive protection for the remainder of the pregnancy
Postnatal Prophylaxis
- When: A standard dose (e.g., 300 µg) is given to the RhD-negative, unsensitized mother within 72 hours after the delivery of an RhD-positive infant
- Why: Delivery is associated with the highest risk and potentially largest volume of FMH. The 72-hour window is critical to intercept any fetal cells before the mother’s immune response is fully initiated
- Confirmation: The infant’s RhD type is determined from cord blood testing at birth
After Other Potential Sensitizing Events: RhIG is also recommended for RhD-negative, unsensitized women following any event during pregnancy that could potentially cause FMH, including:
- Amniocentesis, Chorionic Villus Sampling (CVS), Fetal Blood Sampling (FBS)
- Miscarriage, therapeutic abortion, ectopic pregnancy
- Significant abdominal trauma
- External cephalic version (attempting to turn a breech baby)
- Antepartum bleeding
- Dose Consideration: Smaller doses (e.g., 50 µg) may be sufficient for events occurring in the first trimester

Detecting and Managing Excessive Fetal-Maternal Hemorrhage (FMH)

The Problem: The standard 300 µg dose of RhIG reliably protects against sensitization from an FMH of approximately 15 mL of fetal RBCs (or about 30 mL of fetal whole blood)
The Risk: Occasionally, a much larger FMH can occur, especially during a complicated delivery or trauma. If the FMH exceeds 15 mL of fetal RBCs, the standard dose of RhIG may be insufficient, and the mother could still become sensitized
The Solution: Screening and Quantification
- Screening Test (Qualitative): Rosette Test
  - Performed on a post-delivery maternal blood sample (from an RhD-negative mother delivering an RhD-positive infant)
  - Principle: Maternal sample is incubated with reagent anti-D. If D-positive fetal cells are present, the anti-D coats them. Indicator D-positive cells are added, which bind to the antibody-coated fetal cells, forming visible clusters or “rosettes” around the fetal cells when viewed microscopically
  - Interpretation: A positive rosette test suggests an FMH potentially larger than 15 mL of RBCs and indicates the need for a quantitative test
- Quantitative Test: Kleihauer-Betke (KB) Stain or Flow Cytometry
  - Kleihauer-Betke (KB) Stain: Classic method. Based on the principle that fetal hemoglobin (HbF) is resistant to acid elution, while adult hemoglobin (HbA) is not. A maternal blood smear is treated with acid buffer, lysing adult cells but leaving fetal cells intact. The smear is stained, and fetal cells (containing HbF) appear bright pink/red, while adult “ghost” cells are pale. The percentage of fetal cells is counted microscopically
  - Flow Cytometry: More accurate, objective, and reproducible method. Uses fluorescently labeled antibodies against HbF (or sometimes anti-D) to quantify the proportion of fetal cells in the maternal sample
- Calculating Additional RhIG Dose
  1. Determine the percentage of fetal cells (from KB or flow)
  2. Calculate the volume of FMH (whole blood): % Fetal Cells x Maternal Blood Volume (est. 5000 mL)
  3. Calculate the volume of fetal RBCs: Volume of FMH (WB) x Fetal Hct (estimated or use 0.5) [Simpler calculation often used: `% Fetal Cells x 50 = mL of Fetal WB]
  4. Determine the number of 300 µg RhIG vials needed: Volume of Fetal RBCs / 15 mL per vial (or Volume of Fetal WB / 30 mL per vial)
  5. Always round up to the next whole vial and add one extra vial as a safety margin Example: KB shows 1% fetal cells. FMH = 0.01 x 5000 mL = 50 mL WB. Vials needed = (50 mL WB / 30 mL per vial) = 1.67 -> Round up to 2 vials, add 1 extra = 3 vials total
- Administration: The calculated dose of RhIG should be given within 72 hours of the sensitizing event (delivery)

Prevention of HDFN Due to Other Blood Group Antibodies

The Challenge: Currently, there is NO equivalent immune globulin prophylaxis available to prevent sensitization to other clinically significant red cell antigens like K, c, E, Fy^a, Jk^a, etc
Preventative Strategies (Limited)
- Transfusion Practices: The most effective strategy is careful transfusion practice for women of childbearing potential:
  - Avoid unnecessary transfusions
  - If transfusion is required, provide antigen-negative units whenever possible, especially for highly immunogenic antigens like K (i.e., give K-negative blood to K-negative individuals) and potentially Rh antigens (c, E) for RhD-positive women. This reduces the chance of primary alloimmunization via transfusion
- Early Detection: While not prevention of sensitization, early detection of these antibodies during routine prenatal antibody screening allows for appropriate monitoring and management of the pregnancy if the fetus is at risk

Key Terms

Rh Immune Globulin (RhIG): A solution of concentrated IgG anti-D antibodies used to prevent RhD alloimmunization in RhD-negative individuals
Alloimmunization (Sensitization): The development of antibodies against foreign antigens from the same species
Prophylaxis: Action taken to prevent disease
Passive Immunization: Providing temporary immunity by administering pre-formed antibodies (like RhIG)
Fetal-Maternal Hemorrhage (FMH): Leakage of fetal blood into the maternal circulation
Rosette Test: A qualitative screening test used to detect FMH exceeding the coverage of a standard RhIG dose
Kleihauer-Betke (KB) Stain: A quantitative test using acid elution to determine the percentage of fetal red blood cells (containing HbF) in a maternal blood sample
Flow Cytometry (for FMH): An automated quantitative method using fluorescent antibodies to detect and quantify fetal cells in maternal blood

Treatment

Cytopenias