Keir Pickard (Haematology ST4 in the North-East) and Alex Langridge (Haematology ST6 in the North-East), reviewed by Andy Charlton, (Consultant Haematologist)

Introduction: red cell antigens

Transfusion medicine is fascinating! There are over 700 known red blood cell antigens. Every time someone is exposed to a red cell antigen they don’t express, for example through blood transfusion, they may produce a corresponding antibody. This antibody can then ‘attack’ and lead to a breakdown of the foreign red blood cell. Certain red cell antigens are more ‘immunogenic’, meaning they are more likely to provoke an immune response resulting in antibody formation. It is these antigens which are more clinically relevant as they can result in haemolysis.

We pay particular attention to red cell antigens in pregnancy, and not just because it is a common exam fodder. Rather than through transfusion, pregnant women can be exposed to a foreign red blood cell from the fetus they are carrying when it spontaneously, or through a trigger (see below), enters the maternal circulation. The maternal immune system could then produce an antibody which subsequently attacks fetal red blood cells, causing haemolysis and fetal anaemia. The most widely recognised and highly immunogenic antigen in this context is part of the Rh system: D.

When a woman of childbearing potential or a pregnant woman is D -ve and exposed to D +ve red blood cells, either through a blood transfusion or through contact with fetal red blood cells, the D antigen will stimulate the production of the corresponding antibody in her plasma: anti-D. The anti-D, as an IgG antibody, can cross the placenta and break down fetal red blood cells (if the fetus is D +ve), causing haemolytic disease of the fetus or newborn (HDFN). The reason that ABO antibodies rarely cause this problem is that they are mostly IgM antibodies and too big to cross the placenta.

Initial investigations

All pregnant women should have their blood grouping and a full antibody screen performed at booking (10 – 12 weeks) and also at 28 weeks. If there are no clinically significant antibodies detected at the 28 week screen, no further routine tests are required.

We will now cover 2 scenarios:

1. A pregnant woman is found to be D -ve but without evidence of anti-D
2. A pregnant woman is found to be D -ve and has anti-D antibodies

Scenario 1: D -ve women without anti-D

Around 20% of D -ve women carrying a D +ve fetus will produce anti-D. Anti-D production can be prevented by Routine Antenatal Anti-D Immunoglobulin (Ig) Prophylaxis (RAADP) whereby anti-D immunoglobulins are given as a single dose of 1500 IU between 28 and 30 weeks gestation (an alternative dosing schedule is 2 separate doses of 500 IU around that time). 

HANG ON A MINUTE – I thought we wanted to prevent anti-D production as this can cause HDFN, why are we deliberately giving anti-D?! It seems counter-intuitive, but the idea is that this ‘passive’ anti-D, which is prepared from pooled plasma donations, will rapidly clear any D +ve red blood cells from the fetus in the maternal circulation, therefore preventing the mother from producing her own anti-D. It seems simple but it does work (HDFN incidence has hugely reduced since introduction of RAADP). The amount of Anti-D given in RAADP is also not sufficient to ever cause HDFN itself. 

we do worry about so-called “sensitising events”, where an event results in lots of fetal RBCs entering the maternal circulation

BUT WAIT. If the fetus is D -ve, then isn’t all of this a waste of time and resources? You are right – the frequency of Rh D -ve in Europe is around 16% therefore lots of women are receiving anti-D unnecessarily. There is no perfect way to prevent this at the moment but the balance of risk is to give RAADP to all D -ve mothers. There are a few options which may develop in the future:

1. Check paternal Rh group – if they are D -ve, the child will be too. However you need to be absolutely sure of paternity. 
2. Cell free fetal DNA (cffDNA) can be detected in maternal blood to determine Rh D type from around 11 weeks – this is often used in Europe to guide anti-D Ig and appears very sensitive (false negative rate of ≤ 0.1%) and is being increasingly adopted in the UK

RAADP is given at 28 weeks because the likelihood of clinically relevant quantities of fetal RBCs in the maternal circulation is very low before this. But we do worry about so-called “sensitising events”, where an event results in lots of fetal RBCs entering the maternal circulation. Extra doses of anti-D Ig will be required within 72 hours of these events to reduce the risk of the production of maternal immune anti-D. Sensitising events are listed below along with the recommended dose of anti-D depending on gestation:

Sensitising events

• Amniocentesis
• Antepartum haemorrhage 
• Per vaginal bleeding
• External cephalic version
• Abdominal trauma
• Ectopic pregnancy
• Molar pregnancy
• Intraunterine death/stillbirth
• Miscarriage
• Therapeutic termination of pregnancy
• Delivery

Gestation	Minimum anti-D dose	Fetomaternal haemorrhage estimation
Less than 12 weeks	250 IU	Not required
12 – 20 weeks	250 IU	Not required
20 weeks – term	500 IU	Required
Birth	500 IU	Required

Note that from 20 weeks gestation it is suggested to check for the volume of fetomaternal haemorrhage (FMH). In simple terms, the fetus is now mature enough for a bleed to send enough RBCs into the maternal circulation that extra doses of anti-D Ig may be needed to ‘mop up’ all the fetal RBC’s. The most sensitive test to determine this is by flow cytometry: fetal D +ve cells in maternal blood can be identified using antibody-labelled markers, allowing estimation of the quantity of fetal RBCs to guide further dosing.

Scenario 2: D -ve woman with anti-D antibodies

As discussed before, anti-D can cross the placenta and destroy D +ve red blood cells, causing HDFN in a D +ve fetus. The concentration of the antibody is important: the higher the concentration, the higher the clinical risk. 

Monitoring should be done under the guidance of feto-maternal specialist units and the precise investigation and management is outside the scope of this article. Broadly speaking, the mother is monitored with repeat antibody quantification every 4 weeks up to 28 weeks gestation, and every 2 weeks thereafter. Once the anti-D level reaches 4 IU/ml, it is recommended to monitor the fetus with serial non-invasive middle cerebral artery (MCA) doppler to detect and monitor for fetal anaemia. If significant fetal anaemia is present, intra-uterine transfusion can be considered. Delivery will also be planned at 37 weeks to minimise fetal exposure to the antibody, and cord blood will be obtained to look for anaemia or haemolysis and to confirm the D status of the baby. Poorer outcomes appear related to high MCA peak systolic velocity, hydrops and early gestational age at first transfusion.

Useful links to resources:

• Buku Haematology – a free App for doctors, nurses and other healthcare professionals to help with the interpretation of haematology results and investigation of haematological disorders, with educational content available from their Twitter @bukuhaematology
• British Society for Haematology guidelines
• Royal College of Obstetrics and Gynaecology guidelines
• SHOT – Serious Hazards of Transfusion
• Blood Bank Guy – an educational resource by a transfusion medicine physician

References:

1. Contreras, M. (1998). The prevention of Rh haemolytic disease of the fetus and newborn – general background. BJOG, 105, 7-10
2. Kent, J., et al. (2014), Routine administration of Anti-D: the ethical case for offering pregnant women fetal RHD genotyping and a review of policy and practice. BMC Pregnancy Childbirth, 14, 87
3. Qureshi, H., et al. (2014), BCSH guideline for the use of anti‐D immunoglobulin for the prevention of haemolytic disease of the fetus and newborn. Transfusion Med, 24, 8-20
4. Sanchez-Duran, MA., et al. (2019), Management and outcome of pregnancies in women with red cell isoimmunization: a 15-year observational study from a tertiary care university hospital. BMC Pregnancy Childbirth, 19, 356
5. Accuracy of non-invasive fetal blood group genotyping and sexing at the International Blood Group Reference Laboratory, NHS Blood and Transplant – information sheet for service users

Disclaimer: this article is intended as a summary of theory and key concepts in management of anti-D in pregnancy and should not be taken as clinical guidance. Always check the relevant BSH and Green-top guidelines.