Blood Grouping and Red Cell Antibody Testing in Pregnancy

(BCSH 2015, GTG 2014)

 

15% of UK population Rh D-, therefore 60% of Rh D- mothers will carry a Rh D+ baby

 

Intro

 

G&S should be tested at booking and at 28 weeks in all pregnant women.

 

1% of pregnant women are found to have a clinically significant antibody (most anti-D)

 

Effects of antibodies

-       Fetal and neonatal anaemia (hydrops)

-       Neonatal jaundice (kernicterus)

-       Perinatal loss (Anti-D, anti-K, anti-c)

-       Delays in timely provision of blood due to difficulty in performing screen

 

Sampling

-       Patient consent

-       Hand-labeled at patient bedside

 

Lab Tests

 

Testing Process

-       ABO and D grouping

-       Fetal D Group by cffDNA from 11+2 weeks

o   High throughput screening for Rh D- mothers to D type the fetus

o   Recommended by NICE 2016. 99.9% sensitivity for excluding D positivity

-       Red cell antibody screen (no need to use enzyme technique in the screen)

-       Antibody Identification

o   Once detected, antibody concentration should be monitored throughout pregnancy

-       Antibody Quantification

o   Only available for Anti-D and Anti-c

o   Reported as IU per milliliter

-       Antibody Titration

o   Used to assess concentration of antibodies other than anti-D or anti-c

o   Doubling dilutions of plasma tested by IAT using heterozygous test cells (e.g. Kk)

o   If a rise in titre is detected, ideally re-test previous sample in parallel with current

-       Paternal Testing

o   If Ab detected, consider paternal testing to predict likelihood of HDN

o   Beware partner not being the biological father

-       Fetal Genotyping by cffDNA from 16 weeks

o   RHD, RHCE and KEL*01 genotypes

o   Avoids fetal sampling and pitfalls of partner testing

o   cffDNA is useful when either a, b or both are present with c:

§  a. Pregnant woman has a clinically significant antibody

§  b. Pregnant woman has a history of HDN

§  c. Father’s antigen status is unknown, or known to be corresponding antigen

 

Red Cell Antibodies Detected in Pregnancy

 

Anti-D, Anti-K and Anti-c are the Ab most frequently implicated in HDN severe enough to warrant antenatal intervention.

Women with a history of HDN should be referred to fetal medicine specialist before 20 weeks, regardless of the antibodies detected.

 

Distinguishing between immune and passive Anti-D

-       Passive and immune anti-D cannot be separated serologically

-       Ab concentration should be quantified by continuous flow analyser (CFA)

-       Passive anti-D

o   Concentration will fall with time

o   Concentration rarely exceeds 0.4 IU/ml

o   Detectable over 8 weeks after a 500 IU dose of Anti-D

-       Immune anti-D

o   Concentration will remain stable (or rise if there is re-stimulation)

o   First detectable 4 weeks after exposure to D+ cells

 

Procedure when anti-D is detected in pregnancy

-       Confirm whether anti-D was administered (not just issued)

o   And its indication, both asking patient and in written notes

-       Quantification of Anti-D (unless immediately prior to delivery, in which case monitor baby)

-       Monitor the antibody 4 weekly to 28 weeks, and 2 weekly thereafter until delivery (or until MCA Doppler has been instituted or anti-D is no longer detected) in any of the following situations:

o   Level ≥0.2 IU/ml

o   Anti-D detected at or before 28 week RAADP dose

o   There is no definite record of prior anti-D administration

o   Anti-D was present before first administration of anti-D

-       If Anti-D level is ≤0.4 IU/ml after up to 1500 IU od anti-D has been given, then prophylactic anti-D should continue to be offered in all appropriate situations, unless it is beyond doubt that this is immune anti-D

-       If doses >1500 IU have been given then passive anti-D can cause levels >0.4 IU/ml and so anti-D should continue to be offered.

-       After 28 weeks, if all of the following apply, no further antibody testing is required:

o   Anti-D was not detectable in sample at 28 weeks prior to RAADP

o   And There is a written record of anti-D administration in the previous 8 weeks

o   And Level of detected anti-D is now <0.2 IU/ml

 

Pregnant women with immune Anti-D

-       Anti-D is most frequent cause of serious HDN

-       Test anti-D level monthly to 28 weeks and every 2 weeks thereafter until delivery

-       An increase in concentration >50% is a significant increase

-       Prev history of HDN + Concentration à predicted outcome

o   <4 IU/ml – HDN unlikely

o   4-15 IU/ml – moderate risk, refer to fetal medicine unit

o   >15 IU/ml – high risk, refer to fetal medicine unit

-       MCA Doppler to detect fetal anaemia may be used at the fetal medicine unit

-       Fetuses at risk of HDN should be delivered at 37 weeks to minimize exposure to maternal blood group antibodies

-       D status of fetus should be determined using cffDNA grouping (from 11+2 weeks)

 

Pregnant women with apparent anti-C+D, possible anti-G

-       A proportion of antibodies apparently typing as anti-C+D (often with high anti-C titre) are actually found to be anti-G or anti-C+G

-       These patients are at higher risk of developing immune anti-D

 

Pregnant women with immune anti-c

-       May cause delayed anaemia in the baby

-       Test anti-c level monthly to 28 weeks and every 2 weeks thereafter until delivery

-       Prev history of HDN + Concentration à predicted outcome

o   <7.5 IU/ml – HDN unlikely

o   7.5-20 IU/ml – moderate risk, refer to fetal medicine unit

o   >20 IU/ml – high risk, refer to fetal medicine unit

o   Co-existence of an Anti-E increases severity of fetal anemia. Refer at lower levels.

-       MCA Doppler to detect fetal anaemia may be used at the fetal medicine unit

-       Fetuses at risk of HDN should be delivered at 37 weeks to minimize exposure to maternal blood group antibodies

-       c status of fetus should be determined using cffDNA genotyping from 16 weeks

 

Pregnant women with immune anti-K or other Kell antibodies

-       Other Kell antigens (k, Kpa, Kpb, Jsa, Jsb) should be treated in the same way as anti-K

-       Majority of cases of anti-K in pregnant women are consequence of previous K+ transfusion

o   Female <50 y.o. who are K- or unknown should receive K- blood

-       HDN due to anti-K is characterized by low Hb with normal amniotic/cord bilirubin levels

-       Test anti-K level monthly to 28 weeks and every 2 weeks thereafter until delivery, unless father is confirmed to be negative for the relevant antigen.

-       Prev history of HDN + Concentration à predicted outcome

o   Refer to fetal medicine unit once anti-K detected. Anaemia can occur at low titres

o   Titre 1:32 or greater –->  HDN more likely

-       KEL*01 status of fetus using cffDNA if father is heterozygous or unknown.

 

Pregnant women with other red cell antibodies

-       Only IgG antibodies cross placenta, and red cell antibodies with a significant IgG component are detectable by IAT.

-       Other involved antibodies include

o   Anti-C, -e, -E, -Fya, -Jka

o   Anti-M in Japanese, anti-Ge3 in Hispanic

o   But many others have been reported

-       In the absence of a history of HDN, the risk is much lower than for D, c or Kell

-       So frequency of monitoring assessed on basis of history

-       A titre of >32 would suggest increased risk of HDN

 

Action at time of Birth

 

D typing of cord samples from D-negative women with no immune anti-D

-       Maternal and cord blood sample for D grouping if baby group not already known

-       Post-delivery anti-D as required

 

DAT on cord samples

-       Not required routinely for D+ babies born to D- mothers

-       DAT, Hb and bilirubin should be tested on cord blood of babies born to women with IAT-reactive red cell antibodies and the baby observed for jaundice.

-       If DAT+ and baby shows signs of HDN then a red cell eluate may help confirm the red cell antibody specificity.

-       IgG ABO antibodies can cause severe HDN so in cases of baby having major ABO mismatch with mother, the eluate should also be tested with A1 and B cells.

-       Babies who received IUT with D- cells as prevention of HDN may type as D- for several months until their own D+ cells are released into circulation.

 

Perform FMH at delivery on D- women with no immune anti-D to inform anti-D requirements