Anti-D Haemolytic Disease of the Fetus and Newborn (HDFN)
D- females who are exposed to fetal D+ red cells during pregnancy may develop anti-D
Circulating anti-D (which is usually IgG) can cross the placenta during subsequent pregnancies, leading to haemolytic disease of the newborn in D+ fetuses
Anti-D Prophylaxis
Who to give it to?
D- women carrying a D+ fetus
Women with indeterminate D typing should be treated as D negative till confirmatory typing results are available.
Women with anti-D detected should have further investigations to determine if it is passive or immune.
If it is unclear if the anti-D is passive or immune, it should be treated as passive unless proven otherwise, to prevent alloimmunisation.
To determine fetal blood group:
Phenotype father: if father is D-, then fetus is D- and no further action is necessary (beware issues of non-paternity)
If father is D+, then father will need to be genotyped to determine if he is heterozygous or homozygous for RHD.
If he homozygous, then fetus will be D+ and prophylaxis is required.
If he his heterozygous, then fetus has a 50% chance of being D+ and fetal genotyping should be considered. If fetal genotyping is not available, then treat as if the fetus is D+.
Fetal genotype can be determined by cell-free DNA sequencing of the RHD gene after 16 weeks
May proceed directly to fetal genotyping to avoid issues of non-paternity.
Sensitising events
Before 12 weeks, anti-D prophylaxis is not indicated unless there is:
Ectopic pregnancy
Uterine bleeding which is repeated, heavy or associated with abdominal trauma
In utero therapeutic interventions (IUT, surgery, shunts, lasers etc)
Miscarriage or threatened abortion after 12 weeks’ gestation
Recurrent uterine bleeding (requires prophylaxis every 6 weeks)
Any new symptoms to suggest different pathology (e.g. new abdominal pain) should be managed as a separate sensitising event with a separate dose of anti-D.
Termination of pregnancy
Delivery (by any modality)
For interuterine death, a dose should be given immediately upon diagnosis of IUD and a second dose given at delivery, as there may be a significant delay between diagnosis of IUD and delivery.
Intra-operative cell salvage
Timing of anti-D prophylaxis:
ASAP and within 72h following sensitising events
If administration is not possible within 72hrs, some protection may be afforded with immunisation is given up to 10 days from the sensitising event.
Every 6 weeks at least for women with continual uterine bleeding after 12 weeks gestation
250 IU every 6 weeks between 12 and 20 weeks
500 IU every 6 weeks after 20 weeks
Dose of anti-D
Before 12 weeks, if there is ectopic / molar pregnancy, termination of pregnancy or uterine bleeding with abdominal trauma: 250IU
12-20 weeks: 250 IU (this vial size is no longer available)
After 20 weeks: 500 IU with FMH testing
Intra-operative cell salvage: 1500 IU with FMH testing at 30-45mins after reinfusion.
Positive Kleihauer: 125IU/ml of FMH if IM anti-D or 100IU/ml of FMH if IV anti-D (minimum dose 500 IU)
Routine anti-D prophylaxis: single dose (1500IU) at 28 weeks or two doses (500IU) at 28 and 34 weeks
Routine anti-D prophylaxis should be given regardless of whether there has been previous anti-D administration for prior sensitising events.
When to do FMH testing:
FMH testing is not required:
For any sensitising event occurring before 20 weeks gestation
FMH testing is required:
For any sensitising event occurring after 20 weeks gestation
For patients with recurrent uterine bleeding after 20 weeks gestation, if there is further intermittent uterine bleeding, then estimation of FMH should be carried out every 2 weeks.
ASAP from a sensitising event and within 2 hours of delivery
For any FMH > 4ml (the volume that is covered by a standard 500IU dose of anti-D)
48 hours after IV anti-D
72 hours after IM or SC anti-D
NICE guidelines recommend that pregnant D- women should be given routine anti-D prophylaxis
This is either a single dose at 28 weeks, or
A two-dose regimen at 28 and 34 weeks
These doses should be given regardless of whether anti-D was given for a pregnancy-sensitising event
Testing for fetal-maternal haemorrhage should be done for any pregnancy-sensitising event that occurs after 20 weeks gestation. In general, if the event occurs before 20 weeks, the fetal blood volume is considered insufficient to exceed that covered by the minimum anti-D Ig dose of 25IU (25μg)
The Kleihauer test relies on the different properties of HbA and HbF
HbF is more resistant to acid elution and alkaline denaturation
When a blood film from the maternal sample is fixed and immersed in an acid buffer solution, HbA is denatured and eluted
This leaves red cell ghosts. Fetal cells containing HbF stand out after staining
The percentage of fetal cells gives an estimation of the fetal blood volume
This can be used to guide further anti-D dosing (1ml FMH = 125IU anti-D via IM route, minimum dose 500 IU in the UK)
Standard 300mcg dose of Rhogam in Singapore contains 1500IU of anti-D immune globulin (1mcg = 5IU)
If further doses are given, the Kleihauer should be repeated after 72hrs to confirm clearance of fetal cells from the maternal circulation
FMH quantification:
Acid elution is best used as a screening technique.
At least 100 adult cells should be seen on a high powered field.
A minimum o 25 low power fields should be examined, counting fetal cells in all low power fields.
If >10 fetal cells seen, quantification must be performed.
If <10 fetal cells, then the volume of FMH can safely be assumed to be <2ml and quantification is not required.
Quantification
Recommended that 10,000 maternal cells are examined with a x40 objective.
Count using a Miller square (see below).
Volume of FMH = [(Number of fetal cells/hpf)/(number of maternal cells/hpf)] x2400.
All FMH >2ml should be referred for quantification of D-positive fetal cells by flow cytometry.
Other Indications for Anti-D
Transfusion of D+ platelets to D- women of childbearing age
Prophylaxis against Rh alloimmunisation from RBC contaminating platelet product
Dose of 250IU (25μg) usually sufficient to cover up to five adult therapeutic doses of D+ platelets over a 6 week period
One adult therapeutic dose = 4 random units from whole blood donations, or one single-donor apheresis unit
Inadvertent transfusion of D+ blood to D- women of childbearing age
Consider IV anti-D if a whole unit transfused
If > one unit, can consider red cell exchange transfusion to reduce load of D+ RBCs, as well as dose of anti-D Ig required to suppress immunisation
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