Estimation of FetoMateranl haemorrHage (FMH) (bsh 2009)
Who to test
FMH estimation should be performed on:
Rh D- women, following delivery of an Rh D+ baby
Babies should be typed with routine saline reacting, IgM reagents that do not detect DVI. Weak D or Variant D should be treated as D+ for purpose of anti-D administration.
Following all potentially sensitizing events in Rh D- women after 20 weeks gestation
Flow cytometry for minor D+ population may be required:
Following Rh D+ RBC transfusion to a Rh D- woman of childbearing potential to estimate or confirm the dose of anti-D required.
In solid organ transplant when D+ donor into D- recipient
FMH testing is not required for:
Non-severe sensitizing event prior to 20 weeks gestation
When woman is known to have immune anti-D
When fetus/baby known to be D-
When woman is D+
Should be typed with routine saline reacting, IgM reagents that do not detect DVI.
If testing D+ with these reagents then unlikely to make an anti-D that will adversely affect the baby
In D+ women with unexplained abdominal pain in late pregnancy, FMH by acid elution is of limited diagnostic use. Better tests are available for suspected placental abruption.
Mother – EDTA for FMH, separate G&S tube (FMH underestimated after centrifugation of G&S samples)
Baby – cord blood for ABO and D group
Maternal EDTA for FMH estimation following sensitizing events after 20 weeks gestation
Up to 28 weeks, maternal G&S should be performed prior to each anti-D dose
After 28 weeks, anti-D is still required even if RAADP given, but antibody screening is not.
Methods of FMH Estimation
Acid Elution (Kleihauer Test)
Based on HbF vs HbA
Best for initial quantification of FMH
HbF is more resistant to alkali denaturation and acid elution than HbA
Fixed, dry blood film placed in acid buffer, HbA is denatured and eluted leaving behind ghost cells. Cells containing HbF are stainable and stand out in a sea of maternal ghost cells.
Thin, freshly prepared films easier to read (a 1:2 dilution may help)
Modified test – elute only half the slide (allows comparison)
Negative control (normal adult FBC)
Positive control (cord blood added to adult whole blood in a 1:100 dilution
A low power field using a x10 eyepiece and x10 objective will show a minimum of 1600 red cells (figure extrapolated from 100 cells seen with a x10 eye, x40 obj)
25 low power fields should be screened
If 10 or more fetal cells seen then quantification must be performed
Less than 10 cells can be considered <2ml FMH
Counted with aid of a Miller Square, counting minimum of 10,000 cells
Mollinson formula then calculates the ml of FMH
Fluorochrome conjugated with IgG monoclonal anti-D
Best as a reference test after positive acid elution finds >2ml FMH
Mix sample thoroughly
Wash cells to remove leukocytes and platelets
Two samples should be tested in tandem; discrepancy suggests error in sample preparation or flow counting.
Inert control should be used to determine the proportion of background non-specific uptake of fluorochromes, which can be subtracted for the count of anti-D fluorochrome cells
Reporting results to clinicians
Reports need to be:
Timely – within 72 hours
Clear – report result in ‘mL of fetal red cells’ rounded up to nearest 1ml
Whether supplementary anti-D is required if a standard dose has already been administered.
The Anti-D dose required to cover the reported bleed
Advice on follow-up testing to ensure clearance of D+ cells