HDFN

1. Importance of Antenatal
Antibody Screening
Dr.Biplabendu Talukdar.
MBBS,MD(IHBT),M.Phil(RMTS),PhD scholar
State Program Officer, State Blood Cell, WB
Consultant Narayana Super Speciality Hospital,Andul,Howrah
Consultant Peoples Blood Bank

2. Introduction
It is a procedure to determine the presence of
irregular antibodies in the serum of a
pregnant female during the antenatal period.

3. Pre-natal antibody
screening!
Is it essential ?

4. Objectives of Antenatal Testing
• To identify antigen negative women
• To identify women with potentially significant
alloantibodies
• To assist in diagnosis and management of HDFN, both
during pregnancy and at delivery.

5. Alloantibodies and pregnancy

6. Alloantibodies and pregnancy
• Hydrops Fetalis

7. What we wants ?

8. Prevalence
• Unexpected antibodies- any Red Cell alloantibodies
other than naturally occurring anti-A or anti-B
• Detected by performing an antibody screen test
• Prevalence: 0.3-38%
• depends on group of patient or Donor studied
• sensitivity of the test methods used
• Reacts only with allogeneic red cells vs. auto-antibodies
that reacts with red cells from other individuals

9. Incidence of maternal allo –antibody
mediated HDFN
Incidence of ABO HDFN 1 to 4 %
(depending of ethnicity of population)
In Medical College Hospital Kolkata 2.06 %
(most of them against RhD)

10. Causes of allo-immunization
• Pregnancy
• Feto maternal haemorrhage (FMH)
3% in 1st trimester
12% in 2nd trimester
45% in 3rd trimester
• Blood component transfusion
• Transplantation

12. How it is occurred ?

13. •Anti-Fya, Fyb
, Jkb
Maternal antibodies in HDFN

14. It should be remembered that antibodies against other
red blood cell antigens, such as S, s, K, Fya, Fyb, Jka,
Jkb can cause severe and even fatal HDN.
• Issit PD, ANSTEE DJ.
• The MN Blood Group System.
• In: Applied Blood Group Serology 4th ed.
• Montgomery Scientific Publications, USA, 1998. p.461-557

15. Blood group system Antibody
frequency
(/1000) 1967
Antibody
frequency
(/1000) 1996
Association with HDF / N
Rhesus (Rh)
Anti-D 43.3 2.7 Mild to severe HDF/N, Hydrops
Anti-C 0.1 0.7 Mild to severe HDF/N, Hydrops
Anti-c 1.2 0.9 Mild to severe HDF/N, Hydrops
Anti-E 1.3 2.0 Mild to severe HDF/N, Hydrops
Anti-e 0.05 0 Mild to severe HDF/N, Hydrops rare
Kell
Anti-K1 (anti-K) 2.1 3.0 Mild to severe HDF/N, Hydrops
Anti-K 2 (anti-k) 0 0.03 Rare cause of mod to severe HDN
Kidd
Anti-Jka 0.2 0.2 Mild to severe HDF/N
Anti-Jkb 0 0 Mild to severe HDF/N (rare)

16. Blood group system Antibody
frequency
(/1000) 1967
Antibody
frequency
(/1000) 1996
Association with HDF / N
Duffy
Anti-Fya 0.4 0.8 Mild to severe HDF/N. Hydrops
Anti-Fyb 0.02 0.03 No HDF, No or mild HDN
MNSs
Anti-M 0.7 0.5 Rare cases of moderate to severe HDF
Anti-N 0.1 0.03 No HDF, No or mild HDN
Anti-S 0.2 0.1 Rare cases of moderate to severe HDF
Others 3.42 3.26

18. International Guidelines for Antenatal testing
 All pregnant women should be ABO and D typed and screened for
the presence of red cell antibodies early in pregnancy and at 28 weeks
gestation.
[National Collaborating Centre for Women’s and Children’s Health, 2003]
&
[AABB’s Guidelines for Prenatal and Perinatal Immunohematology 18th edition page 599 & ACOG]
• Advocate that pregnant women must be tested for unexpected
antibodies to red blood cell (RBC) antigens at the first-trimester visit.
“Management of Alloimmunization during Pregnancy” -

19. Antenatal testing scenario in Developed
countries
• Many developed nations have national screening programs for
pregnant women.
• Universal screening of all antenatal women, including D antigen-
positive pregnant ones, is mandatory in these screening programs of
developed countries.
• Routine testing in these countries involves ABO + D testing along
with Antibody screening irrespective of the Rh status.

20. Why developed countries do it?
Testing done well in advance ensures knowing the blood
group of the mother (in case there is any requirement of
blood during the delivery of the baby).
Screening and identification for any underlying
alloantibodies ensures adequate measures to be taken so
as to prevent Hemolytic Disease of the Newborn

21. Antenatal screening- current practice
Developed countries
• Standardized antenatal screening
protocols
• Screening to start at 12th weeks
• No alloantibody – repeat at 28-32
weeks
• Alloantibody – monthly follow up till
32 weeks, then 2 weekly till term
• All antenatal pregnancies screened
irrespective of Rh status
Developing countries
• No standardized protocols /
national recommendation
• Screening restricted only to Rh (D)
negative pregnancies.
• Most centers screen using pooled
‘O’ cells only.

22. Antenatal testing current scenario in India
• No policies or guidelines available for routine tests which
should be performed in the antenatal period.
• Currently ABO grouping and confirmation of the Rh
status (D+ or D-) is done routinely.
• Antibody Screening not done even in the major metros as
a policy, therefore the presence of irregular clinically
significant antibodies other then Anti-D remain
undetected and may lead to HDN.

23. What would be appropriate in the Indian
context ??
 Although there are no formal healthcare guidelines for this
testing in India, it should be made essential to do a basic
Blood group profile (Forward and reverse grouping) during
the first trimester to determine the blood group and Rh
status of the mother.
 Antibody screening should be carried out in the first
trimester for all pregnant women for detection of clinically
significant alloantibodies

24. Scenario at Medical College
Hospital, Kolkata
ICT positive – 534 antenatal mother ( 2015-2016)
Developing HDFN -11
Most of them against anti Rh D (10)
Another Against anti Rh D with C

25. ALGORITHM OF ROUTINE ANTIBODY SCREENING
First antibody screen
IAT positive IAT negative
Anti-D
Anti-c
Anti-K
Other IAT positive
Test 4 weekly (quantification/titre)
until 28 weeks
Second antibody screen
28 weeks
Anti-D
Anti-c
Anti-K
Other IAT positive
Titrate
IAT negative
Test 2 weekly (quantification/titre)
until delivery
Test cord blood for DAT not routinely required
DAT, Hb and bilirubin
DAT not routinely required

26. Repeat maternal
ab. titer at 8-10
wks interval
Heretozygous
Confirm fetal ag. positivity at 24th wk
Serial MCA doppler every 1-2
wks from 24th wk
< 4 fold titer
Delivery at term
MCA ≥ 1.5 MOM
> 4 fold titer
Homozygous
Paternity unsure
Fetus antigen -ve
Fetus antigen +ve
Yes
Serial assesssment
weekly
At 35 wk, consider
phenobarbital
Cordocentesis
for Hct
IUT if Hct < 30%
No
Immuno-hematological evaluation
Induction of delivery in
1 wks

27. All Content © Immucor, Inc.

28. ABO HDFN
• HDFN caused by ABO incompatibility is clinically mild, with
jaundice most frequently developing within 24 hours of birth.
• Profile
• T&S on mom – Mom group O, screen neg
• DAT on cord cells, weakly positive
• ABO/D type on cord cells is A or B
• Elution testing on cord cells – anti-A or –B &-A,B eluted.
• Bilirubin on infant – 12 mg/dL or more

29. Antenatal testing guidelines
Test Situation Timing
ABO typing Pregnancy Initial visit 10-16 weeks
Rh D typing Pregnancy Initial visit and 26-28 weeks
Antibody screen
All pregnancies Initial visit and 28 weeks
D neg. pregnancy Before RhIg therapy
D pos. pregnancy 3rd trimester if history of Abs or Tx
Antibody ID Positive antibody
screen
Upon detection
Antibody titer
Rh or other clinically
significant Ab
Upon initial detection
Repeat at 18-20 weeks
Rep at 2-4 weeks if below critical titer

30. Why do we need Follow Up serological Tests
• To identify the fetuses that may need treatment before
term.
• Predict which infants might require treatment and should
be monitored more carefully after birth.
• Detect new antibodies, since those who develop one
antibody may also develop additional antibodies.

31. Serologic testing
• Tests to be carried out on the Mother…
• ABO and Rh typing
• Detailed Antigen typing (in case an antibody apart from the Rh system identified)
• Antibody Screen
• If negative, repeat before RhIg therapy and/or if patient is transfused or has history of
antibodies (3rd trimester)
• Antibody Identification
• Tests to be carried out on the Father
• ABO and Rh Typing
• Detailed Antigen typing (in case an antibody apart from the Rh system identified)
• Amniocyte testing to determine blood group of baby
• Can be done as early as 10 to 12 weeks of gestation.

32. Antenatal Serology
• Test for mother ABO and Rh (weak D) and antibody
screening
• Group O mother:
• no problem if the fetus is group O
• Rh negative mother:
• take history for Rh Ig administration.

33. Women with apparent anti-C + D, possible
anti-G
Proportion of antibodies with apparent anti-C+D specificity but with
disproportionately high anti-C titres may be demonstrated, by
advanced serological techniques, to be anti-G.
[Maley et al, 2001].

34. Women with anti-G
• Anti-G is present on red blood cells carrying the D ag or C ag
• Red blood cells do not have to be both D+ and C+ to be G
• Only absent when red blood cells lack both D and C antigens
Important to differentiate between Ab and provide RhIg when the Ab is identified as
Anti-G (without Anti-D) to prevent alloimmunization to the D Ag

35. Rh D typing: weak D/Partial D
• Weak D testing is no longer necessary for obstetric patients
a few women who actually have weak expression of the
D antigen will be classified as Rh negative and will be
candidates for Rh immune globulin
• Anti globulin test should not be used to D type maternal
samples

36. [ BCSH considers > 32]
Antibody Titration

37. BCSH Guidelines
• Screening and identification of red cell alloantibodies
• to detect clinically significant antibodies
• to highlight possible transfusion problems
• Follow-up tests when clinically significant red cell antibodies are
present:
• monitor the strength of antibodies to identify those pregnancies which
are at risk of HDN & to predict fetuses /infants who are likely to require
treatment for HDN.

38. BCSH Guidelines
• Follow up tests
• to identify additional maternal allo-antibodies. Women who have developed
one or more antibodies may go on to form further antibodies of different
specificities
• All women who have previously had an infant affected by HDN
should be referred before 20 weeks to a specialist unit for advice
and for assessment of fetal haemolysis, irrespective of antibody
level.

39. What can be done in the current
scenario????
 Testing needs to be incorporated as a routine in large hospitals and reference
centres.
 Studies need to be done on a large scale to identify the need to carry out antibody
screening.
◦ Whether we should test all pregnant females or only in D neg cases.
 A lot of ambiguities regarding occurrence of HDN would get resolved and better
management of cases would be possible with better antenatal testing practices.
 Guidelines need to be formulated at a National level to bring about an awareness
in this field.
 Government funding needs to be extended in this direction as part of the National
Programme for Women and Child Health

40. Conclusion
• A variety of non-anti-D red cell antibodies can cause a degree
of neonatal haemolysis.
• The frequency of antibody testing should be individualised.
• Management of non-anti-D alloimmunisation should be
aimed at minimising perinatal morbidity.
• Anti-D prophylaxis has reduced the incidence of haemolytic
disease of the new born

41. Thank you