UOG Journal Club: Reassessing critical maternal antibody threshold in RhD alloimmunization: a 16-year retrospective cohort study C. A. Walsh, B. Doyle, J. Quigley, F. M. McAuliffe, J. Fitzgerald, R. Mahony, S. Higgins, S. Carroll and P. McParland Volume 44, Issue 6, pages 669–673, December 2014 http://onlinelibrary.wiley.com/doi/10.1002/uog.13383/abstract

1. UOG Journal Club: December 2014
Reassessing critical maternal antibody threshold in
RhD alloimmunization:
a 16-year retrospective cohort study
C.A. Walsh, B. Doyle, J. Quigley, F.M. McAuliffe, J. Fitzgerald,
R. Mahony, S. Higgins, S. Carroll, and P. McParland
Volume 44, Issue 6, Date: December 2014, pages 669–673
Journal Club slides prepared by Dr Aly Youssef
(UOG Editor for Trainees)

2. Reassessing critical maternal antibody threshold in RhD alloimmunization:
a 16-year retrospective cohort study
Walsh et al., UOG 2014
Introduction
• Monitoring of women with RhD antibodies consists of serial
antibody level measurements, with ultrasound surveillance for
those whose levels exceed a certain critical threshold.
• Whereas serial antibody evaluation can be performed in a routine
antenatal care setting, ultrasound surveillance takes place
usually in specialist centers.
• Therefore, the maternal antibody level that mandates specialist
referral becomes critically important
• An antibody threshold > 15 IU/mL was incorporated into clinical
practice for many years. This cut-off has been recently
questioned, and lower cut-offs have been suggested.

3. Reassessing critical maternal antibody threshold in RhD alloimmunization:
a 16-year retrospective cohort study
Walsh et al., UOG 2014
Objective
To determine the critical maternal antibody
threshold for specialist referral in cases of
RhD alloimmunization

4. Reassessing critical maternal antibody threshold in RhD alloimmunization:
a 16-year retrospective cohort study
Walsh et al., UOG 2014
Methods
• All intrauterine transfusion (IUT) cases performed at the National
Maternity Hospital, Dublin, between January 1996 and
December 2011 were extracted
• The Irish Blood Transfusion Service, which offers a national
referral service for red-cell antibody quantification in pregnancy,
provided maternal serum antibody levels (IU/mL) over a 10-year
period (2002–2011) for RhD alloimmunized women who did not
require IUT.

5. Reassessing critical maternal antibody threshold in RhD alloimmunization:
a 16-year retrospective cohort study
Walsh et al., UOG 2014
RhD alloimmunization screening and diagnosis in the study period
• All patients underwent routine blood group antibody screening at the
first antenatal visit, with a further serology screening at 28 weeks’
gestation for women who were RhD negative.
• Following detection of maternal RhD antibodies, serial antibody levels
were checked every 4 weeks until 28 weeks and fortnightly thereafter.
• An anti-D antibody level of ≥ 4 IU/mL warranted referral to the
institution’s fetal medicine department for subspecialist surveillance
• The current study was restricted to fetal Hb levels and maternal
antibody levels in women undergoing their first IUT.
• The maternal antibody level used in the current analysis was from the
sample taken at the first IUT, or if unavailable, from the most recent
antibody level, provided it was within 4 weeks of the first transfusion.

6. Reassessing critical maternal antibody threshold in RhD alloimmunization:
a 16-year retrospective cohort study
Walsh et al., UOG 2014
Definition of fetal anemia
Degree of anemia Hb MoM at cordocentesis
Severe ≤ 0.54
Moderate 0.55–0.64
Mild 0.65–0.84
No anemia ≥ 0.85
To allow meaningful statistical analysis, maternal anti-D antibody levels
from 208 RhD-alloimmunized women attending the same institution who
did not require IUT were excluded

7. Reassessing critical maternal antibody threshold in RhD alloimmunization:
a 16-year retrospective cohort study
Walsh et al., UOG 2014
Results
• In the study period, 66 women underwent a first transfusion
for RhD alloimmunization
• At the time of first transfusion, the degree of fetal anemia was
classified as moderate to severe, mild, or normal in 74%,
15% and 11% of cases, respectively
• There were no cases of undetected moderate to severe
anemia secondary to RhD alloimmunization in non-transfused
fetuses over the 16-year period; i.e. no case of moderate to
severe anemia alloimmunization was missed.

8. Reassessing critical maternal antibody threshold in RhD alloimmunization:
a 16-year retrospective cohort study
Walsh et al., UOG 2014
Results: Comparison of critical anti-D antibody thresholds for the detection
of fetal anemia
Threshold (study)* Degree of anemia
Sensitivity
(%)
Specificity
(%)
PPV NPV
≥ 4 IU/mL
(Gooch 2007)
Any 100 58 39 100
Moderate to severe 100 55 33 100
≥ 6 IU/mL
(current study)
Any 100 64 43 100
Moderate to severe 100 61 36 100
> 15 IU/mL
(Nicolaides 1992)
Any 81 78 50 94
Moderate to severe 80 75 41 94
The optimum maternal anti-D antibody level that warranted specialist
referral was ≥ 6 IU/mL (6.2 IU/mL)
*Only the first author is given. NPV, negative predictive value; PPV, positive predictive value

9. Reassessing critical maternal antibody threshold in RhD alloimmunization:
a 16-year retrospective cohort study
Walsh et al., UOG 2014
Discussion
• A maternal anti-D antibody threshold of ≥ 4 IU/mL for referral for
specialist evaluation is associated with 100% sensitivity for detection of
moderate to severe anemia, but with a 45% false-positive rate
• A threshold of >15 IU/mL for referral lacks sufficient sensitivity (20% of
cases with moderate to severe fetal anemia would have been missed)
• The present study suggests that the optimal maternal anti-D antibody
threshold for predicting moderate to severe fetal anemia is ≥ 6 IU/mL
• Raising the critical anti-D antibody level from 4 to 6 IU/mL would have
potential benefits for cost and workload in busy regional fetal medicine
centers (10% reduction of referrals), without compromising fetal
outcomes.

10. Reassessing critical maternal antibody threshold in RhD alloimmunization:
a 16-year retrospective cohort study
Walsh et al., UOG 2014
Strengths
• Large single-institution study at a national referral center for
alloimmunization
• Consistent antenatal screening and management over the 16-
year study period
Limitations
• Retrospective design
• Due to large numbers involved, it was not possible to extract and
analyze every neonatal Hb level in RhD alloimmunized women
• As there is no reliable conversion from mIU/mL to Ab titers, used
in many countries, this may limit the applicability of the present
findings in some obstetric populations.

11. Reassessing critical maternal antibody threshold in RhD alloimmunization:
a 16-year retrospective cohort study
Walsh et al., UOG 2014
Future perspectives
• Further studies should be undertaken to determine whether
women with anti-D antibodies within the 4.0–15.0 IU/mL
range indeed require specialist referral or would benefit
from a modified program of ultrasound surveillance.

12. Reassessing critical maternal antibody threshold in RhD alloimmunization:
a 16-year retrospective cohort study
Walsh et al., UOG 2014
Discussion points
• What is the best cut-off of maternal anti-D antibody levels to
be used for referral to specialist surveillance?
• Could the interval between antibody testing and fetal blood
sampling influence the referral cut-off used?
• Which cut-off should be used for other antibodies (e.g. anti-
Kell)?
• Should universal testing for fetal Rh genotype using cell-free
DNA techniques be implemented?