Topic 17: Rhesus incompatibility / isoimmunization /

Alloimmunization
***is a type of hemolytic disease of the newborn (HDN). Was first described in Rh negative women with a Rh-
positive fetus, but it can occur with many other blood type incompatibilities.
***Fetal Rh-antigen are present by 38th day after conception.
Etiology for Hemolytic disease:
1) ABO incompatibility: present in ∼ 20% of all pregnancies however only ∼ 5–10% are clinically affected.
2) Rhesus incompatibility: rare following implementation of routine anti-D prophylaxis
Normally, the fetal red cells containing the Rh-antigen enter the maternal circulation during first trimester in 5%
cases, during third trimester in 46% cases. Maximum occurs at the time of delivery. There are a few conditions
which predispose to fetomaternal hemorrhage.
•Trauma• Cordocentesis • Antepartum hemorrhage• Amniocentesis• Vaginal delivery• Chorionic villous
samplingQ, • Cesarean section• Attempted versionQ • Forceps delivery• Manual removal of placenta • Placental
abruption• Blood transfusion
3) Risk factors:
 Maternal exposure to fetal blood during pregnancy
 Antenatal procedures (e.g., amniocentesis, cesarean section, termination of pregnancy)
 Pregnancy-related complications (e.g., ectopic pregnancy, placental abruption)
Pathogenesis:
1) In a Rh-negative mother and Rh-positive child: maternal exposure to fetal blood → production of maternal IgM
antibodies against the Rh antigen → over time, seroconversion to Rh-IgG (able to cross the placenta)
2) In next pregnancy with an Rh-positive child: rapid production of maternal IgG anti-D antibodies to
fetal RhD antigens → Rh-IgG agglutination of fetal RBCs with hemolytic anemia → risk of “Hemolytic diseases”
with possible hydrops fetalis ( fetal condition characterized by generalized edema and accumulation of fluid in
serous cavities) that characterized by destruction of the fetal erythrocytes by maternal antibodies due to blood
group incompatibility.
3) Fetal Affection by the Rh-Antibody: The antibody formed in the maternal system (IgG) crosses the placental
barrier and enters into the fetal circulation:
 Rh-negative fetus: antibody will not have any effect on.
 Rh-positive fetus: the antibody becomes attached to the antigen sites on the surface of the fetal
erythrocytes. The affected cells are rapidly removed from the circulation by the reticuloendothelial system.
Depending upon the degree of agglutination and destruction of the fetal red cells, various types of fetal
hemolytic diseases appear.
 Fetal Problems in Rh-Negative Pregnancy
As a result fetal blood carrying Rh-antigen enters maternal circulation and stimulates production
of antibodiesImmunization is unlikely to occur unless at least 0.1 mL of fetal blood enters the maternal
circulationDetectable antibodies usually develop after 6 months following larger volume of feto-maternal bleed.
***Antibodies once formed remain throughout life.
TYPES OF ANTIBODIE: Two types of antibodies are formed :
1) IgM: is the first to appear in the maternal circulation and agglutinates red cells containing D when
suspended in saline. IgM being larger molecules cannot pass through the placental barrier and is not harmful to
the fetus.
2) IgG: It is also called incomplete or blocking antibody. Because of its small molecule, it can cross the placental
barrier and cause damage to the fetus. It appears at a later period than does the IgM antibody. This is the reason
why first pregnancy is not affected in Rh-negative females.

Clinical features
1) Prenatal (before birth)
 Hydrops fetalis (only expected in cases of Rh incompatibility): defined as abnormal accumulation of fluid in
two or more fetal compartments, including ascites, pleural effusion, pericardial effusion, and skin edema
2) Postnatal:
 Neonatal anemia
 Hepatosplenomegaly
 Neonatal jaundice
 Hypoxia
 Prematurity
Diagnosis:
1) Maternal blood
The Kleihauer–Betke test or flow cytometry: on a postnatal maternal blood sample can confirm that fetal blood
has passed into the maternal circulation and can also be used to estimate the amount of fetal blood that has
passed into the maternal circulation.
The indirect Coombs test: is used to screen blood from antenatal women for IgG antibodies that may pass
through the placenta and cause hemolytic disease of the newborn.
Cell-free DNA: can be run on certain antigens. Blood is taken from the mother, and using PCR, can detect the K,
C, c, D, and E alleles of fetal DNA. This blood test is non-invasive to the fetus and is an easy way of checking antigen
status and risk of HDN.
 Automated measurement of antibody (specific anti-D) is a more accurate test. The safe level of antibody in the
maternal serum is < 4 IU/ml.
 Those females with levels of < 4 IU/mL should have antibody measurement monthly till 28 weeks and then
every 2 weeks.
 Those with levels > 10 IU/ml: should have amniotic fluid optical density measurements at 450 nm wave
length and also have weekly ultrasound assessment to detect fetal hydrops.
Amniotic fluid evaluation: When fetal blood cells undergo hemolysis, breakdown pigments, mostly bilirubin, are
present in the supernatant of amniotic fluid. The amount of amniotic fluid bilirubin correlates roughly with the
degree of hemolysis and thus indirectly predicts the severity of the fetal anemia. Because the amniotic fluid
bilirubin level is low compared with serum levels, the concentration is measured by a continuously recording
spectrophotometer and is demonstrable as a change in absorbance at 450 nm, referred to as DOD450, and the
value is plotted on Liley’s graph:
 Zone 1 (i.e. fetus is mildly affected): Repeat amniocentesis at 4 weeks and delivery at term 38 weeks
 Zone 2 (i.e. fetus is moderately affected) In the lower zone 2-anticipated Hb level is 11-13.9 mg/dl, in the
upper zone 2 it is 8-10.9g/dl: Repeat amniocentesis at 1–2 weeks
 Zone 3 (i.e. fetus is severely affected) anticipated Hb = < 8 g/dl: Intrauterine transfusion if preterm or delivery
at 34 weeks if fetus is salvageable
2) Paternal Blood: Blood is generally drawn from the father to help determine fetal antigen status. If he is
homozygous for the antigen, there is a 100% chance of all offspring in the pairing to be positive for the antigen and
at risk for HDN. If he is heterozygous, there is a 50%
3) Prenatal diagnosis:
Imaging
 Ultrasound: to determine hydrops fetalis
 Doppler sonography of fetal blood vessels → increased flow rate indicates fetal anemi
4) Fetal Rh-status: Do amniocentesis or chorionic villi (CVS) and obtain uncultured amniocytes or trophoblasts →
PCR for fetal DNA testing to detect fetal blood group. Fetal DNA, present in maternal plasma can also be
genotyped. This method has replaced amniocentesis which is an invasive method.
Prevention:
1) Anti-D immunoglobulin (RhoGAM): is given to all pregnant Rh-negative mothers, Anti-D prophylaxis protects
children in future pregnancies
How it works??? if anti D is given to a Rh-negative mother and if due to any reason fetal blood with Rh
antigen enters mothers circulation, this Anti D will lead to an antigen antibody reaction and fetal RBC will be
destroyed before it could stimulate mothers immune system to produce Rh antibodies.
 Thus the usefulness of administering Anti D is only before maternal Rh-antibodies are formed.
-In other words, Anti D should only be given if Indirect Coombs test is negative.
 If maternal antibodies are already formed then there is no point in giving Anti D.
- If Indirect Coombs test is positive then donot give Anti D.
2) Screening: ABO and Rh(D) typing of the mother
Rh-positive mothers do not need further screening
Rh-negative mothers → screening for anti-D antibodies
 No anti-D antibodies (unsensitized mothers) → antibody screening repeated at 28 weeks of gestation and
at delivery → see indications for anti-D prophylaxis
 Anti-D antibodies present (sensitized mothers) → further monitoring for evidence
of hemolysis required: amniocentesisand imaging
Management of Rh-Negative Pregnancies
Rh-negative women presenting for obstetrical care can be categorized in two different groups:
1) Rh-negative nonimmunized women.
2) Rh-negative immunized women.
Rh-negative immunized women:
*** Management in this category requires the determination of whether it is a first affected pregnancy or woman
already had a previously affected pregnancy.
First Affected Pregnancy
If this is a female first affected pregnancy that means first time ever in her obstetric history she is getting indirect
combat test (ICT) positivein, antibody titre should be done.
 Antibody titre of 1:16 is called as critical titre which means fetus is definitely affected.

Levels of the titer:The critical titer is taken as 10 IU/L in India and in Europe and UK as 15 IU/L.
Serum antibody titers are done in these women every 4 weeks till 28 weeks and then every 2 weeks until the
titers are found to be at or above the critical level (1:16).If titres are above critical level, there is no further use of
antibody titer and the pregnancy is further monitored by middle cerebral artery peak systolic velocity (MCA-PSV) or
amniotic fluid bilirubin concentration.
If antibody titers remains below critical level up to 36 weeks, the patient should be delivered by elective
induction
between 38-40 weeks and the birth of unaffected or mildly affected fetus should be anticipated.
If there is sudden rise of antibody titers above the critical level after 34 weeks but before 37 weeks of gestation,
amniocentesis is done to assess the fetal lung maturity. Pregnancy should be terminated if the lungs are mature,
but if the lungs are immature and the bilirubin level is low (less than 0.5 mg/dl), the pregnancy should be allowed
to continue as long as weekly amniocentesis shows fetal pulmonary immaturity and a low bilirubin concentration.
Delivery is contemplated as soon as these fetuses achieve lung maturity.
Women with Previous Affected Pregnancy
After the first affected pregnancy, the ability to predict fetal anemia from the maternal anti-D antibody titers is lost
and now these pregnancies should be monitored by MCA-PSV and amniotic fluid bilirubin concentration.
Indication and Recommended Dose of Anti-D:
It is a IgG antibody that is given by i.m. route.
It binds to fetal RBCs and prevents stimulation of maternal immune system.
300 mg will protect the mother from fetal hemorrhage of up to 15 ml of D-positive red cells or 30 ml of fetal
whole blood.
The dose of anti D is calculated by assessing the fetomaternal haemorrhage by performing kleihauer betke test
 It should be given at 28 weeks if indirect coombs test is negative to all unsensitized Rh-negative mother and
postpartum within 72 hours if the baby’s blood group is Rh-positive and direct Coombs test is negative.
 It is seen without administration of Anti D-immunoglobulin, an Rh-negative woman has 7.2% risk of developing
rhesus antibodies within 6 months of giving birth.
It should also be given after abortion, MTP, and ectopic pregnancy and all those events where feto maternal
mixing of blood is possible.