What is the role of amniocentesis in the treatment of hemolytic disease of the newborn (HDN)?

Updated: Dec 28, 2017
  • Author: Sameer Wagle, MBBS, MD; Chief Editor: Muhammad Aslam, MD  more...
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Answer

When indicated, amniocentesis can be performed as early as 15 weeks' gestation (rarely needed in first affected pregnancy before 24 weeks' gestation) to determine fetal genotype and to assess the severity. Maternal and paternal blood samples should be sent to the reference laboratory with amniotic fluid sample to eliminate false-positive results (from maternal pseudogene or Ccde gene) and false-negative results (from a rearrangement at the RHD gene locus in the father).

Fetal Rh-genotype determination in maternal plasma has become routine in many European countries and is being offered in the United States. [57] Fetal cell-free DNA accounts for 3% of total circulating maternal plasma DNA, is found as early as 38 days of gestation, and is derived from apoptosis of the placental cytotrophoblast layer. The mean half-life of circulating fetal DNA is on average less than 30 minutes, and maternal plasma is subjected to filtration and microcentrifugation to remove all cellular elements before testing. This eliminates false-positive results from engrafted fetal cells of previous pregnancies in maternal lymphoid organs.

Cell-free fetal DNA is subjected to real-time polymerase chain reaction (PCR) for the presence of RHD gene–specific sequences and has been found to be accurate in 99.5% of cases. The SRY gene (in the male fetus) and DNA polymorphisms in the general population (in the female fetus) are used as internal controls to confirm the fetal origin of the cell-free DNA. [20] A panel of 92 single-nucleotide polymorphisms (SNPs) is compared between maternal sample from buffy coat and plasma. A difference of more than 6 SNPs confirms presence of fetal DNA and the validity of the test in a female fetus. [57]  False-negative results being most undesirable and consequential are due to partial or weak D phenotypes. They are detected by using at least two RHD-specific exon primers and run in duplicates. [58]

Fortunately, cell-free fetal DNA testing for determining the genotype for other red blood cell antigens such as c,C, e, E and Kell is also now found to be highly reliable and accurate. [58]

Serial amniocentesis is begun at 10-14 day intervals to monitor the severity of the disease in the fetus. All attempts should be made to avoid transplacental passage of needle which can lead to fetomaternal hemorrhage (FMH) and a further rise in antibody titer. Serial delta-OD 450 values are plotted on the Queenan chart or the extended Liley chart to evaluate the risk of fetal hydrops. Early ultrasonography is performed to establish correct gestational age. Frequent ultrasonographic monitoring is also performed to assess fetal well-being and to detect moderate anemia and early signs of hydrops.


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