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The prevailing concept is that gestational alloimmune liver disease (GALD) is caused by maternal antibodies targeting a currently unknown antigen on the liver of the fetus. This leads to deposition of complement on the fetal hepatocytes and death of the fetal hepatocytes and extensive liver injury. In many cases, the newborn dies. In subsequent pregnancies early treatment of the woman with intravenous immunoglobulin can be instituted, and the prognosis for the fetus will be excellent. Without treatment the prognosis can be severe. Crucial improvements of diagnosis require identification of the target antigen. For this identification, this work was based on two hypotheses: 1. The GALD antigen is exclusively expressed in the fetal liver during normal fetal life in all pregnancies; 2. The GALD antigen is an alloantigen expressed in the fetal liver with the woman being homozygous for the minor allele and the father being, most frequently, homozygous for the major allele. We used three different experimental approaches to identify the liver target antigen of maternal antibodies from women who had given birth to a baby with the clinical GALD diagnosis: 1. Immunoprecipitation of antigens from either a human liver cell line or human fetal livers by immunoprecipitation with maternal antibodies followed by mass spectrometry analysis of captured antigens; 2. Construction of a cDNA expression library from human fetal liver mRNA and screening about 1.3 million recombinants in Escherichia coli using antibodies from mothers of babies diagnosed with GALD; 3. Exome/genome sequencing of DNA from 26 presumably unrelated women who had previously given birth to a child with GALD with husband controls and supplementary HLA typing. In conclusion, using the three experimental approaches we did not identify the GALD target antigen and the exome/genome sequencing results did not support the hypothesis that the GALD antigen is an alloantigen, but the results do not yield basis for excluding that the antigen is exclusively expressed during fetal life., which is the hypothesis we favor., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Rieneck et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The KEL1 antigen can give rise to immunization of KEL2 mothers. Maternal antibodies can be transferred to the fetus and destroy fetal red blood cells and their stem cell precursors and give rise to serious fetal disease. It is important to be able to predict the fetal KEL status in order to intervene in those pregnancies where the fetus is at risk, and to ascertain when the fetus is not at risk. Technically it can be demanding to predict KEL1 status from a maternal blood sample. The KEL1 allele is based on a single SNP present in about 1-10 % of cell-free maternal DNA after gestation week 10. Here we describe our protocol for antenatal prediction of fetal KEL1 status by NGS analysis of maternal DNA on a MiSeq instrument.

Objective: To evaluate two different multiplex real-time PCR assays detecting fetal RHD for screening of RhD negative women in relation to antenatal RhD prophylaxis., Methods: We designed a duplex assay for the detection of RHD exon 7 and 10 and a triplex assay for the detection of RHD exon 7, 10 and 5. We used the same fluorescent dye for the exon 7 and 10 probes to increase sensitivity; exon 5 was VIC labeled. We evaluated possible inhibition of DNA amplification with dilution experiments. We then tested the two multiplex assays with DNA extracted from 97 plasma samples from 38 RhD negative women in gestational weeks 6-37., Results: Dilution experiments revealed no inhibition of amplification in the multiplex assays. For plasma samples, the duplex assay was significantly more sensitive than the triplex assay (p < 0.0001). For the duplex assay (exon 7/10), accuracy was 99.0%. For the triplex assay (exon 7/10), accuracy was 94.2%. Detection of exon 5 was less reliable., Conclusion: The duplex assay using exon 7/10 was the most reliable for prenatal prediction of fetal RhD type as a candidate assay for screening of RhD negative women in relation to antenatal RhD prophylaxis. The triplex assay needs further optimization., (Copyright © 2010 S. Karger AG, Basel.)

We present a reliable test for prenatal prediction of fetal RhD type using maternal plasma from RhD-women. This test is needed for a future antenatal RH prophylaxis. A new real time PCR based assay targeting RHD exon 7 and a published assay for RHD exon 10 were used to determine the fetal RHD status in DNA extracted from plasma from 56 pregnant women in 15th-36th week of gestation. Prediction of fetal RhD type was compared with the phenotype determined after birth, and showed 100% concordance. This setup will be of value in antenatal RH prophylaxis and in the management of immunised women.

Objectives: The objective of this study was to establish a reliable test for prenatal prediction of fetal RhD type using maternal plasma from RhD negative women. This test is needed for future prenatal Rh prophylaxis., Methods: A novel real-time PCR-based assay targeting RHD exon 7 combined with a published assay for RHD exon 10 were used to determine the fetal RHD status in DNA extracted from plasma, sampled from 56 pregnant RhD negative women in 15th-36th week of gestation. Thirty-eight samples were from ongoing pregnancies of Danish women and 21 samples from 18 pregnant women were stored anonymized samples from the International Blood Group Reference Laboratory, Bristol, United Kingdom. Prediction of fetal RhD type was compared with the serological result obtained after birth., Results: The prediction of the fetal RhD type was in 100% concordance with the serological RhD type from the 16th week of gestation. One sample from the 15th week of gestation was inconclusive. The number of copies of fetal RHD DNA was found to increase with gestational age. Low levels of DNA were found to follow the Poisson distribution (p = 1.0000)., Conclusion: Our set-up was very reliable for determination of fetal RhD genotype, and thus will be of value in prenatal Rh prophylaxis and in the management of immunized women., (Copyright 2005 John Wiley & Sons, Ltd.)