Non-Invasive Detection of Fetal Rhesus D Status: A Comparison between Polymerase Chain Reaction and Flow Cytometry

• Published in: Fetal diagnosis and therapy Vol. 21; no. 5; pp. 404 - 409
• Main Authors: Di Simone, Nicoletta, Lai, Marco, Rumi, Carlo, Riccardi, Patrizia, D’Asta, Marco, Leone, Giuseppe, Mancuso, Salvatore, Caruso, Alessandro
• Format: Journal Article
• Language: English
• Published: Basel, Switzerland Karger 01.01.2006; S. Karger AG
• Subjects: Antibodies, Monoclonal; Antigens, CD - analysis; Biological and medical sciences; Blood Grouping and Crossmatching - methods; Erythroblasts - chemistry; Female; Fetal Blood - cytology; Flow Cytometry; Gestational Age; Glycophorin - analysis; Gynecology. Andrology. Obstetrics; Humans; Management. Prenatal diagnosis; Medical sciences; Pregnancy; Pregnancy. Fetus. Placenta; Prenatal Diagnosis; Receptors, Transferrin - analysis; Reverse Transcriptase Polymerase Chain Reaction; Rh-Hr Blood-Group System - blood; Rh-Hr Blood-Group System - classification; Rh-Hr Blood-Group System - genetics; Prenatal diagnosis; Rhesus D; Fetal cells; DNA; Pregnancy; Polymerase chain reaction; Flow cytometry; Fetal cell; Prenatal Diagnosis; Fetus; Rh-System; Molecular biology; Comparative study
• ISSN: 1015-3837; 1421-9964
• DOI: 10.1159/000093880

Objective: A non-invasive prenatal determination of the fetal RhD status might be useful for the management of pregnancies in RhD-negative women whose partners are RhD positive. Methods: Maternal peripheral blood of 32 RhD-negative women (17–24 weeks of gestation) was collected, and circulating fetal cells were enriched by CD71 mini-magnetic activated cell sorting. The RhD status of the fetuses was assessed using multiparametric flow cytometry, and results were compared to those of reverse transcriptase (RT)-polymerase chain reaction (PCR), or PCR, which acted as control. Flow-cytometric study of fetal cells employed monoclonal antibodies directed against CD71, glycophorin A (GPA) and RhD antigens. Results: The median percentage of CD71- and RhD-positive cells was 0.83% (range 0.14–6.44%), and that of CD71 and GPA-positive cells was 10.07% (range 0.52–45.84%). Flow-cytometric analysis correlated with RT-PCR results of RNA obtained from whole maternal blood. In 1 case, an incorrect result was due to the failure of the amplification of the specific RhD band on RNA extracted from the CD71-positive fraction. In two instances, we observed false-positive results for RhD in PCR of DNA obtained from maternal plasma. Conclusion: Based on our results, flow-cytometric analysis might be proposed as a clinical tool for the non-invasive prenatal determination of the fetal RhD status independently of fetal gender.