title: Prenatal genetic diagnosis from maternal blood: simultaneous immunophenotyping and FISH of fetal nucleated erythrocytes isolated by negative and positive magnetic activated cell sorting.
abstract: Fetal nucleated red blood cells (nRBCs) are rare in maternal circulation, but their presence constitutes a potential source of non-invasive prenatal genetic diagnosis. This study was undertaken to establish a non-invasive prenatal genetic diagnosis method using isolated fetal nRBCs. A multi-step method including triple density gradient and magnetic activated cell sorting (MACS) using CD45 and CD71, cytospin centrifugation, K-B staining, and glycophorin A-immuno fluorescence in situ hybridization (GPA-immuno FISH) was performed. The study population included 65 patients from 8 to 41 weeks of gestation, and fetal nRBC was separated from all cases. The number of fetal nRBCs retrieved was 12.8 +/- 2.7 in 8 to 11 gestational weeks, 15.2 +/- 6.5 in 12 to 18 gestational weeks, 16.4 +/- 6.5 in 19 to 23 gestational weeks, 10.6 +/- 3.2 in 24 to 28 gestational weeks, and 5.5 +/- 1.9 in 35 to 41 gestational weeks: the mean number of nRBCs collected from 20 ml of maternal peripheral blood was 13.7 +/- 6.2. The highest value of yield was 45.6% from 12 to 18 weeks gestation. The fetal sex determination confirmed by amniocentesis or chorionic villus sampling showed 100% sensitivity and 91.7% specificity for males; 91.7% sensitivity and 100% specificity for females. We showed that fetal cells can be reliably enriched from maternal blood and that they can be used for detecting specific chromosomes by FISH with a specificity superior to current non-invasive methods.
authors.affiliation: Department of Obstetrics and Gynecology, Yonsei University College of Medicine, Seoul, Korea. obgy@yumc.yonsei.ac.kr
authors.forename: Y H
authors.initials: YH
authors.lastname: Yang
authors.forename: K J
authors.initials: KJ
authors.lastname: Jee
authors.forename: S K
authors.initials: SK
authors.lastname: Kim
authors.forename: Y W
authors.initials: YW
authors.lastname: Park
authors.forename: I K
authors.initials: IK
authors.lastname: Kim
authors.forename: D H
authors.initials: DH
authors.lastname: Cha
authors.forename: J E
authors.initials: JE
authors.lastname: Chung
authors.forename: S H
authors.initials: SH
authors.lastname: Kim
mesh_terms: D004912:Erythrocytes / Q000276:immunology*; D005260:Female; D005312:Fetal Blood / Q000276:immunology*; D005865:Gestational Age; D006021:Glycophorins; D006801:Humans; D018189:Immunomagnetic Separation; D016130:Immunophenotyping; D017404:In Situ Hybridization, Fluorescence; D011247:Pregnancy; D011296:Prenatal Diagnosis
chemical_list: D006021:Glycophorins