Comparison of fetal cell recovery from maternal blood using a high density gradient for the initial separation step: 1.090 versus 1.119 g/ml

• Published in: Prenatal diagnosis 2000-04, Vol.20 (4), p.281-286
• Main Authors: Samura, Osamu, Sekizawa, Akihiko, Zhen, Dong Kai, Falco, Vincent M., Bianchi, Diana W.
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Blood Cells - cytology; Cell Separation - methods; Centrifugation, Density Gradient; Erythrocytes - cytology; Female; fetal cells in maternal blood; Flow Cytometry; fluorescence-activated cell sorting (FACS); Gestational Age; Gynecology. Andrology. Obstetrics; high density gradient method; Humans; Immunomagnetic Separation; In Situ Hybridization, Fluorescence; magnetic activated cell sorting (MACS); Male; Management. Prenatal diagnosis; Medical sciences; non-invasive prenatal diagnosis; Polymerase Chain Reaction; Pregnancy; Pregnancy. Fetus. Placenta
• ISSN: 0197-3851; 1097-0223
• DOI: 10.1002/(SICI)1097-0223(200004)20:4<281::AID-PD812>3.0.CO;2-P

The purpose of this study was to improve recovery of fetal nucleated erythrocytes (NRBCs) from maternal blood for non‐invasive prenatal diagnosis. Peripheral blood samples were obtained from 27 women who had just undergone pregnancy termination at 6 to 23 weeks. Samples were split and mononuclear cells were isolated using Histopaque gradient at densities of 1.090 g/ml and 1.119 g/ml. CD45 depletion using magnetic activated cell‐sorting, followed by flow‐sorting with antibody to γ‐globin and fluorescence in situ hybridization (FISH) analysis, were used to evaluate the number of fetal NRBCs recovered. In samples separated with the 1.119 g/ml density gradient, the yield of true anti‐γ haemoglobin positive cells (median, 14.9; range, 0–717.5) was significantly higher than that with the 1.090 g/ml density gradient (median, 4.9; range, 0–532.5). After FISH analysis, in the 14 samples in which the fetal karyotype differed from the mother, the median number of fetal NRBCs separated by the 1.119 g/ml density gradient was 22.9 (2–717.5), which was significantly higher than that by the 1.090 g/ml gradient (median, 11.5; range, 0–532.5, p=0.022). Increased density of the gradient used for the initial enrichment of fetal cells results in improved fetal cell recovery in fresh post‐termination blood samples, which may permit better non‐invasive detection of fetal cells in maternal blood. Copyright © 2000 John Wiley & Sons, Ltd.