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

Abstract

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 gamma-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-gamma 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.