Reading J P, Huffman J L, Wu J C, Palmer F T, Harton G L, Sisson M E, Keyvanfar K, Gresinger T H, Cochrane W J, Fallon L A
Genetics and IVF Institute, Fairfax, VA 22031, USA.
Hum Reprod. 1995 Sep;10(9):2510-5. doi: 10.1093/oxfordjournals.humrep.a136332.
The discovery of nucleated erythrocytes in maternal circulation provides a potential source for non-invasive prenatal diagnosis. We have evaluated the use of a three-stage procedure to determine the number of cells that are of fetal rather than maternal origin. First, monoclonal antibodies specific for CD45 and CD14 were used in conjunction with a magnetic (MACS) column to deplete unwanted leukocytes from maternal blood. This was followed by a positive MACS enrichment for nucleated erythrocytes, using an anti-CD71 (transferrin receptor) monoclonal antibody. To discriminate between fetal nucleated erythrocytes and those of maternal origin, enriched fractions were simultaneously stained with an anti-fetal haemoglobin (HbF) antibody and hybridized with probes specific for X and Y chromosomes. Samples were then subjected to blind analysis along with negative control samples from non-pregnant volunteers. Using this dual analysis, we were able to determine that less than one nucleated erythrocyte per ml of maternal blood was of fetal origin. Small numbers of these fetal cells were found in 87.5% of pregnancies, ranging from 6 to 35 weeks gestational age. Comparison of HbF and X/Y probe data also suggests that the fetal cells are less suitable for fluorescence in-situ hybridization (FISH) analysis than similar preparations from other sources.
母血循环中核红细胞的发现为无创产前诊断提供了一个潜在来源。我们评估了一种三阶段程序的应用，以确定胎儿来源而非母亲来源的细胞数量。首先，将针对CD45和CD14的单克隆抗体与磁性（MACS）柱结合使用，以去除母血中不需要的白细胞。接下来，使用抗CD71（转铁蛋白受体）单克隆抗体对核红细胞进行阳性MACS富集。为了区分胎儿核红细胞和母亲来源的核红细胞，对富集部分同时用抗胎儿血红蛋白（HbF）抗体染色，并与X和Y染色体特异性探针杂交。然后将样本与非妊娠志愿者的阴性对照样本一起进行盲法分析。通过这种双重分析，我们能够确定每毫升母血中胎儿来源的核红细胞少于一个。在87.5%的妊娠中发现了少量此类胎儿细胞，妊娠孕周为6至35周。HbF和X/Y探针数据的比较还表明，与其他来源的类似样本相比，胎儿细胞不太适合荧光原位杂交（FISH）分析。
