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目的运用不连续密度梯度离心及免疫磁珠分离法分选孕妇外周血中的胎儿有核红细胞以进行人巨细胞病毒-DNA(HCMV-DNA)的检测。方法取20名人巨细胞病毒抗原血症(pp65)阳性的孕妇外周EDTA抗凝血,经不连续密度梯度离心初步富集、免疫磁珠法分选及胎儿细胞特异性抗体-HbF标记、识别的胎儿有核红细胞经全基因组扩增后,对产物进行HCMV-DNA的检测。结果20名PP65阳性孕妇外周血中分选到胎儿有核红细胞的有16例,将此16例胎儿有核红细胞进行HCMV-DNA的检测,检出阳性2例,10份未妊娠妇女外周血细胞涂片中均未发现胎儿有核红细胞;10份正常妊娠妇女外周血细胞涂片中发现胎儿有核红细胞数9例,其量不等。结论不连续密度梯度离心及免疫磁珠分离法结合特异性抗体-HbF标记、识别能有效地分选出母血中胎儿有核红细胞,应用于进一步的无创性产前诊断,将有着良好的应用前景。
Objective To detect human cytomegalovirus-DNA (HCMV-DNA) by sequencing fetal nucleated red blood cells from peripheral blood of pregnant women using discontinuous density gradient centrifugation and immunomagnetic bead separation. Methods EDTA anticoagulants from 20 pregnant women with positive cytomegalovirus antigens (pp65) were initially collected by discontinuous density gradient centrifugation, sorted by immunomagnetic beads and labeled with fetal cell-specific antibody-HbF Fetal nucleated red blood cells after whole genome amplification, the product of HCMV-DNA detection. Results Totally 16 fetuses with fetal nucleated erythrocytes were isolated from peripheral blood of 20 PP65-positive pregnant women. HCMV-DNA was detected in 16 fetuses with nucleated erythrocytes, and 2 positive cases were detected. Peripheral blood cells were smeared on 10 non-pregnant women Fetal nucleated erythrocytes were not found in all the 10 fetuses. The number of fetal erythrocytes in peripheral blood smears of 10 normal pregnant women was found in 9 cases with varying amounts. Conclusion Discontinuous density gradient centrifugation and immunomagnetic bead separation combined with specific antibody-HbF labeling can effectively identify fetal nucleated erythrocytes in maternal blood and be used in further noninvasive prenatal diagnosis. prospect.