产前诊断样本中母体细胞污染对染色体微阵列技术检测的影响

Interfering effect of maternal cell contamination on invasive prenatal chromosome microarray analysis

目的评估产前诊断样本中母体细胞污染(MCC)对染色体微阵列技术(CMA)检测拷贝数变异(CNV)的影响。方法前瞻性研究。挑选2016年12月至2018年8月于杭州市妇产科医院产前诊断中心行羊水产前诊断且经CMA检测出拷贝数变异的DNA标本5份,加入正常女性基因组DNA以模拟不同比例母体细胞污染。采用Agilent微阵列染色体芯片180K CGH(Agilent 180K CGH)对模拟标本进行检测,检测结果通过Agilent CytoGenomics软件进行分析。结果当MCC>38.4%(95%可信区间:36.6%~40.2%)时重复CNV无法检出,MCC>41.3%(95%可信区间:39.9%~42.7%)时缺失CNV无法检出,且随MCC比例增高,CNV检出率逐步降低;大片段的CNV较小片段CNV对MCC耐受程度高;相同大小CNV,缺失型CNV的检出能力较重复型略微升高。在男性标本中,当MCC>10%时,可通过微阵列检测到X/Y染色体发生位移。结论MCC高于一定比例时可以对CMA检测CNV产生影响。基于Agilent 180K CGH对MCC模拟物的检测结果及CNV检测特异性原则,鉴于保守原则,本中心Agilent 180K CGH的MCC阈值设定为30%。

Objective To access the effect of maternal cell contamination(MCC)on the detection of copy number variation(CNV)by chromosome microarray analysis(CMA)in prenatal diagnostic samples.Methods Amniotic fluid DNA samples were collected from Department of Prenatal Diagnosis(Screening)Centre of Hangzhou Women′s Hospital from December 2016 to August 2018.Copy number variations(CNVs)were identified in these DNA samples by CMA and normal female genomic DNA was added to simulate different proportions of maternal cells contamination.The simulated samples were tested using an Agilent microarray chromosome chip 180K CGH(Agilent 180K CGH)and the results were analyzed by Agilent CytoGenomics software.Results The results showed that duplications of CNV could not be detected at>38.4%MCC.Deletions of CNV could not be detected at>41.3%of MCC.With the increase of MCC ratio,the detection rate of CNV decreased gradually.The array and software allowed detection of longer copy-number variations at higher levels of maternal cell contamination than shorter copy-number variations.For the same size CNV,the detection ability of the deleted CNV was slightly higher than that of duplication.In the male sample,the observable shift of the X/Y chromosome ratio at>10%MCC can be detected by the microarray.Conclusions When the MCC is higher than a particular ratio,it can affect the CMA detection of CNV.Based on the Agilent 180K CGH detection results for MCC mimics and the CNV detection specificity principle,the MCC threshold of the Agilent 180K CGH is set to 30%in view of conservative principles.