The non-invasive prenatal test (NIPT) is based on next generation sequencing (NGS) and is used for screening for fetal trisomy. However, it is time-consuming and technically difficult. Recently, peptide nucleic acid (PNA) probe-based real-time polymerase chain reaction (RT-PCR) was developed. This study aimed to examine the performance of the RT-PCR-based NIPT for screening of common fetal trisomies From stored maternal plasma, RT-PCR was performed using Patio™ NIPT Detection Kit. In melting curve analysis, the height of melting peaks of target chromosome and reference chromosome was calculated as a peak ratio. The adjusted peak ratio of 8 markers with correction factors in each target chromosome was summated and calculated to z-score. The cut-off value for each target chromosome was established for classification (low risk vs. high risk for trisomy) whose performance was obtained in the validation phase. 330 plasma samples from pregnant women with normal fetus and 22 trisomy cell-line samples were used to establish the optimal cut-off values for z-score of each target chromosome. In the validation phase, 1023 samples from pregnant women including 22 cases with fetal trisomy and 1001 cases of normal control were used. The RT-PCR-based NIPT showed 95.45% sensitivity [95% confidence interval (CI) 77.16–99.88%], 98.60% specificity (95% CI 97.66–99.23%), and 98.53% accuracy (95% CI 97.59–99.18%) for the identification of trisomy 21, 18, or 13. Of 1023 samples, fifteen cases were mismatched for classification [one case as a false negative (false negative rate: 4.5%) and 14 cases as false positives (false positive rate: 1.4%)]. The RT-PCR-based NIPT showed high sensitivity and specificity for the detection of common fetal trisomies and it could be a feasible alternative to NGS-based NIPT.

中文翻译：

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基于实时 PCR 的常见胎儿三体性筛查新方法


无创产前检测（NIPT）基于下一代测序（NGS），用于筛查胎儿三体性。然而，这既耗时又技术困难。最近，开发了基于肽核酸（PNA）探针的实时聚合酶链反应（RT-PCR）。本研究旨在检查基于 RT-PCR 的 NIPT 在筛查常见胎儿三体性方面的性能 使用 Patio™ NIPT 检测试剂盒对储存的母体血浆进行 RT-PCR。在熔解曲线分析中，计算目标染色体和参考染色体的熔解峰的高度作为峰比。将每条目标染色体中带有校正因子的 8 个标记的调整峰值比率求和并计算为 z 分数。建立每个目标染色体的截止值用于分类（三体性的低风险与高风险），其性能在验证阶段获得。使用来自具有正常胎儿的孕妇的 330 个血浆样本和 22 个三体细胞系样本来建立每个目标染色体 z 得分的最佳截止值。在验证阶段，使用了1023例孕妇样本，其中包括22例胎儿三体性病例和1001例正常对照。基于 RT-PCR 的 NIPT 显示出 95.45% 的敏感性 [95% 置信区间 (CI) 77.16–99.88%]、98.60% 的特异性 (95% CI 97.66–99.23%) 和 98.53% 的准确性 (95% CI 97.59–99.18%) ）用于鉴定21、18或13三体。1023个样本中，有15例分类不匹配[1例为假阴性（假阴性率：4.5%），14例为假阳性（假阳性率：1.4） %)]。 基于 RT-PCR 的 NIPT 对常见胎儿三体性的检测显示出高灵敏度和特异性，它可能是基于 NGS 的 NIPT 的可行替代方案。