Novel method of real-time PCR-based screening for common fetal trisomies

So Yeon Kim, Seung Mi Lee, Sun Min Kim, Byoung Jae Kim, Ja Nam Koo, Ig Hwan Oh, Sohee Oh, Chan Wook Park, Jong Kwan Jun, Ji Hyae Lim, Hyun Mee Ryu, Joong Shin Park

Research output: Contribution to journalArticlepeer-review


Background: 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 Methods: 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. Results: 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%)]. Conclusion: 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.

Original languageEnglish
Article number195
JournalBMC Medical Genomics
Issue number1
StatePublished - Dec 2021


  • Fetal trisomy
  • Non-invasive prenatal test
  • Peptide nucleic acid
  • Prenatal diagnosis
  • Real-time polymerase chain reaction


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