Neonatal cytogenetic validation demonstrates high accuracy of single-nucleotide polymorphism-based non-invasive prenatal testing: a 4466-case single-center study

Non-invasive prenatal testing (NIPT) is widely used for fetal aneuploidy screening; however, most evaluations of test accuracy rely on pregnancy outcomes rather than cytogenetic confirmation after birth, and evidence regarding the real-world performance of single-nucleotide polymorphism (SNP)-based NIPT remains limited. We analyzed prospectively collected clinical data from pregnant women at high risk of fetal aneuploidy who underwent SNP-based NIPT between 2013 and 2022 to evaluate its analytical accuracy using neonatal buccal mucosal fluorescence in situ hybridization (FISH). Positive NIPT results were followed by amniocentesis, whereas negative results were systematically validated by postnatal buccal mucosal FISH. Sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were calculated for trisomies 13, 18, and 21. Among 4466 women undergoing initial testing, a total of 4502 tests were performed, including 36 repeat tests after redraw. Of these tests, 4366 (97.0%) were negative, 73 (1.6%) were positive, and four (0.1%) yielded “no call” results after redraw. Sensitivity for trisomies 13, 18, and 21 was 94.6%, 100.0%, and 80.0%, respectively, with a specificity of 99.9% for all three conditions. The overall PPV was 89.1% and the NPV was 99.9%. Two false-negative cases (0.06%) were identified, both attributable to low-level mosaicism. Of the initial “no call” cases, 88.9% yielded reportable results after redraw. These findings demonstrate that SNP-based NIPT shows high analytical performance when validated with neonatal cytogenetic confirmation, while rare false-negative results due to mosaicism highlight the biological limitations of cfDNA-based screening and the importance of cytogenetic confirmation when ultrasound abnormalities are present.