Improving the accuracy of noninvasive prenatal testing through size‐selection between fetal and maternal cfDNA

OBJECTIVES: In general, fetal cfDNA is shorter than maternal cfDNA, and accuracy of noninvasive prenatal testing (NIPT) results can be improved by selecting shorter cfDNA fragments to enrich fetal-derived cfDNA. This study investigated potential improvements in the accuracy of NIPT by performing classification and analysis based on differences in cfDNA size. METHODS: We performed paired-end sequencing to identify size ranges of fetal and maternal cfDNA from 62,374 pregnant women. We then developed a size-selection method to isolate and analyze both fetal and maternal cfDNA, defining fetal-derived cfDNA as less than 150 bp and maternal-derived cfDNA as greater than 180 bp. RESULTS: By implementing size-selection method, the accuracy of NIPT was improved, resulting in an increase in the overall positive predictive value for all aneuploidies from 89.57% to 97.1%. This was achieved by enriching both fetal and maternal-derived cfDNA, which increased fetal DNA fraction while the number of false positives for all aneuploidies was reduced by more than 70%. CONCLUSIONS: We identified the differences in read length between fetal and maternal-derived cfDNA, and selectively enriched both shorter and longer cfDNA fragments for subsequent analysis. Our approach can increase the detection accuracy of NIPT for detecting fetal aneuploidies and reduce the number of false positives caused by maternal chromosomal abnormalities.