Cell-Free RNA Is a Reliable Fetoplacental Marker in Noninvasive Fetal Sex Determination

Elke Mersy, Brigitte H. W. Faas, Sabine Spierts, Leonie M. H. Houben, Merryn V. E. Macville, Suzanna G. M. Frints, Aimee D. C. Paulussen, Joris A. Veltman*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review


BACKGROUND: Noninvasive genetic tests that use cell-free fetal DNA (cffDNA) are used increasingly in prenatal care. A low amount of cffDNA can have detrimental effects on the reliability of these tests. A marker to confirm the presence of fetal nucleic acids is therefore required that is universally applicable and easy to incorporate. METHODS: We developed a novel multiplex, single-tube, noninvasive fetal sex determination assay by combining amplification of AMELY cffDNA with one-step reverse transcription (RT)-PCR of trophoblast-derived cell-free RNA (cfRNA), which functions as a sex-independent fetoplacental marker. We tested plasma samples from 75 pregnant women in duplicate in a blinded fashion. The fetus was considered to be male in the case of a positive result for AMELY and cfRNA amplification in both RT-PCRs. The fetus was considered to be female in the case of negative AMELY and positive cfRNA result in both RT-PCRs. In other cases, the test was repeated. We compared the results with invasive prenatal testing and pregnancy outcomes. RESULTS: The AMELY cffDNA amplification and cfRNA result was unambiguous and identical in duplicate in 71 of 75 plasma samples (95%). Four samples (5%) required an extra replicate because of an absent fetoplacental marker. Thereafter, fetal sex was correctly determined in all 75 plasma samples. CONCLUSIONS: Amplification of trophoblast-derived cfRNA is a reliable marker for the confirmation of the presence of fetoplacentally derived nucleic acids in noninvasive fetal sex determination.
Original languageEnglish
Pages (from-to)1515-1523
JournalClinical Chemistry
Issue number12
Publication statusPublished - Dec 2015


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