Cell-free DNA (cfDNA) testing on maternal blood samples is now used for fetal aneuploidy screening. This testing examines cfDNA fragments circulating in the maternal plasma that originate primarily from cells of the mother and from placental cells. DNA fragments from particular chromosomes are identified by their nucleic acid sequence. Assuming that the maternal cells are euploid, quantitative analysis of the cfDNA fragments is used to predict whether the cells of the pregnancy have the normal or abnormal copy number of specific chromosomes. Recently, parental single-nucleotide polymorphisms have been used as a means of predicting the copy number of specific chromosomes in placental cells. Validation studies in high-risk patients have shown a sensitivity of greater than 98% for detection of trisomy 21 and trisomy 18, with false-positive rates of less than 0.5%, but lower sensitivity for trisomy 13. It is recommended that positive results be confirmed by invasive prenatal diagnosis before important parental decisions are made regarding the pregnancy because the positive predictive value of the test is imperfect. Although the test methods demonstrate excellent discrimination between euploid pregnancies and those with trisomy 13, 18, or 21, the need to establish a quantitative cutoff will result in some false-positive and false-negative results. In this article, data were provided on 8 patients in whom discordant results were found between an abnormal cfDNA test and the normal cytogenetic testing of the pregnancy. The cfDNA testing in these patients was performed in different laboratories, but massively parallel shotgun sequencing was used to identify the chromosomal origin of cfDNA fragments in all 8 cases.
The 8 women were aged 20 to 41 years, and the cfDNA testing was done between 12.3 and 16 weeks 5 days’ gestation. A normal 46,XY chromosome karyotype was found in 6 cases and 46,XX in 2. Five had cfDNA results that were positive for trisomy 18, whereas 3 were false positive for trisomy 13. Whereas all 8 cases had a normal karyotype, 1 had evidence of placental mosaicism on chorionic villus sampling (CVS).
The discordance between the cfDNA testing and the cytogenetic tests indicates the importance of offering invasive diagnostic testing after an abnormal cfDNA test result. Before cfDNA testing can become routine, the possible causes of false-positive results must be determined. A systematic reporting registry for gathering information on false-positive and false-negative cfDNA testing will be important before the test can be widely used in low-risk pregnant women. Clinicians should discuss the possibility of false-positive test results during pretest counseling for screening, which would include cfDNA testing.
Department of Obstetrics and Gynecology (M.T.M., L.D.), Prenatal Cytogenetic Laboratory (M.T.M.); Department of Pathology and Laboratory Medicine and Clinical Cancer Cytogenetics (J.J.D.M.), Perelman School of Medicine, University of Pennsylvania, Philadelphia PA; and Department of Pathology and Cytogenetics Laboratory, Stanford Hospital and Clinics, Stanford University School of Medicine, Palo Alto, CA (A.M.C.)