Down syndrome and cell-free fetal DNA in archived maternal serum

OBJECTIVE: Increased levels of cell-free fetal DNA (f-DNA) in the maternal circulation are a potential noninvasive marker for fetal Down syndrome. Our objectives were to (1) determine whether f-DNA could be quantified by using archived serum and amniotic fluid, (2) examine whether serum f-DNA levels are elevated in Down syndrome pregnancies in a case-control series matched for gestational age and duration of sample storage, and (3) determine whether f-DNA levels are elevated in the amniotic fluid of Down syndrome fetuses. STUDY DESIGN: Eleven serum and six amniotic fluid samples previously collected and stored at -20°C from gravid women carrying a 47,XY,+21 fetus were each paired with five matched control samples of identical specimen type from gravid women carrying a presumed euploid male fetus. f-DNA concentration was quantified blindly by real-time polymerase chain reaction amplification for a Y-chromosome sequence. Matched rank-sum analysis and analysis of variance were used for analysis. RESULTS: The mean observed rank of 5.0 in the Down syndrome group was significantly higher than expected (P ≤ .005). Adjusted mean serum f-DNA concentrations were 41.2 genomic equivalents (GE) per milliliter for the Down syndrome cases and 24.2 GE/mL for the euploid controls (P = .002). Differences among amniotic fluid samples were not statistically significant. There was a suggestion of a sample storage effect on f-DNA concentration on the order of -0.66 GE/mL per month (P = .071). CONCLUSION: Down syndrome pregnancies exhibit 1.7-fold higher levels of maternal serum cell-free f-DNA compared with matched controls. No such association is observed in amniotic fluid. Archived serum appears to be a useful source of clinical material for retrospective analyses but may require controlling for the duration of sample storage.