Library Concentration of Cell-Free DNA in Spent Culture Medium: a Potential Indicator for Clinical Outcomes of Blastocyst Transplantation

Research Question Can the library concentration of cell-free DNA in spent culture medium be a potential indicator for the pregnancy outcomes of blastocyst transfer? Design 341 infertile couples undergoing intracytoplasmic sperm injection (ICSI) with female partners aged more than 35 years were included in the retrospective observational study. The blastocysts obtained from the enrolled patients were cultured individually with the associated SCM collected. The single-cell whole genome amplification (WGA) and DNA library preparation were performed afterward. The cfDNA concentration and the copy number variation (CNV) results for each sample were analyzed. A follow-up study was conducted after the blastocyst transfers. The correlation between library concentration and clinical outcome was analyzed. Results A total of 644 single blastocyst transfers were performed. In 35.25% of cycles, a clinical pregnancy was achieved by patients, with an ongoing pregnancy rate of 27.80%, resulting in a live birth rate of 27.48%. The blastocysts were divided into low- and high-concentration groups according to the DNA library concentration with a threshold of 1 ng/µL in SCM. The clinical pregnancy rate (43.65% vs. 31.54%, p=0.003), ongoing pregnancy rate 37.56% vs. 23.49%, p < 0.001), and live birth rate (37.56% vs. 23.04%, p < 0.001) were found to be significantly higher in the low-concentration group than in the high-concentration group. Conclusion Library concentration may be a potentially important predictive indicator for blastocyst transfer. Given its shorter testing duration and potential to improve pregnancy outcomes, using library concentration to guide the transfer priority of embryos in fresh cycles could be considered.