Proof of concept: preimplantation genetic screening without embryo biopsy through analysis of cell-free DNA in spent embryo culture media Mousa I. Shamonki,

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Proof of concept: preimplantation genetic screening without embryo biopsy through analysis of cell-free DNA in spent embryo culture media Mousa I. Shamonki, M.D., Helen Jin, Ph.D., Zachary Haimowitz, B.S., Lian Liu, M.D. Fertility and Sterility Volume 106, Issue 6, Pages 1312-1318 (November 2016) DOI: 10.1016/j.fertnstert.2016.07.1112 Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

Figure 1 Illustration of preparation and collection of spent IVF media for noninvasive genetic screening. 1. On day 3 of embryonic development, each embryo was removed from G1 media; 2. the embryo underwent assisted hatching where an opening in the zona pellucida (ZP) was created using a laser; 3. the embryo was placed in isolation in a 15-μL droplet of G2 media; 4. the embryo was incubated from day 3 to the blastocyst stage by day 5 or 6. Once the embryo was removed to undergo trophectoderm biopsy for preimplantation genetic screening/comprehensive chromosomal screening; 5. the spent media droplet was collected, frozen, then eventually tested for presence of DNA and genetic screening performed to compare with the corresponding trophectoderm biopsy result. Fertility and Sterility 2016 106, 1312-1318DOI: (10.1016/j.fertnstert.2016.07.1112) Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

Figure 2 Initial four samples tested with the highest levels of DNA after a 2-hour whole genomic amplification. Only sample AB16, outlined in red, had a derivative log ratio SD (DLRSD) of <0.85, an indication of appropriate DNA quality and quantity to allow for an accurate genetic screening result. The DNA from this sample yielded a result of 45 XY, -13, identical to the blinded preimplantation genetic screening result from the corresponding trophectoderm biopsy. After an additional hour of amplification, an additional sample yielded a DLRSD <0.85 and a result identical to trophectoderm biopsy. Overnight amplification did not produce improved sample accuracy in this cohort of samples. Fertility and Sterility 2016 106, 1312-1318DOI: (10.1016/j.fertnstert.2016.07.1112) Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions