PurposeTo compare clinical outcomes following transfer of euploid blastocysts of varying quality biopsied on day 5 versus day 6.MethodsRetrospective cohort study to evaluate embryo transfer outcomes for women undergoing autologous cryopreserved next generation sequencing euploid single embryo transfer from 10/2015 to 2/2022 at an academic IVF program. The primary outcome was live birth rate (LBR). Secondary outcomes included ongoing pregnancy rate (OPR), implantation rate (IR), and miscarriage rate (SAB rate).ResultsFive hundred and fifty-five transfers from 418 patients were analyzed. Euploid embryos biopsied on day 5 resulted in higher LBR compared to those biopsied on day 6 (62.3% vs. 49.6%; aRR 0.81 95% CI 0.65–0.996). When stratified by biopsy day and blastocyst quality, there was no difference in IR, OPR, and SAB rate for good, fair, and poor quality blastocysts biopsied on day 5 versus day 6. However, day 5 good quality embryos were associated with a higher LBR compared to day 6 good quality embryos (74.3% vs. 51.3%; aRR 0.69; 95% CI 0.48–0.999). There were no significant differences in LBR for fair and poor quality embryos biopsied on day 5 versus day 6.ConclusionOverall LBR are higher for euploid embryos biopsied on day 5 versus day 6. When stratified by embryo quality and day of biopsy, LBR are significantly higher for good quality day 5 versus day 6 embryos. When choosing between multiple euploid embryos, day 5 biopsied good quality embryos should be preferentially selected for transfer over day 6 embryos of the same quality.