Validation of a Universal Warming/Dilution Protocol for Frozen Embryo Transfers Independent of Vitrification Device/Cryoprotectant

Abstract

Objective Validate a universal warming/dilution approach for all vitrification (VTF) solutions associated with various device systems. Design Using a 2 × 2 factorial arrangement of treatments, 124 blastocysts derived from 363 research consented slow frozen embryos were vitrified in Glycerol/ethylene glycol (EG) solution (≥7.9M; I.C.E.) or a 15%DMSO/15%EG solution, and subsequently rapidly warmed and diluted by standard operating procedures (SOP) or a universal sucrose step-down dilution procedure (n = 31 embryos/group). Furthermore, we performed a retrospective analysis of our routine success with microSecure VTF (µS-VTF with Glycerol/EG)/warming to applying a universal sucrose dilution protocol, independent of vitrification device/solution, to blastocysts shipped in from outside facilities between 2015-2017. In validating our clinical approach, we contrasted live birth rates achieved in the latter groups (A and B, respectively; see Table) to outcomes reported in the 2015 CDC data, including the national average (C) and four local IVF labs/clinics (D-H). Materials and Methods All research blastocysts were vitrified by µS-VTF using solutions and dilution treatments described above. Conversely, all internal FET cycles implemented a standardized sucrose step-down dilution using I.C.E. diluents (T1 -T4; estimated to possess 1.0M, 0.5M, 0.25M and 0.125M sucrose concentrations; 3 min/step), independent of the VTF device/cryoprotectant, upon rapid warming and blastocyst isolation. All blastocysts underwent isotonic equilibration in Hepes buffered media for 5 min at 37°C before being in-vitro cultured until ET or research developmental assessment. Our live birth pregnancy rates for FETs (A,B; autologous oocytes, Results No statistical differences in survival or sustained development was observed between the combined dilution treatments of vitrified research blastocysts. SOP thawing (93.5%) or standard sucrose dilution (90.3%) outcomes were similar. Mean live births (see Table) following our universal sucrose dilution approach (B) revealed similar or improved outcomes to those reported in the 2015 CDC summary (C-G). The apparent reduced live birth success compared to µS-VTF (A) and a local group (H) is likely due to differences in embryo production (i.e., lab effect), not to a warming-dilution effect post-VTF. Conclusion A universal sucrose, step-down dilution approach has proven to be a simple, cost-effective and highly successful procedure in our treatment for vitrified blastocysts derived from outside IVF programs, thus alleviating the need to purchase and maintain various commercial thaw solutions. Disclosures MCS developed µS-VTF without any commercial interests. Funding None.