Large-volume vitrification of human biopsied and non-biopsied blastocysts: a simple, robust technique for cryopreservation

Abstract

Purpose To evaluate the transition from a proven slowcooling cryopreservation method to a commercial largevolume vitrification system for human blastocysts. Methods Retrospective analysis of de-identified laboratory and clinical data from January 2012 to present date for all frozen embryo replacement (FET) cycles was undertaken. Cryopreservation of trophectoderm-biopsied or non-biopsied blastocysts utilized during this time period was logged as either slow-cooling, small-volume vitrification, or largevolume vitrification. Blastocyst survival post-warm or postthaw, clinical pregnancy following FET, and implantation rates were identified for each respective cryopreservation method. Results Embryo survival was highest for large-volume vitrification compared to micro-volume vitrification and slowcooling; 187/193 (96.9 %), 27/32 (84.4 %), and 244/272 (89.7 %), respectively. Survival of biopsied and nonbiopsied blastocysts vitrified using the large-volume system was 105/109 (96.3 %) and 82/84 (97.6 %), respectively. Survival for micro-volume biopsied and non-biopsied blastocysts was 16/30 (83.3 %) and 2/2 (100.0 %) respectively. Slow-cooling post-thaw embryo survival was 272/244 (89.7 %). Clinical pregnancy and implantation rates outcomes for non-biopsied embryos were similar between large-volume and slow-cooling cryopreservation methods, 18/39 (46.2 %) clinical pregnancy and 24/82 (29.3 %) implantation/embryo, and 52/116 (44.8 %) clinical pregnancy and 67/244 (27.5 %) implantation/embryo, respectively. Comparing outcomes for biopsied embryos, clinical pregnancy and implantation rates were 39/67 (58.2 %) clinical pregnancy and 50/105 (47.6 %) implantation/embryo and 4/16 (25 %) clinical pregnancy and 6/25 (24.0 %) implantation/embryo, respectively. Conclusions The LifeGlobal large-volume vitrification system proved to be very reliable, simple to learn and implement in the laboratory. Clinically large-volume vitrification was as, or more effective compared to slow-cooling cryopreservation in terms of recovery of viable embryos in this laboratory.

Freeze all-first versus biopsy-first: A retrospective analysis of frozen blastocyst transfer cycles with preimplantation genetic testing for aneuploidy

“…The authors claimed that the blastocysts re-expanding and vitrified later than 3h from biopsy show the highest chance to implant. Lastly, Reed et al [ 21 ] in a study designed to compare different cryopreservation protocols reported no difference in the cryo-survival rate between biopsied and non-biopsied blastocysts. We found that there were no significant differences in embryo survival rates between freeze all-first and biopsy-first groups.…”

Human sperm vitrification: the state of the art

“…Human embryo cryopreservation which has been well established may provide some clues for sperm cryopreservation. Reed et al [17] reviewed laboratory and clinical data for all frozen embryo replacement cycles from 2012 to 2015 and concluded that vitrification was more effective, as indicated by higher survival in vitrified embryos compared to slow-cooling cryopreservation. Furthermore, vitrification is also reliable and simple to learn and implement in the laboratory while clinical pregnancy and implantation rates outcomes are similar [17], which is supported by other studies have [18][19][20].…”

“…If the cooling speed during vitrification is flexible, aseptic vitrification with double straw containing higher volumes of 0.1 to 0.5 mL would allow the use of these techniques in human ART [18,19]. Interestingly, in human embryo vitrification, embryo survival is higher for large-volume vitrification, which has lower cooling rate comapred to micro-volume vitrification [17].…”

Human Embryo Cryopreservation—Methods, Timing, and other Considerations for Optimizing an Embryo Cryopreservation Program

“…Permeating CPAs (commonly a combination of EG and sucrose along with PROH, glycerol, or DMSO) are introduced with a two-step approach, exposing embryos to 50% concentration of the final CPA concentration for 5 to 15 minutes to permit intracellular water to exit the cell and CPA permeation to establish an equilibrium and limiting exposure to the final 30 to 40% solution of CPAs to the minimum amount of time (<60 seconds) required to achieve cell shrinkage. 18,28,47,54,56,57 The embryo is then plunged into liquid nitrogen and, depending on the carrier, sample volume, and solution composition, is cooled at a rate of 2.5 to 30 Â 10 3°C / min. 17,57 Rapid warming is achieved by quickly removing the embryo from liquid nitrogen (À196°C) and immersing it in a solution that has been warmed to core temperature.…”