P-182 Validation of single-step warming for human blastocysts shows successful results are independent of the sucrose concentration used
Authors: Abu ali K , Ahlström A 
Published: June 2023

Abstract

Study question
Can warming of blastocysts in a single step be successfully performed using different sucrose concentrations?

Summary answer
Blastocyst warming in a single step using sucrose concentrations between 0.25 M and 1.0 M shows similar rates of survival, re-expansion and development in vitro.

What is known already
Since the introduction of human blastocyst vitrification, excellent survival rates have been reported. Warming procedures usually involve three to five steps with exposure to solutions with decreasing concentrations of non-penetrating cryoprotectants. Successful blastocyst warming procedures are described using high as well as low starting levels of sucrose. Recent data show that a single step warming procedure can be equally effective in terms of survival and pregnancy rates. Moving towards simpler procedures can support further optimization of laboratory procedures.

Study design, size, duration
Blastocysts donated and consented by patients were used to evaluate a shorter warming procedure. Warming was performed in a single step using different sucrose concentrations. After warming, embryos were assessed for morphological survival and cultured in a time-lapse incubator to monitor re-expansion and development in vitro for 24 h. Three series of tests were performed. A control group using standard warming procedure was included in the first test

Participants/materials, setting, methods
For warming, carriers containing a single blastocyst were plunged into a warming solution at 37 °C, containing 0.25, 0.5 or 1.0 M sucrose and stayed in the solution for 1 or 2 minutes. Three series of experiments were performed, testing different sucrose concentrations or volume of warming solution. Warmed blastocysts were cultured following several rinsing steps and assessed for morphological survival. Re-expansion after 2 h, embryo characteristics and development were monitored until 24 h post warming.

Main results and the role of chance
Results of the different tests are summarized in the table, including information on sucrose levels, exposure times, volume of warming medium, numbers of blastocysts warmed, recovered, meeting criteria for embryo transfer, full re-expansion after 2 hours, viable after 24 h culture.
No differences were observed between the different groups. Overall, 98 % of warmed blastocysts would be considered transferable after single step warming and 96 % were viable after 24 h culture. Results confirm blastocysts can be warmed at 37 °C in a single step using 0.25 M, 0.5 M or 1 M sucrose with no effect on survival or development in vitro.

Limitations, reasons for caution
This is a preclinical validation on use of a reduced warming time for vitrified blastocysts. Further evaluation and clinical validation of the findings is required to confirm safety of a simplified warming procedure. Working temperature rather than warming medium volume may be critical when aiming for shorter warming procedures.

Wider implications of the findings
Reducing the warming time and number of handling steps minimizes exposure of blastocysts to a suboptimal environment and operator or handling related stresses. It also allows further optimization of laboratory protocols and workflow. When confirmed clinically, these findings encourage investigating similar changes for other reproductive cells.

Trial registration number
not applicable