Abstract
Optimization cryopreservation protocols require loading/unloading cryoprotectants (CPAs) while minimizing negative osmotic effects on embryos, particularly in vitrification when high concentration of CPAs are employed. The current study applies microfluidic technology to minimize osmotic effects and improve overall outcome. Experimental lab study. A computer-controlled multi-channel microfluidic device and associated embryo trapping vitrification straw was developed to minimize osmotic effects. Donated frozen patient cleavage embryos were randomly divided into two groups. In conventional vitrification (n = 39) we used a stepwise method, the experimental group (n-34) used a computer controlled automatic microfluidic system which systematically mixing CPAs. Final vitrification CPAs loading solution is 30%CPAs (15% EG+15% DMSO+ 0.5M sucrose in 20% SSS (v/v in PBS) and initial unloading (thawing) from 1.0 M Sucrose to PBS with 20% SSS. Freezing and thawing solution samples were collected in 60 second intervals to examine osmotic change surrounding embryos for both groups. Day 4 and 5 development was assessed. Our data indicate that the automated vitrification system generated a more gradual osmolarity changes in comparison to the more abrupt osmolarity changes in the manual group during the both CPAs loading and unloading process. Post thawing, microfluidic vitrified/thawed embryos shows similar (P>0.05) embryo survival rates (85.3% vs 89.7%), morula formation rate (50.0% vs 46.2%), cavitation (41.2% vs 38.5%) and blastocyst formation rate (32.4% vs 30.8) compared to the control group. This computer-controlled automated vitrification system provides reduced osmotic effect evidenced by smoother osmotic changes for both CPAs loading /unloading procedures. This is the first automatic vitrification platform for with consistent and reproducible outcomes. It requires less, labor consuming and minimal inter-embryologist variability for human embryos cryopreservation.
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