Assisted Hatching and Removal of Degenerate Material Significantly Improves Implantation of Frozen/Thawed Blastocysts

Abstract

It has been suggested that freezing and thawing of embryos could cause artificial hardening of the zona pellucida. (Carroll et al., 1990) The objective of this study is to determine whether Assisted hatching could mechanically facilitate the hatching process in those embryos which have theoretically been compromised by the freezing / thawing process. Prospective, randomized study. All embryos were cultured in a sequential G1 / G2 culture system at 6% CO2, 5 % O2 and 89 % Nitrogen until day 5 or day 6. Only blastocysts demonstrating a clearly defined inner cell mass (ICM) and cellular trophectoderm were frozen. Embryos were frozen using a modified Menezo protocol with 2 steps of glycerol and sucrose. The embryos were thawed on the morning of transfer by passing them through 4 consecutive solutions of decreasing concentrations of glycerol and sucrose. Embryos were cultured for at least 1 hour prior to assisted hatching being performed. Assisted hatching was performed using acidic tyrodes. After the zona was breached, any degenerate material was removed using the hatching tool. Embryos exhibiting the greatest percentage of surviving cells and re-expansion were transferred. Using a computer generated randomization list, patients were randomized into two groups. The “hatching” group had their embryos hatched as described. The “control” group were treated identically, but did not have assisted hatching or removal of degenerate material. Results are summarized in the table below. Tabled 1 This study demonstrates that assisted hatching of frozen/thawed blastocysts increases implantation rates. Continued analysis is needed to determine if pregnancy rates could be significantly improved.