Changing the start temperature and cooling rate in a slow-freezing protocol increases human blastocyst viability

Abstract

Objective: To determine the effect of start temperature and cooling rate of a slow freezing protocol on human blastocyst viability.

Design: Controlled-rate freezing of human blastocysts using different start temperatures and cooling rates.

Setting: Private assisted reproductive technology unit.

Patient(s): Patients donated with consent cryopreserved pronucleate embryos.

Intervention(s): Culture of thawed pronucleate embryos in G III series media, containing hyaluronan, followed by cryopreservation of 36 blastocysts with subsequent noninvasive analysis of embryo metabolism.

Main outcome measure(s): Pyruvate and glucose consumption and blastocyst reexpansion and quality.

Result(s): Glucose consumption and blastocyst reexpansion after thaw were significantly higher when a start temperature of -6 degrees C and a cooling rate of 0.5 degrees C/min to -32 degrees C were used compared with a start temperature of 20 degrees C and a cooling rate of 2 degrees C to -6 degrees C, followed by cooling at 0.3 degrees C to -35 degrees C. Pyruvate uptake after thaw was not affected by the freezing procedure. Clinical use of the lower start temperature and quicker cooling rate, combined with culture in hyaluronan-based media, has led to the establishment of a 30% implantation rate.

Conclusion(s): Human embryos cultured to the blastocyst stage in hyaluronan-based sequential media are readily cryopreserved and maintain their viability after thaw.