Influence of the Cryopreservation Protocol on the Results of Femtosecond Laser Microsurgery of Zona Pellucida

The object of this study is the zona pellucida of a mouse embryo subjected to vitrification and thawing (a cryo-embryo). Zona pellucida dissection was performed by femtosecond laser pulses in the visible spectral range (wavelength 514 nm, duration 280 fs, pulse repetition rate 2.5 kHz). The dependence of the width D of the cut formed on the zona pellucida by the laser beam on the laser pulse intensity I and beam motion velocity υ is investigated. The obtained results are compared with the data on the embryos that were not subjected to cryopreservation (“fresh” embryos). The applied method of statistical analysis (ANCOVA, or covariance analysis) demonstrated the absence of statistical difference in the dependences D(I, υ) for two embryo groups: fresh and cryo-embryos. Thus, the parameters of the laser radiation used in the fresh embryo microsurgery are suitable for dissecting the zona pellucida of embryos subjected to vitrification and thawing.