Optimal Exposure Parameters for Microsurgery of Embryo Zona Pellucida Using Femtosecond Laser Pulses

Abstract

We applied femtosecond laser pulses for microsurgery of the zona pellucida (ZP) of mouse embryos in terms of assisted reproductive technologies. The embryos were subjected to a series of laser pulses (wavelength of 514 nm, pulse duration of 280 fs, repetition rate of 2.5 kHz). Optical microscopy was used to study the dependence of the ZP cut width on the pulse energy E and velocity v of the laser beam. It is shown that the same value of the cut width can be obtained for different combinations of these parameters. The boundaries of admissible values were found to be E = (19–52) nJ, v = (0.001–0.03) mm/s; recommendations on their proper choice are given. An analytical expression of the cut width of the ZP for a given combination of laser-pulse energy, beam velocity, and pulse repetition rate is proposed. This simple and easy-to-use equation allows for quick prediction of ZP cut width in embryo microsurgery procedures of laser-assisted hatching and embryo laser tagging using femtosecond laser pulses.