Flow cytometric assessment of morphology, viability, and production of reactive oxygen species ofCrassostrea gigasoocytes. Application to Toxic dinoflagellate (Alexandrium minutum) exposure : FCM for Viability and ROS of C. Gigas Oocytes

To investigate the effect of vitrification and beta-mercaptoethanol (beta-ME) on reactive oxygen species (ROS) activity and in vitro development of oocytes vitrified before or after in vitro fertilization (IVF). Randomized prospective study. University-based assisted reproductive technology laboratory. ANIMALS(S): Abattoir-derived porcine ovaries. Oocytes were vitrified either before or 4 hours after the end of IVF by solid surface vitrification (SSV) without centrifugation and/or delipation procedure. beta-ME was used to inhibit ROS activity. Viability was evaluated by membrane integrity and esterase enzyme activity using fluorescein diacetate staining while ROS activity was assessed by 2',7'-dichlorofluorescein assay. Vitrification increased the ROS activity and decreased the viability and in vitro development of vitrified oocytes. Addition of beta-ME to vitrification and culture medium partially annihilated the ROS activity but did not improve the viability of vitrified-warmed oocytes. Furthermore, beta-ME had no effect on improving the fertilization ability of oocytes vitrified at metaphase II stage but significantly increased their ability to cleave. beta-ME also increased the rate of cleavage and blastocyst formation ability of oocytes vitrified 4 hours after the end IVF. Vitrification increases ROS activity in oocytes that can be partially annihilated by beta-ME to obtain enhanced embryonic development. Copyright 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.