title: Extreme rapid warming yields high functional survivals of vitrified 8-cell mouse embryos even when suspended in a half-strength vitrification solution and cooled at moderate rates to -196°C.
abstract: To cryopreserve cells, it is essential to avoid intracellular ice formation during cooling and warming. One way to do so is to subject them to procedures that convert cell water into a non-crystalline glass. Current belief is that to achieve this vitrification, cells must be suspended in very high concentrations of glass-inducing solutes (i.e., ≥6 molal) and cooled at very high rates (i.e., ≫1000°C/min). We report here that both these beliefs are incorrect with respect to the vitrification of 8-cell mouse embryos. In this study, precompaction 8-cell embryos were vitrified in several dilutions of EAFS10/10 using various cooling rates and warming rates. Survival was based on morphology, osmotic functionality, and on the ability to develop to expanded blastocysts. With a warming rate of 117,500°C/min, the percentages of embryos vitrified in 1×, 0.75×, and 0.5× EAFS that developed to blastocysts were 93%, 92%, and 83%, respectively. And the percentages of morphological survivors that developed to expanded blastocysts were 100%, 92%, and 97%, respectively. Even when the solute concentration of the EAFS was reduced to 33% of normal, we obtained 40% functional survival of these 8-cell embryos.
authors: Fundamental and Applied Cryobiology Group, Department of Biochemistry and Cellular and Molecular Biology, The University of Tennessee, Knoxville, TN 37996, USA; Present address: Tokyo University of Marine Science and Technology, Tokyo 108-8477, Japan.
authors: Shinsuke
authors: S
authors: Seki
authors: Fundamental and Applied Cryobiology Group, Department of Biochemistry and Cellular and Molecular Biology, The University of Tennessee, Knoxville, TN 37996, USA.
authors: Bo
authors: B
authors: Jin
authors: Fundamental and Applied Cryobiology Group, Department of Biochemistry and Cellular and Molecular Biology, The University of Tennessee, Knoxville, TN 37996, USA. Electronic address: pmazur@utk.edu.
authors: Peter
authors: P
authors: Mazur
mesh_terms: D000818:Animals; D002470:Cell Survival / Q000187:drug effects; D015925:Cryopreservation; D004622:Embryo, Mammalian / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D047108:Embryonic Development; D005260:Female; D008297:Male; D051379:Mice; D019188:Organ Preservation Solutions / Q000494:pharmacology*; D009995:Osmosis; D013997:Time Factors; D058989:Vitrification / Q000187:drug effects
chemical_list: D019188:Organ Preservation Solutions
keywords: Cooling rates; Embryos; Mouse; Oocytes; Solute concentration in media; Survival; Vitrification; Warming rates