ULTRARAPID FREEZING OF MOUSE EMBRYOS
type: CreativeWork
description: The "ultrarapid" technique for murine embryo cryopreservation involves a brief exposure (2.5-3 minutes) of embryos to a high concentration (3.0-4.5 M) of dimethyl sulfoxide (DMSO) and 0.25 M of sucrose at room temperature before direct plunging into liquid nitrogen. The objective of this study was to determine the efficacy of this technique for the cryopreservation of two-cell mouse embryos under various conditions. Three bicarbonate-based culture media namely T6, Medicult and Human Tubal Fluid (HTF) medium were found to result in high rates of survival (96.2%-98.4%) and in vitro development (77.9%-87.8%) when employed to make up the 3.5 M DMSO freezing solution. The elevated pH of these solutions in the absence of a 5 % CO2 atmosphere did not have a detrimental effect on the cryosurvival and developmental capacity of the frozen-thawed embryos. Upon comparing the use of phosphate-, Hepes- and bicarbonate-buffered T6 medium containing 3.5 M or 4.5 M DMSO and 0.25 M sucrose for cryopreservation, phosphate-buffered T6 containing 3.5 M DMSO was found to result in a reduced in vitro blastocyst formation rate (72 % ) compared to bicarbonate- and Hepes-buffered T6 (84.5% and 86.5% respectively). Upon differential staining of the developing blastocysts, it was followed that blastocysts derived from the phosphate-buffered T6 group possessed a significantly smaller inner cell mass (mean cell number of 21. 6) and a reduced mean total cell number (86.6) compared to blastocysts of tile other media groups. With 4.5 M of DMSO, although all three media yielded consistently low blastocyst formation rates (69.2%-75.6%), freeze-thaw in Hepes-T6 yielded blastocysts with a significantly higher mean ICM (26.0) and total blastocyst (97.3) cell number compared to freeze-thaw in the other two media. On transferring the developing blastocysts from the bicarbonate-, Hepes- and phosphate-buffered T6 groups frozen with 3.5 M DMSO into pseudopregnant females, similar implantation (64.0%-76.5%) a…
abstract: The "ultrarapid" technique for murine embryo cryopreservation involves a brief exposure (2.5-3 minutes) of embryos to a high concentration (3.0-4.5 M) of dimethyl sulfoxide (DMSO) and 0.25 M of sucrose at room temperature before direct plunging into liquid nitrogen. The objective of this study was to determine the efficacy of this technique for the cryopreservation of two-cell mouse embryos under various conditions. Three bicarbonate-based culture media namely T6, Medicult and Human Tubal Fluid (HTF) medium were found to result in high rates of survival (96.2%-98.4%) and in vitro development (77.9%-87.8%) when employed to make up the 3.5 M DMSO freezing solution. The elevated pH of these solutions in the absence of a 5 % CO2 atmosphere did not have a detrimental effect on the cryosurvival and developmental capacity of the frozen-thawed embryos. Upon comparing the use of phosphate-, Hepes- and bicarbonate-buffered T6 medium containing 3.5 M or 4.5 M DMSO and 0.25 M sucrose for cryopreservation, phosphate-buffered T6 containing 3.5 M DMSO was found to result in a reduced in vitro blastocyst formation rate (72 % ) compared to bicarbonate- and Hepes-buffered T6 (84.5% and 86.5% respectively). Upon differential staining of the developing blastocysts, it was followed that blastocysts derived from the phosphate-buffered T6 group possessed a significantly smaller inner cell mass (mean cell number of 21. 6) and a reduced mean total cell number (86.6) compared to blastocysts of tile other media groups. With 4.5 M of DMSO, although all three media yielded consistently low blastocyst formation rates (69.2%-75.6%), freeze-thaw in Hepes-T6 yielded blastocysts with a significantly higher mean ICM (26.0) and total blastocyst (97.3) cell number compared to freeze-thaw in the other two media. On transferring the developing blastocysts from the bicarbonate-, Hepes- and phosphate-buffered T6 groups frozen with 3.5 M DMSO into pseudopregnant females, similar implantation (64.0%-76.5%) a…
author: SHIVANI VASUTHEVAN
date published: 1993
date modified: 2026-07-24T03:30:47Z
provider: National University of Singapore
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: National University of Singapore