STUDIES REGARDING THE CRIOPROTECTIVE PROPRIETIES OF THE VITRIFICATION MEDIA, WITH DMSO, SUCROSE, FICOLL 70 AND GALACTOSE USED IN EMBRYO CRYOPRESERVATION
Keywords:
crioprotectors, vitrification, DMSO, sucrose, Ficoll 70, galactoseAbstract
The aim of our paper was to make a series of experiments in order to determine the
concentration at which four cryoprotectants (DMSO, sucrose, Ficoll 70 and
galactose) singly and in pairs would vitrify on plunging into liquid nitrogen and
remain vitreous when thawed in water bath. As penetrating cryoprotector we used
DMSO (MW=78.13 Da, Sigma D5879) and as nonpenetrating cryoprotectors we
used sucrose (MW=342.3 Da, Sigma S7903), Ficoll 70 (MW= 60,000-80,000 Da,
Sigma F4375) and Galactose (MW = 180,16 Da; Sigma G 6152). For DMSO there
were tested concentrations from 1M to 6.5M, with concentrations step of 0.5M. For
the nonpenetrating cryoprotectors there were tested concentrations of 5%, 10%,
15% and 20%. There were a total number of 168 solutions tested. The solutions
vitrification ability on freezing was tested by direct plunging into liquid nitrogen at -
196°C. Three thawing temperatures were tested 20°C, 25°C and 37°C. The
concentration at which DMSO solutions passed into vitreous state was 5M, but at
thawing none of them remained vitreous at thawing. When pairs of cryoprotectors
were tested 67 solutions vitrified at freezing (23 for DMSO-sucrose, 23 for DMSOFicoll 70 and 21 for DMSO-galactose) and 26 of them remained vitreous at
thawing. The DMSO and galacose pair give the best results on thawing (11 solution
remained vitreous on warming) at 37°C.
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