How much damage--and whether it's "too much" for the intended purpose of the archived nucleic acid--will depend on a number of factors, including purity of the extract and buffer components, endogenous polysaccharides or proteins able to act as cryoprotectants
, and rate of freezing.
In this experiment, six concentrations of cryoprotectants
in the equilibration and vitrification solutions were used: 5% EG in DPBS with 0.
Fahy and his colleagues were able to develop a vitrifying cryoprotectant
solution that is nontoxic enough for preserving large organs.
Dr Sharif said vitrification uses high concentrations of cryoprotectants
(the substance used to protect tissue from freezing damage) and an extremely high cooling rate of -20,000 to -25,000 degrees Celsius per minute.
Vitrification uses high concentrations of cryoprotectants
and an extremely high cooling rate (-20,000- 25,000 [euro]centsC per minute) that solidify without the formation of ice crystal by direct immersion into liquid nitrogen in order for the oocytes to be transitioned from 37[euro]centsC to -196[euro]centsC in less than one second and can be stored for future use.
After centrifugation, the pellet was divided in three parts, one with and one without cryoprotectants
respectively (2% w/w glycerol or 5% w/w maltodextrine) [3, 4]) and freeze-dried in a low freeze-drier (LOUW KOELTECHNIEK BVBA) with a standard program by increasing the temperature gradually from - 25[degrees]C to 25[degrees]C at 0,9 mbar pressure during 48h.
Effect of cryoprotectants
on release of various enzymes from buck spermatozoa during freezing.
This two hours process was designed so that the cryoprotectants
would infuse into the embryo's cells and protect it from ice.
This differs from freezing in that 60 percent of the water inside the cells is replaced with protective chemicals called cryoprotectants
before the temperature is dropped.
Although initial survival, fertilisation and cleavage rates for cryopreserved oocytes were relatively poor, the advancements made in cryoprotectants
and method protocols in the recent past have caused these rates to improve steadily.
In cryopreservation, cryoprotectants
such as sucrose, DMSO, Ethylene glycol, glycerol are used to protect the plant material from damaging effects due to freezing.
Upon removal of the cryoprotectants
, the embryos were incubated for 72 hours and assessed for blastocyst development based on morphology.