stock preparation

[aftabac]

Hi
During stock preparation of bacterial cells we most oftenly use 70% glycerol, what is role of glycerol in cell stock preparation?
is there is some alternative of glycerol?
if we have bacterial cells that have some antibiotic resistance gene on plasmid, then is it necessary to add some antibiotic along with glycerol or we can use simply glycerol for stock preparation?

waiting for answer

aftab

[darkwng]

Glycerol is known to be a CryoProtectant.
This prevents crystal formation and damage of cells during freezing.

http://www.iisc.ernet.in/academy/res...2002p59-63.pdf

Lyophilization Best Storing Method

and there is no need to add antibiotics as the cells are frozen and cannot lose the plasmid as they are not growing (ie do not experience any selective pressure to lose the plasmid).


Ok looked up the following, they also act as CryoProtectants:

cryoprotectant solutions include DMSO, glycerol or skim milk/glycerol.

[darkwng]

Looking more into this it seems that CryoProtectants control the freezing process (how???)

Controlled Freezing with Cryoprotectants:

Control of the rate of freezing is critical to optimize the ice nucleation process and minimize cell / tissue injury. Quick freezing results in intracellular ice nucleation. which is lethal to the cell or tissue.

Very slow freezing rates also causes cell death is also excessive due the ‘pickling’ effects of long periods of exposure to hypertonic solutions. At optimal rates of freezing the exposure time to hypertonic conditions is minimized while intracellular ice formation is prevented.

[darkwng]

Cryoprotectants:

Cryoprotectants


Cryoprotective additives include glycerol, dimethyl sulfoxide (dimethyl sulphoxide, DMSO), and propanediol. Cell membranes are permeable to these compounds which protect cells from damage during freezing. Cryoprotectants reduce damage following freezing and thawing, by increasing the unfrozen fraction and thereby reduce the ionic composition of the tissues.

There can be significant differences in the protective efficiency of different cryoprotectants. This may be the result of differing cellular toxicities and/or permeability to specific compounds.

At UCR they have demonstrated no difference in recovery with either a skim-milk/glycerol mixture or DMSO used as cryoprotectants.


Read more about Controlled Freezing and Cryoprotectants here:
http://www.geocities.com/sulawesipri...servation.html