Problem with mouse embryonic stem cells - contamination? - (Mar/17/2008 )
I've been trying to grow up mouse embryonic stem cells from frozen stocks, and I've been having some issues. The first batch that I thawed out all became contaiminated with yeast and bacteria. Since then, I've used pen/strep in the medium (1%) and have had no more problems with bacteria. In order to prevent yeast infection, I've been using Fungizone in the plates at a concentration of 0.1mcg/mL (below the minimum recommended level). The few plates that I didn't use Fungizone in quickly became infected with yeast. The one plate that I did use Fungizone was okay until about 5 days post-thawing. Then the medium started turning yellow and cell colonies began detaching from the plate surface. The remaining adherent colonies began to show signs of differentiation (?). Finally, white specks - visible to the naked eye - appeared on the plate. At this stage, I discarded the cells.
Does anyone have any idea what might be going on here? There are some photos of the cells, both of the detached colonies and of the differentiated (?) colonies, at this address; http://www.geocities.com/lizanne18/ESC.html
Any help would be very much appreciated!
Does anyone have any idea what might be going on here? There are some photos of the cells, both of the detached colonies and of the differentiated (?) colonies, at this address; http://www.geocities.com/lizanne18/ESC.html
Any help would be very much appreciated!
i am not an expert but what i think is , its better to find the source of contamination than using different reagents.
is only your cells getting contaminated? if yes then better to start from new cells. if other people are also geting contamination then better to sterilized the place. we shifted our clean bench from a crowded common romm to another place. usually the thick paper boxes used for carrying glasswares are sources of yeast contamination. better to keep these kind of boxes far from the clean bench.