Etchevers:Notebook/Genomics of hNCC/2009/04/09
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Yeast in cultures
The yeast seems to come from the neural medium. I don't know if it was the B27 supplement or something else (possibly the EGF which is always up in the rim of the centrifuge tube) but as it had not been post-filtered, and I put it on the trunk cultures only yesterday afternoon, there is no doubt. Chains of up to three yeast cells each, and lots of them. Threw away both the culture and that medium. So it may not have been the collagenated flasks that were contaminated on Monday.
Filtered the 20% SVF/DMEM for Stephane's rabbit EC cells. The 0.2% fungizone seemed to work well for any yeast (nothing suspicious visible) but there are some dead cells floating (as well as plenty of live ones, still). The older cultures from 9/3/09 are still slowly dying out - more carcasses than live cells in bottom, now. Filtered medium using Stericup and diluted so only 0.1% fungizone.
Removed 100 μL from one of the old cultures, and from each of the three new EC cultures. Placed in bacteria incubator at 37°C until tomorrow, then can test mycoplasma on them (and see how well the Eppie caps keep out bacteria when closed). Changed the medium on all for the 1/1000 dilution of fungizone (0.1% of 250 μg/mL original = 0.25 μg/mL rather than the 0.5 μg/mL or 0.1 μg/mL I might have written).
Also added 0.1% fungizone (15 μL) to 15 mL of Rich medium for what is left of the hNCC - R1066 cephalic. These are not quite confluent. May be worth splitting tomorrow into a flask.