I´ve thawed a cell line (hESCs on MEF) recently and this morning I found this floating in my media. It is not a contamination I´ve ever seen (fungi or bacteria). Does maybe someone have an idea what this might be? It looks like some tetracoccus, but then again it seems to be too square. I´m a bit confused.
Thank you all for your help!
I suspect that is yeast, which is technically a fungus, but does not form filaments like Pencillium or aspergillus. Yeast is notoriously difficult to get rid of. I would toss those cultures, refilter my culture medium, and thaw new stocks of MEF and hESC.
if the culture is important and if you suspect yeast -harvest and pass the harvested cell suspension through ficoll gradient (google the protocol, it's an old protocol). Then add 1x anti-anti for 10 days. Because your cells attach, and yeast cells tend to float around -you will get rid of them by ficoll (95%) and then by maintaining in anti-anti and washing the cultures each time (before media change) with PBS 2-3 times. This works great. Also, hESCs may look like this when coming out of the suboptimal freezing (excessiver death may be observed). Use 10 microM Rock inhibitor for the 1st 2-4 days. Dont keep it forever. Last, there is possibility that your culture was plated on the old MEFs, you need to use day 2-3 MEFs and MEFs start to degenerate and die at day 7-10. You may have a bad MEF prep, and then your hESC culture degenerates with it. Try plating 1/2 of thawed aliquot on vitronectin or Matrigel. When cells are shifted from feeders to feeder-free culture, they tend to differentiate but some pluripotent colonies still pop up and you expand from them. If you really think you have a contamination -take conditioned medium from "contaminated" cells and add it to you other culture(s) to test. Without showing more images it is close to impossible to have a better guess -you need to show overview pictures of your cultures. Check our protocols, MEFs and MEF-free Article Long-Term, Stable Differentiation Of Human Embryonic Stem Ce...
,Article Characterization of Three-Dimensional Retinal Tissue Derived...
,Article Comparison of Developmental Dynamics in Human Fetal Retina a...
. I am happy to comment if you send more pictures of your culture (zoom out)- Badges
- Science topic
- Similar topics
- Counseling Psychology
- Confusion