these are the DAPI images which we checked for mycoplasma contamination.
1.how to identify mycoplasma contamination
2. what possibility steps can be taken to get rid of these if it is mycoplasma contamination .
please help me in this regard .
-Thank you
-Govind
Hi,
I'm afraid that indeed are positive stainings, notice the granular structures, primarily located on cell membranes, but also in the medium, dependend on the particular mycoplasma species. See the attached pictures for reference.
Do not confuse overstained cells for true positives (second attached, negative picture), overstaining gives a much more uniform staining of the cytosol (only observed if stained for too long or at too high dye concentrations).
How to manage this? My advice is to throw them out along with all already opened reagents (media, supplements, ...) and get a fresh stock, ideally from a reputable cell bank.
We tried to cure cells with sparfloxacin according to established protocols, however, the infection reappeared after several weeks, indicating that a few mycos survived the treatment. No problems with fresh stocks yet (touching wood...).
Hi,
You could also verify by using PCR based detection.
However, it does look positive by DAPI staining.
Thanks
Hello Ellur.
Sorry, the cell are pretty bad by seeing the stained cell boundaries and diffused cytoplasm. Get rid of the cells and stocks stored. Get a new batch of cells from the company. Prepare all the solutions involved with proper sterile work condition.
Good luck.
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