Hi Nidia

Its Best to get rid of contaminated cultures.  fungal spores from the culture may contaminate your incubator and can contaminate other cultures.  Once you remove the contaminated culture, sterilize your lab equipments including your incubator. You can wipe it with hypochlorite and ethanol.

If you need to rescue the contaminated culture replace medium with medium containing antibiotic/antimycotic cocktails and also add gentamycin. You can use fungizone for eradicating fungus.

good luck!!!

Hello Nidia, I would also advise you to take a look at the water bath inside your incubator, and a water bath that is used for warming up media (if you are using one). Those can be sources of repeated contamination. You can use special reagents that are added to the water and prevent growth of bacteria and fungi (e.g., https://www.applichem.com/en/shop/product-detail/as/incuwater-cleantrade/ ). I also agree to the suggestion above: trash all the contaminated cell lines. It feels like losing a lot of work, but in the long run, it might be the only safe option. Also, check all the solutions you are using for cell culture, and autoclave all the plastic that can be autoclaved.

Ola' Nidia!

If your cells are too precious, besides the advice given before I'd recommend treatment with nystatin 100U/ml and slow weaning, as amphotericin B (fungizone) is not as effective against yeasts.

More tips in

https://link.springer.com/protocol/10.1385/1-59259-406-9:309/fulltext.html

Best of luck!

Thank you so much for all your advice! We checked all our reagents and started from the bottom, by using new ones. And we obviously discarded all contaminated cells. We are now using thawed new ones and things are working well, without any contamination in sight. At least for now!

I really appreciate your help. Thank you!