May be, its not a contamination, but a protein precipitate from FBS?

(When frozen serum is thawed small amounts can precipitate out. The more freeze thaw cycles, the more chance of precipitates forming. These are for the most part cosmetic and have no ill effects on cell culture. It is not recommended to try and filter precipitates out as they will clog most lab scale filter apparatuses. If necessary, allow the serum to stand and the precipitates will eventually fall to the bottom of the bottle and the serum can be decanted.

https://atlasbio.com/faq/precipitates-serum-bad/)

Hi Moresi Fabiana

To me it looks like cell clumping - I had a similar issue a few months ago and if your cells are susceptible to clumping, it would explain why you have been unable to get rid of it.

Here is an article published my Sigma: https://www.sigmaaldrich.com/technical-documents/articles/biology/cell-culture-troubleshooting-cell-clumping.html

You may have to reconsider your culturing protocols to remove.

I hope you get your cell culture sorted out!

When you restarted the culture twice Was it from single cell in a 12/24 well plate and scaling up from one cell per well dilution, the way its done for hybridoma cells, after treating with antifungal, and gram +/=?

please use Ivermectin its antiparasitic drug dosage will depend on the cell line higher. it seems to show some antiviral properties. so depending on use this has to be removed (Ivermectin) after testing if the growth has gone.

Also as Solovyev is telling if its a FBS precipitate its not expected to grow. and precipitate dont appear after say 2-3 weeks.

The possibility of the source cell line being contaminated ( so can another cell line be cultured in the same conditions. )