Hi Mohit, These precipitates appear to be protein and some crystals. This type of artifact appears more when you inactivate the serum or when it freezes or thaws again and again. They are toxic to cells when they are in high concentration. In general I try to centrifuge the serum at high speed (5000 rpm, for 30 min) and discard the pellet. Good luck Cynthia.

PS: We was having this problem with GIBCO FBS in the pas ...We decide to change to Hyclon

Hi Mohit,

I agree with Cynthia, it can be a protein precipitation due to repeated freeze and thaw of the FBS. I suggest to make aliquots of FBS in 50 ml Falcon.

As per the pictures you share it doesn't seems to be contaminated. The haziness in media may be due to dead cells and there debris (not due to contamination). Are the cells from continuous passage or cells were cryopreserved, and used later? If they were cryopreserved I would like to know your freezing protocol. Sometimes cells not stored in optimum condition may also lead to such problems. I would also suggest you to grow your cells in 6 well plates (as few are attached to surface). Remove the non attached cells after 12 hours of seeding (healthy cells should be attached by the time). Then sub culture the cells when confluent.

Hope this works,

Regards

Shubham

Hi Mohit,

It's difficult to make out from the pictures, however if you suspect contamination you may want to give this a read: Article Got black swimming dots in your cell culture? Identification...

I bought this cell line from ATCC, after about a week culture, cells were suddenly dead, came off flasks and floated in medium, it apparently a problem of mycoplasma or something reached its point to kill cells, contacted ATCC for a replacement but refused, eventually received a gift from a research and now the cell line grows very well.