They are proliferating but does not attach to the T75 and T25 cell culture flasks. I also used 6 well plate to attach the cells but they did not attach. I tried to use different combinations of media solution, such as DMEM/F12 + %10 FBS + 2mM L-glutamine; DMEM/F12 + %20 FBS + 2mM L-glutamine; DMEM/F12 + %10 FBS. Then I incubate the cells at 37oC in a humidified atmosphere of 5% CO2 incubator.
Do you have any ideas ? What is wrong that I do?
Hello
The cells may be under stress. Do you notice any contamination in culture?
Moreover, BeWo cells show high glycolytic activity and so the culture medium has to be changed every 24-48 hours in order to prevent glucose exhaustion. Subculture the cells when they are 70-80 %confluent.
Avoid using 20% FBS in culture medium as it is not recommended for this cell line, and it could be toxic to the cells.
Best Wishes.
Dear Beyza!
the reason for the non-attachment of cells may be in the vials themselves. Look at the brand and brand of your vials. There are vials with relatively weak adhesion, and there are high ones. It is better to use vials with high adhesion for such cells.
Also pay attention to the density (number) of cells. If there are few cells (low seeding density), they may not adhere well.
Article The use of BeWo cells as an in vitro model for placental iro...
Cell culture.
BeWo cells subclone b30 (kindly donated by Dr. Alan Schwartz, Washington University, St. Louis, MO) were routinely cultured in 75-cm2 flasks (Fisher Scientific, Loughborough, UK) at 37°C and 5% CO2 in high-glucose DMEM containing glutamax1 (Invitrogen, Paisley, UK), penicillin (100 U/ml), streptomycin (100 μg/ml), and 10% vol/vol fetal calf serum (Sigma, Poole, UK). Cells were subcultured every 7 days, and the medium was replenished every 2 days. For transport experiments, cells were seeded onto permeable membrane supports (Thincerts, Greiner Bio-One, Stonehouse, UK) at a density of 8 × 104 cells/cm2.
Hello
Have you ever coated a culture flask with 0.1% gelatin?
BeWo cells are thought to have low adhesion. gelatin helps cells adhere.
I just wanted to update.
I solved the problem by putting the medium in the incubator 30 min-1 hour before adding the cells to the medium. Thus, the temperature and CO2 balance of the medium was maintained. Also, when I thawed the cells, I placed them in a 11 or 22 cm3 petri dish so that they were close to each other.