hello,
I culture FaDU ( oral cancer cell) for few time but i facing problem with their confluence and attachment, even i used standard procedure for culturing? i use DMEM with 10% FBS and desire amount of antibiotic too.. i cultured 2.0 X 10E5 cell but after overnight i didn't get 70% confluency and attachment. any one did work with these cell please suggest me some critical points about it.
Hi, Are you using the required supplements that would be needed for this particular cell line. I have seen some people add L-Glutamine which is a basic supplement. Another interesting thing with this cell line is that they need to seeded at a higher density for optimal growth properties. Depending on the tissue culture flask volume you are using, you will need to seed these cells at a higher density. I feel 200,000 cells is too less seeding density for these cells. Hope this helps.
Dulbecco´s MEM (bei uns üblich: Fa. Biochrom) mit
10 % FCS,
2 % HEPES,
1% Non Essential Amino Acids,
1 % Sodium – Pyruvat und
1 % Penicillin/Streptomycin.
Adding insulin would be a good option in addition to the supplements you already have. Plus taking care of seeding density.
In many ways u can achieve your desire cell confluence by preparing your medium, monitoring cell condition and so on. First step u may increase the FBS concentration!, at present you are using 10%, is it, so you may increase gradually to 20% and antibiotics too, then, if you are using cells directly from stored liquid nitrogen, you just wait two or three days for adapting cells new environmental behavior, sometime it will take more than 12-24 hrs, its based on the cell types etc. one more thing is, if your cell are very older (more than a year) it will take time to grow, actually it may affected by genetic drift. (if possible, try to purchase new). thanking you
I would suggest to do a mycoplasma testing, immediately!! I have worked with them extensively for xenografts and face these problems. Normally they are good till they reach a confluency of 50-60%. However as soon as they reach to getting confluet they came off. Its know fact that many times when you recieve the cell line from ATCC , for eg. , they are mycoplasma+ these has to be rectify.
This question is a few months old so I hope you have sorted out your problem now. But, thought I'd mention that we grow them in RPMI-1640 with L-glutamine plus 10% FBS and antibiotics, and find them to be very robust. In my experience they will stay very well attached even if left to grow to overconfluence.
So it sounds like it could be a problem with your particular batch of cells, and the recommendation for mycoplasma testing was certainly a good one. We do test ours, and another check is to test them for spheroid formation (which we do before using them for a xenograft experiment) because FaDu should do so readily.
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