I have been culturing VCaP cells now for some time and learned a few tricks. At first I struggled getting the cells to passage properly and was killing flask regularly. Since I  can grow them pretty quickly. Here's how...I grow all cells in T-25 flasks and in DMEM with 10% FBS and 1% Anti-anti. Once I add trypsin to the flask I allow them to sit at room temperature until they begin to lift off the flask. At this point I gently swish the medium back and forth and once most cells are off the flask I add full medium back to the flask. I have since learned that VCaP cells are sensitive to residual trypsin, so now I take the trypsin/medium mix and spin the cells down and remove all trypsin/medium mix and flick the cell pellet to remove all clumps of cells so its ~500 uL of cell slurry in the tube. I then place 1/3 or 1/2 (no less) of those cells back into the flask with 5 mL of full medium and 1 mL of the old medium. I have found VCaP cells grow best with some of the old cultured medium (might be some cytokines or something to signal growth). With this method I have no problem with the flasks becoming completely confluent in 4-5 days time.  Hope this helps.

VCaP Cells may take 24 to 48 hours to attach during recovery and after subcultures. The cells grow very slowly. For more detailed information, see below.

http://www.atcc.org/products/all/CRL-2876.aspx#documentation

Thanks but do you guys use any coated cell culture flasks other than the regular cell culture treated polystyrene flasks.

Cell culture treated T-25 or T75 flask from BD works just fine.

Plz, check this:

http://capcelllines.ca/details.asp?id=159

I am having the same problem with VCaP cell growth.

I am growing them in a T25 from BD (vented tap) in DMEM (sigma D5796) + 1/100 Sodium pyruvate + 1/100 MEM non-essential amino acids (sigma M7145) + 1% Penicillin/Streptomycin. The problem is that I thaw them 8 days ago, and anthough the number of cells had incresead, only few (up to 20) cells have attached. I am adding fresh media every 2 days (I remove the media, centrifuge, ressuspend cells in fresh media and put them back in the T25). I don't have trypan blue positive cells.

Rahul, did you overcome your problem? Do you have any suggestion for what might be happening? Should I use different media/culture dishes?

Thanking you in advance,

Isabel

I have been culturing VCaP cells now for some time and learned a few tricks. At first I struggled getting the cells to passage properly and was killing flask regularly. Since I  can grow them pretty quickly. Here's how...I grow all cells in T-25 flasks and in DMEM with 10% FBS and 1% Anti-anti. Once I add trypsin to the flask I allow them to sit at room temperature until they begin to lift off the flask. At this point I gently swish the medium back and forth and once most cells are off the flask I add full medium back to the flask. I have since learned that VCaP cells are sensitive to residual trypsin, so now I take the trypsin/medium mix and spin the cells down and remove all trypsin/medium mix and flick the cell pellet to remove all clumps of cells so its ~500 uL of cell slurry in the tube. I then place 1/3 or 1/2 (no less) of those cells back into the flask with 5 mL of full medium and 1 mL of the old medium. I have found VCaP cells grow best with some of the old cultured medium (might be some cytokines or something to signal growth). With this method I have no problem with the flasks becoming completely confluent in 4-5 days time.  Hope this helps.

Hi Dan,

Thanks for your help! I will try what you say... I was already remoning the trypsin, but its a good sugestion to add a bit of the previous medium...

Thnaks!

@Dan Binzel, thanks for your suggestion. Do you use the same procedure to add some old conditioned medium while doing experiments such as cytotoxic assay or transfection assay.

Thank you!

Hi Chuangzhoung. Sorry for the late reply. Hopefully you have solved any issues with the culture since you asked. But yes anytime I am culturing the cells and need them to be adhered and healthy I will use a small amount of the old medium to keep them happy. Hope this helps.