Establishing human neural stem cells - (Feb/24/2014 )

Hi

A week ago I defrosted and seeded a new vial of human neural stem cells. I have made up the media with appropriate growth supplements and followed the suppliers protocols, changing the media daily. The stem cells are taking longer than anticipated to become 90% confluent. The suppliers protocol says that they should be confluent within 4 - 7 days. Today is day 7 and they are still about 40% confluent. I have no infection in the flask and as I say I have followed the suppliers protocol exactly. Ideas anyone?

Thanks

So it is an adherent culture, are your flasks coated with something ?

Which medium do you use ?

Are the cells checked for mycoplasma ?

Yes its an adherent cell culture, flask coated with poly l ornithine and laminin as per protocol. I haven't tested for micoplasma, but the cells are direct from manufacturer so one would hope that they weren't infected!

This may be a stupid question, but when you say you followed the recommendations exactly, does this include the CO2 percentage ? I used to work with glioma stem cells (suspension, though) and the CO2 percentage can make quite a difference in growth.