How long to wait until cell activation after trypsine use? - (Nov/29/2007 )
I'm culturing CD34+ cells. Now I'm wondering for how long I should wait after I have detached them by trypsine and seeded them in a new plate before I start my activation assay? 1 day, or more?
after trypsinization you have to check regularly whether your cells attached to the plate surface, and once they attached to about more than 50%, then change the medium of the cells to confluent, change every three day. when u see your flask fully confluent about 75% then detach your cell and use for your required assay.
Sorry, but I don't quite follow ...
I just wanted to know for how many days you should wait before activating or treating cells with something for ex. TNFa, after you detached them by using trypsine.
I have to detach them because I have to change the plate for my experience ...
Generally speaking you must leave the cells at least 48 hours to recover after subculturing. BUT you have to optimise this for yourself using your cells, the activation marker you are wanting to measure and growth conditions.
Cell Surface Receptor expression can take more than the time above. However cellular metabolic activity can be recovered within hours. ALSO remember that in general terms cell lines will recover quiker than primary cells that have been freshly isolated.
i leave them overnight.
ensure that all the cells are attached.
however, i don't work with CD34+, so i would suggest going into pubmed, and looking through the methods sections to see what other researchers using this cell line do.
Thank you very much for your help!
According to what you suggest in general, I think I'm not too wrong with what I do. Until now I always left them for 48h and then activated my cells.
when we want to treat our cells with drug, hormone, growth factor,... we seed the cells in plates and let them attach for 24 hours, then we wash the cells with PBS and add the new medium containing the required concentration of the hormone, etc.. . and then incubate them for the desired time.