Low passage HEK293?? - (Sep/19/2012 )
Some protocols suggest using "low passage cells" for transfection. What is a 'low passage' (p0-5??) and when is it advisable to stop using these HEK293 cells for transfection?
I find that HEK293 lifts easily from plastic during media change, so is that technically considered as ONE passage?
A passage is usually defined by reducing the number of cells - so the lifting off wouldn't normally be considered a passage. TO prevent the cells being washed off the flask/plate, dribble the medium down the sides of the flask.
Low passage is tricky with immortal cells - in this instance they mean cells that havn't been thawed for very long. After about 6-10 passages the cells tend to get more difficult to transfect,and also start to show characteristics of getting "old" - slower growth rates, ragged cells, more senescent cells in the population.
Having said that, 293 are very very easy to transfect, so I doubt you will have a problem.
bob1 on Thu Sep 20 09:32:45 2012 said:
A passage is usually defined by reducing the number of cells - so the lifting off wouldn't normally be considered a passage. TO prevent the cells being washed off the flask/plate, dribble the medium down the sides of the flask.
Low passage is tricky with immortal cells - in this instance they mean cells that havn't been thawed for very long. After about 6-10 passages the cells tend to get more difficult to transfect,and also start to show characteristics of getting "old" - slower growth rates, ragged cells, more senescent cells in the population.
Having said that, 293 are very very easy to transfect, so I doubt you will have a problem.
Say p>20?
That should be fine, though I generally recommend that cells only be kept up for about 10 passages to minimise the amount of "aging" that they undergo and to prevent selective pressures becoming obvious.