Low passage HEK293?? (View forum version)



science noob

Posted 19 September 2012 - 06:39 PM

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?

bob1

Posted 20 September 2012 - 01:32 AM

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.

science noob

Posted 21 September 2012 - 02:06 AM

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?

bob1

Posted 23 September 2012 - 12:48 PM

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.