weird problem in transfecting 293 cells in 24-well plate

I am doing transfection in 293 cells in 24-well plate. The transfection
reagent is Lipofectamine2000 (Invitrogen). The problem is after adding
DNA-Lipofectamine complex, in the some of the late wells (such as in Line D
and sometimes also in Line C) of the plate, the cells always die up to
10-30%. But in the wells of Line A and Line B, the cells always do not die
at all. The DNA problem is ruled out. I wonder what the reason is and how
to avoid the problem.

On Mon, 16 Apr 2007 11:40:23 +0800, "??????" <[Only registered users see links. ]>
wrote:


Well, it is hard to know what the problem is, but I can suggest a
general strategy for trying to find out.

For example, it might be something in the filling or something in the
incubation. To distinguish... Fill two plates together, then turn one
around 180 degrees (reverse front/back) before putting in the
incubator. This tells you whether the "back" that is causing problems
is filling or incubating.

Break the process down to as many steps as you can, and do a similar
test for each.


You are lucky if the problem is fairly reproducible. Imagine trying to
track down an erratic problem.

bob