Too Happy Cells - Cells are rapidly growing 7 affecting transfection (Jul/23/2007 )

I ordered and received new medium (DMEM 1X) and added 10% FBS as always, and ever since then, my cells seem to get confluent much faster than before. I am trying to transfet these A549 cells with control and experimental siRNA, but after three days, when we usually notice EMT, I am seeing no drastic changes between the control and experimental cells, except that they are both heavily confluent. Would this affect the transfection efficiency, even if I transfect when the cells are about 50% confluent??

Any responses would help, thanks!

As a first guess: did U order the exact same DMEM than before? There are many differnet DMEM-s, it is not one standard medium (unfortunately). There are different flavours for US and Europe, there are w/wo Glutamine, pyruvate, high- and low-glucose, etc.

Rapidly dividing cells should transfect well on my hands. It has to be other reasons for what you are experiencing.

Kupac is most likely correct. I see changes in my cells from lot to lot of media and especially FBS. I doubt your actual initial transfection efficiency is changed but rather the siRNA is being split between the cells as they divide and therefor you aren't going to get as efficient of a knockdown. I actually transfect my cells (U2OS) twice with the siRNA, 24 hours apart (my desired knockdown protein is very stable). You can also bring the FBS down to 5% or even 2.5% to slow the cells down.