problem when doing transfection for the first time - (May/20/2006 )
Hi,everyone, I used SuperFect Transfection reagents from Qiagen to tranfect 293T cells, and I wonder at two point:
1)after the formation of tansfection complex, I will add the complex(1ml) dropwise into the cell monolayer, how can I make the complex distribute evenly across the monolayer?should I move the dish circurly or back and forth?
2)after the addition, the mannual says culture the cells with the complex for 2-3hrs before addition of medium, does it mean I shouldn't add medium until 2-3hrs after the addition of complex?
......I'm a beginner
1) try to add your drops as evenly as possible on your plate and afterwards gently stir your plate a little. should be enough.
2) mostly you should do transfection in a smaller amount of medium than you would culture your cells, or with a smaller amount of FCS. So afterwards you need to compensate for these (more medium + more FCS). Other people just remove the medium with the transfection agent and add fresh medium to the cells.