COS-7 Cell Electroporation

Frank O. Fackelmayer Frank at Fackelmayer.de
Tue Jul 17 03:02:16 EST 2001


For quicker electroporation, without the need to prepare special
transfection medium, we do it the following way: (optimized for 100mm plate)

* harvest subconfluent (e.g. 1 day after splitting) cells by trypsination
* wash once in 10ml complete medium
* for each electroporation, prewarm 10ml complete medium to 37°C in a
plastic tube
* resuspend cells in 800ul complete medium and add 10ug DNA (cesium
chloride purified)
* transfer mix to a cooled 4mm-electroporation cuvette
* pulse: 400V, 925-1000uFa, 0 Ohm (works well for COS7)
* IMMEDIATELY after the pulse, gently resuspend cells in the cuvette
with a pasteur pipette and transfer into the prewarmed medium; DO NOT
incubate the cells in the cuvette! Every additional second decreases
cell viability by 5-10%!
* mix the suspension and pour it onto a new 100mm plate
* put into the incubator for at least 24h
* analyse cells

COS7 cells are easy to transfect, and will yield at least 30% of
transfected cells with the method above. Carefully optimized conditions
(e.g. voltage) will allow for efficiency up to 70%. Some other cells do
not perform that well, and do not stand that high a voltage. In all
cases, it is important that the cells be healthy and well growing. 


Frank




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