Cell lines have different properties and are sensitive or insensitive to different manipulations. This is certainly true for transfections and different cell lines require the development of an efficient transfection protocol to optimize efficient delivery while minimizing toxicity. Here we provide our esiRNA transfection protocols for some cell lines used in our laboratory. The values provided here can only serve as guidelines and a detailed optimization should be carried out in the individual laboratories. We recommend using an eGFP expressing cell line in conjunction with an eGFP and a control esiRNA to optimize the transfection protocol.

Cell line	Origin	Confluency	Amount esiRNA	Plate format	Transfection method/reagent
R1/E	129Sv Mouse ES	15-25%	50 ng	96 well	Lipofectamine 2000
E14Tg2a	129Sv Mouse ES	15-25%	50 ng	96 well	Lipofectamine 2000
E14Tg2a	129Sv Mouse ES	15-25%	20 ng	384-well	Lipofectamine 2000
E14Tg2a	129Sv Mouse ES	15-25%	500 ng	6-well	Lipofectamine 2000
E14Tg2a	129Sv Mouse ES	15-25%	7500 ng	10-cm	Lipofectamine 2000
SW48	Human Colorectal carcinoma	20-30%	40 ng	96- well	Oligofectamine
DLD-1	Human Colorectal carcinoma	20-30%	50 ng	96- well	Lipofectamine 2000
HCT116	Human Colorectal carcinoma	20-30%	50 ng	96- well	Oligofectamine
HCT116	Human Colorectal carcinoma	20-30%	1200 ng	6-well	Oligofectamine
RKO	Human Colorectal carcinoma	20-30%	50 ng	96- well	Oligofectamine
HeLa	Human cervical cancer	20-30%	15 ng	384-well	Oligofectamine
HeLa	Human cervical cancer	20-30%	30 ng	96-well	Oligofectamine
HeLa	Human cervical cancer	20-30%	1000 ng	6-well	Oligofectamine
U2OS	Human osteosarcoma	25-35%	60 ng	384-well	Effecten
NIH3T3	Immortalized mouse embryonic fibroblasts	10%-20%	50 ng	384 well	Lipofectamine RNAiMAX
NIH3T3	Immortalized mouse embryonic fibroblasts	10%-20%	200 ng	96-well	Lipofectamine RNAiMAX
NIH3T3	Immortalized mouse embryonic fibroblasts	10%-20%	1600 ng	12-well	Lipofectamine RNAiMAX
HEK293	Human embryonic kidney	20-30%	25 ng	384-well	Effectene