( ll plates we recommend to seed 0.94.0 ...)Transfection Reagent Selector Kit Handbook

ll plates, we recommend to seed 0.94.0 x 105 adherent cells in 2 ml of medium per well the day prior to transfection, and 1.03.5 x 106 suspension cells in 1.6 ml of medium on the day of transfection. The actual number of cells depends on cell type and size. If you prefer to use a different culture format for your determination, please contact QIAGEN Technical Services for the recommended number of adherent or suspension cells to seed.

The optimal confluency at the time of transfection complex addition is normally 4080%. The optimal confluency should be determined for every new cell line to be transfected, and kept constant in future experiments. This is achieved by counting cells prior to seeding and by keeping the time period between seeding and transfection constant. For suspension cells, split the cells the day prior to the transfection experiment. This will ensure that the cell density is not too high and that the cells are in optimal physiological condition on the day of transfection.

Amount of DNA

The optimal quantity of plasmid DNA used for transfection is determined by the properties of the transfected plasmid which include: type of promoter, origin of replication, and plasmid size. Toxic effects may arise if too much plasmid with a high expression rate is used. Conversely, if insufficient plasmid with a low expression rate is used, gene expression may be too low. Therefore, optimization of plasmid DNA concentration should be performed for every new plasmid and/or new cell line used. A pipetting scheme for optimizing the amount of DNA for transfection in 6-well plates is provided in Figure 2.

Amount of Enhancer (Effectene Reagent
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