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I’m having problems with the transfection of my siRNA. What should I do?

Please visit our Transfection Support Center to find technical resources, tips and tricks, and troubleshooting information on transfection.

How long can the diluted transfection lipid sit out at room temperature before losing transfection efficiency?

This is cell line and reagent dependent but we have found that diluted Lipofectamine® RNAiMAX reagent is quite stable. We tested up to 2 hours following transfections into HeLa cells, with good knockdown. However, we noticed a difference if the lipid was diluted in a tissue culture plate versus a conical tube, but if the transfections occur within an hour there isn‘t a problem.

How long can the siRNA and lipid complex before losing efficiency?

This depends on the cell line and reagent, but we found this complex is stable. We tested up to 3 hours, and there was no decline in knockdown in HeLa.

What are important parameters to consider for transfection optimization?

The goal of transfection optimization is to determine the conditions that will provide maximum gene knockdown while maintaining cell viability for the particular cell type. There is no single transfection parameter that by itself ensures efficient uptake by cells in culture. However, there are several critical variables to consider in systematically addressing the optimal siRNA uptake into cells: health of cultured cells, cell type, transfection method choice, volume of transfection agent, exposure time of cells to transfection agent, and cell density.

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