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I am getting very low transfection efficiency. Can you please provide some troubleshooting tips?

Here are some reasons why you may be getting low transfection efficiency, along with suggested solutions. 

Why do I see cytotoxicity after performing transfection? Can you please help?

Here are possible causes for reduced viability following transfection, along with suggested solutions. Please note that as per recent findings, antibiotics can be used in media during transfection.We have compared transfecting cells in medium with and without antibiotics in multiple cell lines, assessed both the transfection efficiency and toxicity, and found no difference. For stable transfections, wait at least 72 hours after transfection before adding selective antibiotics.

Why are my transfections not reproducible?

Please review these possible causes for non-reproducible transfections, along with suggested solutions. Also, we recommend setting up one master mix of the DNA/lipid complex for the number of transfections planned to reduce multiple pipetting errors. When adding your complexes, we recommend changing tips between wells since re-used tips could bring carryover, especially for the 96- or 384-well format with small-volume formats.

I see a small granular precipitate on my cells (microscopically) following addition of the transfection reagent:DNA complexes to my cells. Will it decrease the transfection efficiency?

A common reason for seeing precipitate on your cells following lipid-based transfection is if there is excess EDTA or cationic lipid present. We recommend diluting the DNA in water or, if TE is preferred, use EDTA concentrations of <0.3 mM. Also ensure that concentrations of cationic lipid reagents do not exceed recommended amounts during complex formation. The presence or absence of this precipitate is not indicative of the transfection performance. 

I have tried several different transfection reagents and have failed to transfect my gene into my cell line of interest. Do you have any suggestions?

We recommend that you try electroporation as a method of delivering your plasmid of interest. We offer the Neon® Transfection System for highly efficient transfection of primary cells, stem cells, and difficult-to-transfect cells.

You may also consider using a viral-based system to deliver your gene into your mammalian cell line of interest. 

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