I’m seeing cell death after CRISPR RNA transfection. What can I do to prevent this?
Transfection reagents can be cytotoxic, and the extent of cell death can vary from one cell type to the next. If you are seeing excessive cell death, the first step should be to optimize the transfection reaction using our CRISPR Human HPRT Positive Control crRNA so you are using the least amount of lipid transfection reagent, without significantly affecting transfection efficiency.
You may also try a different transfection reagent, as minor differences in chemistry can impact toxicity. Make sure to look for transfection reagents optimized for RNA to transfect the Alt-R™ CRISPR-Cas9 RNAs. We have delivered these reagents successfully into HEK293 cells using TransIT-X2 (Mirus Bio) for transfection of the S.p.
Cas9 Expression Plasmid and RNAiMAX (Thermo Fisher Scientific) for the Alt-R CRISPR-Cas9 RNAs. You should empirically optimize transfection reagents for use in other cell lines.
It is also possible that your target gene is vital to your cells, and thus its knockout results in cell death. If the Alt-R CRISPR HPRT Positive Control crRNA
works and cell death is not observed, it is likely you are working with a vital gene. In this case you may have to try different cell lines. Learn more about our Alt-R CRISPR-Cas9 System products