Does anyone has good protocol/reagent for transfecting U937 cells with siRNA? It is a human macrophages and looks like they are quite sensitive and float/die after, even with very low amount of lipid based reagent and oligos on forward transfection.
I am looking to transfect after 3 days PMA differentiation and induced with interferon after siRNA for 2 days.
I believe U937 is a suspension cell line, so electroporation is generally gonna be the way to go. Look for electroporation reagents that are for AML cell lines (like K562: https://altogen.com/product/k562-electroporation-kit-aml-leukemia-cells-ccl243/) and those should be applicable to your transfection. I've noticed that lipofection of suspension cell lines only gets around 10% transfection efficiency, but electroproation can get up to 80% efficiency, especially for siRNA transfection.
hi Ted, thanks for the input, they are monocytes in suspension, but we normally differentiate them into macrophages, meaning that they attach at the time I did the transfection though.
Interesting, I haven't personally conducted such kinds of suspension-turned-adherent cell transfections, but if your cells are adherent then lipofection should work. Given the issues you've been facing, it seems to me on a surface level that your concentration might be too high or that your medium isn't suitable for the cells. Maybe do an optimization round of different concentrations, and that might give a direction to head in. Also you might want to read this study (Article Transfection of adherent murine peritoneal macrophages with ...
) which seems to deal with the same kind of transfection you are.- Badges
- Science method