Hi everyone,

I have a question about the transfection of my RAW264.7 cells.

I have produced GFP fusions with my protein of interest (POI) on both the C- and N-terminal side. I have validated these fusions in HEK293T cells, they localize like the full protein and have similar luciferase activity and look similar on a western blot. I also transfected them with electroporation in Jurkat cells which worked.

However, I would now like to transfect them into RAW264.7 cells. By using Lipofectamine LTX, I have been able to transfect RAW264.7 cells with CMV-GFP and CMV-POI. However, GFP-POI and POI-GFP cannot be expressed in these cells using this system, which I do not understand! The cells will not become green and cannot be seen on a Western Blot.

I also cloned them into a pLenti vector to transduce the RAW264.7 cells. But also in this way the cells do not become green (but do with pLenti CMV-GFP).

Does anyone have an idea why these plasmids do not work in RAW cells but do in other cell types?

Thanks in advance!