I tried several times using the Lipofectamine 2000 reagent, extracted the proteins but couldn't detect on Western blot. My plasmids were constructed with PcDNA 3.1+ Please does anyone have any suggestions? Thank you.
Noted, thank you Sir Tom Masi Will try that. I had a positive control, though not GFP but I'll try a different transfection set up as you've suggested.
Any positive control should do, GFP is a prefferred one for most. Are you measuring the cells or the media? Could the protein be secreted? What promoter is driving expression of your protein? And how long after transfection are you collecting? It could easily be a problem with the antibody you are using to detect it as well.
Thank you Sir, Bruno Salomone Gonzalez de Castejon, we changed the method and got positive results. We still used the HEK 293 cells but with the PEI 40K reagent this time.
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