I am using Lipofectamine 2000 to transfect my cells and it did work. Now I want to know whether my cells for real are transfected or not? ( just to confirm)
what kind of assay I should do? or does check the protein by doing westren blot will be enough to answer that.
Thanks
whatever is the easiest assay to run in your lab. eg you can: quickly measure protein levels for fLuc or bGal using fluorometer (if you transfected plasmid). or measure siRNA-induced knockdown by measuring target mRNA levels by qRT-PCR, with TaqMan (if you transfected siRNA). Using positive controls is a good idea- many companies are advising to use specific positive controls, for which easy readout is widely available, to confirm that delivery worked well
Alexander Sasha Vlassov's suggestion will work equally as well for miRNA as siRNA - look at the knockdown of your target by measuring the target mRNA levels. A western blot for the target protein would work as well, as long as you have a negative control (vector with non-target sequence) and a housekeeping gene to monitor RNA and/or protein loading.
A more complete explanation:
https://www.thermofisher.com/us/en/home/references/ambion-tech-support/microrna-studies/tech-notes/conveniently-monitor-mirna-transfection-and-function-at-the-prot.html
If you want to prove equal transfection efficiencies throughout several experiments I suggest to co-tranfect another plasmid encoding for a fluorescent protein. So by checking the amount of fluorescent cells vs. all cell you have in your culture, you get some quantification of the efficiency. If you use always the same amount of fluorescence-encoding plasmid, it will be possible to compare different experiments for their efficiencies.
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