I have been doing wb since I started the phd and they always were ok, but now I'm having problems with a new set of samples. I thought the problem was the amount of protein because I could not see any protein, even the actin, but I repeated the western and the signal was right. Yesterday I did a stripping of the membrane which has also workd always and I didn't get any signal. It is not that I don't have signal, I have a weak signal or in one case I had no signal.
I have done the tbstween 1x and the milk tha same day as I did the western and the antybodies are recycled so they work.
Could you help me?