Hello, everyone,
I've been doing Western-blocks for years.
For a few days now I have had problems, even with "easy" antibodies such as GAPDH. According to my findings, proteins are in good condition (Ponceau shows defined bands) and the transfer is efficient, but I don't detect any signal.
Looks like something stopped working.
On Wednesday I got signal by changing all the reagents, but the next day I repeated it and got no signal.
This problem has coincided with an increase in room temperature (summer is coming). It seems I can't recycle first antibodies, when I usually recycle GAPDH several times. And the reagents also seem to have to be prepared fresh the same day.
Has anything similar happened to anyone?
Thank you very much.