Hi, I am new to this forum.
In our laboratory we experience some weird problems regarding the Western blot protocoll we used for years.
It all started about 3 month ago. We didn´t get any signal with the normal ECL (GE) we use.
With the enhanced ECL (Millipore) we mostly get excessive signals but not as long as normal.
When we put a film on, 5 min after appling the ECL there is no signal.
In some occasions we see „light“ areas (like clouds).
And the keep staying at the same place everytime we try another ECL (so first standard ECL, then wash with TBS-T, then enhanced ECL -> Same clouds)
We got a fresh Nitrocellulose (Millipore), ECL (another Lot), non-fat milk (changed from noname to Bio-Rad, which seemed to work better… but not for long).
I tryed reducing the milk concentration from 5% to 3% with the old milk – that appeared to be a little bit better. But with the new Bio-Rad milk we continued to use 5% since it normaly should work. It appeared to work with the new milk.
But the problems continue… Anyone any tips?