I completed a western blot and during detection, there were black dots all over the membrane on the output. What could have caused this?
What transfer buffer or method do you use for Western blotting?
I have removed SDS from the transfer buffer recipe for wet (tank) blotting because the presence of SDS tended to cause black dots.
In the case of IRS and ACC which have relatively large molecular weights, the blotting efficiency was somewhat lower, but transfer was also possible without problems when blotted for a long time at low voltage with well-chilled conditions. (30-45V, 3h)
For transfer from SDS-PAGE to the membrane , SDS left on the gel should be sufficient.
For semi-dry, please wait for answers from others.
Good luck!
Yasuhiro Nishida
I also agree with the suggestions given by Yang Wang. BSA dissolution is particularly prone to problems.
However, I feel that the overall background tends to be black/dark when blocking is problem. When dissolving the antibody, if the BSA or skim milk is not completely dissolved, you will get unspecific spotty or speckled noise on the detection.
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