I've had issues with my secondary antibodies for ICC. They seem to form complexes which then result in bright artificial spots in the images which I cannot attribute to protein complexes rather than antibodies clustering together.
Did anyone have the same issue and found a way to resolve it? Just using the "supernatant" of the antibody aliquot doesn't do the trick.
Thanks!
Yulia Panina
Milena Korneck 1) Use less secondary antibodies (the amount may be too high), 2) wash samples VERY thoroughly to get rid of "dot" stains, especially the final wash, after you incubated with secondary antibodies. Wash like 4 times, as much as possible.
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