I am staining tissue sections and I did a mistake, I was supposed to have mixed 3 primary antibodies but I stained only with one of them. After the 1h incubation I washed 10 min with PBS and then I did another hour incubation with the 2 antibodies I forgot to add before. Can that affect my staining? I am afraid I wanted to test different dilution of the first antibody I added alone, and then I added another hour incubation with 2 more antibodies.


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