Hi All,

I am trying to detect phosphorylation of a protein in cell lysates, I do immunoprecipitation of the target protein then look for Pan-phosphotyrosine by western blot.

I did precipitate the target protein however when I IB for P-tyro I just see these very bright bands that give no signal with chemiluminescence developing.

I am wondering what is the reason for that?

I tried diluting the immunoprecipitated lysates and lowered the dilution of the secondary antibody, by which I was able to see some bands but still not able to visualize all bands!!

How to overcome that?

Any help would be appreciated