My protein size is 240kDa and I am interested in identifying the interacting target protein at endogenous level. I have earlier tried using the whole cell lysate but 4-5 mg of MEF cell lysate do not give any result. Instead overnight binding with 20 mg of lysate gave a diminished band. I suppose my protein appears to be low in cell.
Does cytosolic lysate interferes in the Immunoprecipitation Asaay?
Going for Nuclear lysate Immunoprecipitation assay can predict the binding? As other articles have shown the succesful interacting assays even at 60ug of lysate.
Any suggestions are welcome. Thanking you.