Raw (264.7) cell extracts (1mg protein) were immunoprecipitated with increasing amounts of sheep anti-mouse ABIN1 (s345c) antibody (lanes 1-3) or pre-immune serum (ane 4) for 2h along with 10 ul of protein G. Immunoprecipitated proteins were denaturd in SDS and subjected to SDS-PAGE on 8% gels. Lanes 5-8 indicated teh amount of mouse ABIN1 protein remaining in the supernatant after immunoprecipitation , the amounts loaded being equivilent to 20 ug of cell extract protein. Proteins were trasferred to PVDF membrane and immunoblotting was carried out for 2h at room temperature using 0.3 ug/ml anti-mouse ABIN1 antibody (s345c).