I take the cells trypsinize the cells and wash them with cold PBS two times.
Then i use RIPA lysis buffer according to the pellet and add protease inhibitors 1/20th volume of lysis buffer. Pipet it for mixing and then incubate on ice for 15 mins. then i sonicate it for 3 cycles of 20 seconds each.
Further i centrifuge at 14000g for 30 mins at 4C.
and then take the supernatant and make aliquots.
and i am getting these type of gels.
Thanks