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Ken's Lysis Buffer

19.5 mL L-RIPA
100 μL 100mM NaVO3
200 μL 100X PMSF
100 μL PIC I
100 μL PIC II

Total Volume of 20 mL

1. Rinse cells with cold PBS, add 1 mL of the lysis buffer, scrape plates and transfer cells to a new tube.
2. Vortex on high for 10 seconds and then incubate on ice for 15 minutes.
3. Boil tube at 100 ̊C for 5 minutes, put on ice.
4. Spin in the cold room for 5 minutes on high (use this supernatant for Bradford assay)
5. Add 4x loading buffer to the supernatant, boil for 10 minutes and place and ice.
6. Spin again for 5 minutes on high.
**Use this supernatant to load the gel**