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** |
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