p53 immunocapture kit (ab154470) is specific to the p53 protein. Control proteins HSP60 and actin are present in the Hek293 and MCF7 lysate inputs, but are not found in p53 immunoprecipiations. Hek293 cells were extracted with lauryl maltoside (lanes 2, 9), RIPA buffer (lanes 3, 10) or SDS (lanes 4, 11). MCF7 vehicle-treated cells were extracted with lauryl maltoside (lanes 5, 12) or RIPA buffer (lanes 7, 14). MCF7 cells treated with 1 µM camptothecin were extracted with lauryl maltoside (lanes 6, 13) or RIPA buffer (8, 15). Extracts of whole cells (20 µg, lanes 2-8), and one-fifth of immunoprecipitation samples using the p53 immunocapture beads (1 mg extract per 10 µL beads, lanes 9-15), were analyzed by Western blot using ab46798 anti-Hsp60 and ab46805 anti-muscle actin.
Western blot with ab154470 and MCF7 camptothecin-treated cells All lanes: ab32389 anti-p53 1/1000 Lane 1: MW marker Lane 2: MCF7 cells extracted with lauryl maltoside (LM). Lane 3: MCF7 cells treated with 1 µM camptothecin for 6 hrs, extracted with LM. Lane 4: MCF7 cells extracted with RIPA buffer. Lane 5: MCF7 cells treated with 1 µM camptothecin for 6 hours, extracted with RIPA buffer. Lane 6: IP with 10 µL p53 IP beads and 1 mg MCF7 cells, LM extract. Lane 7: IP with 10 µL p53 IP beads and 1 mg MCF7 cells treated with 1 µM camptothecin for 6 hrs, LM extract. Lane 8: IP with 10 µL p53 IP beads and 1 mg MCF7 cells, RIPA extract. Lane 9: IP with 10 µL p53 IP beads and 1 mg MCF7 cells treated with 1 µM camptothecin for 6 hrs, RIPA extract. Lane 10: IP with 10 µL ab135397 beads and 1 mg MCF7 cells, RIPA extract. Lane 11: IP with 10 µL ab135397 beads and 1 mg MCF7 cells treated with 1 µM camptothecin for 6 hours, RIPA extract.
