Effect of PFTα on Bax conformation change induced by p53-elevating agents. Cells from patient 26 were incubated in the presence of ZVAD. Chlorambucil, nutlin and PFTα were added as indicated. Following 18-hour incubation, cells were lysed in CHAPS buffer. (A) Lysates were analyzed by immunoprecipitation with conformation-specific Bax 6A7 antibody. Unprecipitated lysate from cells incubated with nutlin + PFTα was loaded on the right hand lane to define the migration positions of Bax, Hsp60 and actin. The nutlin + PFTα lysate was also used to generate the negative control in which the 6A7 antibody was omitted from the immunoprecipitation reaction (penultimate lane). (B) Aliquots of the lysates were analyzed directly by Western blotting to document equal inputs into the corresponding immunoprecipitation reactions shown in panel A. Similar results were obtained using cells from 2 additional patients (patients 8 and 9).