Anti aif antibody

Following treatment, the cells were washed with 1 mL of prewarmed PBS and then fixed in 4% formaldehyde in PBS for 15 min at room temperature. The coverslip was washed in 1x PBS and then incubated in 2% Triton X-100 in PBS for 15 min at room temperature. To prevent nonspecific binding the coverslip was incubated in blocking buffer (3% BSA, 1% Triton X-100 in PBS) for 1 h at room temperature. Anti-AIF antibody (0.5 μg/mL; 2 mL) (Abcam, Cambridge, MA, USA) was added and the mixture was incubated for 4°C overnight. The coverslip was washed before incubating in 2 mL donkey anti-rabbit IgG (1 : 1000) (Abcam, Cambridge, MA, USA) for 1 h at room temperature. After washing the coverslip, it was incubated in Hoechst 33258 stain and prepared and analyzed as described above.