Fig. 2

Immunocytochemistry. Sub-confluent cells were fixed by 10% formalin and were permeabilized by 0.1% triton X-100 in PBS. The endogenous peroxidase activity was blocked by incubation in 5% H₂O₂ in methanol for 20 min. The cells were pre-blocked using Ultra V block (Lab Vision) for 10 min and incubated for overnight at 4°C with respective antibodies. The cells were washed in PBS then incubated with anti-goat biotinylated secondary antibody (Santa Cruz Biotech) for 30 min followed by another wash and incubation with HRP-conjugated streptavidin. Finally, reactions were visualized by incubation with DAB (substrate and chromogen) and counterstaining with Harris hematoxylin. For negative control, cells were incubated overnight with dilution buffer (no primary antibody). Representative examples of expression of Bcl₂ in untreated TSUPr1 (a). In 250 nM 5-azadC-treated TSUPr1 cells, the amount of expression of Bcl₂ was very high (b). Caspase-3: there was no expression in control/untreated TSUPr1 (c). The amount of expression in 5-azadC 250 nM 5-azadC-treated TSUPr1 cells was very high (d). Bax: there was no expression in control/untreated TSUPr1 (e), reasonable in 5-azadC 250 nM 5-azadC-treated TSUPr1 cells (f)