Paraformaldehyde-fixed, paraffin embedded (Human brain glioma); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (Phospho-Caspase-9 (Tyr153)) Polyclonal Antibody, Unconjugated (SL3083R) at 1:400 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.
Blank control(blue): Hela(fixed with 2% paraformaldehyde (10 min) and then permeabilized with ice-cold 90% methanol for 30 min on ice)).
Primary Antibody: Rabbit Anti-Phospho-Caspase-9 (Tyr153)/FITC Conjugated antibody (SL3083R /FITC), Dilution: 1μg in 100 μL 1X PBS containing 0.5% BSA.
Isotype Control Antibody: Rabbit IgG/FITC(orange),used under the same conditions.