Perforin Antibody [CB5.4]

Application Note:

Perforin antibody recognizes only membrane bound active (non-cleaved) human FasL and not the inactive soluble FasL. This product can be used for detection of active human FasL by flow cytometry and western blot. Method: A mouse T cell clone was plated onto polylysine treated glass slides, fixed and permeabilized in methanol at -20oC for 5 min, then in acetone at -20oC for 30 sec. After 3 washings in PBS, 0.1% BSA, slides were incubated with 5 μg/mL of CB5.4 antibody, washed in PBS, incubated in anti-rat IgG-HRP, washed in PBS, and developed in appropriate substrate solution (DAB or AEC). Method: Cell extracts from the T cell clone B6.1 (2x106) were resolved by SDS-PAGE under reducing conditions, transferred to nitrocellulose and probed with the CB5.4 antibody at 1 μg/mL. Proteins were visualized using a peroxidase-conjugated antibody to rat IgG and a chemiluminescence detection system.