anti-Perforin (human), mAb (deltaG9) (APC)

Figure 1: Binding of anti-Perforin/APC to fixed permeabolized stimulated human PBL. Methods: Ficoll-prepared human peripheral blood mononuclear cells were stimulated by incubating 2 days at 5 x 10^6 cells/ml in RPMI 10% FBS media including 5µg/ml Phytohemagglutinin-P. Cells were then fixed for 30 min with 2% buffered formaldehyde after which they were washed two times. Subsequent reagent incubations and washes were done using a buffer containing 0.3% Saponin to permeabolize cells. Five x 10^5 cells per tube were washed and incubated 45 min on ice with 80µl of anti-Perforin/APC at a 1:50 dilution (5µg/ml). Cells were washed three times, fixed and analyzed by FACS. A net 16.1% subpopulation of the cells shifted positively with a mean shift of 1.49 log10 fluorescent units when compared to an IgG2b/R-PE isotype Control (#ANC-284-060). Binding was blocked when cells were pre-incubated 10 min with 20µl of 0.5mg/ml unlabeled anti-perforin antibody (#ANC-358-020).