Western blot analysis of extracts from NK-92 cells, untreated (-) or treated (+) with TPA #4174 (80 nM, 5 hr), Ionomycin #9995 (3 μM, 5 hr), and Brefeldin A #9972 (300 ng/mL, last 4 hr of stimulation), using IFN-γ (D3H2) XP ® Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).
Immunoprecipitation of IFN-γ from NK-92 cell extracts, treated with TPA #4174 (80 nM, 5 hr), Ionomycin #9995 (3 μM, 5 hr), and Brefeldin A #9972 (300 ng/mL, last 4 hr of stimulation), using Rabbit (DA1E) mAb IgG XP ® Isotype Control #3900 (lane 2) or IFN-γ (D3H2) XP ® Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using IFN-γ (D3H2) XP ® Rabbit mAb.
Confocal immunofluorescent analysis of NK-92 cells, treated with TPA #4174 and Ionomycin #9995 (80 nM and 3 μM, 4 hr; left) or untreated (right), using IFN-γ (D3H2) XP ® Rabbit mAb (green). Blue pseudocolor = DRAQ5 ® #4084 (fluorescent DNA dye).
Flow cytometric analysis of human peripheral blood cells, untreated (left) or treated (right) with TPA #4174 (40 nM, 4 hr), Ionomycin #9995 (2 μM, 4 hr), and Brefeldin A #9972 (1 μg/mL, last 3 hr of stimulation), using a CD3 antibody and IFN-γ (D3H2) XP ® Rabbit mAb. Anti-rabbit IgG (H+L), F(ab') 2 Fragment (Alexa Fluor ® 647 Conjugate) #4414 was used as a secondary antibody. Analysis was performed on cells in the lymphocyte gate.