Anti-Cyclophilin F antibody (ab126573)

Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
Lane 2: Cyclophilin F knockout HAP1 whole cell lysate (20 µg)
Lane 3: HeLa whole cell lysate (20 µg)
Lane 4: HEK293 whole cell lysate (20 µg)              

Lanes 1 - 4: Merged signal (red and green). Green - ab126573 observed at 24 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab126573 was shown to specificaly recognize Cyclophilin F in wild-type HAP1 cells along with additional cross-reactive bands. No bands was observed when Cyclophilin F knockout samples were examined. Wild-type and Cyclophilin F knockout samples were subjected to SDS-PAGE.  Ab126573 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1 ug/ml and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging. 

ICC/IF image of ab126573 stained A431 cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab126573, 5µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.