Our Abpromise guarantee covers the use of ab54764 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||Use a concentration of 1 - 5 µg/ml.|
|Flow Cyt||Use 1µg for 106 cells.
ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
|ICC/IF||Use a concentration of 10 µg/ml.|
Lane 1: Wild type HAP1 whole cell lysate (20 µg)
Lane 2: BIN1 (KO) knockout HAP1 whole cell lysate (20 µg)
Lane 3: HeLa whole cell lysate (20 µg)
Lane 4: U-87MG whole cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab54764 observed at 45-65 kDa. Red - loading control, ab181602, observed at 37 kDa.
ab54764 was shown to specifically react with BIN1 when BIN1 knockout samples were used. Wild-type and BIN1 knockout samples were subjected to SDS-PAGE. Ab54764 and ab181602 (Rabbit anti GAPDH loading control) were incubated overnight at 4°C at 1 µg/ml and 1/10000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed ab216772 and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed ab216777 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.
ICC/IF image of ab54764 stained SKNSH 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 (ab54764, 10µg/ml) overnight at +4°C. The secondary antibody (green) was ab96879, DyLight® 488 goat anti-mouse 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.
ab54764 has not yet been referenced specifically in any publications.