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Our Abpromise guarantee covers the use of ab692 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
|IHC-P||Use at an assay dependent concentration.|
|IHC-Fr||Use at an assay dependent concentration.|
|IP||Use at an assay dependent concentration.|
|ICC/IF||Use a concentration of 5 µg/ml.|
Lane 1: Wild type HAP1 whole cell lysate (20 µg)
Lane 2: BCL2 knockout HAP1 whole cell lysate (20 µg)
Lane 3: HeLa whole cell lysate (20 µg)
Lanes 1 - 3: Merged signal (red and green). Green - ab692 observed at 26 kDa. Red - loading control, ab181602, observed at 37 kDa.
ab692 was shown to specifically react with BCL2 when BCL2 knockout samples were used. Wild-type and BCL2 knockout samples were subjected to SDS-PAGE. Ab692 and ab181602 (Rabbit anti GAPDH loading control) were incubated overnight at 4°C at 500 dilution 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.
ab692 staining Bcl-2 in SK-N-SH cells treated with (R)-(-)-Deprenyl hydrochloride (Selegiline hydrochloride) (ab120604), by ICC/IF. Increase of Bcl-2 expression correlates with increased concentration of (R)-(-)-Deprenyl hydrochloride (Selegiline hydrochloride), as described in literature.
The cells were incubated at 37°C for 3h in media containing different concentrations of ab120604 ((R)-(-)-Deprenyl hydrochloride (Selegiline hydrochloride)) in DMSO, fixed with 4% formaldehyde for 10 minutes at room temperature and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab692 (5 μg/ml) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A anti-mouse DyLight 488 polyclonal antibody (ab96879) at 1/250 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI and are shown in blue.
Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling Bcl-2 with ab692 at 1/100 dilution. Samples were incubated with primary antibody for 30-45 minutes at RT.
Overlay histogram showing Jurkat cells stained with ab692 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab692, 1/10 dilution) for 30 min at 22°C. The secondary antibody used was a goat anti-mouse DyLight® 488 (IgG; H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in Jurkat cells fixed with methanol (5 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.
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