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Our Abpromise guarantee covers the use of ab14713 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|IHC-P||Use at an assay dependent concentration.|
|WB||Use a concentration of 1 µg/ml. Detects a band of approximately 36 kDa (predicted molecular weight: 40 kDa).|
|Flow Cyt||Use 1-2µg for 106 cells. ab170190-Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.|
Lane 1: Wild-type HAP1 cell lysate (20 µg)
Lane 2: NDUFA9 knockout HAP1 cell lysate (20 µg)
Lane 3: WI38 cell lysate (20 µg)
Lane 4: NIH3T3 cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab14713 observed at 40 kDa. Red - loading control, ab176560, observed at 52 kDa.
ab14713 was shown to specifically react with NDUFA9 when NDUFA9 knockout samples were used. Wild-type and NDUFA9 knockout samples were subjected to SDS-PAGE. ab14713 and ab176560 (loading control to alpha tubulin) were diluted at 1 μg/mL and 1/10 000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.
Overlay histogram showing HepG2 cells stained with ab14713 (red line). The cells were fixed with 80% methanol (5 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 (ab14713, 2µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. 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.