Anti-ATPase Inhibitory Factor 1/IF1 antibody [5E2D7]

• WB

Lab

Western blot - Anti-ATPase Inhibitory Factor 1/IF1 antibody [5E2D7] (AB110277)

Lanes 1 - 3 : Merged signal (red and green). Green - ab110277 observed at 12 kDa. Red - loading control, ab181602, observed at 37 kDa.

ab110277 was shown to recognize ATPase Inhibitory Factor 1 / IF1 in wild-type HAP1 cells as signal was lost at the expected MW in ATPase Inhibitory Factor 1 / IF1 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and ATPase Inhibitory Factor 1 / IF1 knockout samples were subjected to SDS-PAGE. ab110277 and ab181602 (Rabbit anti-GAPDH loading control) were incubated overnight at 4°C at 1 μg/ml and 1/20000 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 1/20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-ATPase Inhibitory Factor 1/IF1 antibody [5E2D7] (ab110277) at 1 µg/mL

Lane 1:

Wild-type HAP1 whole cell lysate at 20 µg

Lane 2:

ATPase Inhibitory Factor 1 / IF1 knockout HAP1 whole cell lysate at 20 µg

Lane 3:

HeLa whole cell lysate at 20 µg

Predicted band size: 12 kDa

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• Flow Cyt

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Flow Cytometry - Anti-ATPase Inhibitory Factor 1/IF1 antibody [5E2D7] (AB110277)

Overlay histogram showing HepG2 cells stained with ab110277 (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 (ab110277, 1μg/1x10⁶ cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab91353) 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.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-ATPase Inhibitory Factor 1/IF1 antibody [5E2D7] (AB110277)

ICC/IF image of ab110277 stained HepG2 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 (ab110277, 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

• ICC

AbReview80686****

Immunocytochemistry - Anti-ATPase Inhibitory Factor 1/IF1 antibody [5E2D7] (AB110277)

Immunocytochemistry analysis of paraformaldehyde-fixed rat primary cardiac fibroblasts permeabilized with 1% Triton for 4 min staining with ab110277 at 1/50 dilution. Secondary antibody was Dylight™ 488 donkey anti mouse at 1/100 dilution. Samples were incubated with the primary antibody with PBST for 30 minutes at 25°C

This image is courtesy of an anonymous Abreview

• WB

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Western blot - Anti-ATPase Inhibitory Factor 1/IF1 antibody [5E2D7] (AB110277)

All lanes:

Western blot - Anti-ATPase Inhibitory Factor 1/IF1 antibody [5E2D7] (ab110277) at 1 µg/mL

Lane 1:

Human heart mitochondria at 10 µg

Lane 2:

Bovine heart mitochondria at 4 µg

Lane 3:

Rat heart mitochondria at 10 µg

Lane 4:

Mouse heart mitochondria at 10 µg

Predicted band size: 12 kDa

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• WB

CiteAb

Western blot - Anti-ATPase Inhibitory Factor 1/IF1 antibody [5E2D7] (AB110277)

ATPase Inhibitory Factor 1/IF1 western blot using anti-ATPase Inhibitory Factor 1/IF1 antibody [5E2D7] ab110277. Publication image and figure legend from Kahancová, A., Sklenář, F., et al., 2020, Sci Rep, PubMed 32005857.

ab110277 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab110277 please see the product overview.

Effect of glucose concentrations on relative IF1 protein levels in INS-1E cells. Cells were incubated for 8 h with modified culturing medium with adjusted glucose levels. As non-treated controls, we used INS-1E cells incubated at standard 11 mM glucose. (a) Representative western-blot, (b) quantification of protein levels. (c) Relative mRNA levels in INS-1E cells incubated for 8 h with modified culturing medium with adjusted glucose levels. As non-treated controls, we used INS-1E cells incubated at standard 11 mM glucose. Significance was determined by the Kruskal–Wallis's test followed by posthoc Dunnett's multiple comparisons to control. p values are indicated as follows : *p < 0.05, **p < 0.005, ***p < 0.0005.

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