Anti-UQCRC2 antibody [13G12AF12BB11] (ab14745) at 1/1000 dilution + H23 whole cell lysate at 20 µg

Secondary
HRP-conjugated goat anti-mouse IgG polyclonal at 1/2000 dilution

Developed using the ECL technique.

Performed under reducing conditions.

Observed band size: 48 kDa why is the actual band size different from the predicted?


Exposure time: 1 minute

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IHC image of ab14745 staining in human heart formalin fixed paraffin embedded tissue section, performed on a Leica Bond^TM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab14745, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

Overlay histogram showing HepG2 cells stained with ab14745 (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 (ab14745, 1µg/1x10⁶ cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse 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.

Mitochondrial localization of complex III visualized by immunofluorescence using anti-complex III subunit Core 2 mAb 2E3GC12FB2AE2 (ab14745). Cultured human embryonic lung-derived fibroblasts (strain MRC5) were fixed, permeabilized and then labeled with ab14745 (0.5 µg/ml) followed by an AlexaFluor® 488-conjugated-goat-anti-mouse IgG1 isotype specific secondary antibody (2 µg/ml).

All lanes : Anti-UQCRC2 antibody [13G12AF12BB11] (ab14745) at 0.5 µg/ml

Lane 1 : Human skeletal muscle tissue lysate
Lane 2 : Ramos (human Burkitt's lymphoma cell line) whole cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat polyclonal to mouse IgG - H&L - pre-adsorbed (HRP) at 1/3000 dilution

Observed band size: 48 kDa why is the actual band size different from the predicted?
Additional bands at: 30 kDa. We are unsure as to the identity of these extra bands.