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Bovine heart mitochondria
Our Abpromise guarantee covers the use of ab110321 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|IP||Use at an assay dependent concentration.|
|ICC/IF||Use a concentration of 1 µg/ml. for 2 hours.|
|WB||Use a concentration of 1 µg/ml. Predicted molecular weight: 85 kDa.|
|In-Cell ELISA||Use a concentration of 1 µg/ml.|
|Flow Cyt||Use a concentration of 1 µg/ml.
ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
|IHC-P||Use a concentration of 5 µg/ml.|
Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
Lane 2: Aconitase 2 knockout HAP1 whole cell lysate (20 µg)
Lane 3: HeLa whole cell lysate (20 µg)
Lane 4: Jurkat whole cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab110321 observed at 90 kDa. Red - loading control, ab181602, observed at 37 kDa.
ab110321 was shown to specifically react with Aconitase 2 in wild-type HAP1 cells as signal was lost in Aconitase 2 knockout cells. Wild-type and Aconitase 2 knockout samples were subjected to SDS-PAGE. ab110321 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.
IHC image of Aconitase 2 staining in Human normal colon formalin fixed paraffin embedded tissue section, performed on a Leica BondTM 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 ab110321, 5µg/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.