This antibody gave a positive signal in Mouse Liver tissue lysate as well as the following whole cell lysates: HeLa; HepG2; MEF1; NIH3T3; Raw264.7; PC12.
This antibody gave a positive result in IHC in the following FFPE tissue: Human normal heart muscle.
This antibody gave a positive result when used in the following methanol fixed cell lines: HepG2.
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
pH: 7.40 Preservative: 0.02% Sodium azide Constituent: PBS Note: Batches of this product that have a concentration < 1mg/ml may have BSA added as a stabilising agent. If you would like information about the formulation of a specific lot, please contact our scientific support team who will be happy to help.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml.
Use a concentration of 1 µg/ml. Detects a band of approximately 56 kDa (predicted molecular weight: 56 kDa).
Mitochondrial membrane ATP synthase (F(1)F(0) ATP synthase or Complex V) produces ATP from ADP in the presence of a proton gradient across the membrane which is generated by electron transport complexes of the respiratory chain. F-type ATPases consist of two structural domains, F(1) - containing the extramembraneous catalytic core, and F(0) - containing the membrane proton channel, linked together by a central stalk and a peripheral stalk. During catalysis, ATP synthesis in the catalytic domain of F(1) is coupled via a rotary mechanism of the central stalk subunits to proton translocation. Subunits alpha and beta form the catalytic core in F(1). Rotation of the central stalk against the surrounding alpha(3)beta(3) subunits leads to hydrolysis of ATP in three separate catalytic sites on the beta subunits.
ab128743 stained HepG2 cells. The cells were 100% methanol fixed (5 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 ab128743 at 5µg/ml overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti- rabbit (ab96899) 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.
Western blot - Anti-ATPB antibody (ab128743)
All lanes : Anti-ATPB antibody (ab128743) at 1 µg/ml
Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate Lane 2 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate Lane 3 : MEF1 (Mouse embryonic fibroblast cell line) Whole Cell Lysate Lane 4 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate Lane 5 : RAW 264.7 (Mouse leukaemic monocyte macrophage cell line) Whole Cell Lysate Lane 6 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate Lane 7 : Liver (Mouse) Tissue Lysate
Lysates/proteins at 10 µg per lane.
Secondary All lanes : Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution
Developed using the ECL technique.
Performed under reducing conditions.
Predicted band size: 56 kDa Observed band size: 56 kDa
Exposure time: 10 seconds
This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 5% Bovine Serum Albumin before being incubated with ab128743 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution.
IHC image of ATPB staining in human normal heart muscle 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 ab128743, 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.
Western blot - Anti-ATPB antibody (ab128743)This image is courtesy of an anonymous Abreview
All lanes : Anti-ATPB antibody (ab128743) at 1/5000 dilution
Drerup CM et al. Regulation of mitochondria-dynactin interaction and mitochondrial retrograde transport in axons. Elife6:N/A (2017).
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Sawhney S et al. Alpha-enolase is upregulated on the cell surface and responds to plasminogen activation in mice expressing a ?133p53a mimic. PLoS One10:e0116270 (2015).
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