This antibody gave a positive signal in the following human whole cell lysates:
TE 671, Ramos, SK N BE, Hela - Staurosporine Treated (24hr, 500nM), Hela - Hydroxyurea Treated (48hr, 2uM), Jurkat - Staurosporine Treated (24hr, 500nM) and Y79.
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.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Use a concentration of 1 µg/ml.
Use a concentration of 1 µg/ml. Detects a band of approximately 54 kDa (predicted molecular weight: 58 kDa).
Involved in the unfolded protein response (UPR) during ER stress. Co-chaperone of HSPA8/HSC70, it stimulates its ATPase activity. May inhibit both the autophosphorylation of EIF2AK2/PKR and the ability of EIF2AK2 to catalyze phosphorylation of the EIF2A. May inhibit EIF2AK3/PERK activity.
Widely expressed with high level in the pancreas and testis. Also expressed in cell lines with different levels.
ICC/IF image of ab70840 stained HeLa 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 (ab70840, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 Goat anti-Rabbit IgG (H+L) used at a 1/1000 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. This antibody also gave a positive result in 100% Methanol fixed (5 min) Hek293, HepG2, MCF-7 cells at 1µg/ml.
IHC image of ab70840 staining in human testis 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 ab70840, 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.
Lin Y & Sun Z In vivo pancreatic ß-cell-specific expression of antiaging gene Klotho: a novel approach for preserving ß-cells in type 2 diabetes. Diabetes64:1444-58 (2015).
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