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Synthetic peptide within Human Claudin 4 aa 160-209. The exact sequence is proprietary.
Database link: O14493
Our Abpromise guarantee covers the use of ab53156 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|IHC-P||Use a concentration of 4 µg/ml. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.|
|ICC/IF||Use a concentration of 5 µg/ml.|
|WB||1/500 - 1/1000. Detects a band of approximately 22 kDa (predicted molecular weight: 22 kDa).|
ab53156 staining Claudin 4 in human colon tissue. Staining is localized to the cell membrane.
Left panel: ab53156 at 4 µg/ml.
Right panel: Isotype control.
Sections were stained using an automated system at room temperature. Sections were rehydrated and antigen retrieved with EDTA pH 9.0. Slides were blocked in 3% H2O2 in methanol for 10 minutes. They were then blocked again for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 minutes and detected for 30 minutes. Colorimetric detection was completed with diaminobenzidine for 5 minutes. Slides were counterstained with hematoxylin and coverslipped. Please note that for manual staining we recommend to optimize the primary antibody concentration and incubation time (overnight incubation), and amplification may be required.
ICC/IF image of ab53156 staining Claudin 4 (green) in MCF7 (Human breast adenocarcinoma cell line) cells. The cells were fixed in 4% formaldehyde (10 minutes) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1 hour to permeabilize the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab53156, 5 µg/ml) overnight at +4°C. The secondary antibody was an Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1 hour. An Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1 hour. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43 µM.