The immunogen used to raise this antibody has 78% homology with the Mouse Occludin protein and the Rat Occludin protein. Some customers have successfully used ab31721 with Mouse/Rat lysates/tissue, however we have not been successful detecting Occludin in these species in our own testing. Please contact Abcam Scientific Support for more information.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use at an assay dependent concentration. PubMed: 20978075
Use at an assay dependent concentration. PubMed: 22689949
Use a concentration of 1 - 2 µg/ml.
1/100. PubMed: 19828778
ab171870 - Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.
May play a role in the formation and regulation of the tight junction (TJ) paracellular permeability barrier. It is able to induce adhesion when expressed in cells lacking tight junctions.
Localized at tight junctions of both epithelial and endothelial cells. Highly expressed in kidney. Not detected in testis.
Defects in OCLN are the cause of band-like calcification with simplified gyration and polymicrogyria (BLCPMG) [MIM:251290]; also known as pseudo-TORCH syndrome. BLCPMG is a neurologic disorder with characteristic clinical and neuroradiologic features that mimic intrauterine TORCH infection in the absence of evidence of infection. Affected individuals have congenital microcephaly, intracranial calcifications, and severe developmental delay.
Belongs to the ELL/occludin family. Contains 1 MARVEL domain.
The C-terminal is cytoplasmic and is important for interaction with ZO-1. Sufficient for the tight junction localization. Involved in the regulation of the permeability barrier function of the tight junction (By similarity). The first extracellular loop participates in an adhesive interaction.
Phosphorylated upon DNA damage, probably by ATM or ATR. Dephosphorylated by PTPRJ. The tyrosine phosphorylation on Tyr-398 and Tyr-402 reduces its ability to interact with TJP1.
IHC image of Occludin staining in human normal kidney formalin fixed paraffin embedded tissue section*, performed on a Leica Bond™ 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 ab31721, 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.
*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre
Immunocytochemistry/ Immunofluorescence - Anti-Occludin antibody (ab31721)Image is courtesy of an anonymous AbReview
ab31721 staining Occludin in Human Breast cancer cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with methanol/acetone (1:1) and blocked with 0.1% milk for 30 minutes at 25°C. Samples were incubated with primary antibody (1/200 in PBS + 0.1% milk) for 16 hours at 4°C. An Alexa Fluor® 594-conjugated Goat anti-rabbit polyclonal (1/1500) was used as the secondary antibody.
ICC/IF image of ab31721 stained Hek293 cells. The cells were 4% formaldehyde fixed (10 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 (ab31721, 5µg/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.