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Anti-Claudin 5, clone 2B12, Cat. No. MABT1528, is a mouse monoclonal antibody that detects Claudin 5 and has been tested for use in ELISA, Flow Cytometry, Inhibition/Function Analysis, and Western Blotting.
More>>Anti-Claudin 5, clone 2B12, Cat. No. MABT1528, is a mouse monoclonal antibody that detects Claudin 5 and has been tested for use in ELISA, Flow Cytometry, Inhibition/Function Analysis, and Western Blotting. Less<<
Claudin 5 (UniProt: O00501; also known as Transmembrane protein deleted in VCFS, TMDVCF) is encoded by the CLDN5 (also known as AWAL, TMVCF) gene (Gene ID: 7122) in human. Claudin 5 is a multi-pass membrane protein that plays a major role in tight junction-specific obliteration of the intercellular space. It is localized specifically to the endothelial cell layer in the brain, lung, liver, kidney, and skin. In peripheral tissues, it displays greater heterogeneity in tissue expression. It is one of the most enriched tight junction proteins and its dysfunction has been linked to several neurodegenerative diseases, including Alzheimer s disease and multiple sclerosis. Cldn5-null mice are reported to die shortly after birth. Claudin 5 contains four transmembrane domains (aa 8-28, 82-102, 123-143, 160-180), two extracellular loops (aa 29-81, 144-159), and three cytoplasmic domains (aa 1-7, 103-122, 181-218). (Ref.: Hashimoto, Y., et al. (2018). Sci. Rep. 8; 8383; Greene, C., et al. (2019). Fluids Barriers CNS. 16;3).
References
Product Information
Format
Purified
Presentation
Purified mouse monoclonal antibody IgG2a in PBS without azide.
Applications
Application
Anti-Claudin 5, clone 2B12, Cat. No. MABT1528, is a mouse monoclonal antibody that detects Claudin 5 and has been tested for use in ELISA, Flow Cytometry, Inhibition/Function Analysis, and Western Blotting.
Key Applications
ELISA
Flow Cytometry
Inhibits Activity/Function
Western Blotting
Application Notes
Western Blotting Analysis: A representative lot detected Claudin 5 in Western Blotting applications (Hashimoto, Y., et. al. (2018). Sci Rep. 8(1):8383).
Inhibits Activity/Function Analysis: A representative lot significantly decreased Trans-epithelial/trans-endothelial electrical resistance (TEER) in the hCLDN-5-expressing MDCKII cell monolayer. (Hashimoto, Y., et. al. (2018). Sci Rep. 8(1):8383).
Flow Cytometry Analysis: A representative lot detected Claudin 5 in Flow Cytometry applications (Hashimoto, Y., et. al. (2018). Sci Rep. 8(1):8383).
Enzyme Immunoassay (ELISA) Analysis: A representative lot detected Claudin 5 in ELISA applications (Hashimoto, Y., et. al. (2018). Sci Rep. 8(1):8383).
Biological Information
Immunogen
An artificial membrane protein consisting of the extracellular region of human Claudin 5.
Epitope
extracellular domain
Clone
2B12
Concentration
Please refer to lot specific datasheet.
Host
Mouse
Specificity
Clone 2B12 is a mouse monoclonal antibody that detects human Claudin 5. It targets an epitope within nine amino acids in the second extracellular domain.
Evaluated by Flow Cytometry in human umbilical vein endothelial cells (HUVEC).
Flow Cytometry Analysis: 1 µg of this antibody detected Claudin 5 one million human umbilical vein endothelial cells (HUVEC).
Usage Statement
Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
Storage and Shipping Information
Storage Conditions
Stable for 1 year at -20°C from date of receipt. Handling Recommendations: Upon receipt and prior to removing the cap, centrifuge the vial and gently mix the solution. Aliquot into microcentrifuge tubes and store at -20°C. Avoid repeated freeze/thaw cycles, which may damage IgG and affect product performance.
The production of antibodies against the extracellular regions (ECR) of multispanning membrane proteins is notoriously difficult because of the low productivity and immunogenicity of membrane proteins due to their complex structure and highly conserved sequences among species. Here, we introduce a new method to generate ECR-binding antibodies utilizing engineered liposomal immunogen prepared using a wheat cell-free protein synthesis system. We used claudin-5 (CLDN-5) as the target antigen, which is a notoriously difficult to produce and poorly immunogenic membrane protein with two highly conserved extracellular loops. We drastically improved the productivity of CLDN-5 in the cell-free system after suppressing and normalizing mRNA GC content. To overcome its low immunogenicity, two engineered antigens were designed and synthesized as proteoliposomes: a human/mouse chimeric CLDN-5, and a CLDN-5-based artificial membrane protein consisting of symmetrically arranged ECRs. Intraperitoneal immunization of both engineered CLDN-5 ECR antigens induced ECR-binding antibodies in mice with a high success rate. We isolated five monoclonal antibodies that specifically recognized CLDN-5 ECR. Antibody clone 2B12 showed high affinity (<10 nM) and inhibited CLDN-5-containing tight junctions. These results demonstrate the effectiveness of the methods for monoclonal antibody development targeting difficult-to-produce membrane proteins such as CLDNs.
