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Application: Study Mesothelin:CD3 complex formation and assess the binding of BsAbs to their dual-antigen targets simultaneously for drug discovery and high throughput screening (HTS). Note: Suitable for use with human serum, cell culture supernatants, and with purified proteins.
Background: Bispecific antibodies (BsAbs) are antibodies that have binding sites directed against different antigens or different epitopes of the same antigen. They are composed of heavy and light chains, and the appropriate chain matching is required for efficacy. To enhance the chance of chain matching and different applications, more than 30 different technologies have been developed, such as knob-into hole, ART-Ig, BiTE (bispecific cell engager) and others. They have gained attention for the treatment of cancer and other disorders due to their superior cytotoxicity effects and less development of resistance to their use. BsAbs can act as immune cell-cancer cell bridges, enhancing the killing potential of the immune cells. Current BsAbs have a CD3, CD16 or CD47 as one of the binding domains, and target CD19, PD-1, LAG-3, PSMA, mesothelin or other tumor antigens with the other. Mesothelin (MSLN) is a glycophosphatidylinositol (GPI) linked cell-surface protein that is produced as a ~70 kDa precursor protein and cleaved by Furin protease to generate the ~40 kDa mature form. MSLN is frequently overexpressed in mesothelioma, ovarian, pancreatic, and non-small cell lung cancers, while its expression in normal tissues is restricted to the mesothelial lining. MSLN is a tumor-associated antigen and has been an attractive target for targeted immunotherapy approaches, including drug-conjugated antibodies and chimeric antigen receptor T cells (CAR-T Cells).
Contraindications: This kit is compatible with up to 1% DMSO.
Description: The Bispecific Mesothelin:CD3 Bridging Colorimetric ELISA Kit is an ELISA designed to analyze the ability of Bispecific Antibodies (BsAbs) to bridge Mesothelin (also known as MSLN, MPF, SMRP) and CD3 (cluster of differentiation 3) for screening and profiling applications. This assay kit can determine if an anti-Mesothelin-anti-CD3 bispecific antibody binds to both targets simultaneously and bridges Mesothelin to CD3. The Bispecific Mesothelin:CD3 Bridging Colorimetric ELISA Assay Kit comes with enough recombinant human Mesothelin (amino acids 296–580), CD3-Containing Detection Reagent, and assay buffer for 100 enzyme reactions. This kit also includes Anti-Mesothelin-Anti-CD3 Bispecific Antibody as a positive control.A 96-well plate is coated with Mesothelin protein. After coating, an anti-Mesothelin-anti-CD3 bispecific antibody is added in an optimized assay buffer. The plate is washed to remove the unbound antibody. The plate is then incubated with CD3-Containing Detection Reagent. Finally, HRP Colorimetric Substrate is added to produce absorbance that can be measured using a UV/Vis spectrophotometer microplate reader. The absorbance signal is proportional to the bridging efficacy.
Storage Stability: This assay kit will perform optimally for up to 6 months from date of receipt when the materials are stored as directed.
Target: CD3, Mesothelin
Uniprot: Q13421
Warnings: Avoid freeze/thaw cycles.
Biosafety Level: Not applicable (BSL-1)
References: Ma J., et al., 2021 Front Immunol. 12:626616. Asgarov K., et al., 2017 MAbs 9(3): 567–577.Ye L., et al., 2019 Experimental and Therapeutic Medicine 17: 739-747.
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