United Kingdom (UK)
Recombinant monoclonal antibody to Mesothelin. Manufactured using AbAb’s Recombinant Platform with variable regions (i.e. specificity) from the hybridoma MN-1.
UniProt Accession Number of Target Protein: Q13421
Alternative Name(s) of Target: MSLN; CAK1 antigen; Pre-pro-megakaryocyte-potentiating factor; MN
Immunogen: The original antibody was generated in mesothelin-deficient mice by immunization with plasmid cDNA corresponding to the mesothelin extracellular domain followed by a single boost with a recombinant human mesothelin-rabbit IgG Fc fusion protein.
Specificity: This antibody can recognize and bind human mesothelin. This antibody binds a different antibody on the human mesothelin than another antibody called MB. Mesothelin (MSLN) is a glycophosphatidylinositol (GPI)-linked cell surface protein highly expressed in several types of malignant tumors sometimes in association with increased tumor aggressiveness and poor clinical outcome. The membrane-anchored forms of this molecule may play a role in cellular adhesion.
Application Notes: The binding characterization of this antibody was done using ELISA and SPR. The binding affinity of the original IgG2a antibody for mesothelin-Fc protein was determined by BIACore and reported to be Kd = 1.0 nM. This antibody can recognize membrane bound mesothelin on the surface of mesothelin positive H226, A1847, Hela and Panc3.014 cells (mesothelin-positive) in a flow cytometry. This antibody can also bind denatured human mesothelin in a western blot assay. This antibody was also used in the immunohistochemical staining of cancerous tissue like ovarian adenocarcinoma and normal tissues like peritoneal mesothelium on uterus, pleural mesothelium on lung and fallopian tube luminal fluid (PMID: 16115924). This antibody was also used in an immunohistochemical analysis of 1562 tumors for mesothelin expression. It was reported that mesothelin was highly expressed in ovarian serous carcinoma, pancreatic adenocarcinoma and malignant pleural mesothelioma (PMID: 28460459). This antibody was also used in the immunoprecipitation of mesothelin from cell lysates (PMID: 23694968).
Antibody first published in:
Onda et al. New monoclonal antibodies to mesothelin useful for immunohistochemistry, fluorescence-activated cell sorting, Western blotting, and ELISA. Clin Cancer Res. 2005 Aug 15;11(16):5840-6. PMID:16115924
Note on publication:
Describes the generation of two antibodies against different epitopes of human mesothelin.
Flow cytometry using anti-Mesothelin antibody MN-1 (Ab03251). HeLa cells were stained with the anti-unknown specificity antibody (Ab00178-23.0; isotype control, black line) or the rabbit IgG version of MN-1 (Ab03251-23.0, blue line) at a dilution of 1:100 for 1h at RT. After washing, the bound antibody was detected using a goat anti-rabbit IgG AlexaFluor® 488 antibody at a dilution of 1:1000, and the cells were analyzed using a FACSCanto flow-cytometer.
Western blot using anti-Mesothelin antibody MN-1 (Ab03251). HeLa cells lysate (35µg protein in RIPA buffer) was resolved on an SDS-PAGE gel, and blots were probed with the chimeric rabbit version of MN-1 (Ab03251-23.0) at 0.3µg/ml before detection using an anti-rabbit secondary antibody. A primary incubation of 1 hour was used, and protein was detected by chemiluminescence.
Immunofluorescence staining of HeLa cells with anti-Mesothelin antibody MN-1 (Ab03251). Immunofluorescence analysis of paraformaldehyde fixed HeLa cells on Shi-fix™ coverslips stained with the chimeric rabbit IgG version of MN-1 (Ab03251-23.0) (1:100 dilution) for 1h followed by Alexa Fluor® 488 secondary antibody (1:1000 dilution), showing membrane staining. The nuclear stain is DAPI (blue). Panels show, from left-right, top-bottom, Ab03251-23.0, DAPI, merged channels, and an isotype control. The isotype control was an unknown specificity antibody (Ab00178-23.0) followed by staining with Alexa Fluor® 488 secondary antibody.