Mesothelin antibody

• Catalog Number: orb344445
• Category: Antibodies
• Description: Mesothelin antibody
• Species/Host: Mouse
• Clonality: Monoclonal
• Clone Number: MN-1
• Tested applications: ELISA, FC, IHC, IP, WB
• Reactivity: Human
• Isotype: IgG2a
• Immunogen: This antibody was produced in mesothelin-deficient mice by immunizations with plasmid cDNA encoding human MSLN full length protein followed by a single boost of a recombinant human mesothelin-Fc fusion protein.
• Concentration: 1.0 mg/mL
• Dilution range: ELISA: 1:10,000 - 1:50,000, IHC: 1:100, IP: 1:800, WB: 1:1,000
• Form/Appearance: Liquid (sterile filtered)
• Purity: This antibody is directed against human mesothelin protein. This product was purified from tissue culture supernatant fluid by Protein A chromatography. Cross reactivity with homologues from other sources has not been tested.
• Conjugation: Unconjugated
• UniProt ID: Q13421
• NCBI: 53988378
• Storage: Store vial at -20° C or below prior to opening. This vial contains a relatively low volume of reagent (25 µL). To minimize loss of volume dilute 1:10 by adding 225 µL of the buffer stated above directly to the vial. Recap, mix thoroughly and briefly centrifuge to collect the volume at the bottom of the vial. Use this intermediate dilution when calculating final dilutions as recommended below. Store the vial at -20°C or below after dilution. Avoid cycles of freezing and thawing.
• Buffer/Preservatives: 0.01% (w/v) Sodium Azide
• Alternative names: mouse anti-Mesothelin Antibody, Mesothelian, MN, M
• Note: For research use only
• Application notes: This antibody has been tested for use in immunohistochemistry, flow cytometry, and western blotting. Specific conditions for reactivity should be optimized by the end user. Expect a band approximately 40 kDa in size corresponding to mature mesothelin by western blotting in the appropriate cell lysate or extract. For immunohistochemistry, archival PEFF human tissues were deparaffinized followed by hydration. Antigen-retrieval is recommended. Block tissues with 1% BSA in PBS for 30 min at 23° C. Antibodies are diluted in 1% BSA and reacted with tissue for 60 min at room temperature.
• Expiration Date: 12 months from date of receipt.

(A) Tumor processing flow-chart of 235 samples, which were processed for RNA extraction, cell culture and embedding either in PFA or OCT. Samples showing attributes which are grayed in the chart, were not used in the live cell biobank. (B) Circos whole genome copy number variations (CNVs) view of tumor and primary cell culture in patients malignant pleural mesothelioma (MPM) 236 and MPM95. The quilt plot highlights the CNV and SNVs in genes that are part of MPM landscape (2). (C) Immunofluorescence analysis of selected markers in primary culture from patient MPM236. Scale bar 200 µM. (D) Selected genes expression analysis at mRNA (upper panel) and protein (lower panel) level in primary cultures derived from samples from two patients, MPM236 and MPM95. The latter one underwent EMT during disease progression. (E) Selected genes expression analysis at mRNA in tumor samples from patients MPM236 and MPM95. (F) Significant correlation between gene expression changes in tumor and primary culture from patient MPM236 at passage 3.

Flow Cytometry Results of Anti-Mesothelin (MOUSE) Monoclonal Antibody. The green histogram shows NCI-H226 cells and blue histogram shows MCF-7 cells. Both cell lines are stained with a 1:800 dilution Anti-Mesothelin (MOUSE) Monoclonal Antibody. The secondary antibody use was Anti-Mouse IgG (H&L) (GOAT) Antibody DyLight™ 488 Conjugated at the 1:400 dilution.

Immunohistochemistry of Mouse anti-Mesothelin antibody. Tissue: human tonsil. Fixation: formalin fixed paraffin embedded. Antigen retrieval: not required. Primary antibody: anti-Mesothelin antibody at 15 µg/mL for 1 h at RT. Secondary antibody: Peroxidase mouse secondary antibody at 1:10000 for 45 min at RT. Staining: Mesothelin as precipitated red signal with hematoxylin purple nuclear counterstain.

Immunohistochemistry using Biorbyt's anti-mesothelin antibody to react with two epitopes on mesothelin in PEFF human mesothelioma tissue sections treated by antigen retrieval methods. Anti-mesothelin primary antibodies were used at 10 µg/mL to label these sections as follows: C, MAb MB; and D, MAb MN followed by goat anti-mouse IgG conjugated to horseradish peroxidase at 25 µg/mL in 1% BSA/PBS for 30 minutes. (magnification, ×200; bar, 50 µM).

Mesothelin expression in mesothelioma monolayers and spheroids. NCI-H226 cells incubated with an anti-mesothelin mAb (MN) and detected with goat anti-mouse IgG conjugated with Alexa488 by flow cytometry.

Western Blot of Mouse Anti-Mesothelin Antibody. Lane 1: Opal Prestained Molecular Weight Marker. Lane 2: HeLa [10 µg] + Mesothelin-Fc [0.1 µg]. Lane 3: HeLa [10 µg] + Mesothelin-Fc [0.05 µg]. Lane 4: HeLa [10 µg] + Mesothelin-Fc [0.02 µg]. Lane 5: HeLa Whole Cell Lysate (p/n orb348668). Primary Antibody: Anti-Mesothelin at 1 µg/mL overnight at 2-8°C. Secondary Antibody: Rabbit Anti-Mouse IgG HRP conjugated (p/n orb347544) at 1:40000 for 30 mins at RT. Block: BlockOut Buffer (p/n orb348644) 30 mins at RT. Exposure: 15 sec. Predicted MW: 40 kDa Mesothelin + Fc region 30 kDa. Observed MW: ~70-75 kDa.

Western blotting using Biorbyt's anti-mesothelin antibodies to detect mesothelin-Fc at 100 ng (lane 1), 25 ng (lane 2), 6 ng (lane 3), 2 ng (lane 4) and 0.4 ng (lane 5). Lane 6 contains 50 ng of CDC25-Fc. Proteins were separated on 4-20% gradient gel by SDS-PAGE followed by transfer to PVDF membrane. Primary antibody was used at 1 µg/ml followed by reaction with ALP goat anti-mouse IgG and BCIP/NBT substrate.