The 2H5 antibody reacts with mouse and rat monocyte chemoattractant protein (MCP-1), formerly termed JE. This antibody also recognizes human MCP-1, but shows no reactivity with the closely related mouse β chemokines, TCA3 and MIP-1β. The immunogen used to generate the 2H5 hybridoma was heparin-purified CHO-expressed mouse MCP-1. This is a neutralizing antibody.
ELISA: The purified 2H5 antibody (Cat. No. 551217) is useful as a capture antibody for a sandwich ELISA for measuring mouse MCP-1 protein levels. Purified 2H5 antibody can be paired with the biotinylated 4E2/MCP antibody (Cat. No. 554444) as the detecting antibody, with recombinant mouse MCP-1 (Cat. No. 554590) as the standard. Purified 2H5 antibody should be titrated 6 - 10 µg/ml to determine optimal concentration for ELISA capture. To obtain linear standard curves, doubling dilutions of mouse MCP-1 ranging from ~4,000 to 30 pg/ml are recommended for inclusion in each ELISA plate. For maximal sensitivity, an overnight incubation (4°C) of samples/standards with the coated capture antibody is recommended. For specific methodology, please visit the protocols section of the Immune Function Handbook, which is also posted on our web site, www.bdbiosciences.com. Note 1: This reagent is recommended primarily for assay of cytokine from experimental cell culture systems. For detection of MCP-1 in mouse serum or plasma samples, the BD OptEIA™ Mouse MCP-1 ELISA Set is recommended. Note 2: This ELISA pair shows no cross-reactivity with any of the cytokines tested (e.g., mouse IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-9, IL-10, IL-12 p70, IL-15, GM-CSF, IFN-γ, TCA-3, TNF; human IL-1α, IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12 p70, IL-12 p40, IL-13, IL-15, G-CSF, GM-CSF, IFN-γ, lymphotactin, MCP-1, MCP-2, MIP-1α, MIP-1β, NT-3, PDGF-AA, sCD23 , SCF, TNF, TNF-β, VEGF; rat IL-2, IL-4, IL-6, IL-10, GM-CSF, IFN-γ, TNF). Neutralization: The NA/LE™ 2H5 antibody (Cat. No. 554440) is useful for neutralization of mouse MCP-1 bioactivity. A suitable NA/LE™ hamster IgG isotype-matched control is the G235-2356 antibody (Cat. No. 554709). Immunofluorescent Staining and Flow Cytometric Analysis: The 2H5 antibody is useful for immunofluorescent staining and flow cytometric analysis to identify and enumerate MCP-1 producing cells within mixed cell populations. PE conjugated 2H5 antibody (Cat. No. 554443) is especially suitable for these experiments. The use of a specificity control, such as recombinant mouse MCP-1 (Cat No. 554590) or unlabeled 2H5 antibody (Cat. No. 554441) is recommended. For specific methodology, please visit the protocols section or the Immune Function Handbook, which is also posted on our web site, www.bdbiosciences.com.
The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. Store undiluted at 4°C.
1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
2. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
3. Although hamster immunoglobulin isotypes have not been well defined, BD Biosciences Pharmingen has grouped Armenian and Syrian hamster IgG monoclonal antibodies according to their reactivity with a panel of mouse anti-hamster IgG mAbs. A table of the hamster IgG groups, Reactivity of Mouse Anti-Hamster Ig mAbs, may be viewed at http://www.bdbiosciences.com/documents/hamster_chart_11x17.pdf.
4. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
