ELISA Capture: The purified JES5-2A5 antibody (Cat. No. 551215) is useful as a capture antibody for a sandwich ELISA for measuring mouse IL-10 protein levels. Purified JES5-2A5 antibody can be paired with the biotinylated SXC-1 (Cat. No. 554423) antibody or the biotinylated JES5-16E3 (Cat No. 554465) antibody as the detecting antibody, with recombinant mouse IL-10 protein (Cat. No. 550070) as the standard. Purified JES5-2A5 antibody should be titrated (2 - 6 µg/ml) to determine the optimal concentration for ELISA capture. An appropriate range of concentrations of the mouse IL-10 recombinant protein for obtaining a linear standard curve is 2 ng/ml to 15 pg/ml mouse IL-10. For maximal sensitivity, an overnight incubation (4°C) of samples/standards with the coated capture antibody is suggested. For specific methodology, please visit the protocols section of the website or the chapter on ELISA in the Immune Function Handbook, both of which are posted on our web site, www.bdbiosciences.com. This reagent is recommended primarily for assay of cell culture supernatants. For assay of mouse IL-10 in serum or plasma, our OptEIA™ Mouse IL-10 ELISA Set (Cat. No. 555252) is recommended.
Note: This ELISA pair shows no cross-reactivity with any of the cytokines or chemokines tested (e.g., mouse IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-9, IL-12 p70, IL-15, GM-CSF, IFN-γ, MCP-1, 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, LT-α, VEGF; rat IL-2, IL-4, IL-6, IL-10, GM-CSF, IFN-γ, TNF).
Western blot: The JES5-2A5 antibody (Cat. No. 551215) has been found useful for western blotting. A concentration of 1-5 µg/ml (in conjunction with the AKP-conjugated goat anti-rat Igs) has been found to enable visualization of ≤100 ng/lane of recombinant mouse IL-10 (Cat. No. 550070) under reducing conditions. Please note that this application is not routinely tested at BD Biosciences.
Neutralization: The no azide/ low endotoxin format (NA/LE) of JES5-2A5 antibody (Cat. No. 554421) is useful for neutralization of mouse IL-10 bioactivity. A suitable NA/LE rat IgG1 isotype-matched control is the R3-34 antibody (Cat. No. 554682).