Description
The MP5-20F3 monoclonal antibody specifically binds to mouse interleukin-6 (IL-6). The immunogen used to generate the MP5-20F3 hybridoma was recombinant mouse IL-6.
ELISA Capture: The purified MP5-20F3 antibody (Cat. No. 554400) is useful as a capture antibody for a sandwich ELISA for measuring mouse IL-6 protein levels. Purified MP5-20F3 antibody can be paired with the biotinylated MP5-32C11 antibody (Cat. No. 554402) as the detecting antibody, with recombinant mouse IL-6 (Cat. No. 554582) as the standard. Purified MP5-20F3 antibody should be titrated 2-6 µg/ml to determine optimal concentration for ELISA capture. To obtain linear standard curves, doubling dilutions of mouse IL-6 ranging from ~2000 to 15 pg/ml are recommended for inclusion in each ELISA plate. For specific methodology please visit the protocols sections or the chapter on ELISA in the Immune Function Handbook, both of which are posted on our website, www.bdbiosciences.com. Note: This ELISA pair is recommended primarily for measuring cytokine from experimental cell culture systems. These ELISA reagents are not recommended for assaying serum or plasma samples. For measuring mouse IL-6 in serum or plasma our mouse IL-6 BD OptEIA™ set (Cat. No. 555240) or BD OptEIA™ Kit (Cat. No. 550950) are specifically formulated and recommended. Blocking Control for Intracellular Staining: The purified MP5-20F3 antibody (Cat. No. 554400) can be used as a blocking control to demonstrate specificity of IL-6 staining by the PE-conjugated MP5-20F3 antibody (Cat. No. 554401). To perform this control, the fixed/permeabilized cells (~1 million) can be incubated with 1 - 10 µg of purified MP5-20F3 antibody (Cat. No. 554400) for 20 minutes at 4°C, prior to staining with the PE-MP5-20F3 antibody (e.g., 0.1 - 0.5 µg mAb/1 million cells). The intracellular cytokine staining technique and use of blocking controls are described in detail by C. Prussin and D. Metcalfe. For specific methodology, please visit the protocols section or chapter on intracellular staining in the Immune Function Handbook, both of which are posted on our website, www.bdbiosciences.com. Neutralization: The BD NA/LE™ MP5-20F3 antibody (Cat. No. 554398) is useful for neutralization of mouse IL-6 bioactivity. A suitable BD NA/LE™ rat IgG1 isotype-matched control is the R3-34 antibody (Cat. No. 554682). Western Blot: The MP5-20F3 antibody has been reported to be useful for Western blotting. Please note that this application is not routinely tested at BD Biosciences Pharmingen.
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. 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.
