Western Blotting (WB), ELISA, Blocking Antibody (Inhibition)
Brand
BD Pharmingen™
Characteristics
1. Since applications vary, each investigator should titrate the reagent to obtain optimal results. 2. Please refer to us 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.
Purification
The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography.
Blocking Control for Intracellular Staining: The purified MP6-XT22 antibody (ABIN967462) can be used as a blocking control to demonstrate specificity of TNF protein staining by directly conjugated -MP6-XT22 antibody. To perform this control, the fixed/permeabilized cells (~ 1 million) can be incubated with 1 -10 µg of unlabeled MP6-XT22 antibody (ABIN967462) for 20 minutes at 4°C, prior to staining with directly conjugated 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.
ELISA Capture: The purified MP6-XT22 antibody (ABIN967462) is useful as a capture antibody for a sandwich ELISA for measuring mouse TNF protein levels. Purified MP6-XT22 antibody can be paired with the biotinylated polyclonal rabbit anti-mouse/rat TNF antibody as the detecting antibody, with recombinant mouse TNF as the standard. This pair measures total TNF, free TNF as well as TNF bound by soluble receptors. 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.
WB: The MP6-XT22 antibody has been reported to be useful for Western blotting. Please note that this application is not routinely tested.
This product contains Sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.
Storage
4 °C
Storage Comment
Store undiluted at 4° C.
Abrams: "Immunoenzymetric assay of mouse and human cytokines using NIP-labeled anti-cytokine antibodies." in: Current protocols in immunology / edited by John E. Coligan ... [et al.], Vol. Chapter 6, pp. Unit 6.20, (2008) (PubMed).
Prussin, Metcalfe: "Detection of intracytoplasmic cytokine using flow cytometry and directly conjugated anti-cytokine antibodies." in: Journal of immunological methods, Vol. 188, Issue 1, pp. 117-28, (1996) (PubMed).
Litton, Sander, Murphy, OGarra, Abrams: "Early expression of cytokines in lymph nodes after treatment in vivo with Staphylococcus enterotoxin B." in: Journal of immunological methods, Vol. 175, Issue 1, pp. 47-58, (1994) (PubMed).
Abrams, Roncarolo, Yssel, Andersson, Gleich, Silver: "Strategies of anti-cytokine monoclonal antibody development: immunoassay of IL-10 and IL-5 in clinical samples." in: Immunological reviews, Vol. 127, Issue 9-10, pp. 5-24, (1992) (PubMed).
The MP6-XT22 antibody reacts with mouse tumor necrosis factor (TNF, also known as TNF-alpha). The immunogen used to generate this hybridoma was recombinant mouse TNF. This antibody is routinely tested by flow cytometric analysis.