Preparation And Storage
Recommended Assay Procedures
Purified I87-1162 as capture antibody can be paired with the biotinylated I87-419 anti-mouse C3a (Cat. No. 558251) as the detection antibody, using purified mouse C3a protein as standard. The purified I87-1162 antibody should be titrated between 1-4 µg/ml to determine the optimal ELISA plate-coating concentration. For plate coating, we recommend to use a pH 6.5 phosphate buffer. To obtain linear standard curves, doubling dilutions of purified mouse C3a protein (Cat. No. 558618), ranging from 781 pg/ml to 50 ng/ml are recommended for inclusion in each ELISA plate. Addition of FUT-175 (Futhan) to plasma samples at the time of sample collection provides additional protection from ex-vivo activation, and therefore ensures more accurate measurements that reflect the circulating levels of complement activation products. For specific methodology, see Chapter 7: ELISA for specifically measuring the levels of cytokines, chemokines, and inflammatory mediators and their receptors. 2003. Techniques for Immune Function Analysis Application Handbook 1st Edition. BD Biosciences. This ELISA antibody pair shows no cross-reactivity with the following: recombinant mouse C5a, native human C3a, C4a and C5a, and rat C5a. This ELISA antibody pair shows a limited cross-reactivity with rat C3a at ~0.2% as compared to the corresponding mouse signal.
Product Notices
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- 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.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
Companion Products
The purified I87-1162 antibody reacts with the mouse C3a protein. The I87-1162 antibody is specific for a neoepitope exposed in mouse C3a/C3adesArg and does not cross-react with mouse C3. This capacity makes antibody I87-1162 a preferential capture antibody for direct detection of mouse C3a from plasma or serum samples in an ELISA sandwich assay. Anaphylatoxin C3a is a bioactive cleavage product released from plasma component C3 during complement activation and is involved in mediation of a variety of cellular immune responses, as well as being a potent pro-inflammatory agent. The release of this cleavage product is a reliable indicator of in vivo or in vitro complement activation.
Development References (4)
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Ember J.A, Jagels M.A., Hugli T.E. Characterization of complement anaphylatoxins and their biologic responses. In: Volenakis JE and Frank MM, ed. The human complement system in health and disease. New York: Marcel Dekker Inc; 1998:241-284.
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Hugli T.E. Structure and function of C3a anaphylatoxin . Curr Top Microbiol Immunol. 1990; 153:181-208. (Immunogen).
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Hugli T.E. Structure and function of the anaphylatoxins . Springer Semin Immunopathol. 1984; 7:193-219. (Immunogen).
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Volanakis J.E. Overview of the complement system . In: Volenakis JE and Frank MM, ed. The human complement system in health and disease. New York: Marcel Dekker Inc; 1998:9-32.
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Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims. Comparisons are not made against non-BD technologies, unless otherwise noted.
For Research Use Only. Not for use in diagnostic or therapeutic procedures.