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Submitted to: Book Chapter
Publication Type: Book / Chapter Publication Acceptance Date: 4/4/2015 Publication Date: 7/14/2015 Citation: Hnasko, R.M., McGarvey, J.A. 2015. Affinity purification of antibodies. In: Hnasko, R., editor. ELISA Methods and Protocols. New York, NY: Humana Press. p. 29-41. doi: 10.1007/978-1-4939-2742-5. Interpretive Summary: This Manuscript describes methods used in the purification of antibodies from different sources. Technical Abstract: Antibodies are provided in a variety of formats that includes antiserum, hybridoma culture supernatant or ascites. They can all be used successfully in crude form for the detection of target antigens by immunoassay. However, it is advantageous to use purified antibody in defined quantity to facilitate assay reproducibility, economy, and reduced interference of non--specific components as well as improved storage, stability and bio--conjugation. Although not always necessary, the relative simplicity of antibody purification using commercially available protein--A, --G or --L resins with basic chromatographic principles warrants purification when antibody source material is available in sufficient quantity. Here we define three simple methods using immobilized 1) protein--A, 2) protein--G, and 3) protein--L agarose beads to yield highly purified antibody. |
