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Monoclonal antibodies versus polyclonal antisera , Polyclonal antisera

المؤلف:  Sue Jickells , Adam Negrusz (Editors)

المصدر:  Clarkes Analytical Forensic Toxicology

الجزء والصفحة:  379

2026-11-04

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Monoclonal antibodies versus polyclonal antisera

Polyclonal antisera

Immunisation of a rabbit or a sheep usually proceeds with 20 to 100 lg of the protein conjugated hapten, which can be mixed with an adjuvant to stimulate the immune system. This produces an immune response that consists mainly of IgM followed by IgG. This response is further stimulated by additional immunisations of a similar or lower dose at regular intervals– typically every 3–4 weeks for 20 weeks or longer. Test bleed samples of approximately 1–2 mL are taken at 10–14 days post injection, at which point the immune response (and therefore antibody titre in the serum) is at its highest. After analysis of the test bleeds, a larger anti serum sample can be drawn. This can be as much as several hundred millilitres when using larger host animals (e.g. sheep). Antisera yield can vary from hundreds to many thousands of tests per millilitre depending on the success of the immunisation programme and the type of assay in which they are employed. It is not the absolute volume of serum that is important, but rather the amount and quality of antibodies contained therein.

 Using standard separation methods, such as ammonium sulfate precipitation, Protein A separation and/or ion-exchange chromatography, antisera can be purified for further use. Affinity chromatography can be beneficial in certain circumstances and involves passing the antiserum over a column that contains an immobilised form of the drug of interest. In this way it is possible to fractionate a complex polyclonal antiserum. In some cases purification is not necessary and the antiserum (i.e. the serum itself) can be used with simple dilution.