Purifying Monoclonal Antibodies and Producing Antibody Fragments

Abstract

The intended use of a monoclonal antibody (MAB) determines the degree of purity required in the antibody preparation. If, for example, a sufficiently specific anti-MAB conjugate is used as second reagent in indirect immunofluorescence or immunocytochemistry, then further purification of the concentrated culture supernate or ascites fluid is generally unnecessary. If, on the other hand the MAB is to be labeled directly (see Chap. 9 ff.), then contamination of the mouse ascites or the culture medium with foreign proteins (such as insulin or transferrin) must be excluded, since by binding to the specific membrane-bound receptors these could mimic binding of the MAB to cells. For similar reasons one cannot use insufficiently purified materials in any biological test system in which contaminants could cause effects that might be falsely attributed to the MAB. In such cases one can use as a control an antibody of the same immunoglobulin subclass which does not react with the antigen in question, but which is derived from the same fusion and prepared under the same conditions. MAB preparations which are intended for therapeutical uses in vivo (see Chap. 1.3) must satisfy particularly stringent criteria.