Determination of epitope specificities and affinities of monoclonal antibodies in solution phase using biotin-labeled carcinoembryonic antigen and avidin as precipitating agent |
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Authors: | U Krüger L Wickert C Wagener |
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Affiliation: | Institute of Clinical Chemistry and Pathobiochemistry, University of Technology, Aachen, F.R.G. |
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Abstract: | A generally applicable method for the selective precipitation of antigens from solution phase is presented. The method is based on the rapid and quantitative binding of avidin to biotin-labeled antigen in the presence of biotinylated carrier protein. Using carcinoembryonic antigen (CEA) as a model antigen biotin-labeled CEA was precipitated to greater than 97% at amounts of up to 5000 ng/tube. The method was applied to the determination of epitope specificities and affinities of monoclonal anti-CEA antibodies in solution phase. Non-specific precipitation did not exceed 2% for radiolabeled Fab fragments and 6% for radiolabeled IgG. Because antigen-antibody binding takes place in solution, artifacts introduced by the immobilization of reactants to solid supports are omitted. |
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