Abstract
Since the introduction of hybridoma technology, monoclonal antibodies have proven to be very useful tools for the study of specific sites on proteins (1, 2, 3 and 4). The site specificity of monoclonal antibodies offers new ways to link areas of protein function with structure. For some proteins, this mapping has been a simple matter of digesting the antigen and analyzing which fragment(s) the antibodies interact with (5, 6, 7 and 8). However, a large population of monoclonal antibodies (MAbs) are highly dependent on the protein conformation of the antigen (9, 10). These MAbs often lose their reactivities with their antigenic determinants when the target protein is digested or during protein preparation for cleavage. This has been a serious barrier for studying many complex proteins.
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Problems with Protein A binding my mouse IgG1 type MAbs can be circumvented by binding the MAbs to Protein A sepharose in Biorad’s MAPS buffer followed by 2 rinses in 0.2 M triethanolamine, pH 8.2 and covalently cross-linking as previously described. Biorad MAPS buffer contains components which inhibit cross-linking by DMP and so it must be removed completely. Even with these modifications, some IgG1 MAbs cannot be bound to Protein A sepharose in adequate amounts.
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© 1987 Plenum Press, New York
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Bricker, B.J., Wagner, R.R., Fox, J.W. (1987). Immunoprotection — A Novel Approach for Mapping Epitopes on an Antigen. In: L’Italien, J.J. (eds) Proteins. Springer, Boston, MA. https://doi.org/10.1007/978-1-4613-1787-6_48
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DOI: https://doi.org/10.1007/978-1-4613-1787-6_48
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