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Updated: Sep 23, 2026
Capillary Electrophoresis Separation of Monoclonal Antibody Isoforms Using a Neutral Capillary
Published on: January 16, 2017
Monoclonal antibodies to human tumor necrosis factors alpha and beta: application for affinity purification,
1Department of Molecular Immunology, Genentech, Inc., South San Francisco, CA 94080.
Monoclonal antibodies were produced against two structurally related tumor necrosis factors (TNFs), TNF-alpha (previously called tumor necrosis factor) and TNF-beta (previously called lymphotoxin). The potential of these antibodies for the purification of TNFs, the development of specific immunoassays, and for defining the antigenic and functional domains of these cytokines was investigated. None of the monoclonal antibodies cross-reacted with both TNF-alpha and TNF-beta, or reacted with synthetic peptides which represented several of the regions of homology between these cytokines. Neutralizing monoclonal antibodies were utilized as immunoadsorbents to purify recombinant TNF-alpha and TNF-beta from E. coli lysates. TNFs purified by this method were greater than 98 percent pure by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and exhibited specific activities that were the same as TNFs isolated from natural sources using conventional chromatographic techniques. In addition, specific ELISA assays were developed that could detect less than 1 ng/ml of TNF-alpha or TNF-beta, and in contrast to bioassays, could discriminate between these related cytokines.
Monoclonal antibodies were produced against two structurally related tumor necrosis factors (TNFs), TNF-alpha (previously called tumor necrosis factor) and TNF-beta (previously called lymphotoxin). The potential of these antibodies for the purification of TNFs, the development of specific immunoassays, and for defining the antigenic and functional domains of these cytokines was investigated. None of the monoclonal antibodies cross-reacted with both TNF-alpha and TNF-beta, or reacted with synthetic peptides which represented several of the regions of homology between these cytokines. Neutralizing monoclonal antibodies were utilized as immunoadsorbents to purify recombinant TNF-alpha and TNF-beta from E. coli lysates. TNFs purified by this method were greater than 98 percent pure by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and exhibited specific activities that were the same as TNFs isolated from natural sources using conventional chromatographic techniques. In addition, specific ELISA assays were developed that could detect less than 1 ng/ml of TNF-alpha or TNF-beta, and in contrast to bioassays, could discriminate between these related cytokines.
