Monoclonal Antibodies to Intermediate Filament Proteins of Human Cells: Unique and Cross-Reacting Antibodies

• Published in: The Journal of cell biology 1982-11, Vol.95 (2), p.414-424
• Main Authors: Gown, Allen M., Vogel, Arthur M.
• Format: Article
• Language: English
• Subjects: Antibodies; Antibodies, Monoclonal - immunology; Carcinoma, Hepatocellular; Cell Line; Cell lines; Cross Reactions; Epidermis; Epithelial cells; Epitopes; Fibroblasts; Fluorescent Antibody Technique; Glial Fibrillary Acidic Protein; Humans; Hybridomas; Intermediate filament proteins; Intermediate Filament Proteins - analysis; Intermediate Filament Proteins - immunology; Intermediate filaments; Keratins; Keratins - immunology; Liver Neoplasms; Molecules; Peptides - analysis; Reactivity; Vimentin
• ISSN: 0021-9525; 1540-8140
• DOI: 10.1083/jcb.95.2.414

Monoclonal antibodies were generated against the intermediate filament proteins of different human cells. The reactivity of these antibodies with the different classes of intermediate filament proteins was determined by indirect immunofluorescence on cultured cells, immunologic identification on SDS polyacrylamide gels ("western blot" experiments), and immunoperoxidase assays on intact tissues. The following four antibodies are described: (a) an antivimentin antibody generated against human fibroblast cytoskeleton; (b), (c) two antibodies that recognize a 54-kdalton protein in human hepatocellular carcinoma cells; and (d) an antikeratin antibody made to stratum corneum that recognizes proteins of molecular weight 66 kdaltons and 57 kdaltons. The antivimentin antibody reacts with vimentin (58 kdaltons), glial fibrillary acidic protein (GFAP), and keratins from stratum corneum, but does not recognize hepatoma intermediate filaments. In immunofluorescence assays, the antibody reacts with mesenchymal cells and cultured epithelial cells that express vimentin. This antibody decorates the media of blood vessels in tissue sections. One antihepatoma filament antibody reacts only with the 54 kdalton protein of these cells and, in immunofluorescence and immunoperoxidase assays, only recognizes epithelial cells. It reacts with almost all nonsquamous epithelium in intact tissue, but fails to recognize squamous epithelium. Thus this antibody identifies an epithelial-specific "cytokeratin" molecule present in only nonsquamous epithelium. The other antihepatoma filament antibody is much less selective, reacting with vimentin, GFAP, and keratin from stratum corneum. This antibody decorates intermediate filaments of both mesenchymal and epithelial cells. The antikeratin antibody recognizes 66-kdalton and 57-kdalton proteins in extracts of stratum corneum and also identifies proteins of similar molecular weights in all cells tested. However, by immunofluorescence, this antibody decorates only the intermediate filaments of epidermoid carcinoma cells. When assayed on tissue sections, the antibody reacts with squamous epithelium and some, but not all, nonsquamous epithelium. Therefore this antistratum corneum antibody and the anti-54-kdalton antibody identify unique epitopes present in the various cytokeratin molecules of epithelial cells. None of the hybridoma antibodies react with neurofilament proteins. The different patterns of reactivity of these antibodies suggest that ma