Department of Radiology, Division of Cell Biology and Imaging
Amino Acids, Peptides, and Proteins | Cancer Biology | Cell Biology | Enzymes and Coenzymes | Immunology and Infectious Disease | Neoplasms
Supervillin isoforms have been implicated in cell proliferation, actin filament-based motile processes, vesicle trafficking, and signal transduction. However, an understanding of the roles of these proteins in cancer metastasis and physiological processes has been limited by the difficulty of obtaining specific antibodies against these highly conserved membrane-associated proteins. To facilitate research into the biological functions of supervillin, monoclonal antibodies were generated against the bacterially expressed human supervillin N-terminus. Two chimeric monoclonal antibodies with rabbit Fc domains (clones 1E2/CPTC-SVIL-1; 4A8/CPTC-SVIL-2) and two mouse monoclonal antibodies (clones 5A8/CPTC-SVIL-3; 5G3/CPTC-SVIL-4) were characterized with respect to their binding sites, affinities, and for efficacy in immunoblotting, immunoprecipitation, immunofluorescence microscopy and immunohistochemical staining. Two antibodies (1E2, 5G3) recognize a sequence found only in primate supervillins, whereas the other two antibodies (4A8, 5A8) are specific for a more broadly conserved conformational epitope(s). All antibodies function in immunoblotting, immunoprecipitation and in immunofluorescence microscopy under the fixation conditions identified here. We also show that the 5A8 antibody works on immunohistological sections. These antibodies should provide useful tools for the study of mammalian supervillins.
Immunoprecipitation, Cloning, Immunohistochemistry techniques, Hybridomas, Monoclonal antibodies, Enzyme-linked immunoassays, Immunoblotting, Cell staining
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Copyright: © 2018 Smith et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
DOI of Published Version
PLoS One. 2018 Oct 17;13(10):e0205910. doi: 10.1371/journal.pone.0205910. eCollection 2018. Link to article on publisher's site
Smith TC, Saul RG, Barton ER, Luna EJ. (2018). Generation and characterization of monoclonal antibodies that recognize human and murine supervillin protein isoforms. Open Access Publications by UMass Chan Authors. https://doi.org/10.1371/journal.pone.0205910. Retrieved from https://escholarship.umassmed.edu/oapubs/3628
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This work is licensed under a Creative Commons Attribution 4.0 License.