Impact of protein shedding on detection of Mycobacterium avium subsp paratuberculosis by a whole-cell immunoassay incorporating surface-enhanced Raman scattering

TitleImpact of protein shedding on detection of Mycobacterium avium subsp paratuberculosis by a whole-cell immunoassay incorporating surface-enhanced Raman scattering
Publication TypeJournal Article
Year of Publication2008
AuthorsYakes BJ, Lipert RJ, Bannantine JP, Porter MD
Journal TitleClinical and Vaccine Immunology
Volume15
Pages235-242
Date PublishedFeb
Type of ArticleArticle
ISBN Number1556-6811
Accession NumberISI:000258666600009
KeywordsCATTLE, DIAGNOSIS, FECAL CULTURE, INFECTION, JOHNES-DISEASE, MEDIATED CAPTURE PCR, nanoparticles, RAPID DETECTION, spectroscopy, TESTS
Abstract

The etiological agent of Johne's disease is Mycobacterium avium subsp. paratuberculosis. Controlling the spread of this disease is hindered by the lack of sensitive, selective, and rapid detection methods for M. avium subsp. paratuberculosis. By using a recently optimized sandwich immunoassay (B. J. Yakes, R. J. Lipert, J. P. Bannantine, and M. D. Porter, Clin. Vaccine Immunol. 15: 227-234, 2008), which incorporates a new monoclonal antibody for the selective capture and labeling of M. avium subsp. paratuberculosis and surface-enhanced Raman scattering for sensitive readout, detection limits of similar to 630 and similar to 740 M. avium subsp. paratuberculosis cells/ml are achieved in phosphate-buffered saline and whole milk samples, respectively, after spiking with heat-treated M. avium subsp. paratuberculosis. Surprisingly, these detection limits are 3 orders of magnitude lower than expected based on theoretical predictions. Experiments designed to determine the origin of the improvement revealed that the major membrane protein targeted by the monoclonal antibody was present in the sample suspensions as shed protein. This finding indicates that the capture and labeling of shed protein function as a facile amplification strategy for lowering the limit of detection for M. avium subsp. paratuberculosis that may also be applicable to the design of a wide range of highly sensitive assays for other cells and viruses.

DOI10.1128/cvi.00335-07
Alternate JournalClin. Vaccine Immunol.