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Journal of Clinical Microbiology, July 2001, p. 2418-2424, Vol. 39, No. 7
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.7.2418-2424.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Combinatorial Use of Antibodies to Secreted Mycobacterial Proteins in a Host Immune System-Independent Test for Tuberculosis

Christopher P. Landowski,^1, Henry P. Godfrey,^2,* Stuart I. Bentley-Hibbert,² Xinyan Liu,² Zhishan Huang,² Ricardo Sepulveda,³ Kris Huygen,⁴ Maria L. Gennaro,⁵ Fred H. Moy,² Scott A. Lesley,^1, and Mary Haak-Frendscho¹

Immunology and Neurobiology R & D, Promega Corporation, Madison, Wisconsin 53711¹; Department of Pathology, New York Medical College, Valhalla, New York 10595²; National Chest Institute, Santiago, Chile³; Pasteur Institute of Brussels, Brussels, Belgium⁴; and Public Health Research Institute, New York, New York 11021⁵

Received 7 December 2000/Returned for modification 26 February 2001/Accepted 24 April 2001

Laboratory diagnosis of tuberculosis is often difficult. Immunodetection of circulating Mycobacterium tuberculosis proteins shed during active infection would not depend on an intact host immune response and could take advantage of the speed and low costs afforded by antibody-based assays. We previously showed that patients with active tuberculosis had increased levels of circulating antigen 85 (Ag85) proteins independent of their tuberculin skin test status (S. I. Bentley-Hibbert, X. Quan, T. Newman, K. Huygen, and H. P. Godfrey, Infect. Immun. 67:581-588, 1999). To extend these observations to a Mycobacterium bovis BCG-vaccinated population and to another secreted mycobacterial protein, Ag85 and PstS-1 (protein antigen B, p38 antigen) were quantified in sera from 97 Chilean tuberculosis patients and healthy controls (many of whom had received BCG as children) using dot immunobinding, mouse monoclonal anti-BCG Ag85 complex antibody, and chicken antipeptide antibodies reactive with M. tuberculosis Ag85B and PstS-1. The latter antibodies had been raised to peptide-derived immunogens expressed on a novel proprietary protein carrier in Escherichia coli. Median serum Ag85 levels measured by using either anti-Ag85 antibody were significantly higher in patients with active tuberculosis than in healthy controls (P, <0.001 to 0.01); the median serum PstS-1 levels were similar in patients and controls. The sensitivity of significantly elevated circulating Ag85 levels in patients with pulmonary tuberculosis measured by anti-Ag85 complex or anti-Ag85B antibodies was 60 and 55%, respectively, but increased to 77% when results obtained with both anti-Ag85 antibodies were considered jointly (P < 0.02). The corresponding specificities for individual and joint consideration were 95, 85, and 80%, respectively. These results indicate that elevated Ag85 levels can be detected in patients with active tuberculosis even after BCG vaccination and suggest that combinatorial use of antibodies directed at different epitopes of this protein could provide a viable strategy for developing new host immune response-independent diagnostic tests for tuberculosis.

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^* Corresponding author. Mailing address: Department of Pathology, New York Medical College, Basic Science Building, Valhalla, NY 10595. Phone: (914) 594-4160. Fax: (914) 594-4163. E-mail: hgodfrey{at}nymc.edu.

Present address: Department of Pharmaceutical Sciences, University of Michigan, Ann Arbor, MI 48108.

Present address: Genomics Institute, Novartis Research Foundation, San Diego, CA 92121.

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Journal of Clinical Microbiology, July 2001, p. 2418-2424, Vol. 39, No. 7
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.7.2418-2424.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

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