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Clinical and Diagnostic Laboratory Immunology, July 2000, p. 600-606, Vol. 7, No. 4
1071-412X/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Cloning and Expression of Immunoreactive Antigens from Mycobacterium tuberculosis

Renee Lay Hong Lim,1,* Li Kiang Tan,1 Wai Fun Lau,1 Maxey Ching Ming , Chung,1,2 Roseanne Dunn,1,dagger Heng Phon Too,2 and Lily Chan1

Bioprocessing Technology Centre1 and Department of Biochemistry,2 The National University of Singapore, Singapore

Received 1 December 1999/Returned for modification 8 February 2000/Accepted 4 April 2000

Four immunoreactive proteins, B.4, B.6, B.10, and B.M, with molecular weights ranging from 16,000 to 58,000, were observed from immunoblots of Mycobacterium tuberculosis total lysates screened with sera from individuals with active tuberculosis. These proteins were identified from microsequence analyses, and genes of proteins with the highest homology were PCR amplified and cloned into the pQE30 vector for expression studies. In addition, a 37.5-kDa protein, designated C17, was identified from a phage expression library of M. tuberculosis genomic DNA. Preliminary immunoblot assays indicated that these five resultant recombinant proteins could detect antibodies in individuals with active pulmonary and extrapulmonary tuberculosis. The overall ranges of sensitivities, specificities, positive predictive values, and negative predictive values for the recombinant antigens were 20 to 58, 88 to 100, 69 to 100, and 56 to 71%, respectively. The B.6 antigen showed preferential reactivity to antibodies in pulmonary compared to nonpulmonary tuberculosis serum specimens. All of these recombinant antigens demonstrated potential for serodiagnosis of tuberculosis.

* Corresponding author. Mailing address: Bioprocessing Technology Centre, National University of Singapore, 5th Floor, MD11, 10 Kent Ridge Crescent, Singapore 119260. Phone: 65-874-6222. Fax: 65-775 4933. E-mail: btclimr{at}nus.edu.sg.

dagger Present address: Department of Cell and Molecular Biology, University of Technology, Sydney, NSW, Australia.

Clinical and Diagnostic Laboratory Immunology, July 2000, p. 600-606, Vol. 7, No. 4
1071-412X/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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