Development of a quantitative sandwich enzyme-linked immunosorbent assay for detecting the MPT64 antigen of Mycobacterium tuberculosis.
OBJECTIVE
Tuberculosis (TB) is a major infectious disease and is responsible for two million deaths annually. For the identification and quantitation of Mycobacterium tuberculosis (M. tuberculosis), a causative agent of TB, a sandwich enzyme-linked immunosorbent assay (ELISA) against the MPT64 protein of M. tuberculosis, an antigen marker of the M. tuberculosis complex, was developed.


METHODS
The MPT64 protein was expressed, and anti-MPT64 monoclonal antibodies were prepared. A sandwich ELISA was established using recombinant MPT64 protein and anti-MPT64 monoclonal antibodies. The sandwich MPT64 ELISA was evaluated using reference and clinical mycobacterial strains.


RESULTS
The sandwich MPT64 ELISA detected MPT64 protein from 2.1 ng/mL to 250 ng/mL (equivalent to 1.7×10⁴ CFU/mL and 2.0×10⁶ CFU/mL). All 389 clinical M. tuberculosis isolates tested positive in the sandwich MPT64 ELISA (sensitivity, 100%), and the assay showed no cross reactivity to any tested nontuberculous mycobacterial strain (specificity, 100%).


CONCLUSIONS
The sandwich MPT64 ELISA is a highly sensitive and quantitative test for MPT64 protein, which can identify M. tuberculosis.
Department of Life Science, College of Natural Science, Kyonggi University, Suwon, Korea. ; Biotech Laboratory, Standard Diagnostics., Yongin, Korea.
Mijung
M
Ji
Biotech Laboratory, Standard Diagnostics., Yongin, Korea.
Byungki
B
Cho
Biotech Laboratory, Standard Diagnostics., Yongin, Korea.
Young Shik
YS
Cho
Division of Biological Science and Technology, College of Science and Technology, Yonsei University, Wonju, Korea.
Song-Yong
SY
Park
Department of Microbiology, Yonsei University College of Medicine, Seoul, Korea.
Sang-Nae
SN
Cho
Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University, Wonju, Korea.
Bo-Young
BY
Jeon
Department of Life Science, College of Natural Science, Kyonggi University, Suwon, Korea.
Byoung-Su
BS
Yoon
D000942:Antigens, Bacterial / Q000032:analysis* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D009169:Mycobacterium tuberculosis / Q000276:immunology*
D000942:Antigens, Bacterial; C510537:MPT64 protein, Mycobacterium tuberculosis
MPT64 protein; Mycobacterium tuberculosis; sandwich ELISA