Efficient ELISA for Diagnosis of Active Tuberculosis Employing a Cocktail of Secretory Proteins of <i>Mycobacterium tuberculosis</i>

Tiwari, D.; Tiwari, R. P.; Chandra, R.; Bisen, P. S.; Haque, Shafiul

doi:10.14712/fb2014060010010

Folia Biologica > Archive > 2014, Vol. 60 > Issue 1 > Efficient ELISA for Diagnosis of Active…

Fol. Biol. 2014, 60, 10-20

https://doi.org/10.14712/fb2014060010010

Efficient ELISA for Diagnosis of Active Tuberculosis Employing a Cocktail of Secretory Proteins of Mycobacterium tuberculosis

D. Tiwari^1,2,3, R. P. Tiwari⁴, R. Chandra², P. S. Bisen⁵, Shafiul Haque⁶

¹Catalysis and Peptide Research Unit, School of Health Sciences, University of KwaZulu-Natal, Durban, South Africa
²Dr. B.R. Ambedkar Centre for Biomedical Research, University of Delhi, Delhi, India
³School of Chemistry & Physics, University of KwaZulu-Natal, Durban, South Africa
⁴Department of Biotechnology, Immunodiagnostic Division, Aspen Laboratories Pvt. Ltd., Delhi, India
⁵School of Studies in Biotechnology, Jiwaji University, Gwalior, MP, India
⁶Department of Biosciences, Jamia Millia Islamia (A Central University), New Delhi, India

Received July 2013
Accepted September 2013

Rapid and accurate diagnosis is important for preventing transmission of Mycobacterium tuberculosis. Currently available tuberculosis (TB) diagnostic methods lack desired sensitivity and specificity, and require sophisticated equipment and skilled workforce including weeks’ long duration to yield results. In this study, extracellular proteins or secretory protein antigens of M. tuberculosis H37Rv have been isolated using ion exchange chromatography, immunocharacterized and exploited for the development of efficient enzyme-linked immunosorbent assay (ELISA) for diagnosis of active TB with enhanced specificity and sensitivity. Apparent molecular masses for purified proteins were found to be 6, 27, 30, 38 and 64 kDa. Out of five purified proteins, one (64 kDa) was found to be novel. Of the five proteins, four (6, 27, 30 and 38 kDa) were found significant to be used in the development of ELISA for pulmonary and extra-pulmonary TB. The immune responses of serum samples of TB patients and other healthy subjects against the above-mentioned antigens’ cocktail were evaluated. Critical parameters of newly developed ELISA were optimized and it was observed that the cocktail antigens have a greater specificity (98.06 %) and sensitivity (98.67 %) as compared to other commercially available diagnostic tests. The present findings suggest that the developed ELISA is an effective tool for routine screening and early-stage diagnosis of TB.

Keywords

tuberculosis, culture filtrate proteins, ELISA, MDR, secretory antigens, serodiagnosis.

Funding

The financial support for the research work was jointly provided by the Zydus Research Centre, Ahmedabad, Gujarat, India, Dr. B.R. Ambedkar Centre for Biomedical Research, University of Delhi, Delhi, India and the School of Studies in Biotechnology, Jiwaji University, Gwalior, MP, India.