Indian Journal of Clinical Biochemistry

, Volume 13, Issue 2, pp 98–105 | Cite as

Mycobacterium tuberculosis H37Ra ESAS-7—An excretory-secretory antigen fraction of immunodiagnostic potential in pulmonary tuberculosis

  • E. Raji Nair
  • Satish Kumar
  • M. V. R. Reddy
  • B. C. Harinath


A mycobacterial excretory-secretory protein fraction ESAS-7 purified by 50% ammonium sulphate precipitation followed by SDS-PAGE fractionation was evaluated by penicillinase enzyme linked immuno-sorbent assay (ELISA) for its sensitivity and specificity in the diagnosis of pulmonary tuberculosis. At a “cut off” serum dilution of 600, 38 (90%) of 42 sera from bacteriologically confirmed tuberculosis cases, 15 (100%) of 15 sera from bacteriogically negative but anti tubercular therapy (ATT) responded cases, 3 (7%) of 43 sera from normal healthy subjects and 4 (8%) of 48 sera from non tuberculous disease control cases gave positive reaction for tubercular antibody to ESAS-7 antigen fraction containing predominantly 33-kDa protein with a sensitivity of 90% in bacteriologically confirmed cases and specificity of 92%. Further, this diagnostic assay using the ESAS-7 antigen is more sensitive requiring as little as one nanogram antigen per test compared to use of 100 nanogram EST-6 antigen reported earlier. Thus use of ESAS-7 antigen for antibody detection has good diagnostic potential with improved specificity in pulmonary tuberculosis.

Key Words

ELISA ESAS—7 antigen Penicillinase 


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  1. 1.
    Murray, C.J.L., Styblo, K. and Rouillon, A. (1990) Tuberculosis in developing countries, burden, intervention and cost. Bull. Int. Union. Tuberc. Lung. Dis. 65, 6–24.PubMedGoogle Scholar
  2. 2.
    CDS. (1991) Nosocomial transmission of MDR-TB among HIV infected persons—Florida and New York. M.M.W.R. 40, 585–591.Google Scholar
  3. 3.
    Benjamin, R.G., Debanne, S.M., Ma, Y. and Daniel, T.M. (1984) Evaluation of mycobacterial antigen in an enzyme linked immunosorbent assay (ELISA) for the serodiagnosis of tuberculosis. J. Med. Microbiol. 18, 309–318.PubMedGoogle Scholar
  4. 4.
    Krombovitis, E. (1986) Detection of antibodies toMycobacterium tuberculosis plasma membrane antigen by enzyme linked immunosorbent assay. J. Med. Microbiol. 21, 257–264.Google Scholar
  5. 5.
    Stroebal, A.B., Daniel, T.M., Lau, J.H.K., Leong, J.C.Y. and Richardson, H. (1982) Serological diagnosis of bone and joint tuberculosis by an enzyme linked immunosorbent assay. J. Infect. Dis. 146, 280–283.Google Scholar
  6. 6.
    Nagai, S., Wiker, H.G., Harboe, M. and Kinomoto, M. (1991) Isolation and partial characterization of major protein antigens in culture fluid ofMycobacterium tuberculosis. Infect. Immun. 59, 372–382.PubMedGoogle Scholar
  7. 7.
    Basak A., Choudhary, S., Lodam, A.N., Gupta, O.P., Narang, P., and Harinath, B.C. (1992) Detection of IgG using Mycobacterium tuberculosis H37Ra by penicillinase stick ELISA. In: proceedings of CSIR Golden jubilee symposium of Tropical Diseases; Molecular Biology and control strategies. Feb. 19–20, Lucknow.Google Scholar
  8. 8.
    Bhaskar, A., Pradhan, P., Chaturvedi, P., Basak, A., Lodam, A.N., Narang, P. and Harinath, B.C. (1994) Immunodiagnosis of childhood pulmonary and extrapulmonary tuberculosis usingMycobacterium tuberculosis ES antigen by penicllinase ELISA. Ann. Trop. Paediatr. 14, 25–30.PubMedGoogle Scholar
  9. 9.
    Kumar, S., Chenthamarkshan, V., Reddy, M.V.R., Narang, P., Gupta, O.P. and Harinath, B.C. (1994) Detection of tuberculous IgG antibodies usingMycobacterium tuberculosis H37Ra excretory-secretory antigen and tuberculin purified protein derivative. Indian. J. Expt. Biol. 32, 163–167.Google Scholar
  10. 10.
    Lodam, A.N., Reddy, M.V.R., Narang, P., Gupta, O.P. and Harinath, B.C. (1996) Fractionation, analysis and diagnostic utility ofMycobacterium tuberculosis H37Ra excretory-secretory antigen in pulmonary tuberculosis. Indian. J. Biochem. Biophy. 33, 66–71.Google Scholar
  11. 11.
    Lodam, A.N., Pramanik, J., Reddy, M.V.R. and Harinath, B.C. (1997) Diagnostic potential of fractionatedMycobacterium tuberculosis H37Ra excretory—secretory (EST-DE1) antigen in pulmonary tuberculosis. Indian. J. Clin. Biochem. 12 (1), 71–77.Google Scholar
  12. 12.
    Pramanik, J., Lodam, A.N., Reddy, M.V.R., Narang, P. and Harinath, B.C. (1997) Increased yield of excretory-secretory antigen with thyroxine supplemented ininvitro culture of tubercle bacilli. Indian. J. Tuberc. 44, 185–190.Google Scholar
  13. 13.
    Lodam, A.N., Reddy, M.V.R. and Harinath, B.C. (1998) Immunodiagnosis of pulmonary tuberculosis by concomitant detection of antigen and antibodies of excretory-secretory protein ofMycobacterium tuberculosis H37Ra. J. Biosci. 1, 19–23.CrossRefGoogle Scholar
  14. 14.
    Lowry, O.H., Rosebrough, N.J., Farr, A.L. and Randall, R.J. (1951) Protein measurement with Folin phenol reagent. J. Biol. Chem. 193, 265–275.PubMedGoogle Scholar
  15. 15.
    Avrameas, S. (1969) Coupling of enzymes to proteins with glutaraldehyde—Use of conjugates for the detection of antigen and antibodies. Immuno. Chemistry. 6, 43–52.CrossRefGoogle Scholar
  16. 16.
    Parkhe, K.A., Prasad, G.B.K.S., Das, A., Harinath, B.C., Roebbar, M. and Hamilton, R.G. (1986) Disc/Stick ELISA for diagnosis of bancroftian filariasis. Indian. J. Expt. Biol. 24, 437–439.Google Scholar
  17. 17.
    Kaplan, M.H. and Chase, M.W. (1980) Antibodies to mycobacteria in human tuberculosis. 1. Development of antibodies before and after antimycobacterial therapy. J. Infect. Dis. 142 (6), 825–834.PubMedGoogle Scholar
  18. 18.
    Radhakrishnan, V.V. and Mathai, A. (1990) Detection of mycobacterial antigen in cerebrospinal fluid in patients with chronic meningitis by inhibition ELISA. Indian. J. Med. Res. 91, 355–359.PubMedGoogle Scholar
  19. 19.
    Daniel, T.M. and Debanne, S.M. (1987) The serodiagnosis of tuberculosis and other mycobacterial disease by enzyme linked immunosorbent assay. Amer. Rev. Respir. Dis. 135, 1137–1151.Google Scholar
  20. 20.
    Mathai, A., Radhakrishnan, V.V. and Sehgal, S. (1990) IgG antibody toM. tuberculosis antigen 5 in cerebrospinal fluid and its diagnostic application in tuberculous meningitis. Indian. J. Exp. Biol. 28, 816–820.PubMedGoogle Scholar
  21. 21.
    Reggiardo, A., Vazques, E. and Schnaper, L. (1980) ELISA test for antibodies against mycobacterial glycolipids. J. Immunol. Methods. 34, 55–60.PubMedCrossRefGoogle Scholar
  22. 22.
    Sada, E., Ferguson, L.E. and Daniel, T.M. (1990) An ELISA for the Serodiagnosis of Tuberculosis using a 30,000-Da Native Antigen ofMycobacterium tuberculosis J. Infect. Dis. 162, 928–931.PubMedGoogle Scholar
  23. 23.
    Thongkrajai, P., Lulitanon, V. and Chamnavanki, C. (1989) Improved ELISA with immunosorbent purified mycobacterial antigens for serodiagnosis of tuberculosis. J. Med. Microbiol. 30, 101–104.PubMedCrossRefGoogle Scholar
  24. 24.
    Das, P.K., Rambukkana, A., Baas, J.G., Groothuis, D.G. and Halperin, M. (1990) ELISA for distingnuishing serological responses of lepramatous and tuberculoid leprosies to the 29/33 kDa doublet and 64 antiges of M.tb. J. Clin. Microbiol. 28(2), 379–382.PubMedGoogle Scholar
  25. 25.
    Alamelu, R., Narayanan P.R., Jawahar, M.S. and Prabakar, R. (1994) Evaluation ofMycobacterium tuberculosis antigen 6 by enzyme linked immunosorbent assay (ELISA). Indian. J. Tub. 41, 245–251.Google Scholar

Copyright information

© Association of Clinical Biochemists of India 1998

Authors and Affiliations

  • E. Raji Nair
    • 1
  • Satish Kumar
    • 1
  • M. V. R. Reddy
    • 1
  • B. C. Harinath
    • 1
  1. 1.Jamnalal Bajaj Tropical Disease Research Centre and Department of BiochemistryMahatma Gandhi Institute of Medical SciencesWardha

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