Development of an enzyme-linked immunosorbent assay (ELISA) combined with a streptavidin-biotin and enzyme amplification method to detect anti-2,3-di-o-acyltrehalose (DAT) antibodies in patients with tuberculosis

J Immunol Methods. 1994 Dec 28;177(1-2):69-77. doi: 10.1016/0022-1759(94)90144-9.


IgG antibodies against the 2,3-di-o-acyltrehalose glycolipid of Mycobacterium tuberculosis were determined in a set of 49 sera from patients with pulmonary tuberculosis and 65 negative control subjects. We compared a conventional ELISA method using a beta-galactosidase anti-human IgG conjugate developed with ONPG, with an amplification ELISA system constituted of an anti-human IgG biotinylated conjugate, a streptavidin-alkaline phosphatase complex, and NADP as a substrate. The resulting NAD was measured by using a redox enzymatic recycling system of alcohol dehydrogenase, diaphorase and iodonitrotetrazolium as chromogen. With specificity set at 92.31% in both methods, we obtained a sensitivity of 42.86% in the conventional method and a sensitivity of 61.22% in the amplified method. We conclude that by using a more sensitive method we can detect cases that otherwise could be identified as false negatives.

MeSH terms

  • Antibodies, Bacterial / analysis*
  • Antigens, Bacterial / immunology*
  • Bacterial Proteins
  • Biotin
  • Dose-Response Relationship, Immunologic
  • Enzyme-Linked Immunosorbent Assay / methods
  • Glycolipids / immunology*
  • Humans
  • Mycobacterium tuberculosis / immunology*
  • Polysaccharides, Bacterial / immunology*
  • Streptavidin
  • Trehalose / immunology*
  • Tuberculosis / immunology*


  • Antibodies, Bacterial
  • Antigens, Bacterial
  • Bacterial Proteins
  • Glycolipids
  • Polysaccharides, Bacterial
  • glycolipid IV, Mycobacterium tuberculosis
  • Biotin
  • Streptavidin
  • Trehalose