Tuberculosis has been declared a global emergency. The mainstay for its control is the rapid and accurate identification of infected individual.
Antibodies to A60, one of the macromolecular
antigen complexes of mycobacteria were commonly used in the rapid detection of Mycobacterium tuberculosis. The aim of this study was to prepare specific
antibodies against A60 for detection of
tuberculosis infection. Specific polyclonal
antibodies against A60, (A60-Ab) were prepared in rabbits using 2 boosted
injections of the
antigen (A60). The
antibodies were purified and treated with normal oral flora to remove any non-specific and cross-reactive
antibodies. These
antibodies were conjugated to CNBr-activated
Sepharose 4B and used to isolate subunits of A60 with more specificity for M.
tuberculosis. A new affinity column was designed to prepare modified (purified) A60
antigen. Purified A60
antigen (PA60-Ag) was used to develop antibody production by Immunoaffinity chromatography. 113 patients with a confirmed diagnosis of pulmonary TB at Pasteur Institute were selected for the study. The specificity of the results was analyzed with TB-rapid test by using PA60-antibodies. TB-rapid test revealed that normal oral flora-absorbed
antibodies could lead to more specific results than that of the non-absorbed
antibodies. The developed, modified A60
antibodies, (PA60-Ab)-rapid test showed higher sensitivity, specificity, Positive Predictive Value (PPV), Negative Predictive Value (NPV) and overall efficiency (93.0%, 86.0%, 90.0%, 91.0%, and 90.0% respectively) for the detection of the Mycobacterium
antigen. Moreover, PA60-Ag showed only two
protein bands of molecular weight 45 and 66kDa in SDS-PAGE while untreated A60 showed multiple bands. Thus, our study helped in the purification of a novel and well characterized A60
antigen and good diagnostic potential for detecting
tuberculosis infection.