In-vitro assessment of cell-mediated immunity by demonstrating effector-t cells for diagnosis of tuberculosis in Nepalese subjects.

The immune response against mycobacterium tuberculosis (MTB) is cell mediated. T-cells become sensitized when they encounter MTB antigens and subsequently activated effector T-cells produce a number of cytokines including interferon- gamma (INF-gamma) to fight the infecting organisms. Demonstration of either production of INF-gamma or presence of effector T-cells sensitized to MTB specific antigens in vitro can be diagnostic for TB infection. Aim of this study was to determine the efficacy of commercially available T-SPOT.TB kit which is used for the in vitro diagnosis of TB infection and to determine if this test has any cross reactivity in leprosy patients. Blood sample was taken from 30 sputum AFB positive, 30 sputum AFB negative healthy controls and 10 cases of paucibacillary leprosy patients. The blood samples were processed to separate peripheral blood mononuclear cells. The final cell suspensions were cultured along with MTB specific antigens namely- Early Secretory Antigenic Target (ESAT-6) and Culture Filtrate Protein (CFP 10) along with negative and positive controls. The production of INF-gamma was demonstrated by enzyme linked immunospot (ELISPOT) assay technique. All AFB positive samples produced INF-gamma after exposure to MTB specific antigens. 4 (16.6%) of healthy controls were also found reactive for INF-gamma. The sensitivity and "specificity" for active disease of the ELISPOT (T-SPOT.TB) in respect to AFB microscopy was 100% and 85.7% respectively. Assessment of CMI against tuberculosis, by demonstrating effector T-cell sensitized to MTB antigens can be use to aid the diagnosis of tuberculosis. T-SPOT.TB has no cross reactivity with leprosy patients.