Objective To investigate the diagnostic value of T cell enzyme-linked immunospot (ELISPOT) assay in tuberculosis patients. Methods 103 patients with Mycobacterium tuberculosis (MTB) infection and 30 controls with non-MTB infection were recruited to detect early secreting antigen target-6(ESAT-6) and culture filtrate protein-10(CFP-10) specific T cells in blood samples by ELISPOT assay, then compare it with serum tuberculosis antibody(TB-Ab) and PPD skin test. Results The sensitivity, specificity, positive prospective value, negative prospective value, positive likelihood ratio and negative likelihood ratio of T cell ELISPOT assay were 95.15%, 93.33%, 98.00%, 84.85 %, 14.27% , and 0.05%, respectively. All these indexes were significantly higher than those of TB-Ab and PPD skin test(P<0.01), meanwhile T cell ELISPOT assay had low agreement with means of TB-Ab and PPD skin test(Kappa＜0.40, P<0.001). Conclusions T cell ELISPOT assay has a higher sensitivity and specificity than conventional test in the diagnosis of tuberculosis; therefore,it has great value and applicability as a screening test.