Abstract: Objective To study the frequency and clinical significance of antigen-specific multifunctional Th1 cells in the peripheral blood and pleural effusion of tuberculosis (TB) patients.Methods Ninety-three patients with active TB(including 27 patients with tuberculous pleurisy), 30 latent TB infection individuals (LTBI) and 66 healthy controls (HD) were collected in this study. Separate the peripheral blood monouclear cells (PBMCs) from all of the individuals and also the pleural fluid mononuclear cells (PFMCs) from the tuberculous pleurisy patients. The PBMCs and PFMCs were co-cultured with the specific antigen of M. tuberculosis (Mtb), and then detected the frequency of multifunctional CD4⁺ T cells (including of 7 subgroups: IL-2⁺IFN-γ⁺TNF-α⁺,IL-2⁺IFN-γ⁺,IL-2⁺ TNF-α⁺,IFN-γ⁺TNF-α⁺,IL-2⁺,TNF-α⁺,IFN-γ⁺ CD4⁺ T cells) using intracellular cytokine staining and flow cytometric analysis. The results were showed as (x±s) and all data were analyzed by using GraphPad Prism 4.0. The Wilcoxon matched pair t test was used to analyze the antigen-specific multifunctional Th1 cells in the peripheral blood and pleural effusion. The ANOVA was used to analyze the differences of antigen-specific multifunctional Th1 cells in HD,TB and LTBI. Differences were considered significant when the P value was less than 0.05.Results The frequency of antigen-specific multifunctional Th1 cell subsets (IL-2⁺IFN-γ⁺TNF-α⁺CD4⁺T cells) in TB patients was (0.107±0.278)%, which was higher than that in LTBI(0.019±0.032)% and HD(0.008±0.016)%(F=5.675, P=0.004). The frequency of IL-2⁺TNF-α⁺ CD4⁺ T cells was (0.049±0.123)% in TB patients and (0.046±0.050)% in LTBT, those were higher than that in HD (0.003±0.014)% (F=5.435, P=0.005). The frequency of IFN-γ⁺TNF-α⁺ CD4⁺ T cells and IL-2⁺IFN-γ⁺ CD4⁺ T cells in TB patients were (0.136±0.256)% and (0.146±0.347)%, respectively, which were higher than those in HD (0.052±0.082)% and (0.029±0.042)% (F=3.774, P=0.024;F=4.912, P=0.008). The frequency of antigen-specific multifunctional Th1 cells, IL-2⁺TNF-α⁺ CD4⁺ T cells, IFN-γ⁺TNF-α⁺ CD4⁺ T cells, IL-2⁺ CD4⁺ T cells, IFN-γ⁺ CD4⁺ T cells in PFMCs were (0.719±0.996)%, (0.628±1.284)%, (0.704±0.829)%, (0.955±1.538)%, (1.188±1.924%), respectively, which were higher than those in PBMCs (0.033±0.065)%, (0.095±0.174)%, (0.137±0.317)%,(0.285±0.434)%, (0.318±0.598)% (t=3.700, P=0.001;t=2.125, P=0.043;t=3.216, P=0.003;t=2.144, P=0.041;t=2.412, P=0.023).Conclusion Antigen-specific multifunctional Th1 cells were correlative with Mtb infection, and may play a protective immune role in TB.