Superior T cell activation by ESAT-6 as compared with the ESAT-6–CFP-10 complex

Using intracellular cytokine staining we show herein that T cells will respond to short-term (6 h) activation with phorbol ester plus ionomycin by production of tumor necrosis factor (TNF), IFN-c or both. Here CD4 T cells preferentially produce TNF and CD8 cells IFN-c. The same pattern is seen when T cells are activated with the Mycobacterium tuberculosis protein early secretory antigenic target-6 (ESAT-6). Responses with >0.02% IFN-c 1 CD3 cells were seen in 8 of 10 patients diagnosed with tuberculosis and in 12 of 14 healthy individuals selected for likely exposure to M. tuberculosis. T cell responses to the 1:1 complex of ESAT-6 and culture filtrate protein-10 (CFP-10) were inferior to ESAT-6 alone, and only reached the level of T cell response achieved with CFP-10 alone. Extending the time of incubation to 18 h leads to an increased response to the complex, but it still reached only the level of CFP-10 alone. In vitro digestion with lysosomal enzymes cathepsin L and S at 2000:1 protein to enzyme ratio demonstrates rapid digestion of the individual proteins while the ESAT-6–CFP-10 complex is resistant. The data suggest that the natural complex of ESAT-6–CFP-10 is less amenable to antigen processing leading to a lower T cell response as compared with the individual proteins.