Objective To study the expression and significance of Th17 cells from peripheral blood of patients with rheumatoid arthritis (RA). Methods Intracelluar flow cytomete detection of IL-17/IFN-γ and IL-17/IL-6 was established using anti-CD3/Anti-CD28/IL-23 as stimulators after isolation of untouched human CD4~+T cells from PBMC. There were three groups in the present study: ①healthy controls group; ② RA stable group; ③RA active group. Results The isolation of untouched human CD4~+T cells from PBMC was effective and its purity was over 90%. The percentage of intracelluar IL-17 in CD4~+ T cells from RA patients was increased significantly. Such percentage in active group (1.54±0.41) was higher than that of stable group (0.70±0.21, P<0.01) and both of them were higher than those of healthy controls (0.42±0.12, P<0.01). Under anti-CD3/Anti-CD28/IL-23 stimulation, the percentage of intracelluar IL-17 was also increased significantly(P<0.01). The porcentage of intracellular IFN-γ was similar to that of IL-17, while that of IL-6 was not significantly different. There is an correlation between IL-17 and IFN-γ or IL-6. Conclusion There is an abnormal expression of IL-17 and IFN-γ in human CD4~+T cells in RA patients, which is related to disease activity . Th17 cells may be used as a new marker for the assessment of RA activity.