T cell acute lymphoblastic leukemia (T-ALL) is a hematological disease including malignancy of T cell precursors. There are some T-ALL patients that are drug-resistant. A major cause of treatment failure in cancers can be associated with the existence of cancer stem cells. The identification of these cell populations helps us to clarify resistance mechanisms and rely on special markers for recognizing cancer stem cells. CD133 is one of the markers that is used for the identification of cancer stem cells. In this study, we evaluated CD133(+) and CD133(-) characteristic cells in Jurkat cells by assay proliferation, invasion, and apoptosis. CD133(+) and CD133(-) Jurkat cells were separated and immediately analyzed for proliferation, invasion, and doxorubicin-induced apoptosis. Proliferation, invasion, and resistance to chemotherapy of CD133(+) Jurkat cells were significantly more than CD133(-) Jurkat cells. Also, our results showed that CD133(+) Jurkat cells expressed ABCG2 gene more than CD133(-) Jurkat cells. In conclusion, CD133 marker could be introduced as a specific marker of cancer stem cells in Jurkat cell line.