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Glioma-initiating cells (GICs) have been proven to play key roles in initiation and progress of malignant glioma.However, a mass of pervious in vivo studies were based on the heterograft model which is implanting GICs derived from primary human glioma specimens into immunodeficient mice.Whether this in vivo model could duplicate the microenvironment of malignancy and evaluate the biological features of GICs accurately is unclear.Here, we compared the biological features of neurosphere-like tumor cells derived from murine glioma cell line GL261 (GL261-NS) and neurosphere-like tumor cells of primary human glioma specimens (P-NS) in vitro and in vivo.We found that GICs enriched in GL261-NS, GL26t-NS and P-NS both exhibited similar increased GICs potential and drug resistance in vitro.But, GL261-NS in syngeneic graft model showed different enhanced tumorigenicity and better duplicated the primary human glioma histologically relative to P-NS in nude mice which were lack of immunocytes and immunomediators.Additionally, the biological features of P-NS were different among individual largely, although the patients were all diagnosed as glioblastoma.Our studies conclude that implantation of GL261-NS into C57/BL6 mice is a more reliable and stable syngeneic graft model for in vivo study on GICs relative to implantation of P-NS into heterogenous immunodeficient mice.