Objective To establish and verify a mature and stable glioblastoma (GBM) organoid model, so as to provide an accurate and personalized preclinical model for the research and treatment of GBM. Methods Fresh GBM tissues obtained through surgical procedures were initially processed, and then GBM stem cells (GSCs) were isolated using stem cell culture medium and were identified. Subsequently, GSCs were cultured in organoid culture medium for 3D cultivation, and GBM organoids were successfully obtained. The histological morphology of GBM organoids was observed by hematoxylin-eosin (H-E) staining; the stemness and similarity to the parental tumor were identified by immunofluorescence staining; and the in vivo tumorigenic ability of GBM organoids was identified by orthotopic tumorigenesis experiments in nude mice. Results A total of 7 GBM organoids were constructed from 9 human GBM samples, with a morphology resembling “neurosphere”, and the average duration for organoid formation was 1 week. H-E staining results showed that the histological morphology of GBM organoids under high-power microscope was very similar to that of GBM tumor tissues; immunofluorescence staining results indicated that the GBM organoids possessed stemness characteristics and histological cellular similarity; and GBM organoids had a stronger tumorigenic ability compared to ordinary GBM cells in nude mice. Conclusion This study presents a stable and reliable method for constructing GBM organoids retaining the histological characteristics of the original GBM tissue, which providing new insights for future GBM research and clinical practice.