Abstract BACKGROUND Glioblastoma is the most frequent and malignant brain tumor. Immune therapies have had limited effect but not much is known about the frequency and type of immune cells in the transition zone and periphery of the tumors. In these areas, tumor cells migrate into the brain parenchyma and prevent total tumor resection thereby leading to recurrence. This study aims to quantify the type and distribution of immune cells in glioblastomas. MATERIAL AND METHODS A cohort of 67 glioblastomas with strong P53 immunoreactivity in the tumor cells was established. Slides contained tissue from central tumor and/or transition zone and/or tumor periphery. A chromogenic multiplex was established, where tissue sections were stained with P53 (tumor cells), FOXP3 (FOXP3+ regulatory T cells), CD8 (CD8+ cytotoxic T cells) and IBA1 (microglia/macrophages). Cells were counted in tumor core, transition zone and periphery using Visiopharm software based on training of convolutional neural networks. Moreover, the number of tumor cells with immune cells in proximity (30 µm) was quantified, and the distance from a reference cell to another cell type within 30 µm was calculated. RESULTS The densities of CD8+, FOXP3+ and IBA1+ cells were significantly higher in the core (10, 1 and 473 cells/mm2) than in the periphery (3, 0,2 and 179 cells/mm2) (P < 0.001), and in the transition zone (10, 1 and 396 cells/mm2) than in the periphery (P < 0.001). However, the CD8+ and IBA1+ cells/tumor cell ratio increased from core (0,004 and 0,18) to transition zone (0,01 and 0,36) and to periphery (0,015 and 0,8) (P < 0.001). The highest density of tumor cells with CD8+ cells in proximity was in the tumor core (55 cells/mm2 in core, 22 cells/mm2 in transition zone, 3 cells/mm2 in periphery) (P < 0.001), whereas the highest density of tumor cells with IBA1+ cells (1412 cells/mm2 in core, 7412 cells/mm2 in transition zone, 98 cells/mm2 in periphery) and FOXP3+ cells (0 cells/mm2 in core, 3 cells/mm2 in transition zone, and 0 cells/mm2 in periphery) in proximity was in the transition zone (P < 0.001). CONCLUSION Although levels of immune cells are highest in the tumor core - the higher proximity and ratio of immune cells to tumor cells in the transition zone/periphery suggest immune-suppressive mechanisms to be active in these areas.