Protocol to analyze antitumor immunity of orthotopic injection and spontaneous murine high-grade glioma models using flow cytometry and single-cell RNA sequencing

Summary: Despite the recognized importance of antitumor immunity, our understanding of brain tumor immunity is poor. Orthotopic injection models have been widely used for immunological analyses. However, these models have limitations in analysis of antitumor immunity because the approach involves drilling skulls and injecting tumor cells, which can induce adverse effects. We describe a protocol for the induction of spontaneous brain tumor model, isolation of single cells from brain tumor microenvironment, and analysis of the immune responses using scRNA-seq and flow cytometry.For complete details on the use and execution of this protocol, please refer to Park et al. (2021). : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.