Objective Glioma is an invasive malignant primary brain tumor, characterized by its infiltrative growth pattern, making it challenging to differentiate its boundaries from healthy brain tissues during surgical procedures. Our study aims to address the challenge of accurately distinguishing glioma from healthy brain tissues. Based on a homemade highresolution polarizationsensitive optical coherence tomography (PS -OCT) system, we conduct ex vivo imaging of normal mouse brain and human glioma model mouse brain. Further, based on the polarization analysis of the tumor and normal brain tissues, we propose a novel tumor differentiation metric called optic axis standard deviation to distinguish normal and glioma tissues. It is proven by the results that highresolution PS -OCT has great potential in imaging brain tissue and can offer enhanced precision to detect glioma during surgical interventions, which will help to solve the urgent clinical need in neurosurgical practice.Methods A homemade spectral domain PS -OCT system is constructed by utilizing a superluminescent diode as a broadspectrum light source. The system is performed with an axial resolution of 3.4 µm in air and a transverse resolution of 4 µm in the focal plane.