This chapter discusses the role of brain biopsy in diagnosing primary central nervous system lymphoma (PCNSL). PCNSL is a rare but aggressive form of non-Hodgkin lymphoma that affects the brain and spinal cord without systemic involvement. Early and accurate diagnosis is crucial because of the rapid tumor progression and the need for specific treatment. This chapter highlights the importance of brain biopsy, which has been the gold standard for diagnosis despite its invasiveness. Although advancements in imaging techniques, such as MRI, PET scans, and CSF biomarkers, (e.g., IL-10 and MYD88) provide valuable information, they cannot replace brain biopsy for definitive diagnosis. This chapter also emphasizes the risks associated with brain biopsies, such as hemorrhage, especially when performed in deep-seated areas. The chapter concludes that biopsy remains essential for diagnosis, but noninvasive techniques are increasingly useful for the follow-up and detection of relapses.