Background: Molecular characterization of spatially heterogeneous tumors, such as primary brain tumors, requires precise, contamination-free tissue collection. This study develops a novel biopsy instrument enabling efficient, spatially registered tissue collection across tumors and surgical cavities, advancing personalized tumor characterization and treatment. Methods: Iterative modeling and 3D printing were used to develop prototypes, which underwent proof-of-concept and phantom testing. Final device evaluation compared the novel capsule biopsy tool to Yasargil tumor grasping forceps, with six neurosurgeons and six students conducting tests on porcine brain. Additionally, intraoperative samples were collected, with spatial coordinates recorded on preoperative scans. Results: The design features a capsule that attaches to the end of a Frazier suction. When suction is applied, an internal piston secures the sample, which can later be released before or after tissue fixation. The capsule method reduced variability in sample weight and collection time compared to Yasargil forceps, maintaining tissue integrity without contamination or instrument failure. Notably, students demonstrated proficiency comparable to experienced surgeons, highlighting the tool’s ease of use. Conclusions: This low-cost, optically tracked biopsy tool provides an efficient, reliable method for spatially precise tissue collection, meeting the demands of precision medicine and translational research.