Significance Cancer immunotherapies, including checkpoint inhibitor blocking antibodies and antibody drug conjugates, currently revolutionize cancer treatment. However, a remaining challenge is the identification of tumor surfaceome (TS) targets for the design of more rational, individualized treatments. We have developed a procedure for unbiased mapping of TS targets in glioblastoma (GBM), i.e., the most common primary malignant brain tumor that remains among the most aggressive forms of cancer, and for which attempts to find effective treatments have failed so far. The present study provides additional layers of understanding fundamental to the future development of immunotherapy strategies, as well as procedures for proteomics-based target identification aimed at a better understanding of how to harness the TS for personalized immunotherapy.