Updated publication reference for PubMed record(s): 32795414. There is an unmet clinical need for improved tissue and liquid biopsy tools for cancer detection. We investigated the proteomic profile of exosomes in 426 human samples from tissue explants (TE), plasma and other bodily fluids. Among traditional exosome markers, CD9, HSPA8, ALIX, HSP90AB1 represent pan-exosomes/exomeres. Moreover, we identified novel pan-exosome/exomere markers (e.g., ACTB, MSN, RAP1B). As proof of principle that exosomes are ideal diagnostic tools, we analyzed the proteome of TE exosomes (n=151) or plasma-derived exosomes (n=120). Comparison of TE exosomes identified proteins (e.g., VCAN, TNC, THBS2) that distinguish tumors from normal tissues with 90% sensitivity/94% specificity. Machinelearning classification of plasma-derived exosomes revealed 95% sensitivity/90% specificity in identifying cancer-associated exosomes. Finally, we defined a panel of tumor-type specific exosomal proteins in TE and plasma, which may help classify tumors of unknown primary origin. Overall, exosomal proteins can serve as reliable biomarkers for cancer detection and for determining cancer type