Glypican-1 (GPC1) is a cell surface proteoglycan that is specifically enriched on cancer cell-derived exosomes. Using mass spectrometry, the authors identified GPC1 as a specific marker for cancer exosomes. GPC1+ circulating exosomes (crExos) were isolated from the serum of cancer patients and mice with cancer. These crExos were detected in the serum of patients with pancreatic cancer with high specificity and sensitivity, distinguishing healthy subjects and patients with benign pancreatic disease from those with early and late-stage pancreatic cancer. Levels of GPC1+ crExos correlate with tumor burden and survival in patients pre- and post-surgical tumor resection. GPC1+ crExos from patients and mice with spontaneous pancreatic tumors driven by oncogenic KRAS contained RNA with specific KRAS mutations, suggesting their potential as a reliable biomarker for the detection of PanIN lesions despite negative MRI signals in mice. GPC1+ crExos may serve as a non-invasive diagnostic and screening tool for early-stage pancreatic cancer, facilitating curative surgical therapy.Glypican-1 (GPC1) is a cell surface proteoglycan that is specifically enriched on cancer cell-derived exosomes. Using mass spectrometry, the authors identified GPC1 as a specific marker for cancer exosomes. GPC1+ circulating exosomes (crExos) were isolated from the serum of cancer patients and mice with cancer. These crExos were detected in the serum of patients with pancreatic cancer with high specificity and sensitivity, distinguishing healthy subjects and patients with benign pancreatic disease from those with early and late-stage pancreatic cancer. Levels of GPC1+ crExos correlate with tumor burden and survival in patients pre- and post-surgical tumor resection. GPC1+ crExos from patients and mice with spontaneous pancreatic tumors driven by oncogenic KRAS contained RNA with specific KRAS mutations, suggesting their potential as a reliable biomarker for the detection of PanIN lesions despite negative MRI signals in mice. GPC1+ crExos may serve as a non-invasive diagnostic and screening tool for early-stage pancreatic cancer, facilitating curative surgical therapy.