BackgroundSmall extracellular vesicles (sEVs) have great potential as new biomarkers in liquid biopsy. However, due to the limitations of sEVs extraction and component analysis procedures, further clinical applications of sEVs are hampered. Carcinoembryonic antigen (CEA) is a commonly used broad-spectrum tumor marker that is strongly expressed in a variety of malignancies.ResultsIn this study, CEA+sEVs were directly separated from serum using immunomagnetic beads, and the nucleic acid to protein ultraviolet absorption ratio (NPr) of CEA+sEVs was determined. It was found that the NPr of CEA+sEVs in tumor group was higher than that of healthy group. We further analyzed the sEV-derived nucleic acid components using fluorescent staining and found that the concentration ratio of double-stranded DNA to protein (dsDPr) in CEA+sEVs was also significantly different between the two groups, with a sensitivity of 100% and a specificity of 41.67% for the diagnosis of pan-cancer. The AUC of dsDPr combined with NPr was 0.87 and the ACU of dsDPr combined with CA242 could reach 0.94, showing good diagnostic performance for pan-cancer.ConclusionsThis study demonstrates that the dsDPr of CEA+sEVs can effectively distinguish sEVs derived from tumor patients and healthy individuals, which can be employed as a simple and cost-effective non-invasive screening technology to assist tumor diagnosis.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12885-023-10886-3.