Background: Although troponin assays improve the detection of acute myocardial infarction (AMI), troponin elevation is observed in various non-ischemic conditions. Studies have proposed that, when used in combination with a cardiac troponin I (TnI) assay, serum copeptin would increase the diagnostic accuracy for AMI. Therefore, we assessed the utility of copeptin in the diagnosis of non-ST-segment elevation myocardial infarction (NSTEMI). Methods: A total of 180 patients (age, 68.2±13.3 years; M:F, 113:67) were enrolled for the study, who are presented with chest pain and mild TnI elevation (0.04–1.0 ng/mL) in the emergency department, excluding those with ST elevation on the electrocardiogram. Copeptin was measured using an automated immunofluorescent assay, Copeptin proAVP KRYPTOR (Thermo Fisher Scientific, Germany). Results: The subjects included 49 patients (27.2%) who had NSTEMI, 64 (35.6%) patients who had angina, and 67 (37.2%) patients who had other diseases. The median (interquartile range) copeptin level in the NSTEMI group (69.57 [35.56–172.50] pmol/L) was significantly higher than those in the angina group (7.64 [3.36–17.19] pmol/L) and the other diseases group (6.75 [4.33–13.02] pmol/L) (P<0.0001). At the 14.4 pmol/L cutoff for copeptin, TnI plus copeptin had a higher area under the curve than TnI plus CK-MB (0.898 vs. 0.711, P=0.0001) for diagnosing NSTEMI. Conclusions: Non-ischemic mild TnI elevation is common. Copeptin levels provide additional information for differentiating NSTEMI from non-NSTEMI patients with mild TnI elevation. The combination of copeptin and TnI could improve NSTEMI diagnosis by excluding non-ischemic mild TnI elevation.