Objective To evaluate the diagnostic value of copeptin and cancer antigen 125 (Ca-125) in acute heart failure (AHF) patients with atrial fibrillation, and to explore the relationship between copeptin, Ca-125 and short-term cardiovascular events. Methods A total of 376 patients with acute left heart failure or permanent atrial fibrillation admitted to the Department of Cardiology of First Affiliated Hospital of Nanjing Medical University from January 2016 to January 2018 were enrolled as the study group. According to whether having atrial fibrillation or not, 376 patients were divided into atrial fibrillation group (n = 108), AHF group (n = 134) and AHF with atrial fibrillation group (n = 134). 102 healthy persons in the same period were enrolled as healthy control group. Copeptin, Ca-125, N-terminal pro-brain natriuretic peptide (NT-proBNP) within 24 hours after admission or on the day of physical examination were determined, and cardiac function indexes including left atrial diameter (LAD), left ventricular diameter (LVD) and left ventricular ejection fraction (LVEF) at 1 week after admission or on the day of physical examination were determined. Correlation analysis among above indexes was conducted by Pearson correlation analysis. Receiver operating characteristic (ROC) curve was plotted to evaluate the diagnostic value of copeptin and Ca-125 in AHF with atrial fibrillation. Results Compared with the healthy control group, copeptin, Ca-125, NT-proBNP, LAD, and LVD in atrial fibrillation group, AHF group and AHF with atrial fibrillation group showed a tendency of gradual increase [copeptin (pmol/L): 12.43±4.36, 18.77±5.29, 32.82±7.07 vs. 6.68±1.94; Ca-125 (kU/L): 18.82±7.39, 27.97±11.47, 61.37±25.49 vs. 4.43±1.74; NT-proBNP (ng/L): 1 070.87±428.84, 1 734.13±725.09, 2 745.92±709.91 vs. 570.40±213.87; LAD (mm): 37.24±6.35, 41.31±7.94, 46.24±10.96 vs. 33.29±4.53; LVD (mm): 49.46±5.19, 52.51±8.09, 55.96±6.49 vs. 45.99±6.26, all P < 0.05], and LVEF showed a tendency of gradual decrease (0.52±0.11, 0.46±0.10, 0.41±0.09 vs. 0.57±0.08, all P < 0.05), indicating that the deterioration of all indexes in AHF patients with atrial fibrillation was more obvious. Correlation analysis showed that copeptin was positively correlated with LAD (r = 0.479, P = 0.012) and LVD (r = 0.513, P = 0.005), and it was negatively correlated with LVEF (r = -0.626, P < 0.001). Ca-125 was positively correlated with LAD (r = 0.479, P = 0.011) and LVD (r = 0.513, P = 0.028), and it was negatively correlated with LVEF (r = -0.645, P = 0.019). ROC curve analysis showed that the area under ROC curve (AUC) of copeptin, Ca-125, NT-proBNP and copeptin combined with Ca-125 in the diagnosis of AHF with atrial fibrillation was 0.750, 0.623, 0.647 and 0.842, respectively, with diagnostic value on AHF with atrial fibrillation. The diagnostic value of copeptin combined with Ca-125 was the largest, with a sensitivity of 72.64% and a specificity of 92.47%. Compared with the healthy control group, the incidence of cardiovascular events after 3 months of follow-up in the atrial fibrillation group, AHF group and AHF with atrial fibrillation group was significantly increased [6.5% (7/108), 9.0% (12/134), 30.6% (41/134) vs. 1.0% (1/102), χ2 = 56.574, P = 0.000], indicating that patients with AHF and atrial fibrillation were more likely to have cardiovascular events. Copeptin combined with Ca-125 showed a significant positive correlation with short-term cardiovascular events (r = 0.641, P = 0.004). Conclusions The combination of copeptin and Ca-125 has a higher diagnostic accuracy for AHF patients with atrial fibrillation. Copeptin and Ca-125 were positively correlated with short-term cardiovascular events. It may be used to assess the prognosis of AHF patients with atrial fibrillation.