Rationale and objective: This retrospective study was to evaluate the feasibility and accuracy of coronary artery calcium score (CACS) from virtual non-contrast (VNC) images in comparison with that from true non-contrast (TNC) images. Materials and methods: A total of 540 patients with suspected of coronary artery disease (CAD) who underwent a dual-layer spectral detector CT (SDCT) in three hospitals were eligible for this study and 233 patients were retrospectively enrolled for further analysis. The CACS was calculated from both TNC and VNC images and compared. Linear regression analysis of the CACS was performed between TNC and VNC images. Results: The correlation of overall CACS from VNC and TNC images was very strong (r = 0.923, p < 0.001). The CACS from VNC images were lower than that from TNC images (221 versus. 69, p < 0.001). When the regression equation of the overall coronary artery was applied, the mean calibrated CACS-VNC was 221 which had a significant difference from the CACS-TNC (p = 0.017). When the regression equation of each coronary branch artery was applied, the mean calibrated CACS-VNC was 221, which had a significant difference from the CACS-TNC (p = 0.003). But the mean difference between the CACS-TNC and the calibrated CACS-VNC in either way was less than 1. The agreement on risk stratification with CACS-TNC and CCACS-VNC was almost perfect. Conclusion: This multicenter study with dual-layer spectral detector CT showed that it was feasible to calculate CACS from the VNC images derived from the spectral coronary artery CT angiography scan, and the results were in good accordance with the TNC images after correction. Therefore, the TNC scan could be omitted, reducing the radiation dose to patients and saving examination time while using dual-layer spectral detector CT.