Purposes: To investigate the clinical application of CEUS combined with MRI virtual navigation in evaluating the thermal ablation of hepatocellular carcinoma. Methods: From September 2020 to December 2022, patients who were diagnosed with HCC and going to be treated with thermal ablation were enrolled from three hospitals. Patients were divided into two groups randomly, CEUS group and CEUS combined with MRI virtual navigation group separately. Then, patients underwent CEUS examination or CEUS combined with MRI examination 1 day before and 30 days after treatment. All patients underwent Gd-EOB-DTPA MRI after treatment to evaluate the ablative margin and to define whether the lesions were complete ablation. Results: 46 patients with 46 lesions in the CEUS group and 31 patients with 31 lesions in CEUS combined with MRI virtual navigation group were enrolled in our multi-center study. Among 77 lesions, 27 were treated by radiofrequency ablation and 50 by microwave ablation. After treatment, 14 lesions were not reached the ablative margin (5mm) in the CEUS group and 4 lesions in CEUS combined with MRI virtual navigation group (P = 0.063). The number of lesions at specific locations (the distance with liver capsule, portal vein, hepatic vein, bile duct, gallbladder, and diaphragm within 5mm) that didn’t reach the ablative margin was 6 in the CEUS group and 2 in the CEUS combined with MRI group (P < 0.001). The number of iso-echoic lesions at B-mode ultrasound or with an ill-defined margin that didn’t reach the ablative margin was 5 in the CEUS group and 2 in the CEUS combined with the MRI group (P < 0.001). Conclusions: CEUS combined with MRI virtual navigation is helpful in evaluating the ablative margin of HCC, especially for lesions at specific locations, iso-echoic and ill-defined margin lesions.