Objective To evaluate the feasibility and therapeutic efficacy of real-time virtual navigation system (RVS) combined with contrast-enhanced ultrasonography (CEUS) for hepatocellular carcinoma undetectable by conventional ultrasound and CEUS.Methods A total of 213 patients with 265 lesions (undetectable on conventional ultrasound but detectable by other imaging examinations) were enrolled in this study.CEUS was performed,and lesions which were detected with CEUS were given CEUS guided radio-frequency ablation (RFA) (CEUS group).RVS combined with CEUS was performed to locate the rest lesions and guide RFA (RVS + CEUS group).Diagnostic value and therapeutic efficacy of RVS + CEUS were assessed.Results In 256 lesions,174 (65.7％) could be detected with CEUS,and among the rest 91 lesions,82 (90.1％) lesions could be detected by RVS combined with CEUS.The technique effectiveness rate of RVS + CEUS was significantly higher than that of CEUS (x2 =18.85,P ＜0.0001).Complete ablation rate after one month in RVS+ CEUS group was 9t.5％ (75/82),which was significantly higher than that in CEUS group [86.2％ (150/174),P ＜0.05].Local progression in one year in CEUS group and RVS + CEUS group were 13.4％ and 10.9％,respectively.Conclusions RVS combined with CEUS is feasible for hepatocellular carcinoma undetectable by conventional ultrasonography and CEUS,and the therapeutic efficacy of RVS + CEUS guided RFA is satisfactory.