Objective To develop rolling circle amplification (RCA) method combined with specific surface plasmon resonance (SPR) nucleic acid gold-chip for the detection of hepatitis C virus (HCV). Methods According to the specific test sequence of HCV x-tail region, probes and primers for detecting HCV with RCA method were designed and synthesized, and were divided into test group, negative sample group and positive sample group for RCA experiments to detect HCV. The template concentration was diluted into ten gradients, and the detection limit of SPR combined with RCA method was evaluated. Based on the ordinary gold chip, through the surface chemical processing, the nucleic acid chip with high specificity was obtained, and the anti-protein capacity of protein chip was verified by anti-protein experiment. Real-time detection of RCA reaction and signal magnification reaction was conducted with double channel SPR apparatus. Sixty-three blood samples collected from clinics were detected by SPR combined with RCA method, comparisons were made with Real-Time PCR, and the sensitivity and specificity were evaluated. Results The minimum detection concentration of SPR combined with RCA method in HCV testing was 1 pmol/L, which was lower than the detection limit of Real-Time PCR (0.1 nmol/L). SPR chip had the favorable anti-protein absorptive capacity. The signal-to-noise ratio of double channel SPR apparatus in detection of RCA chip system was 100, which achieved the detection of HCV. The sensitivity of SPR combined with RCA method in detection of clinical samples was 90.0%（27/30）, and the specificity was 84.8% （28/33）（χ2=8.10，P=0.004）. Conclusion SPR combined with RCA method combines biological sensing technology with in situ amplification technology, which may detect HCV in a fast, label-free and real-time way.