Objective To establish a method based on polyacrilamide gel electrophoresis (PAGE) and Western blot for detection of hepatoma-specific alpha-fetoprotein (HS-AFP) isoform.Methods Serum AFP was separated by native polyacrylamide gel electrophoresis,and AFP isoforms were subsequently detected by Western blot.The experimental conditions for separation,transfer and staining of AFP isforms were optimized.AFP isoforms in serum samples from 82 cases of primary hepatic cancer and 139 cases of various benign liver diseases were detected.The difference of electrophoretic patterns for AFP isoforms between primary hepatic cancer and benign liver diseases were analyzed.Results HS-AFP in serum was distinctly separated and a method for detection of HS-AFP isoforms was successfully established in this study.The results of detection for clinical samples showed that the positive rate for HS-AFP in the patients with primary hepatic cancer was 74.4%, which was significantly higher than that in benign liver diseases ( P <0.01).Conclusion The method established in this study for detection of HS-AFP in this study is helpful for the diagnosis of primary hepatic cancer.