OBJECTIVE To study the clinical value of hepatoma-specific alpha-fetoprotein (HS-AFP) in the diagnosis and differential diagnosis of hepatocellular carcinoma (HCC).METHODS A method of vertical slab polyacrylamide gel electrophoresis with discontinuous buffer system was developed to separate AFP subtypes. After separation, the AFP subtypes were transferred to nitrocellulose and reacted first with rabbit anti-human AFP and then with goat anti-rabbit IgG-HRP. Finally, AFP subtypes were visualized by reacting with 3,3-diaminobenzidine tetrahydrochloride. A HS-AFP band was determined in 82 cases with HCC and 95 cases with benign liver diseases.The correlations between the positive rates of HS-AFP and serum AFP concentration, tumor size as well as portal vein metastasis were analyzed.RESULTS Serum AFP in the cases with various liver diseases was separated into one to several bands. The fastest band on electrophoresis (FAFP) was found in all patients, while the band at the cathodal site (HSAFP) was detected predominantly in HCC but rarely in benign liver diseases. The positive rate of HS-AFP in HCC was 74.4%, which was significantly higher than that in benign liver diseases (9.1%, 7.3% and 10.0% in liver cirrhosis, chronic hepatitis and acute hepatitis respectively). HSAFP was detected in 3 out of 9 HCC cases with AFP＜50 μg/L , but in none of 22 cases of benign liver diseases with the same AFP concentration. HS-AFP correlated with serum AFP concentration and tumor size to some extent, but not with portal vein metastasis.CONCLUSION HS-AFP increases the sensitivity of diagnosing HCC in patients with negative AFP, and is useful in distinguishing high AFP due to HCC from that caused by benign liver diseases.