approved by the Animal Experiment Committee of Fudan University. Each mouse was injected subcutaneously on days 0, 7 and 14 with normal saline cordycepin(2mg/kg), vaccine(1 ug), or HBsAg(I ug) adjuvanted with cordycepin at 0.2 mg/kg, I mg/kg or 2 mg/kg. Serum samples were obtained on days 7, 14 and 21 after the first injection for the measurement of cytokines and HBV-specific antibodies The body weight of each mouse was measured on days 7, 14, 21 and 28 after the first injection, the liver and spleen were collected on day 28 for routine histology and haematoxylin-eosin staining to evaluate the side effects of the new adjuvant. At 15 days after the last injection, the mice were sacrificed, and splenocytes were isolated to determine the cytokine production and cellular proliferation, as well as the cell differentiation Measurement of serum antibodies. The HBV-specific serum antibody titers were measured by ELISA. Briefly, 96-well plates(Grenier, Frickenhausen, Germany )were coated with 2 ug/ml of HBsAg overnight at 4C. After blocking with 5% bovine serum albumin in phosphate-buffered saline containing Tween-20(PBST), 100 ul/well of serially-diluted serum samples from the immunized animals was added following by incubation with a horseradish peroxidase(HRP)-conjugated goat anti-mouse IgG or IgM antibody(Santa Cruz Biotechnology, Santa Cruz, CA, USA) After washing, 100 ul/well of TMB substrate(BD Biosciences, San Diego, CA, USA) was ad dded, and the reactions were terminated by the addition of 50 ul/well of 2M H2SO4. The absorbance was read at 495 nm on a microplate reader(Molecular Devices, Sunnyvale, CA, USA)14 approved by the Animal Experiment Committee of Fudan University. Each mouse was injected subcutaneously on days 0, 7 and 14 with normal saline, cordycepin(2mg/kg), vaccine (1 μg), or HBsAg (1 μg) adjuvanted with cordycepin at 0.2 mg/kg, 1 mg/kg or 2 mg/kg. Serum samples were obtained on days 7, 14 and 21 after the first injection for the measurement of cytokines and HBV-specific antibodies. The body weight of each mouse was measured on days 7, 14, 21 and 28 after the first injection, the liver and spleen were collected on day 28 for routine histology and haematoxylin-eosin staining to evaluate the side effects of the new adjuvant. At 15 days after the last injection, the mice were sacrificed, and splenocytes were isolated to determine the cytokine production and cellular proliferation, as well as the cell differentiation. Measurement of serum antibodies. The HBV-specific serum antibody titers were measured by ELISA. Briefly, 96-well plates (Grenier, Frickenhausen, Germany) were coated with 2 μg/ml of HBsAg overnight at 4°C. After blocking with 5% bovine serum albumin in phosphate-buffered saline containing Tween-20 (PBST), 100 μl/well of serially-diluted serum samples from the immunized animals was added, following by incubation with a horseradish peroxidase (HRP)-conjugated goat anti-mouse IgG or IgM antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA). After washing, 100 μl/well of TMB substrate (BD Biosciences, San Diego, CA, USA) was added, and the reactions were terminated by the addition of 50 μl/well of 2M H2SO4. The absorbance was read at 495 nm on a microplate reader (Molecular Devices, Sunnyvale, CA, USA)