Fig. 1
Screening and identification of chemokines for fusion with antigens to improve immunogenicity. A Schematic diagram of the cellular immune screening process. Chemokines and E6E7 antigens were assembled into plasmid DNA and immunized animals for evaluation. B-D Verifying the cellular immune response and anti-tumor effect of the six selected chemokines. 2 × 105 TC-1 cells were subcutaneously transplanted, and vaccine injection was initiated at an average 50mm3 tumor volume. B Fourteen days after the vaccination, specific T cell levels in peripheral blood were detected by flow cytometry. Bar graphs (mean ± SEM, n = 5) show the percentage of E7RAHYNIVTF tetramer + of CD8 + T cells. C The tumor size is measured every other day and plotted as a tumor growth curve. Mean volume ± SEM was used to represent the average tumor size for each group of mice (n = 8). The complete remission ratio was indicated. D The survival status of the mice is recorded and plotted as a survival curve. If the tumor volume exceeds 2000mm3 or continues to grow for a week after shrinking, it is considered dead. E Vaccines of different chemokines were administrated 3 days post 5 × 105 B16-OVA cells were subcutaneously transplanted. Bar graphs (mean ± SEM, n = 6) show the percentage of OVA tetramer + of CD8 + T cells from blood 14 days post vaccine immunization. Statistical significance was calculated between CCL11-OVA and OVA groups using a student's t-test. F Average growth curve of implanted B16-OVA tumors in C57BL/6 mice was shown with treatment of different fusion plasmids. Mean volume ± SEM was used to represent the average tumor size for each group of mice (n = 6). G When the tumor volume in the vehicle group grew to an average value of 1400 mm.3, all mice from each group were euthanized. Pictures showed the tumors dissected from OVA, CXCL14-OVA, CXCL6-OVA, CCL11-OVA, CCL13-OVA, CCL7-OVA, and Vehicle from top to bottom. The tumors were arranged in order of size. Scale bar: 1 cm (n = 6). H Five micrograms of GST-tagged HPV16-E6E7 protein suspended in PBS buffer was coated on the Corning ELISA plates. The mouse serum obtained at 21 days after immunization was analysed with a 1:100 dilution. Bar graphs (mean ± SEM, n = 5) show the ELISA reactivity of IgG2c in the serum. Result is expressed as the optical densities at 450 nm (A450)