Peptide analysis and verification

Gene sequences were analysed by the BLAST algorithm for homology to proteins with known or putative cancer correlations. The query was performed using the BLAST search program (BLASTP 2.6.1+) against the Homo sapiens non-redundant protein database using Blastp (http://ncbi.nlm.nih.gov/blast/). The identified sequences were further scanned using the SAROTUP webserver software (http://immunet.cn/sarotup/) and the MimoDB database for already existing mimotopes.

To determine the cell surface expression pattern of Glypican-3 receptor, HCT116 and LoVo cells were fixed with 4% paraformaldehyde for 30 min. After blocking with 5% BSA for 1 h, fixed cells were incubated with sheep anti-human Glypican-3 (1:800, R&D Systems, Minneapolis, MN, USA) at 4 °C overnight. They were subsequently stained with NorthernLights 557 conjugated anti-sheep IgG secondary antibody (1:200, R&D Systems) for 1 h at room temperature. After washing three times with PBS, cells were mounted with DAPI and observed using a fluorescence microscope (Olympus).