Aim: To compare characteristics of polyclonal anti-sera against extracellular domain of hepatoma-associated antigen HAb18G/CD147(HAb18GEF) generated by different immunization schemes.
Methods: BALB/c mice were immunized with GST-HAb18GEF fusion protein expressed in E.coli (routine immunization method), recombinant eukaryotic expression plasmid pcDNA3/HAb18G (intramuscular injection) and pcDNA3/HAb18G plasmid followed by human hepatoma cells booster (DNA-cell booster), respectively. The titers and Ig subclasses of polyclonal anti-sera against denatured and natural HAb18GEF were detected by indirect ELISA and cell ELISA, respectively. The specific binding of polyclonal anti-sera prepared by different immunization schemes to denatured HAb18GEF was analyzed by Western blot. The specific binding of polyclonal anti-sera produced by DNA-cell booster immunization to natural HAb18G antigen on hepatoma cells was detected by immunofluorescence staining.
Results: GST-HAb18GEF immunization could induce polycolonal antibody IgG1 with higher titer mainly against denatured or linear epitopes on HAb18GEF. Antibody induced by pcDNA3/HAb18G intramuscular immunization was IgG2a with lower titer against natural epitopes on HAb18GEF. DNA-cell booster immunization could induce generation of polycolonal antibody IgG2a and IgG1 of moderate titers against the native epitopes on HAb18G.
Conclusion: The polycolonal sera with different titers against different epitopes on HAb18GEF can be induced by different immunization schemes.