Production of Monoclonal Antibodies Specific to FimA of Porphyromonas gingivalis and Their Inhibitory Activity on Bacterial Binding

Abstract

Background: The FimA of Porphyromonas gingivalis is a crucial pathogenic component of the bacteria and has been implicated as a target for vaccine development against the periodontal diseases.

Methods: In this study, the purified fimbriae (FimA subunit polymers) protein was used for immunization in their native form and B hybridoma clones producing antibodies specific to FimA were established.

Results: The monoclonal antibodies prepared from selected two clones, designated #123 (IgG2b/ kappa) and #265 (IgG1/kappa), displayed different patterns of binding activity against the cognate antigen. Both antibodies reacted with conformational epitopes expressed by partially dissociated oligomers, but not with monomer as elucidated by Western blot analysis. Ascites fluid containing the monoclonal antibodies showed the inhibitory activity against P. gingivalis to saliva-coated hydroxyapatite beads, an in vitro model for the pellicle-coated tooth surface.

Conclusion: These results suggest that the monoclonal antibodies could be used as vaccine material against the periodontal diseases through passive immunization.

Keywords: fimbriae; hybridoma clone; monoclonal antibody; passive immunization.

Figure 1

Binding activity of anti-FimA monoclonal antibodies. The two conditions for sample preparation were applied to display FimA antigen in Western blot analysis by incubating the strips with polyclonal antibody (1:5,000) prepared from the mice injected with FimA. (A) Monomerized FimA protein was prepared by treating protein with 100℃ for 10 min in the absence of 2-ME and display through gel electrophoresis. (B) Partially depolymerized FimA protein was prepared by treating protein with 80℃ for 5 min in the presence of 2-ME. Lane PC is purified bacterial FimA. Lane M is Prestained Protein Ladder (Fermentas, Glen Burnie, MD). Lane 1 is culture supernatant from #123 hybridoma clone. Lane 2 is culture supernatant from #256 hybridoma clone. The arrow indicated the size of FimA monomer of about 43 kDa.