Production and characterisation of monoclonal antibodies to the principle sonicate antigens of Porphyromonas gingivalis w50.
Protein antigens from whole cell sonicates of Porphyromonas gingivalis W50, previously shown to be discriminatory antigens for patients with adult periodontitis, were purified using SDS-PAGE. Electroeluted proteins were used to immunize mice for the production of monoclonal antibodies (mAbs). A combination of enzyme-linked immunosorbent assay (ELISA) and Western blotting were used to screen hybridoma supernatants for mAbs. MAbs were successfully raised against M(r) 115,000, M(r) 55,000 and M(r) 47,000 antigens together with a second M(r) 55,000 polypeptide which was a contaminant of the M(r) 55,000 antigen. No immunological cross-reactivity was found between these four proteins. The mAbs were used to examine the distribution of these antigens among fifteen P. gingivalis strains together with related oral bacteria using immunostaining of dot blots and Western blots. The antigens were confined to P. gingivalis with the M(r) 115,000 and M(r) 47,000 antigens being present in all strains tested. The distribution of the M(r) 55,000 antigens were slightly more restricted: one M(r) 55,000 (outer membrane location) was present in nine of the fifteen P. gingivalis strains tested, while the other M(r) 55,000 (location unknown) was only absent from one strain. Whole cell ELISA demonstrated that the M(r) 115,000 and the outer membrane M(r) 55,000 antigen possess epitopes which are located on the surface of the bacterium.
Millar
D J
DJ
MRC Dental Research Unit, Royal London Hospital Medical College, UK.
Scott
E E
EE
Slaney
J M
JM
U
S
S
Benjamin
P
P
Curtis
M A
MA
D000328:Adult; D000818:Animals; D000907:Antibodies, Bacterial / Q000096:biosynthesis / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology*; D000918:Antibody Specificity; D000942:Antigens, Bacterial / Q000032:analysis / Q000276:immunology*; D000954:Antigens, Surface / Q000032:analysis / Q000276:immunology; D015153:Blotting, Western; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D016056:Immunodominant Epitopes / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D010510:Periodontal Diseases / Q000382:microbiology; D017861:Porphyromonas / Q000276:immunology; D016966:Porphyromonas gingivalis / Q000276:immunology*; D012680:Sensitivity and Specificity; D013010:Sonication; D013045:Species Specificity
D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D000954:Antigens, Surface; D016056:Immunodominant Epitopes