Simple and rapid detection of serum antibody to periodontopathic bacteria by dot blotting.
The aim of this study was to detect the specific immunoglobulin G antibodies against periodontopathic bacteria by dot blotting. In the procedure used, bacterial preparations were blotted on a nitrocellulose membrane. After blocking the nonspecific binding sites, the diluted serum was blotted onto the preparations. The membrane was immersed in secondary antibodies and then in substrate buffer. The colored blots were then evaluated. To test the reliability of this procedure, 20 serum samples were examined for antibody: ten for anti-Porphyromonas gingivalis antibody, and the other ten for anti-Actinobacillus actinomycetemcomitans antibody. Five samples out of each set of ten had previously been confirmed as having high enzyme-linked immunosorbant assay (ELISA) titers to the antigen, while the other five had been confirmed as having average titer levels. Both whole-cell sonic extracts and fimbriae of P. gingivalis were used as antigens in the dot blotting, in order to compare their use as antigens in assays of the patients' sera. ELISA was also used to measure anti-P. gingivalis antibody titers. For the measurement of IgG antibodies against A. actinomycetemcomitans, formalin-killed whole cells were used. Fifty serum samples were examined for IgG antibodies against A. actinomycetemcomitans by dot blotting and ELISA. With both antigens, after 4 h, coloration of blots was more clearly visible for the high-titer sera than for the average-titer sera. The intensity of coloration of the blots for P. gingivalis and A. actinomycetemcomitans showed correlation with the ELISA titers. A particularly significant correlation was shown when P. gingivalis fimbriae were used as antigen. These results suggest that this dot blot method is a simple and rapid means of detection of serum antibodies, and that it shows promise as a chair-side assay method.
Kawashima
Yoko
Y
Department of Hard Tissue Engineering, Graduate School, Tokyo Medical and Dental University, Japan. kawaperi@tmd.ac.jp
Ishikawa
Isao
I
D016976:Aggregatibacter actinomycetemcomitans / Q000276:immunology*; D000907:Antibodies, Bacterial / Q000097:blood / Q000302:isolation & purification*; D000937:Antigen-Antibody Reactions; D000942:Antigens, Bacterial; D001431:Bacteriological Techniques; D016022:Case-Control Studies; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D010861:Fimbriae, Bacterial / Q000276:immunology; D006801:Humans; D015151:Immunoblotting / Q000379:methods*; D007074:Immunoglobulin G / Q000097:blood / Q000302:isolation & purification*; D008297:Male; D008875:Middle Aged; D010518:Periodontitis / Q000097:blood / Q000276:immunology / Q000382:microbiology; D016133:Polymerase Chain Reaction; D016966:Porphyromonas gingivalis / Q000276:immunology*; D015203:Reproducibility of Results; D018709:Statistics, Nonparametric
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D007074:Immunoglobulin G