Rapid evaluation of serum and gingival crevicular fluid immunoglobulin G subclass antibody levels in patients with early-onset periodontitis using checkerboard immunoblotting.
A method was developed to evaluate the presence of immunoglobulin G (IgG) subclass (1-4) antibody to Actinobacillus actinomycetemcomitans, serotype b (strain Y4) in patients with early-onset periodontitis on a single nitrocellulose membrane. Sera from 30 early-onset periodontitis patients and gingival crevicular fluid samples from 2 patients were collected and tested with four different preparations of A. actinomycetemcomitans (Y4). The principle steps of the assay are: a) binding of the bacterial antigen (Y4) and the anti-human IgG antibody (capture antibody) in parallel lanes on nitrocellulose membranes; b) incubation of known concentrations of the IgG subclasses 1, 2, 3 and 4, as well as a dilution of serum and/or gingival crevicular fluid from patients in lanes perpendicular to the antigen lanes; c) incubation of the membranes with the corresponding peroxidase conjugated anti-human IgG subclass secondary antibody; d) detection of positive signals by enhanced chemiluminescence. The blots were evaluated by visual comparison to a series of blots containing known concentrations of IgG subclasses. The method was used to rapidly screen a relatively large number of patient sera and gingival crevicular fluid samples for IgG subclasses in a cost-effective assay. The predominant IgG subclass found in early-onset periodontitis was IgG2.
Dibart
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S
Department of Periodontology and Oral Biology, Boston University School of Dental Medicine, Massachusetts 02118, USA.
Eftimiadi
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C
Socransky
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Taubman
M A
MA
Van Dyke
T E
TE
D000293:Adolescent; D000328:Adult; D016976:Aggregatibacter actinomycetemcomitans / Q000276:immunology*; D010520:Aggressive Periodontitis / Q000097:blood / Q000276:immunology* / Q000382:microbiology*; D000907:Antibodies, Bacterial / Q000032:analysis / Q000097:blood / Q000145:classification; D015373:Bacterial Typing Techniques; D005883:Gingival Crevicular Fluid / Q000276:immunology*; D006801:Humans; D015151:Immunoblotting / Q000379:methods; D007124:Immunoenzyme Techniques; D007074:Immunoglobulin G / Q000032:analysis* / Q000097:blood / Q000145:classification; D015203:Reproducibility of Results
D000907:Antibodies, Bacterial; D007074:Immunoglobulin G