DEVELOPMENT OF A CHECKERBOARD IMMUNOBLOT

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Several lines of evidence demonstrate that immune mechanisms are responsible for the tissue destruction characteristic of periodontal disease. Gingival crevicular fluid (GCF) could provide a non-invasive diagnostic means to measure the inflammatory mediators released during disease progression. Several immune mediators have been detected in GCF, and their levels have been associated with disease status. We propose to use the checkerboard immunoblotting (CBIB) technique to quantify multiple components of the GCF in a large number of samples. A similar approach has been applied in our laboratory for the characterization of the microbiota associated with periodontal infection. This technology would allow us to characterize GCF components with microbiological data. No such high throughput technique has been available for cytokines or other markers of host status at periodontal sites. Preliminary data indicate that it will be possible to measure significant numbers of cytokines in routine GCF samples. Therefore, it should be possible to quantify the nature of the bacterial challenge and the nature of the host response on a site-by-site basis within the individual. The levels of some inflammatory mediators can influence the synthesis and secretion of others. The ability to investigate several molecules at once would allow inferences on the mechanisms of modulation of immune response and tissue destruction by these mediators. This approach will fill a gap in our knowledge regarding the cross-regulation of inflammatory mediators and the relationship of the levels of these mediators to subgingival periodontal pathogens.