Detection of biomarker for periodontal disease using a microchip

Objective: Several biomarkers including procollagen type I C-terminal propeptide (PICP), have been reported for objective evaluation of the state of periodontal disease. Although these markers are usually examined by conventional ELISA method with a microplate reader, it is time- and sample- consuming, and it is not adaptable to automated analyzers. Convenient, sensitive, and accurate methods for the measurement of the biomarkers are thus required. Bio-nano technology has received considerable interest in analytical chemistry due to the intrinsic characteristics of high speed, high throughput, easy operation, low consumption of samples and reagents, miniaturization, and automation. To develop the analysis system of biomarkers in periodontal disease in dental clinic, we first developed the quantitative analysis system based on the antigen-antibody reaction in microchannel on microchip.

Methods: After the adsorption of 1st anti-PICP antibody on the surface of microchannel (300 mm width, 100 mm depth) on cyclic olefin copolymers (COC) microchip (Sumitomo Bakelite Co. Ltd./S-BIO, PrimeSurface), sample and peroxidase-conjugated 2nd anti-PICP antibody mixture was introduced into a microchannel for 30 min, followed by washing and peroxidase activity was measured for the determination of PICP level.

Results: A double-logarithm plot analysis revealed a linear relationship between PICP concentration and peroxidase activity in the range of 50 to 600 ng/ml of PICP (r²=0.9745). PICP concentrations in humoris of four subjects determined by the present methods were compared with the results obtained by conventional ELISA method. Good correlations were observed for each method on simple linear regression analysis (P<0.05).

Conclusion: We demonstrated the possible application of microchip for determination of PICP levels with high sensitivity and accuracy, and the potential for the dental clinical diagnosis.