Development of a C-reactive protein quantification method based on flow rate measurement of an ink solution pushed out by oxygen gas generated by catalase reaction.

A novel determination method for protein biomarkers based on on-chip flow rate measurement was developed using a microchip with organic photodiodes (OPDs). This quantitative method is based on the flow rate measurement of an ink solution pushed out by oxygen gas generated through catalase reaction. The amount of oxygen gas generated in the sample reservoir is dependent on the concentration of the analyte; therefore, the flow rate of the ink solution is also dependent on the concentration of the analyte. The concentration of the analyte can thus be estimated by measurement of the ink solution flow rate. The ink solution flow rate was estimated by measuring the migration time of the ink solution between two points using two OPDs placed below the microchannel. The principle of this method was demonstrated by the measurement of catalase using the microchip. In addition, the developed method was applied to the determination of C-reactive protein (CRP), a biomarker of inflammation, based on a catalase-linked immunosorbent assay (C-LISA). The limit of detection for CRP was 0.20 µg/mL. The method was also applied to the determination of CRP in human serum, and the quantitative values obtained by this method were in excellent agreement with those obtained by the conventional enzyme-linked immunosorbent assay (ELISA) method. The developed method does not require a photodetector with high sensitivity and is thus capable of downsizing; therefore, this will be useful for on-site analyses such as point-of-care testing and field measurements.

Development of a fluorescence microplate reader using an organic photodiode array with a large light receiving area.

We developed a small fluorescence microplate reader with an organic photodiode (OPD) array. The OPD array has nine OPDs that have a large light receiving area (9.62 mm2 per one OPD). Since the OPD array is fabricated on a flat glass plate, it can be placed just below microwells and can detect fluorescence emitted through the entire surface of the microwell bottom. The analytical performance of the developed plate reader was evaluated by measuring an aqueous solution of resorufin. The limit of detection (LOD) for resorufin (0.01-0.05 µM) was lower than that obtained with a plate reader equipped with nine inorganic photodiodes developed in a previous study (0.30 µM) and a commercially available microplate reader (0.16 µM). These results indicate that the large light receiving area improves the detection performance of the system. In addition, the developed reader was successfully used to quantify immunoglobulin A (IgA) in human saliva. The LOD for IgA was estimated to be 1.2 ng/mL, which is low enough to objectively evaluate human stress.