1. Colorimetric and fluorometric determination of uric acid by a suspension-based assay using enzyme-immobilized micro-sized particles.
- Author
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Moriiwa Y, Hatakeyama K, Morioka K, Inoue Y, Murakami H, Teshima N, Yanagida A, and Shoji A
- Subjects
- Particle Size, Humans, Suspensions, Oxazines chemistry, Uric Acid blood, Uric Acid chemistry, Uric Acid analysis, Colorimetry, Enzymes, Immobilized chemistry, Enzymes, Immobilized metabolism, Urate Oxidase chemistry, Urate Oxidase metabolism, Horseradish Peroxidase chemistry, Horseradish Peroxidase metabolism, Fluorometry
- Abstract
Daily monitoring of serum uric acid levels is very important to provide appropriate treatment according to the constitution and lifestyle of individual hyperuricemic patients. We have developed a suspension-based assay to measure uric acid by adding a sample solution to the suspension containing micro-sized particles immobilized on uricase and horseradish peroxidase (HRP). In the proposed method, the mediator reaction of uricase, HRP, and uric acid produces resorufin from Amplex red. This resorufin is adsorbed onto enzyme-immobilized micro-sized particles simultaneously with its production, resulting in the red color of the micro-sized particles. The concentration of resorufin on the small surface area of the microscopic particles achieves a colorimetric analysis of uric acid with superior visibility. In addition, ethanol-induced desorption of resorufin allowed quantitative measurement of uric acid using a 96-well fluorescent microplate reader. The limit of detection (3σ) and RSD (n = 3) were estimated to be 2.2 × 10
-2 μg/mL and ≤ 12.1%, respectively. This approach could also be applied to a portable fluorometer., (© 2024. The Author(s), under exclusive licence to The Japan Society for Analytical Chemistry.)- Published
- 2024
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