Glucose oxidase (EC 1.1.3.4) is the most widly employed enzyme as analytical reagent. This is the result of (1) its utility in the determination of glucose, an analyte of wide analytical interest, and (2) its relatively low cost and good stability that make the glucose/glucose oxidase system a very convenient model for method development (particularly in the area of biosensors). This review discusses in detail enzyme structure, biocatalysis, enzymes as analytical reagents, properties of glucose oxidase (including a historical account), and the use of glucose oxidase as an analytical reagent in homogeneous systems as well as an immobilized reagent.