A sensor chip that can detect enzyme electrode reactions with a sample volume as small as 20 μL has been developed using screen-printing technology with porous silica spheres (PSSs). PSSs are spherical particles of silica with mesoporous pores, a material with a large specific surface area. Other material properties of PSSs include good dispersibility, high mechanical strength, and the ability to be synthesized with precisely controlled mesopore sizes. These advantages support the increased adsorption of enzymes to create highly sensitive sensors. The electrode was designed as follows: carbon paste (CP) with PSSs was used as the working electrode, glucose dehydrogenase (GDH) was used as the enzyme, and quinoline-5,8-dione (QD) was used as the mediator of electron transfer to induce a glucose dehydrogenation reaction. The sensitivity of the sensor was demonstrated through the detection of glucose. The glucose sensor showed reliable sensitivity between 1 and 60 mM glucose, which is comparable to the detection range of a traditional working electrode such as platinum. Furthermore, the GDH had minimal reduction in activity over repeated usage. This device demonstrates the ability to create an equally sensitive but less expensive and reusable sensor that has the potential to better suit general applications.