Nanomaterial-based biosensors are a promising fit for portable and field-deployable diagnosis sensor devices due to their mass production, miniaturization, and integration capabilities. However, the fabrication of highly stable and reproducible biosensor devices is challenging. In this work, we grow a vertically oriented architecture of zinc oxide nanorods onto the active working area (i.e., the channel between the source and drain) of a field-effect transistor (FET) using a low-temperature hydrothermal method. The glucose oxidase enzyme was immobilized on the zinc oxide nanorod surface by a physical adsorption method to fabricate the electrolyte-gated FET-based glucose biosensor. The electrical properties of the electrolyte-gated FET biosensor were measured with different glucose concentrations. We found a linear increase in current up to 80 mM glucose concentration with high sensitivity (74.78 μA/mMcm2) and a low detection limit (∼0.05 mM). We illustrate a highly reproducible fabrication process of zinc oxide nanorod-based FETs, where vertically grown nanorods with a higher surface-to-volume ratio enhance the enzyme immobilization, provide a microenvironment for longer enzyme activity, and translate to better glucose sensing parameters. Additionally, our electrolyte-gated FET biosensor showed promising application in freshly drawn mouse blood samples. These findings suggest a great opportunity to translate into practical high-performance biosensors for a broad range of analytes.