AbstractA flexible glucose biosensor was constructed by immobilizing glucose oxidase within a gold‐coated, hydrophilic polytetrafluoroethylene membrane. All reagents and materials used were harmless and nontoxic. Enzyme immobilization was improved using a mesh mask during gold deposition, the grid pattern permitting better access into the membrane. The masked sensors had greatly improved output currents and linearity. The sensor was linear up to 662 mgL−1, the range covering the concentrations encountered in normal and diabetic human tear fluids. The current was proportional to the square of the electrode area, hence an electrode of known dimensions can be used without precalibration. Sterilization using 80% ethanol had a negligible effect on its response. The sensor is well suited to the detection of glucose or other analytes in body fluids, such as tears, airway mucus, sweat, and sal