Paper-based maskless enzymatic sensor for glucose determination combining ink and wire electrodes

In this work we have developed an amperometric enzymatic biosensor in a paper-based platform with a mixed electrode configuration: carbon ink for the working electrode (WE) and metal wires (from a low-cost standard electronic connection) for reference (RE) and auxiliary electrodes (AE). A hydrophobic wax-defined paper area was impregnated with diluted carbon ink. Three gold-plated pins of the standard connection are employed, one for connecting the WE and the other two acting as RE and AE. The standard connection works as a clip in order to support the paper in between. As a proof-of-concept, glucose sensing was evaluated. The enzyme cocktail (glucose oxidase, horseradish peroxidase and potassium ferrocyanide as mediator of the electron transfer) was adsorbed on the surface. After drying, glucose solution was added to the paper, on the opposite side of the carbon ink. It wets RE and AE, and flows by capillarity through the paper contacting the carbon WE surface. The reduction current of ferricyanide, product of the enzymatic reaction, is measured chronoamperometrically and correlates to the concentration of glucose. Different parameters related to the bioassay were optimized, adhering the piece of paper onto a conventional screen-printed carbon electrode (SPCE). In this way, the RE and the AE of the commercial card were employed for optimizing the paper-WE. After evaluating the assay system in the hybrid paper-SPCE cell, the three-electrode system consisting of paper-WE, wire-RE and wire-AE, was employed for glucose determination, achieving a linear range between 0.3 and 15 mM with good analytical features and being able of quantifying glucose in real food samples.