Electrochemical evaluation of screen-printed sensor manufacturing and LOx enzyme immobilization for lactate biomarker detection: influence of reference electrode material.

Electrochemical sensors have gained significant attention in medical diagnostics, with continuous advancements in materials improving their performance. This study focuses on the development of screen-printed electrodes (SPEs) for lactate detection. The electrodes were produced using a carbon/graphene paste, and this material was evaluated as an alternative to the commonly used Ag/AgCl reference electrode (WE). The screen-printing technique enabled scalable, efficient sensor production on polymeric substrates. Cyclic voltammetry (CV) was used to assess the electrochemical properties and reproducibility of the sensors. The results showed that Ag/AgCl WE exhibited a higher Δ , indicating greater charge transfer resistance, but also demonstrated higher current density, which enhances the efficiency of the faradaic process and improves repeatability. To evaluate the impact of the conductive material of the WE on lactate detection, lactate oxidase (LOx) was immobilized on the working electrode using a Nafion polymer membrane, ensuring enzyme stability and minimizing interference. The linear relationship between lactate concentration and measured electric current revealed that carbon/graphene reference electrodes are a viable alternative to Ag/AgCl, offering comparable performance in terms of sensitivity and detection limit. These sensors are unaffected by interferents such as glucose and ascorbic acid; however, when using human plasma, a reduction in the measured electric current was observed at all concentrations, which may impact analyte detection sensitivity. This finding suggests the need for future studies to evaluate other biological interferents.