Paper microfluidics on screen-printed electrodes for simple electrochemical magneto-immunosensor performance

Using magnetic beads (MB) allows producing relatively fast and simple electrochemical magneto-immunosensors. However, this type of sensor implies carrying out serial MB incubation and washing steps in tubes using external magnets, a procedure that requires user training and is tedious and time consuming. In this work, a paper device has been developed that allows performing MB washing and electrochemical detection on-chip with less intervention of the user. In order to achieve this goal, we optimized a single-step magneto-immunoassay for matrix metalloproteinase 9 (MMP-9) detection. Electrochemical detection was then carried out using a customized multiplexed holder that incorporated 8 screen-printed carbon electrodes (SPCE) and a movable multiplexed fluidic module. This paper device provided along the assay sequential MB confinement under static conditions, reagent adsorption and MB washing under flow conditions, and electrochemical detection under static conditions. As we show, the sensor produced in this way achieved detection of MMP-9 in about 10 min, with lineal response spanning 0.03-2 ng mL(-1), detection limit of 0.01 ng mL(-1) and quantification limit of 0.09 ng mL(-1). Furthermore, MMP-9 quantification in 18 plasma samples from patients correlated linearly with the estimates of a reference ELISA that took > 5 h. These results demonstrate that paper microfluidics can be exploited to simplify magneto-immunosensor manipulation, providing fast, simple and sensitive assay formats.