Magnetic bead handling using a paper-based device for quantitative point-of-care testing

Microfluidic paper-based analytical devices (PADs) have been extensively proposed as ideal tools for point-of-care (POC) testing with minimal user training and technical requirements. However, most PADs use dried bioreagents, which complicate production, reduce device reproducibility and stability, and require transport and storage under temperature and humidity controlled conditions. In this work, we propose a PAD produced using an affordable craft-cutter and stored at room temperature, which is used to partially automate a single-step colorimetric magnetoimmunoassay. As a proof-of-concept, the PAD has been applied to the quantitative detection of
Plasmodium falciparum lactate dehydrogenase (Pf-LDH), a biomarker of malaria infection. In this system, detection is based on a single-step magneto-immunoassay that consists of a single 5-min incubation of the lysed blood sample with immuno-modified magnetic beads (MB), detection antibody,
and an enzymatic signal amplifier (Poly-HRP). This mixture is then transferred to a single-piece paper device where, after on-chip MB magnetic concentration and washing, signal generation is achieved by adding a chromogenic enzyme substrate. The colorimetric readout is achieved by the
naked eye or using a smartphone camera and free software for image analysis. This PAD afforded quantitative Pf-LDH detection in <15 min, with a detection limit of 6.25 ng mL􀀀1 when the result was interpreted by the naked eye and 1.4 ng mL􀀀1 when analysed using the smartphone imaging
system. Moreover, the study of a battery of clinical samples revealed concentrations of Pf-LDH that correlated with those provided by the reference ELISA and with better sensitivity than a commercial rapid diagnostic test (RDT). These results demonstrate that magneto-immunoassays can be partly
automated by employing a PAD, achieving a level of handling that approaches the requirements of POC testing.