A reusable PMMA/paper hybrid plug-and-play microfluidic device for an ultrasensitive immunoassay with a wide dynamic range.

Conventional colorimetric enzyme-linked immunosorbent assay (ELISA) is a time-consuming laboratory assay that is not very sensitive and consumes a large amount of samples. Herein, the development of a reusable, cost-effective, and eco-friendly poly(methyl methacrylate) (PMMA)/paper hybrid plug-and-play (PnP) device for high-sensitivity immunoassay by analyte enrichment and efficient passing-through washing has been reported. The PMMA device has multiple slots where a pre-patterned paper substrate can be inserted. The sample flows back-and-forth through a low-cost, 3D paper substrate within the PMMA channels, thereby enhancing the amount of analyte adsorbed and dramatically increasing the sensitivity while decreasing the assay time. After the enrichment assay, the paper substrate can simply be pulled out of the device, and the results can be qualitatively viewed with the naked eye or scanned through a simple desktop scanner for quantitative analysis. The paper substrate can be replaced with a new substrate so that the device can be reused. The limits of detection (LODs) of 200 pg/mL for immunoglobulin G (IgG) and 270 pg/mL for hepatitis B surface antigen (HBsAg) were obtained. This IgG assay is at least 10 times more sensitive than commercial ELISA kits. In addition, the PnP ELISA exhibited a significant increase in the linear dynamic range from 3 orders of magnitude in a common paper-based device to a wide range of six orders of magnitude in the PnP hybrid device. This reusable PnP device has great potential for the low-cost yet high-sensitivity detection of infectious diseases, cancers, and other important biomolecules.