Single-Reservoir Electrophoresis to Facilitate Layer-by-Layer Assembly of Gold Nanoparticles in Lateral Flow Immunoassay

Electrophoretically facilitated layer-by-layer assembly of gold nanoparticles (GNPs) in lateral flow immunoassays (LFIAs) significantly enhances the signal-to-background ratio and, consequently, the diagnostic sensitivity of these tests. However, conventional two-reservoir electrophoresis on paper is limited by counterflow induced by capillary action, which disrupts the electrophoretic migration of GNPs toward the anode. This counterflow necessitates manual intervention to facilitate the movement of GNP-labeled immunocomplexes from the membrane to the absorption pad, complicating the assay workflow. To address this challenge, we propose a nonconventional single-reservoir electrophoresis system on paper, which inherently eliminates counterflow. In this configuration, the loading side of the paper strip and the cathode reside within the buffer reservoir, while the anode is directly affixed to the opposite end of the paper strip. We demonstrate the efficacy of this single-reservoir system in driving layer-by-layer assembly, while presenting favorable spatial temperature profiles as a side benefit. By eliminating the need for manual steps, this design streamlines the electrophoresis process and enhances the usability of electrophoretically facilitated LFIA assays.