Planarized through-hole valves enabling multilayered microfluidic architecture towards pipette-free ELISA

Abstract

Decentralized healthcare systems face significant challenges in adopting gold-standard diagnostics methods such as ELISA, due to their time-consuming steps, reliance on specialized equipment, and the need for trained personnel. In response, this study presents an innovative solution: a platform facilitating 3-D reconfigurable microfluidic networks without relying on clean-room fabrication or complex bonding techniques. This platform revolutionizes conventional approaches by integrating a novel scheme of latching valves (namely, planarized through-hole valves) coupled with compression-based sealing. In this manner, the sequential steps of the ELISA protocol are achieved on vertically interconnected microfluidic modules, avoiding reagent cross-contamination while ensuring complete leakage-free performance, all within a compact footprint. Given the simplicity and efficiency of the design, this device can be readily automated to perform in a straightforward workflow, circumventing lengthy pipetting steps and operator involvement. The reliability of the platform was verified by performing the detection of the cytokine TNF-α, using smartphone colorimetric analysis. The results were directly compared with the traditional spectrophotometer-based ELISA 96-well plates, yielding negligible quantitative differences and a limit of detection of 4.21 pg/mL. Overall, this platform’s ability to streamline complex procedures, ensure reliability, and simplify automation holds promise for significantly improving diagnostic accessibility and accuracy in resource-constrained settings.