Ultrasensitive and Visual Detection of Osteoprotegerin Using a Rotation Microfluidic Device Enables ELISA on Paper

Osteoporosis, characterized by reduced bone strength and density, poses a significant health threat, especially to the elderly, impacting their quality of life. Osteoprotegerin (OPG) acts as a decoy receptor in the RANK/RANKL/OPG system, inhibiting bone resorption. The existing methods, such as dual X-ray absorptiometry (DXA) and enzyme-linked immunosorbent assay (ELISA), have limitations, making them unsuitable for point-of-care testing (POCT). This highlights the need for molecular-level OPG testing, where microfluidic paper-based analytical devices (<inline-formula xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink"> <tex-math notation="LaTeX">$\mu $ </tex-math></inline-formula>PADs) show promise. This study presents an innovative method for OPG detection using a rotating paper-based microfluidic chip housed in a 3D-printed casing, coupled with a modified ELISA colorimetric technique. Integrating paper-based ELISA (P-ELISA) with a rotational design, the platform provides a portable point-of-care (POC) solution for OPG analysis. The system exhibits key features, including low cost (approximately 1 per test), operational simplicity, stability, and high sensitivity (2.5 pg/mL), akin to commercial ELISA kits. Notably, the device incorporates a complex trilayer architecture and an independent rotational mechanism, enabling flexible analysis of 1–12 samples based on specific requirements. This innovation offers a promising avenue for efficient OPG detection in clinical settings, surmounting current diagnostic limitations.