Procion red dye chemically bonded PGMA microspheres towards leakage free and sensitivity improved lateral flow immunoassay†

Lateral flow immunoassay strips (LFIAs) are reliable tools for point-of-care testing. However, their sensitivity is limited by the low extinction coefficient of microspheres with low dye content. To avoid these flaws, we report here a type of Procion red chemical-bonded amino-decorated poly(glycidyl methacrylate) (PGMA) microspheres through a three-step process, including a soap-free emulsion polymerization and carboxyl-functionalization, an amination, and a final colorization process. Ethylenediamine and Procion red are introduced onto PGMA microspheres with high concentrations of 10.2 N wt% and 39.1 wt%, respectively. Due to such heavy introduction of chemically bonded dyes, the molar extinction coefficient reached up to 3.28 × 10¹¹ L mol⁻¹ cm⁻¹, which significantly exceeds that of commercially used colloidal gold nanoparticles (9.44 × 10⁹ L mol⁻¹ cm⁻¹). Based on the colored PGMA microspheres, we propose a drop casting and photo scanning protocol to find the minimum detectable particle density of the microspheres. As a result, the Procion red bonded PGMA microspheres could be easily identified with a density as low as 4.43 × 10⁴ particles per mm². Finally, when applied to COVID-19 antigen detection, the microspheres achieved a sensitivity of 0.025 ng mL⁻¹ and exhibited broad pH (3–11) and long-term (7 months) stability and high specificity. They are better or comparable to other typical bio-label materials, such as colloidal gold nanoparticles and dye-composite polystyrene (PS) microspheres. The Procion red bonded PGMA microspheres offer several advantages for diagnosing major sudden and high-incidence diseases, facilitating rapid screening while maintaining high bio-detection sensitivity and stability without modifying the test format.