Construction of a Core-Satellite Immunoassay with Homogeneous Magnet-Assisted Ratiometric Fluorescence for Highly...

The development of in vitro diagnostics (IVD) has been characterized by a focus on efficiency and convenience in a variety of testing scenarios. Herein, the homogeneous magnet-assisted ratiometric fluorescence (HMRF) method was used to capture, recognize, and convert the signal of target inflammatory factors in the same reaction system, which achieves highly sensitive and visual detection of multiple inflammatory factors (C-reactive protein (CRP), procalcitonin (PCT), and interleukin-6 (IL-6)). The aqueous green quantum dots (QDs)-magnetic microspheres (Fe₃O₄@Tri-layer QDs, three closely stacked QD layers) with high fluorescence and magnetic properties were prepared by the layer-by-layer (LBL) self-assembly method. These were combined with red silica-coated QDs (QDs@SiO₂) to design a core-satellite immunoassay, yielding a visual ratiometric fluorescence signal ranging from green-yellow-red color. Under the optimal conditions, a wide linear range was achieved for CRP, PCT, and IL-6, with the limit of detection (LOD) of 50.0 pg/mL, 35.5 pg/mL, and 18.4 pg/mL, respectively. Furthermore, the core-satellite immunoassay demonstrated resistance to interference from common substances in blood, and the recoveries ranged from 105.8% to 109.7%, showing good specificity, accuracy, and stability. The HMRF assay was verified with 44 human serum samples of CRP, and the results were found to be in good agreement with the clinical method. The core-satellite immunoassay provides a new technical tool for the diagnosis and research of multiple diseases and is of great importance in the field of highly sensitive, rapid, and accurate IVD.