Fluorescence-encoded microspheres are widely used in the detection and analysis of biological molecules, especially in suspension arrays. Here, we report an efficient strategy for the preparation of fluorescence-encoded polystyrene microspheres with desirable optical and surface properties. The micron-sized, monodisperse polystyrene seed beads were first synthesized by dispersion polymerization. Then, dye molecules and carboxyl functional groups were copolymerized on the surface of the seed beads by forming a core-shell structure. Rhodamine 6G (R6G) was used as a model dye molecule to prepare the fluorescent beads, and the fluorescence intensity of the beads can be precisely controlled by adjusting the quantity of R6G. These fluorescent beads were characterized by environmental scanning electron microscopy, laser scanning confocal microscopy, and spectrofluorometry. The differences of the fluorescence spectra between fluorescent beads and R6G in solution were investigated. Twelve kinds of fluorescent beads encoded with different R6G fluorescence intensities were prepared, and they can be clearly distinguished on a conventional flow cytometer. Furthermore, the encoded beads are stable in water and resistant to photobleaching, which is crucial for their potential applications in diagnostic assays and imaging. Detection of human alpha fetoprotein antigen via a sandwich microsphere-based immunoassay yielded a detection limit of 80 pg mL(-1), demonstrating that the fluorescence-encoded microspheres synthesized herein are efficient in serving as the microcarriers in suspension arrays. As both the encoding and functionalizing procedures are made simultaneously, the newly designed technique is extremely simple and time-saving. Moreover, it could be readily applicable to the preparation of a wide size range of fluorescent particles made by polymerization.
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