Single-bead immunoassays using magnetic microparticles and spectral-shifting quantum dots

This paper reports a single-bead immunoassay method based on the combined use of magnetic microparticles (MMPs) for target capturing/enrichment and antibody-conjugated semiconductor quantum dots (QDs) for fluorescence detection. In comparison with organic dyes and fluorescent proteins, QDs exhibit unique optical properties such as size-tunable fluorescence emission (spectral shifting), large absorption coefficients, improved brightness, and superior photostability. Magnetic beads, composed of iron oxide nanoparticles embedded in polymeric matrices, provide a platform for rapid capturing and enrichment of biomolecules and pathogens in dilute biological and environmental samples. However, a major problem in using magnetic beads for fluorescence immunoassays is that the bead's autofluorescence strongly interferes with the target detection signal. This spectral overlapping problem can be overcome by using semiconductor QDs as a new class of spectral-shifting labels. By shifting the QD emission signals away from the bead autofluorescence, it is possible to detect biomolecular antigens such as tumor necrosis factor (TNF-alpha) at femtomolar (10(-15) M) concentrations when the target molecules are captured and enriched on the magnetic bead surface. This sensitivity is almost 1000 times higher that that of traditional immunoabsorbent assays and more than 100 times higher than immunofluorescent assays using organic dyes.