Homogeneous Phase Fluorescence Assay Based on NaYF4∶Er3+, Yb3+ UCNPs as Donors
SONG Kai1, 2, ZHANG Qing-bin2, ZHAO Jun-wei2, KONG Xiang-gui2*
1. School of Life Science, Changchun Normal University, Changchun 130032, China 2. Key Laboratory of Excited State Process, Changchun Institute of Optics, Fine Mechanics and Physics, Chinese Academy of Sciences, Changchun 130033, China
Abstract：Upconversion nanoparticles (UCNPs) of small size have great potential in homogeneous assay based on fluorescence resonance energy transfer (FRET). A novel approach of surface ligand-exchange for preparing water-soluble and amido-functionalized NaYF4∶Er3+, Yb3+ UCNPs with 12 nm was developed. The ligand exchange process was confirmed by Fourier transform infrared spectra. Investigations by scan electron microscopy showed no obvious variations in the size and shape of the UCNPs. The circular dichroism characterization demonstrates that the secondary structure of the avidin remains largely intact after the conjugation. Avidin-biotin served as a bridge to make the short enough distance for FRET between the acceptor biotinylated R-phycoerythrin and the donors avidin-conjugated UCNPs. When the free biotins were added into this system, they competitively combined with avidin on the UCNPs surface and impede the FRET to triggered fluorescence changes. According to the relationship between this change and the addition of the amount of biotin, such FRET-based approach can reach a limit of detection in the nanomolar concentration range.
Key words：Upconversion nanoparticales;Fluorescence resonance energy transfer;Homogeneous phase fluorescence assays
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