Influence of Luminescence Quantum Yield, Surface Coating, and Functionalization of Quantum Dots on the Sensitivity of Time-Resolved FRET Bioassays
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- 作者：K. David Wegner ; Phung Thi Lanh ; Travis Jennings ; Eunkeu Oh ; Vaibhav Jain ; Simon M. Fairclough ; Jason M. Smith ; Emerson Giovanelli ; Nicolas Lequeux ; Thomas Pons ; Niko Hildebrandt
- 刊名：ACS Applied Materials & Interfaces
- 出版年：2013
- 出版时间：April 24, 2013
- 年：2013
- 卷：5
- 期：8
- 页码：2881-2892
- 全文大小：634K
- 年卷期：v.5,no.8(April 24, 2013)
- ISSN：1944-8252
文摘
In clinical diagnostics, homogeneous time-resolved (TR) FRET immunoassays are used for fast and highly sensitive detection of biomarkers in serum samples. The most common immunoassay format is based on europium chelate or cryptate donors and allophycocyanin acceptors. Replacing europium donors with terbium complexes and the acceptors with QDs offers large photophysical advantages for multiplexed diagnostics, because the Tb-complex can be used as FRET donor for QD acceptors of different colors. Water-soluble and biocompatible QDs are commercially available or can be synthesized in the laboratory using many available recipes from the literature. Apart from the semiconductor material composition, an important aspect of choosing the right QD for TR-FRET assays is the thickness of the QD coating, which will influence the photophysical properties and long-term stability as well as the donor鈥揳cceptor distance and FRET efficiency. Here we present a detailed time-resolved spectroscopic study of three different QDs with an emission maximum around 605 nm for their application as FRET acceptors (using a common Tb donor) in TR-bioassays: (i) Invitrogen/Life Technologies Qdot605, (ii) eBioscience eFluorNC605 and iii) ter-polymer stabilized CdSe/CdS/ZnS QDs synthesized in our laboratories. All FRET systems are very stable and possess large F枚rster distances (7.4鈥?.1 nm), high FRET efficiencies (0.63鈥?.80) and low detection limits (0.06鈥?.0 pM) within the FRET-bioassays. Shapes, sizes and the biotin/QD ratio of the biocompatible QDs could be determined directly in the solution phase bioassays at subnanomolar concentrations. Both commercial amphiphilic polymer/lipid encapsulated QDs and self-made ligand-exchanged QDs provide extremely low detection limits for highly sensitive TR-FRET bioassays.