Quantum-dot-based multiplexed fluorescence resonance energy transfer
Quantum-dot-based multiplexed fluorescence resonance energy transfer
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DOI：
10.1117/12.591181
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发表时间：
2005-04
期刊：
影响因子：
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通讯作者：
A. R. Clapp;Igor L. Medintz;H. T. Uyeda;B. Fisher;E. Goldman;M. Bawendi;H. Mattoussi
中科院分区：
文献类型：
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作者：
A. R. Clapp;Igor L. Medintz;H. T. Uyeda;B. Fisher;E. Goldman;M. Bawendi;H. Mattoussi
胶体半导体量子点(QD)具有窄的光电发射带宽和宽的吸收光谱，这是多路应用的理想选择。与有机染料不同的是，有机染料需要复杂的激发源和过滤器来产生多个信号，而许多量子点可以用一个激发源同时激发。在混合样品中，量子点的窄而对称的发射轮廓允许复合信号的简单去卷积以产生单独的量子点光致发光(PL)贡献。我们已经证明，在荧光共振能量转移(FRET)系统中，CdSe-ZnS核壳量子点是有效的能量供体。在这项研究中，我们测试了几种QD-蛋白质生物偶联物，每个生物偶联物都具有作为独立信号通道的独特的PL光谱(或“颜色”)，以评估基于QD FRET的多路复用系统的可行性。几个量子点群体与标记和未标记的蛋白质自组装，混合在溶液中，并在单一波长下激发。利用已知的量子点发射曲线对所得光谱进行去卷积处理，以揭示每个量子点群体的个体贡献。由于近端染料受体的存在，包被染料标记蛋白受体的量子点显示出明显的FRET诱导的荧光猝灭。来自混合样品的时间分辨光谱数据验证了稳态荧光结果，其中缩短的量子点寿命表明在一个或多个量子点群体上存在近端染料猝灭剂。我们将讨论如何利用这些发现来开发基于QD的FRET多路生物传感器，使用类似的策略，其中每个QD种群具有对唯一分子靶标敏感的表面结合蛋白。
Colloidal semiconductor quantum dots (QDs) have narrow photoemission bandwidths and broad absorption spectra that are ideal for multiplexing applications. In contrast to organic dyes, which require a complex arrangement of excitation sources and filters to generate multiple signals, many populations of QDs can be simultaneously excited with a single excitation source. In a mixed sample, the narrow and symmetric emission profile of QDs allows simple deconvolution of the composite signal to generate individual QD photoluminescence (PL) contributions. We have shown that CdSe-ZnS core-shell QDs function as efficient energy donors in fluorescence resonance energy transfer (FRET) systems. In this study, we tested several QD-protein bioconjugates, each having a unique PL spectrum (or "color") functioning as independent signal channels, to assess the feasibility of a QD FRET-based multiplexing system. Several populations of QDs were self-assembled with labeled and unlabeled proteins, mixed in solution and excited at single wavelength. The resulting spectra were deconvoluted using the known QD emission profiles to reveal individual contributions of each QD population. QDs coated with dye-labeled protein acceptors showed distinct FRET-induced PL quenching due to the presence of proximal dye acceptors. Steady-state fluorescence results were verified by time-resolved spectroscopic data from the mixed samples where a reduced QD lifetime indicated the presence of proximal dye quencher on one or more QD populations. We will discuss how these findings are used to develop QD-based FRET multiplexed biosensors using a similar strategy where each QD population has surface-bound proteins that are sensitive to a unique molecular target.