Evaluating the intracellular stability and unpacking of DNA nanocomplexes by quantum dots-FRET
Evaluating the intracellular stability and unpacking of DNA nanocomplexes by quantum dots-FRET
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DOI：
10.1016/j.jconrel.2006.09.005
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发表时间：
2006-11-10
影响因子：
10.8
通讯作者：
Wang, Tza-Huei
中科院分区：
文献类型：
--
作者：
Ho, Yi-Ping;Chen, Hunter H.;Wang, Tza-Huei
我们展示了一种高度灵敏的方法来表征聚合物DNA纳米复合物的结构组成和细胞内的命运，通过静电相互作用将质粒DNA与阳离子聚合物缩合而形成。合理设计更有效的聚合物基因载体将是可能的，只有在非病毒基因转移过程中的限速步骤的机械见解。为了表征纳米复合物的组成和结合动力学，将纳米复合物中的质粒及其聚合物载体分别用量子点（QD）和荧光有机染料标记，作为荧光共振能量转移（FRET）的供体和受体对。量子点介导的FRET中的高信噪比使得能够在单颗粒水平上精确检测纳米复合物状态的离散变化，以对抗各种细胞内微环境。单个纳米复合物在细胞内的分布和解包因此可以通过荧光显微镜明确地进行。QD-FRET是一种高度灵敏和定量的方法，用于确定纳米复合物在细胞内转运过程中的组成和动态稳定性，其中可以识别基因递送的障碍以促进基因载体优化。(c)2006 Elsevier B. V.保留所有权利。
We demonstrate a highly sensitive method to characterize the structural composition and intracellular fate of polymeric DNA nanocomplexes, formed by condensing plasmid DNA with cationic polymers through electrostatic interactions. Rational design of more efficient polymeric gene carriers will be possible only with mechanistic insights of the rate-limiting steps in the non-viral gene transfer process. To characterize the composition and binding dynamics of nanocomplexes, plasmid and its polymer carrier within nanocomplexes were labeled with quantum dots (QDs) and fluorescent organic dyes, respectively, as a donor and acceptor pair for fluorescence resonance energy transfer (FRET). The high signal-to-noise ratio in QD-mediated FRET enabled precise detection of discrete changes in nanocomplex state at the single-particle level, against various intracellular microenvironments. The distribution and unpacking of individual nanocomplexes within cells could thus be unambiguously followed by fluorescence microscopy. QD-FRET is a highly sensitive and quantitative method to determine the composition and dynamic stability of nanocomplexes during intracellular transport, where barriers to gene delivery may be identified to facilitate gene carrier optimization. (c) 2006 Elsevier B.V. All rights reserved.