Nanometer-scale mapping and single-molecule detection with color-coded nanoparticle probes
Nanometer-scale mapping and single-molecule detection with color-coded nanoparticle probes
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DOI：
10.1073/pnas.0712351105
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发表时间：
2008-03-04
影响因子：
11.1
通讯作者：
Nie, Shuming
中科院分区：
文献类型：
--
作者：
Agrawal, Amit;Deo, Rajesh;Nie, Shuming
我们报告了一种基于双色成像和纳米精度生物共轭纳米颗粒探针自动共定位的单分子检测和生物分子结构绘图方法。与有机染料和荧光蛋白相比，荧光能量转移纳米珠和量子点等纳米粒子探针在信号亮度、光稳定性和多色光发射方面具有显着优势。因此，我们利用标准荧光显微镜和廉价的数码彩色相机实现了常规的双色超分辨率成像和单分子检测。通过使用绿色和红色纳米颗粒同时识别单个靶标上的两个结合位点，可以检测和识别单个生物分子（例如核酸），而无需靶标扩增或探针/靶标分离。我们还证明，一种强大的天体物理方法（最初是为分析拥挤的恒星场而开发的）可用于纳米颗粒共定位信号的自动快速统计分析。以纳米精度快速定位明亮纳米颗粒探针的能力不仅对超灵敏的医学检测具有影响，而且对分子复合物的结构绘图也具有影响，其中各个成分用颜色编码的纳米颗粒进行标记。
We report a method for single-molecule detection and biomolecular structural mapping based on dual-color imaging and automated colocalization of bioconjugated nanoparticle probes at nanometer precision. In comparison with organic dyes and fluorescent proteins, nanoparticle probes such as fluorescence energy-transfer nanobeads and quantum dots provide significant advantages in signal brightness, photostability, and multicolor-light emission. As a result, we have achieved routine two-color super-resolution imaging and single-molecule detection with standard fluorescence microscopes and inexpensive digital color cameras. By using green and red nanoparticles to simultaneously recognize two binding sites on a single target, individual biomolecules such as nucleic acids are detected and identified without target amplification or probe/target separation. We also demonstrate that a powerful astrophysical method (originally developed to analyze crowded stellar fields) can be used for automated and rapid statistical analysis of nanoparticle colocalization signals. The ability to rapidly localize bright nanoparticle probes at nanometer precision has implications not only for ultrasensitive medical detection but also for structural mapping of molecular complexes in which individual components are tagged with color-coded nanoparticles.