He Hua, Ren Jicun
College of Chemistry & Chemical Engineering, Shanghai Jiaotong University, 800 Dongchuan Road, Shanghai 200240, PR China.
Talanta. 2008 Oct 19;77(1):166-71. doi: 10.1016/j.talanta.2008.05.059. Epub 2008 Jun 17.
We propose a novel evanescent wave scattering imaging method using an objective-type total internal reflection system to image and track single gold nanoparticles (GNPs) in solution. In this imaging system, only a millimeter-scale hole is employed to efficiently separate GNPs scattering light from the background reflected beam. The detailed experimental realization of the imaging system was discussed, and the effect of the hole size on imaging was investigated. We observed that the hole diameters from 2.5 to 4mm are suitable to perform the scattering imaging by adjusting the incidence angle. The technology was successfully applied to track single gold nanoparticles in solution and on live cell membrane via the anti-epidermal growth factor receptor antibody. Compared to total internal fluorescence microscopy, the resonance light scattering detection has no photobleaching or blinking inherent to fluorescent dyes and quantum dots. Compared to conventional dark-field microscopy, the evanescent wave illumination can be conveniently applied to study membrane dynamics in living cells. Additionally, the objective-based configuration provides a free space above the coverslip, and allows imaging and concomitant manipulation of live cells in culture by microinjection, patch-clamping, AFM and other techniques.
我们提出了一种新颖的倏逝波散射成像方法，该方法使用物镜型全内反射系统对溶液中的单个金纳米颗粒（GNP）进行成像和跟踪。在该成像系统中，仅采用一个毫米级的小孔来有效地将GNP散射光与背景反射光束分离。讨论了成像系统的详细实验实现，并研究了孔径大小对成像的影响。我们观察到，通过调整入射角，2.5至4毫米的孔径适合进行散射成像。该技术已成功应用于通过抗表皮生长因子受体抗体跟踪溶液中和活细胞膜上的单个金纳米颗粒。与全内荧光显微镜相比，共振光散射检测不存在荧光染料和量子点固有的光漂白或闪烁现象。与传统暗场显微镜相比，倏逝波照明可方便地用于研究活细胞中的膜动力学。此外，基于物镜的配置在盖玻片上方提供了一个自由空间，并允许通过显微注射、膜片钳、原子力显微镜和其他技术对培养中的活细胞进行成像和同步操作。
