Kufer Stefan K, Strackharn Mathias, Stahl Stefan W, Gumpp Hermann, Puchner Elias M, Gaub Hermann E
Nat Nanotechnol. 2009 Jan;4(1):45-9. doi: 10.1038/nnano.2008.333. Epub 2008 Nov 23.
Bottom-up assembly at the level of individual molecules requires a combination of utmost spatial precision and efficient monitoring. We have previously shown how to 'cut-and-paste' single molecules, and other groups have demonstrated that it is possible to beat the diffraction limit in optical microscopy. Here we show that a combination of single-molecule cut-and-paste surface assembly, total internal reflection fluorescence microscopy and atomic force microscopy can be used to deposit individual fluorophores in well-defined nanoscale patterns and also to monitor the process in real time with nanometre precision. Although the size of the pattern is well below the optical resolution of the microscope, the individual dyes are identified by localizing the centroids and detecting the photobleaching of the fluorophores. With this combination of methods, individual dyes or labelled biomolecules can be arranged at will for specific functions, such as coupled fluorophore systems or tailored enzyme cascades, and monitored with nanoscale precision.
在单个分子层面进行自下而上的组装需要极高的空间精度和高效的监测手段相结合。我们之前已经展示了如何“剪切粘贴”单个分子，其他研究团队也证明了在光学显微镜中突破衍射极限是可行的。在此，我们表明，单分子剪切粘贴表面组装、全内反射荧光显微镜和原子力显微镜相结合，可用于以明确的纳米级图案沉积单个荧光团，并能以纳米精度实时监测这一过程。尽管图案尺寸远低于显微镜的光学分辨率，但通过定位质心和检测荧光团的光漂白来识别单个染料。通过这种方法的组合，单个染料或标记的生物分子可以根据特定功能随意排列，如耦合荧光团系统或定制的酶级联反应，并以纳米级精度进行监测。
