Atomic Force Microscopy Imaging and Probing of DNA, Proteins, and Protein-DNA Complexes: Silatrane Surface Chemistry
Atomic Force Microscopy Imaging and Probing of DNA, Proteins, and Protein-DNA Complexes: Silatrane Surface Chemistry
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DOI：
10.1007/978-1-60327-015-1_21
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发表时间：
2009-01-01
期刊：
影响因子：
--
通讯作者：
Gall, Alexander A.
中科院分区：
文献类型：
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作者：
Lyubchenko, Yuri L.;Shlyakhtenko, Luda S.;Gall, Alexander A.
尽管原子力显微镜是最近才发明的，但它在商业上已经广泛使用。 AFM 及其特殊改进（轻敲模式和溶液中的非接触操作）已成功用于大量生物物体的形貌研究，包括 DNA、RNA、蛋白质、细胞膜甚至整个细胞。 AFM 还成功应用于核酸和各种蛋白质-DNA 复合物的研究。这一成功的部分原因在于可靠的样品制备程序的开发。本章介绍了一种基于用 1-(3-氨基丙基)硅杂烷 (APS) 对云母表面进行化学功能化的方法。 APS-云母方法最重要的特性之一是，在不存在二价阳离子和宽范围的 pH 条件下，样品可以在宽范围的离子强度下沉积在表面上。除了干燥样品成像之外，APS-云母还可以在水溶液中进行可靠且可重复的延时成像。最后，APS 云母以反应性氨基封端，可用于 AFM 力谱研究中分子的共价和离子附着。概述了 APS 的制备、APS 云母和 AFM 探针功能化、空气和水溶液中成像样品的制备以及力谱研究的方案。所有这些应用程序都通过一些示例进行说明。
Despite their rather recent invention, atomic force microscopes are widely available commercially. AFM and its special modifications (tapping mode and noncontact operation in solution) have been successfully used for topographic Studies of a large number of biological objects including DNA, RNA, proteins, cell membranes, and even whole cells. AFM was also successfully applied to Studies of nucleic acids and various protein-DNA complexes. Part of this Success is due to the development of reliable sample preparation procedures. This chapter describes one of the approaches based on chemical functionalization of mica surface with 1-(3-aminopropyl) silatrane (APS). One of the most important properties of APS-mica approach is that the sample can be deposited Oil the surface in a wide range of ionic strengths, in the absence of divalent cations and a broad range of pH. In addition to imaging of dried sample, APS-mica allows reliable and reproducible time lapse imaging in aqueous solutions. Finally, APS mica is terminated with reactive amino groups that can be used for covalent and ionic attachment of molecules for AFM force spectroscopy studies. The protocols for the preparation of APS, functionalization with APS mica and AFM probes, preparation of samples for imaging in air and in aqueous solutions, and force spectroscopy studies are outlined. All these applications are illustrated with a few examples.