CEA, iBEB, Service de Biochimie et Toxicologie Nucléaire, F-30207 Bagnols sur Cèze, France.
J Mol Recognit. 2011 May-Jun;24(3):503-10. doi: 10.1002/jmr.1118.
The study of high-resolution topographic surfaces of isolated single molecules is one of the applications of atomic force microscopy (AFM). Since tip-induced distortions are significant in topographic images the exact AFM tip shape must be known in order to correct dilated AFM height images using mathematical morphology operators. In this work, we present a protocol to estimate the AFM tip apex radius using tobacco mosaic virus (TMV) particles. Among the many advantages of TMV, are its non-abrasivity, thermal stability, bio-compatibility with other isolated single molecules and stability when deposited on divalent ion pretreated mica. Compared to previous calibration systems, the advantage of using TMV resides in our detailed knowledge of the atomic structure of the entire rod-shaped particle. This property makes it possible to interpret AFM height images in term of the three-dimensional structure of TMV. Results obtained in this study show that when a low imaging force is used, the tip is sensing viral protein loops whereas at higher imaging force the tip is sensing the TMV particle core. The known size of the TMV particle allowed us to develop a tip-size estimation protocol which permits the successful erosion of tip-convoluted AFM height images. Our data shows that the TMV particle is a well-adapted calibrator for AFM tips for imaging single isolated biomolecules. The procedure developed in this study is easily applicable to any other spherical viral particles.
研究孤立单分子的高分辨率地形表面是原子力显微镜（AFM）的应用之一。由于在地形图像中尖端诱导的变形非常显著，因此为了使用数学形态学算子校正膨胀的 AFM 高度图像，必须知道确切的 AFM 尖端形状。在这项工作中，我们提出了一种使用烟草花叶病毒（TMV）颗粒估计 AFM 尖端半径的方案。TMV 的许多优点包括其非研磨性、热稳定性、与其他孤立单分子的生物相容性以及在二价离子预处理云母上的稳定性。与以前的校准系统相比，使用 TMV 的优势在于我们对整个棒状颗粒的原子结构有详细的了解。这一特性使得能够根据 TMV 的三维结构来解释 AFM 高度图像。本研究的结果表明，当使用低成像力时，尖端会感知病毒蛋白环，而当使用更高的成像力时，尖端会感知 TMV 颗粒核心。已知的 TMV 颗粒大小使我们能够开发出一种尖端尺寸估计方案，该方案允许成功地侵蚀尖端卷积的 AFM 高度图像。我们的数据表明，TMV 颗粒是一种非常适合用于成像单个孤立生物分子的 AFM 尖端的校准器。本研究中开发的程序很容易适用于任何其他球形病毒颗粒。
