Delain E, Michel D, Le Grimellec C
Laboratoire de Microscopie Moléculaire et Cellulaire/CNRS UMR 8532, Institut Gustave Roussy, Villejuif, France.
Morphologie. 2000 Jun;84(265):25-30.
Near field (or scanning probe) microscopy is a recent technology which, owing to the huge amount of publications, is becoming a reference method in molecular and cellular imaging. These microscopies consist in the scanning of the sample, line by line, with a very tiny tip and thus providing informations on its surface down to the nanometer scale. These methods gather scanning tunelling microscopy (STM), which measures a current between the tip and the specimen support, atomic force microscopy (AFM), which measures the repulsive and attractive forces of the tip in contact or very close to the specimen, and scanning near field optical microscopies (SNOM), for which a glass tip allows to catch light signals. Atomic force microscopy, which allows the observation of specimens in air or physiological conditions environments, is presently dominant in biology, in complementarity with the classical optical and electron microscopies, which by the way, have also shown considerable improvements during the last years. The complementarity of these microscopies is due to their very different basic principles, which provide them various possibilities and limits. The biological applications of STM is limited by the need of conducting samples, but the different models of SNOM, often still in development, allow to consider very interesting applications, particularly for detecting very faint and tiny fluorescence signals. Different examples will be given concerning the visualization by AFM of isolated DNA molecules, naked or associated with proteins, the observation of intact or decondensed chromosomes, as well as living cells. One of the originality of AFM is its capacity to observe objects in a wide range of enlargements, with fields from a few hundred of nanometers to several micrometers.
近场(或扫描探针)显微镜是一项新兴技术,由于大量的相关出版物,它正成为分子和细胞成像中的一种参考方法。这些显微镜通过用非常微小的探针逐行扫描样品,从而提供其表面直至纳米尺度的信息。这些方法包括扫描隧道显微镜(STM),它测量探针与样品支撑之间的电流;原子力显微镜(AFM),它测量探针与样品接触或非常接近时的排斥力和吸引力;以及扫描近场光学显微镜(SNOM),其玻璃探针可捕获光信号。原子力显微镜能够在空气或生理条件环境中观察样品,目前在生物学领域占据主导地位,与传统的光学显微镜和电子显微镜相辅相成,顺便说一句,传统显微镜在过去几年也有了显著改进。这些显微镜的互补性源于它们截然不同的基本原理,这为它们提供了各种可能性和局限性。STM的生物学应用受到对导电样品需求的限制,但不同型号的SNOM(通常仍在开发中)使得人们可以考虑非常有趣的应用,特别是用于检测非常微弱和微小的荧光信号。将给出不同的例子,涉及通过AFM可视化分离的DNA分子(裸DNA或与蛋白质结合的DNA)、完整或解聚的染色体以及活细胞。AFM的独特之处之一在于其能够在广泛的放大倍数下观察物体,视野范围从几百纳米到几微米。