Scanning ion-conductance microscopy with a double-barreled nanopipette for topographic imaging of charged chromosomes
Scanning ion-conductance microscopy with a double-barreled nanopipette for topographic imaging of charged chromosomes
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使用双管纳米移液器的扫描离子电导显微镜对带电染色体进行形貌成像
DOI：
10.1093/jmicro/dfab009
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发表时间：
2021
期刊：
影响因子：
1.8
通讯作者：
Ushiki Tatsuo
中科院分区：
文献类型：
--
作者：
Iwata Futoshi;Shirasawa Tatsuru;Mizutani Yusuke;Ushiki Tatsuo
扫描离子电导显微镜（SICM）可用于对液体中柔软和易碎的生物样品进行成像，因为它通过在非接触和无力条件下检测离子电流来探测样品的表面形貌。SICM通过检测当填充电解质的玻璃纳米移液管接近样品表面时发生的离子电流降低来获取表面形貌高度。然而，大多数生物材料在液体环境中具有带电表面，这有时会影响SICM检测到的离子电流的行为，特别是使形貌测量变得困难。为了测量这种带电样品，我们提出了一种新的成像方法，使用双管纳米移液管作为SICM探针。纳米移液管的两个孔之间的离子电流降低了表面电荷对成像的影响。在这项研究中，中期染色体的印度赤麂通过这种技术成像，因为，由于其在磷酸盐缓冲液中的强负电荷的表面，它是很难获得的拓扑结构的染色体由传统的SICM与单孔纳米移液管。使用所提出的方法与双筒纳米移液管，染色体的表面被成功地测量，没有任何表面电荷混淆。由于样品形貌的详细成像可以在生理液体条件下进行，而与样品电荷无关，因此预计将用于分析与细胞分裂中的动态变化相关的染色体的高级结构。
Scanning ion conductance microscopy (SICM) is useful for imaging soft and fragile biological samples in liquids because it probes the samples’ surface topography by detecting ion currents under non-contact and force-free conditions. SICM acquires the surface topographical height by detecting the ion current reduction that occurs when an electrolyte-filled glass nanopipette approaches the sample surface. However, most biological materials have electrically charged surfaces in liquid environments, which sometimes affect the behavior of the ion currents detected by SICM and, especially, make topography measurements difficult. For measuring such charged samples, we propose a novel imaging method that uses a double-barrel nanopipette as an SICM probe. The ion current between the two apertures of the nanopipette desensitizes the surface charge effect on imaging. In this study, metaphase chromosomes of Indian muntjac were imaged by this technique because, owing to their strongly negatively charged surfaces in phosphate-buffered saline, it is difficult to obtain the topography of the chromosomes by the conventional SICM with a single-aperture nanopipette. Using the proposed method with a double-barrel nanopipette, the surfaces of the chromosomes were successfully measured, without any surface charge confounder. Since the detailed imaging of sample topography can be performed in physiological liquid conditions regardless of the sample charge, it is expected to be used for analyzing the high-order structure of chromosomes in relation to their dynamic changes in the cell division.