Characterization of Membrane Patch‐Ion Channel Probes for Scanning Ion Conductance Microscopy

Abstract

Integration of dual-barrel membrane patch-ion channel probes (MP-ICPs) to scanning ion conductance microscopy (SICM) holds promise of providing a revolutionized approach of spatially resolved chemical sensing. A series of experiments are performed to further the understanding of the system and to answer some fundamental questions, in preparation for future developments of this approach. First, MP-ICPs are constructed that contain different types of ion channels including transient receptor potential vanilloid 1 and large conductance Ca2 -activated K channels to establish the generalizability of the methods. Next, the capability of the MP-ICP platforms in single ion channel activity measurements is proved. In addition, the interplay between the SICM barrel and the ICP barrel is studied. For ion channels gated by uncharged ligands, channel activity at the ICP barrel is unaffected by the SICM barrel potential; whereas for ion channels that are gated by charged ligands, enhanced channel activity can be obtained by biasing the SICM barrel at potentials with opposite polarity to the charge of the ligand molecules. Finally, a proof-of-principle experiment is performed and site-specific molecular/ionic flux sensing is demonstrated at single-ion-channel level, which show that the MP-ICP platform can be used to quantify local molecular/ionic concentrations.

Application of scanning ion conductance microscope in cell characterizations

“…Tanaka等 [74] 通过SICM持续 观察了大鼠大脑皮层单个神经元细胞凋亡的生化和形 态学变化, 从凋亡早期体积减少到细胞膜起泡和磷脂 酰丝氨酸的易位等过程(图7(c)). [78] . 例如, Gorelik等 [79] 利 用SICM与膜片钳技术联合研究了海胆的精子细胞和 颈上神经节的钙离子通道形貌及钡离子通过的局部离 子电流.…”

Advances and Perspectives in Chemical Imaging in Cellular Environments Using Electrochemical Methods

“…Using a doublebarrel probe for membrane-patching required the pipettes to be fire-polished to minimize capacitative artifacts and facilitate a gigaseal membrane across the opening [99]. This method of membrane patching opens up the range of ion channels that can be incorporated into a ICP [99] [102].…”

Recent development of samples’ surface properties using scanning ion conductance microscopy

“…Exploration of surface properties 2.1. Surface charge: Shi et al [26] integrated dual-barrel membrane patch-ion channel probes (MP-ICPs) into SICM, which was deemed as a revolutionised method of spatially resolved chemical sensing. The setup is shown in Fig.…”