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用于 VGIC 动力学筛选的可记录细胞。

Ready-to-Record Cells for Kinetic Screening of VGICs.

机构信息

Blue Brain Project, Ecole Polytechnique Fédérale de Lausanne, Geneva, Switzerland.

出版信息

Methods Mol Biol. 2024;2796:87-95. doi: 10.1007/978-1-0716-3818-7_5.

Abstract

Voltage-gated ion channels (VGICs) are integral membrane proteins crucial for transmitting electrical signals in excitable cells. Understanding the kinetics of these ion channels requires conducting patch-clamp experiments using genetically modified cell lines that express a single type of ion channel gene. However, this process relies on the continuous maintenance of cell lines to ensure an adequate supply of sample cells for patch-clamp experiments. Advancements in automated patch-clamp methods have enabled researchers to significantly increase the number of patch-clamped cells per experiment, from just a few cells to as many as 384 cells. Despite this progress, the manual task of preparing the cell samples remains a significant bottleneck in the kinetic screening of VGICs. Here we describe a method to address this challenge by generating ready-to-record (RTR) VGIC-expressing cells that can be frozen and stored separately from patch-clamp experiments. This decoupling of the cell sample preparation process from the patch-clamp experiments offers a streamlined approach to studying VGICs on manual or an automated patch-clamp system.

摘要

电压门控离子通道(VGICs)是细胞膜的整合蛋白,对于可兴奋细胞中电信号的传递至关重要。理解这些离子通道的动力学特性需要使用表达单一类型离子通道基因的基因修饰细胞系进行膜片钳实验。然而,这个过程依赖于细胞系的持续维护,以确保有足够数量的样本细胞用于膜片钳实验。自动化膜片钳方法的进步使研究人员能够显著增加每个实验中钳制的细胞数量,从几个细胞增加到多达 384 个细胞。尽管取得了这一进展,但细胞样本制备的手动任务仍然是 VGIC 动力学筛选的一个重大瓶颈。在这里,我们描述了一种通过生成可记录(RTR)表达 VGIC 的细胞来解决这一挑战的方法,这些细胞可以与膜片钳实验分开冷冻和储存。这种将细胞样本制备过程与膜片钳实验解耦的方法为在手动或自动化膜片钳系统上研究 VGIC 提供了一种简化的方法。

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