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在细胞膜中可视化受体动力学的替代方法。

Alternative method to visualize receptor dynamics in cell membranes.

机构信息

Department of Molecular and Translational Medicine, University of Brescia, Brescia, Italy.

The Mechanobiology Research Center, UNIBS, Brescia, Italy.

出版信息

PLoS One. 2024 Jun 11;19(6):e0304172. doi: 10.1371/journal.pone.0304172. eCollection 2024.

Abstract

There is a close relation between membrane receptor dynamics and their behavior. Several microscopy techniques have been developed to study protein dynamics in live cells such as the Fluorescence Recovery After Photobleaching (FRAP) or the Single Particle Tracking (SPT). These methodologies require expensive instruments, are time consuming, allow the analysis of small portion of the cell or an extremely small number of receptors at a time. Here we propose a time-saving approach that allows to visualize the entire receptor pool and its localization in time. This protocol requires an epifluorescence microscope equipped for structured illuminated sectioning and for live cell imaging. It can be applied to characterize membrane receptor and multi protein complex and their response to activators or inhibitors. Image acquisition and analysis can be performed in two days, while cells and substratum preparation require a few minutes a day for three days.

摘要

膜受体动力学与其行为之间存在密切关系。已经开发出几种显微镜技术来研究活细胞中的蛋白质动力学,例如荧光漂白后恢复(FRAP)或单粒子跟踪(SPT)。这些方法需要昂贵的仪器,耗时耗力,一次只能分析细胞的一小部分或受体的极少量。在这里,我们提出了一种节省时间的方法,可以实时可视化整个受体库及其定位。该方案需要配备结构照明切片和活细胞成像的荧光显微镜。它可用于表征膜受体和多蛋白复合物及其对激活剂或抑制剂的反应。图像采集和分析可以在两天内完成,而细胞和基质制备每天需要几分钟,持续三天。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/60b7/11166301/b77ba7956081/pone.0304172.g001.jpg

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