Department of Biology, School of Medicine, University of Patras, Rio, Patras 26505, Greece.
Department of Physiology, School of Medicine, University of Patras, Rio, Patras 26505, Greece.
Nucleic Acids Res. 2018 Jul 2;46(W1):W467-W472. doi: 10.1093/nar/gky508.
Understanding protein dynamics is crucial in order to elucidate protein function and interactions. Advances in modern microscopy facilitate the exploration of the mobility of fluorescently tagged proteins within living cells. Fluorescence recovery after photobleaching (FRAP) is an increasingly popular functional live-cell imaging technique which enables the study of the dynamic properties of proteins at a single-cell level. As an increasing number of labs generate FRAP datasets, there is a need for fast, interactive and user-friendly applications that analyze the resulting data. Here we present easyFRAP-web, a web application that simplifies the qualitative and quantitative analysis of FRAP datasets. EasyFRAP-web permits quick analysis of FRAP datasets through an intuitive web interface with interconnected analysis steps (experimental data assessment, different types of normalization and estimation of curve-derived quantitative parameters). In addition, easyFRAP-web provides dynamic and interactive data visualization and data and figure export for further analysis after every step. We test easyFRAP-web by analyzing FRAP datasets capturing the mobility of the cell cycle regulator Cdt2 in the presence and absence of DNA damage in cultured cells. We show that easyFRAP-web yields results consistent with previous studies and highlights cell-to-cell heterogeneity in the estimated kinetic parameters. EasyFRAP-web is platform-independent and is freely accessible at: https://easyfrap.vmnet.upatras.gr/.
了解蛋白质动力学对于阐明蛋白质功能和相互作用至关重要。现代显微镜技术的进步促进了对活细胞内荧光标记蛋白的流动性的探索。荧光漂白恢复（FRAP）是一种越来越流行的功能活细胞成像技术，可用于研究蛋白质在单细胞水平上的动态特性。随着越来越多的实验室生成 FRAP 数据集，需要快速、交互和用户友好的应用程序来分析这些数据。在这里，我们介绍了 easyFRAP-web，这是一个简化 FRAP 数据集定性和定量分析的网络应用程序。easyFRAP-web 通过直观的网络界面提供快速的 FRAP 数据集分析，该界面具有相互关联的分析步骤（实验数据评估、不同类型的归一化和曲线衍生定量参数的估计）。此外，easyFRAP-web 提供动态和交互式数据可视化以及每步之后的数据分析和图形导出。我们通过分析在培养细胞中存在和不存在 DNA 损伤时细胞周期调节剂 Cdt2 的流动性的 FRAP 数据集来测试 easyFRAP-web。我们表明，easyFRAP-web 产生的结果与先前的研究一致，并突出了估计的动力学参数中的细胞间异质性。easyFRAP-web 是独立于平台的，可免费访问：https://easyfrap.vmnet.upatras.gr/。
