首页> 外文会议>Conference on Optical Diagnostics of Living Cells Ⅴ, Jan 23-25, 2002, San Jose, USA >FRAP model to determine the bi-directional transport rate of GFP across the nuclear membrane and the mobile fraction in the cytoplasm and nucleus
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FRAP model to determine the bi-directional transport rate of GFP across the nuclear membrane and the mobile fraction in the cytoplasm and nucleus

机译:FRAP模型可确定GFP跨核膜的双向传输速率以及细胞质和细胞核中的可移动部分

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A mathematical model was developed to predict the bi-directional transport rate of fluorescent proteins across the nuclear membrane during a fluorescence recovery after photobleaching (FRAP) experiment. The model assumes that the total amount of fluorescent protein remains the same in the cell (i.e. no production, loss or exchange with the outside of the cell) and that the cell is in a state of equilibrium; i.e. proteins are leaving and entering the nucleus at an equal rate. The latter assumption has the advantage of not needing to take into account the method of protein transport (e.g. active or passive). The model includes correction for the photobleaching that happens during image acquisition following the deliberate photobleach. In this study, the green fluorescent protein (GFP) was transfected into cells in order to study its free behavior. In the FRAP experiments, either the entire nucleus and part of the cytoplasm or only part of the cytoplasm was photobleached followed by time-series imaging of the fluorescence redistribution. The model was fitted to the curves of intensity loss or recovery after photobleaching using numerical, nonlinear methods. In addition, the mobile fractions of free GFP in the cytoplasm and the nucleus could be determined.
机译:建立了数学模型以预测光漂白(FRAP)实验后的荧光恢复过程中荧光蛋白跨核膜的双向运输速率。该模型假设荧光蛋白的总量在细胞中保持不变(即不与细胞外部产生,损失或交换),并且细胞处于平衡状态;即蛋白质以相同的速度离开和进入细胞核。后一种假设的优点是不需要考虑蛋白质转运的方法(例如主动或被动)。该模型包括对在故意进行光漂白之后图像采集期间发生的光漂白的校正。在这项研究中,绿色荧光蛋白(GFP)被转染到细胞中以研究其自由行为。在FRAP实验中,将整个细胞核和部分细胞质或仅部分细胞质进行光漂白,然后对荧光重新分布进行时间序列成像。使用数值非线性方法将模型拟合到光漂白后强度损失或恢复的曲线。另外,可以确定游离GFP在细胞质和细胞核中的流动级分。

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