Richard D. Berlin Center for Cell Analysis and Modeling, University of Connecticut School of Medicine, Farmington, Connecticut.
Richard D. Berlin Center for Cell Analysis and Modeling, University of Connecticut School of Medicine, Farmington, Connecticut.
Biophys J. 2023 Sep 19;122(18):3722-3737. doi: 10.1016/j.bpj.2023.06.013. Epub 2023 Jun 23.
Fluorescence redistribution after photobleaching is a commonly used method to understand the dynamic behavior of molecules within cells. Analytic solutions have been developed for specific, well-defined models of dynamic behavior in idealized geometries, but these solutions are inaccurate in complex geometries or when complex binding and diffusion behaviors exist. We demonstrate the use of numerical reaction-diffusion simulations using the Virtual Cell software platform to model fluorescence redistribution after photobleaching experiments. Multiple simulations employing parameter scans and varying bleaching locations and sizes can help to bracket diffusion coefficients and kinetic rate constants in complex image-based geometries. This approach is applied to problems in membrane surface diffusion as well as diffusion and binding in cytosolic volumes in complex cell geometries. In addition, we model diffusion and binding within phase-separated biomolecular condensates (liquid droplets). These are modeled as spherical low-affinity binding domains that also define a high viscosity medium for exchange of the free fluorescently labeled ligand with the external cytosol.
光漂白后荧光再分布是一种常用于了解细胞内分子动态行为的方法。已经针对理想几何形状中特定的、明确的动态行为模型开发了解析解，但在复杂的几何形状或存在复杂的结合和扩散行为时，这些解并不准确。我们展示了使用虚拟细胞软件平台进行数值反应-扩散模拟来模拟光漂白后荧光再分布实验。多个模拟实验采用参数扫描和改变漂白位置和大小的方法，可以帮助确定复杂基于图像的几何形状中扩散系数和动力学速率常数的范围。这种方法应用于膜表面扩散以及复杂细胞几何形状中胞质体积内的扩散和结合问题。此外，我们还模拟了相分离生物分子凝聚物（液滴）内的扩散和结合。这些凝聚物被建模为具有低亲和力结合域的球形结构，这些结合域还定义了一个高粘度介质，用于与外部细胞质交换游离荧光标记配体。
