Department of Mathematics, University of California, Santa Barbara, California, USA.
Biophys J. 2012 Mar 7;102(5):1144-53. doi: 10.1016/j.bpj.2012.01.045. Epub 2012 Mar 6.
The mechanical properties of biopolymers can be determined from a statistical analysis of the ensemble of shapes they exhibit when subjected to thermal forces. In practice, extracting information from fluorescence microscopy images can be challenging due to low signal/noise ratios and other artifacts. To address these issues, we develop a suite of tools for image processing and spectral data analysis that is based on a biopolymer contour representation expressed in a spectral basis of orthogonal polynomials. We determine biopolymer shape and stiffness using global fitting routines that optimize a utility function measuring the amount of fluorescence intensity overlapped by such contours. This approach allows for filtering of high-frequency noise and interpolation over sporadic gaps in fluorescence. We use benchmarking to demonstrate the validity of our methods, by analyzing an ensemble of simulated images generated using a simulated biopolymer with known stiffness and subjected to various types of image noise. We then use these methods to determine the persistence lengths of taxol-stabilized microtubules. We find that single microtubules are well described by the wormlike chain polymer model, and that ensembles of chemically identical microtubules show significant heterogeneity in bending stiffness, which cannot be attributed to sampling or fitting errors. We expect these approaches to be useful in the study of biopolymer mechanics and the effects of associated regulatory molecules.
生物聚合物的力学性能可以通过对它们在热作用力下表现出的形状集合进行统计分析来确定。在实践中，由于低信噪比和其他伪影，从荧光显微镜图像中提取信息可能具有挑战性。为了解决这些问题，我们开发了一套基于生物聚合物轮廓在正交多项式光谱基上的表示的图像处理和光谱数据分析工具。我们使用全局拟合例程来确定生物聚合物的形状和刚度，该例程优化了一个效用函数，该函数测量荧光强度被这些轮廓重叠的量。这种方法允许过滤高频噪声并在荧光的零星间隙上进行插值。我们使用基准测试来通过分析使用具有已知刚度的模拟生物聚合物生成的一组模拟图像来验证我们的方法，并对各种类型的图像噪声进行了分析。然后，我们使用这些方法来确定紫杉醇稳定的微管的持久长度。我们发现，单个微管可以很好地用蠕虫状链聚合物模型来描述，并且化学上相同的微管的集合在弯曲刚度上表现出显著的异质性，这不能归因于采样或拟合误差。我们期望这些方法在生物聚合物力学和相关调节分子的影响的研究中是有用的。
