Appl Opt. 2020 Mar 20;59(9):2914-2923. doi: 10.1364/AO.388265.
A method to measure the dimensions of objects below the optical diffraction limit using diffraction analysis of out-of-focus bright-field images is presented. The method relies on the comparison of the diffraction patterns of an object of unknown size to those of calibration objects of known size. Correlative scanning electron microscope measurements are used to demonstrate the applicability of this method to measure 100 nm microbeads as well as objects with a geometry different from the calibration objects. This technique is important in the context of tethered particle experiments, in which bio-filaments are bound between a substrate and a microbead. This procedure is applied to obtain the diameters of axonal extensions or neurites that are mechanically created in samples of rat hippocampal neurons. The dependence of neurite geometry on mechanical pull speed is investigated, and the diameter is found to be rate independent.
提出了一种利用离焦明场图像的衍射分析来测量小于光衍射极限的物体尺寸的方法。该方法依赖于将未知尺寸的物体的衍射图案与已知尺寸的校准物体的衍射图案进行比较。相关的扫描电子显微镜测量用于证明该方法可用于测量 100nm 微珠以及与校准物体的几何形状不同的物体的适用性。在连接粒子实验的背景下，这种技术很重要，其中生物丝被束缚在基板和微珠之间。该程序用于获得在大鼠海马神经元样本中机械产生的轴突延伸或神经突的直径。研究了神经突几何形状对机械拉伸速度的依赖性，并发现直径与速率无关。
