Small structures, as inside living cells, move on millisecond timescales, which is usually far beyond the imaging rate of superresolution fluorescence microscopes. In contrast, label-free imaging techniques providing high photon densities can operate at $>100\mathrm{Hz}$. For simple structures, an oblique, coherent illumination with a static laser beam increases image contrast and resolution considerably, whereas illumination of complex structures results in an image full of speckles. Remarkably, an artifact-free image is generated by subsequent oblique illumination of the structure from all azimuthal directions. This is the working principle of ROCS microscopy, which currently achieves 150 nm spatial and 10 ms temporal resolution without fluorophore bleaching, and is therefore highly beneficial for live-cell imaging. However, the complicated formation of ROCS images and image spectra during one sweep, i.e., the superposition of different speckle patterns is still unclear. Here, we investigate with experiments and computer simulations the influence of speckle-like interference patterns on the final image contrast and resolution, in darkfield mode and, by adding a reference wave, in brightfield mode. In close comparison to experimental results, we present a theoretical framework, which describes the ROCS image formation in real space and in $k$ space by identifying different spectral components. In addition, we vary the degree of coherence by a rotating diffuser and thereby demonstrate that maximal spatial coherence and maximal speckle interference from multiple scattering provide the best image contrast and resolution. We find that the cross correlations of elementary waves emitted in a distance of several micrometers to each other positively contribute to image formation and do not, as commonly believed, distort image formation. By understanding the composition of image speckles in time and space, future coherent microscopes should provide new insights into the high-speed world of living cells.

中文翻译：

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通过旋转相干散射（ROCS）显微镜中的图像形成实现出色的对比度和分辨率

微小的结构（如活细胞内部）以毫秒为单位移动，通常远远超出超分辨率荧光显微镜的成像速率。相反，提供高光子密度的无标签成像技术可以在$>100\mathrm{赫兹}$。对于简单的结构，使用静态激光束进行倾斜，相干的照明会显着提高图像的对比度和分辨率，而复杂结构的照明会导致图像上充满斑点。值得注意的是，通过随后从所有方位角方向对结构进行倾斜照明，可以生成无伪像的图像。这是ROCS显微镜的工作原理，目前可实现150 nm的空间分辨率和10 ms的时间分辨率而无需荧光团漂白，因此对活细胞成像非常有利。然而，在一次扫描期间ROCS图像和图像光谱的复杂形成，即不同斑点图案的叠加仍然不清楚。这里，我们通过实验和计算机仿真研究了暗场模式下斑点状干涉图样对最终图像对比度和分辨率的影响，并在明场模式下通过添加参考波进行了研究。与实验结果进行比较，我们提供了一个理论框架，该框架描述了在现实空间和太空中ROCS图像的形成。$ķ$通过识别不同的光谱成分来确定空间。另外，我们通过旋转漫射器改变相干程度，从而证明多重散射的最大空间相干性和最大斑点干扰提供了最佳的图像对比度和分辨率。我们发现以几微米的距离相互发射的基本波的互相关对成像形成有积极作用，并且，正如通常认为的那样，不会使成像变形。通过了解时间和空间中图像斑点的组成，未来的相干显微镜应提供对活细胞高速世界的新见解。