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首页> 美国卫生研究院文献>Scientific Reports >Flat-Field Super-Resolution Localization Microscopy with a Low-Cost Refractive Beam-Shaping Element
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Flat-Field Super-Resolution Localization Microscopy with a Low-Cost Refractive Beam-Shaping Element

机译：具有低成本折射光束整形元件的平场超分辨率定位显微镜

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摘要

Super-resolution single-molecule localization microscopy, often referred to as PALM/STORM, works by ensuring that fewer than one fluorophore in a diffraction-limited volume is emitting at any one time, allowing the observer to infer that the emitter is located at the center of the point-spread function. This requires careful control over the incident light intensity in order to control the rate at which fluorophores are switched on; if too many fluorophores are activated, their point-spread functions overlap, which impedes efficient localization. If too few are activated, the imaging time is impractically long. There is therefore considerable recent interest in constructing so-called ‘top-hat’ illumination profiles that provide a uniform illumination over the whole field of view. We present the use of a single commercially-available low-cost refractive beamshaping element that can be retrofitted to almost any existing microscope; the illumination profile created by this element demonstrates a marked improvement in the power efficiency of dSTORM microscopy, as well as a significant reduction in the propensity for reconstruction artifacts, compared to conventional Gaussian illumination.
机译：超分辨率单分子定位显微镜（通常称为PALM / STORM）的工作原理是，确保在任何一次发射的衍射受限体积中的荧光团少于一种，从而使观察者可以推断出发射器位于该位置。点扩展功能的中心。这就需要仔细控制入射光的强度，以控制荧光团的开启速率。如果激活了太多的荧光团，它们的点扩展功能就会重叠，这会阻碍有效的定位。如果激活的次数太少，则成像时间过长。因此，最近人们对构建所谓的“大礼帽”照明配置文件感兴趣，该配置文件可在整个视场上提供均匀的照明。我们介绍了使用单个商业上可获得的低成本折射光束整形元件，该元件可以改装到几乎任何现有的显微镜上。与传统的高斯照明相比，由该元件产生的照明曲线证明了dSTORM显微镜的功效显着改善，并且显着降低了重建伪影的可能性。

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