Pozzi Paolo, Smith Carlas, Carroll Elizabeth, Wilding Dean, Soloviev Oleg, Booth Martin, Vdovin Gleb, Verhaegen Michel
Opt Express. 2020 May 11;28(10):14222-14236. doi: 10.1364/OE.389974.
Inhomogeneities in the refractive index of a biological microscopy sample can introduce phase aberrations, severely impairing the quality of images. Adaptive optics can be employed to correct for phase aberrations and improve image quality. However, conventional adaptive optics can only correct a single phase aberration for the whole field of view (isoplanatic correction) while, due to the highly heterogeneous nature of biological tissues, the sample induced aberrations in microscopy often vary throughout the field of view (anisoplanatic aberration), limiting significantly the effectiveness of adaptive optics. This paper reports on a new approach for aberration correction in laser scanning confocal microscopy, in which a spatial light modulator is used to generate multiple excitation points in the sample to simultaneously scan different portions of the field of view with completely independent correction, achieving anisoplanatic compensation of sample induced aberrations, in a significantly shorter time compared to sequential isoplanatic correction of multiple image subregions. The method was tested in whole Drosophila brains and in larval Zebrafish, each showing a dramatic improvement in resolution and sharpness when compared to conventional isoplanatic adaptive optics.
生物显微镜样本折射率的不均匀性会引入相位像差，严重损害图像质量。自适应光学可用于校正相位像差并提高图像质量。然而，传统的自适应光学只能对整个视场校正单个相位像差（等晕校正），而由于生物组织的高度异质性，显微镜中样本引起的像差在整个视场中经常变化（非等晕像差），这显著限制了自适应光学的有效性。本文报道了一种激光扫描共聚焦显微镜像差校正的新方法，其中使用空间光调制器在样本中生成多个激发点，以完全独立校正的方式同时扫描视场的不同部分，实现样本引起像差的非等晕补偿，与多个图像子区域的顺序等晕校正相比，所需时间显著缩短。该方法在完整的果蝇大脑和斑马鱼幼虫中进行了测试，与传统的等晕自适应光学相比，每种情况在分辨率和清晰度上都有显著提高。
