基于荧光显微镜的各种组织折射率测定方法

IF 1.1 Q4 OPTICS Optics continuum Pub Date : 2023-06-30 DOI:10.1364/optcon.492897
Xiaoming Fan, Lele Tao, Xiaoyu Zhou, Xiao He, Yu Zhang, Haixin Huang, Jiale Yang, Simei Wang, Zhihui Ma, Thomas Gensch, Ruimin Huang
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引用次数: 0

摘要

折射率是表征光与试样相互作用的重要光学常数。试样与浸没介质之间的折射率差异导致了成像像差,导致光学显微镜直接测量试样厚度不准确。然而,这种像差校正仍然需要试样和浸没介质折射率的准确信息。在此,我们提出了一种成像策略来估计一个未知标本的折射率。建立了简化的衍射模型,得到了轴向比例因子与折射率之间的关系。然后用常规荧光显微镜测量小鼠肌肉和肿瘤移植标本的实际轴向缩放系数。根据轴向比例因子与折射率的理论关系图,确定了组织标本的折射率。例如,我们获得正常肌肉组织的平均折射率(n)值为1.36,肿瘤异种移植物的平均折射率(n)值为1.41。我们证明了这种基于衍射模型的估计方法是一种替代现有技术的方法,提高了组织标本折射率的精确测量。这种折射率估计方法的仪器要求简单,样品制备容易,可以提高组织标本的像差较小的图像质量。
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Method for the refractive index of various tissues based on fluorescence microscopy
Refractive index is an important optical constant that characterizes the interaction between light and specimen. A difference in refractive index between specimen and immersion medium introduces the imaging aberration and leads to a problem that the direct thickness measurement of a specimen by optical microscopy is not accurate. However, this aberration correction still requires the exact information of the refractive index of specimen and immersion medium. Herein, we propose an imaging strategy to estimate the refractive index for an unknown specimen. A simplified diffraction model is generated to obtain the relationship between axial scaling factor and refractive index. Then regular fluorescence microscopy is performed to measure the actual axial scaling factors of specimens from mouse muscle and tumor xenograft. Referring to our theoretical plot of axial scaling factor versus refractive index, the refractive index of tissue specimen is determined. For example, we obtain a mean refractive index (n) value of 1.36 for normal muscle tissues, and 1.41 for tumor xenografts. We demonstrate that this diffraction model-based estimation method is an alternative to the current techniques, improving the accurate measurement for refractive index of tissue specimen. The simple instrument requirement with an easy specimen preparation for this estimation method of refractive index may increase the image quality on tissue specimens with less aberration.
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