Experimental test of an analytical model of aberration in an oil-immersion objective lens used in three-dimensional light microscopy.
Experimental test of an analytical model of aberration in an oil-immersion objective lens used in three-dimensional light microscopy.
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DOI：
10.1364/josaa.9.000154
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发表时间：
1991-10
期刊：
影响因子：
--
通讯作者：
S. Gibson;Frederick Lanni
中科院分区：
文献类型：
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作者：
S. Gibson;Frederick Lanni
油浸显微镜物镜经过设计和优化，用于研究薄的二维物体切片，这些物体切片安装在盖玻片正下方的介质中，介质的折射率与浸油相匹配。实验以及几何和物理光学理论都证明，当显微镜未使用正确的盖玻片或浸油时，当探测器未位于图像空间的最佳平面时，或者当物体不满足特定条件时，像差会降低图像的对比度和分辨率。在生物学中，当使用油浸物镜研究厚样本（例如活细胞和组织）时，会引入最严重的像差，其折射率与浸油的折射率显着不同。我们提出了受这种像差影响的荧光光学显微镜的三维成像特性的模型，并将该模型预测的成像特性与实验测量的成像特性进行了比较。该模型可用于理解和补偿在非设计光学条件下引入显微镜系统的像差，从而可以优化共焦激光扫描显微镜和光学串行切片显微镜。
Oil-immersion microscope objective lenses have been designed and optimized for the study of thin, two-dimensional object sections that are mounted immediately below the coverslip in a medium that is index matched to the immersion oil. It has been demonstrated both experimentally and through geometrical- and physical-optics theory that, when the microscope is not used with the correct coverslip or immersion oil, when the detector is not located at the optimal plane in image space, or when the object does not satisfy specific conditions, aberration will degrade both the contrast and the resolution of the image. In biology the most severe aberration is introduced when an oil-immersion objective lens is used to study thick specimens, such as living cells and tissues, whose refractive indices are significantly different from that of the immersion oil. We present a model of the three-dimensional imaging properties of a fluorescence light microscope subject to such aberration and compare the imaging properties predicted by the model with those measured experimentally. The model can be used to understand and compensate for aberration introduced to a microscope system under nondesign optical conditions so that both confocal laser scanning microscopy and optical serial sectioning microscopy can be optimized.