Comparison of objective lenses for multiphoton microscopy in turbid samples.
Comparison of objective lenses for multiphoton microscopy in turbid samples.
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DOI：
10.1364/boe.6.003113
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发表时间：
2015-08
影响因子：
3.4
通讯作者：
Avtar Singh;J. McMullen;E. A. Doris;W. Zipfel
中科院分区：
文献类型：
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作者：
Avtar Singh;J. McMullen;E. A. Doris;W. Zipfel
照明和探测光学系统的优化是高散射组织多光子成像的关键，而物镜是这两个路径的核心部件。为了更好地了解基本透镜参数(NA、放大率、场数)如何影响荧光采集和图像质量，使用了带有专门样品池的双探测器设置，将总激发测量与荧光采集分开。我们的数据证实了早期的发现，即低MAG透镜在收集散射光子方面可能更好，我们比较了一组常用的多光子物镜收集散射荧光的能力，为多光子成像系统的设计提供了指导。例如，我们在存在散射的情况下对表观荧光光束发散的测量显示了最小的光束展宽，这表明通常倡导的超大尺寸收集光学系统并不像之前认为的那样有利。这些实验还通过将我们的测量结果与所用物镜的各种设计参数相关联，为选择多光子成像的物镜提供了一个框架。
Optimization of illumination and detection optics is pivotal for multiphoton imaging in highly scattering tissue and the objective lens is the central component in both of these pathways. To better understand how basic lens parameters (NA, magnification, field number) affect fluorescence collection and image quality, a two-detector setup was used with a specialized sample cell to separate measurement of total excitation from epifluorescence collection. Our data corroborate earlier findings that low-mag lenses can be superior at collecting scattered photons, and we compare a set of commonly used multiphoton objective lenses in terms of their ability to collect scattered fluorescence, providing guidance for the design of multiphoton imaging systems. For example, our measurements of epi-fluorescence beam divergence in the presence of scattering reveal minimal beam broadening, indicating that often-advocated over-sized collection optics are not as advantageous as previously thought. These experiments also provide a framework for choosing objective lenses for multiphoton imaging by relating the results of our measurements to various design parameters of the objectives lenses used.