A multiphoton microscope platform for imaging the mouse eye
A multiphoton microscope platform for imaging the mouse eye
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发表时间：
2012-07
期刊：
影响因子：
2.2
通讯作者：
O. Masihzadeh;Tim C. Lei;D. Ammar;M. Kahook;E. Gibson
中科院分区：
文献类型：
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作者：
O. Masihzadeh;Tim C. Lei;D. Ammar;M. Kahook;E. Gibson
目的证明多光子显微镜在不需要眼球摘除的情况下获得完整小鼠眼前房的全三维高分辨率图像的能力。方法构建并优化定制的多光子显微镜用于深部组织成像。同时进行双光子自发荧光（2PAF）和二次谐波产生（SHG）成像。设计了鼠标保持器和立体定位平台，以进入眼睛的不同部位进行成像。在成像过程中使用了用于保持眼睛湿润的储液器。结果在不需要眼球摘除的情况下，获得了小鼠眼前房深处的非侵入性多光子图像。2PAF和SHG信号显示虹膜、角膜上皮和内皮、小梁网区和结膜。解剖结构的识别是通过天然组织的内在特性实现的，而不需要任何外源性标记。深入眼睛600微米的图像被清晰地展示出来。使用定制软件进行整个前房的全三维图像重建和分析。结论多光子成像技术在眼科研究中具有广阔的应用前景。我们已经证明了在其自然状态下对小鼠眼睛的整个前房进行成像的能力。这些结果为今后的眼部体内研究提供了基础。
Purpose To demonstrate the ability of multiphoton microscopy to obtain full three-dimensional high-resolution images of the intact mouse eye anterior chamber without need for enucleation. Methods A custom multiphoton microscope was constructed and optimized for deep tissue imaging. Simultaneous two-photon autofluorescence (2PAF) and second harmonic generation (SHG) imaging were performed. A mouse holder and stereotaxic platform were designed to access different parts of the eye for imaging. A reservoir for keeping the eye moist was used during imaging sessions. Results Non-invasive multiphoton images deep inside the anterior chamber of the mouse eye were obtained without the need for enucleation. The iris, corneal epithelium and endothelium, trabecular meshwork region and conjunctiva were visualized by the 2PAF and SHG signals. Identification of the anatomy was achieved by the intrinsic properties of the native tissue without any exogenous labeling. Images as deep as 600 microns into the eye were clearly demonstrated. Full three-dimensional image reconstructions of the entire anterior chamber were performed and analyzed using custom software. Conclusions Multiphoton imaging is a highly promising tool for ophthalmic research. We have demonstrated the ability to image the entire anterior chamber of the mouse eye in its native state. These results provide a foundation for future in vivo studies of the eye.