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首页> 外文会议>Conference on Molecular Probes for Biomedical Applications; 20080121-22; San Jose,CA(US) >Imaging elastic and collagen fibers with sulforhodamine B and second-harmonic generation
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Imaging elastic and collagen fibers with sulforhodamine B and second-harmonic generation

机译：用磺基罗丹明B和二次谐波产生成像弹性和胶原纤维

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Since the early nineties, multiphoton microscopy has become a powerful tool to investigate morphological and physiological parameters in vivo or on thick ex vivo sections. To stain structures of interest many dyes have been developed and two-photon properties (cross section, excitation and emission spectra) of existing ones have been characterized. Recently, our team has shown a new property of sulforhodamine B (SRB). This dye has the ability to bind specifically elastic fibers. The observation of elastin using its endofluorescence properties was already widely described but required long exposition delays up to 10s and the imaging depth was limited to approximately 50 μm. With a multiphoton microscope and SRB, it is possible to observe elastic fibers directly in the living animal or on thick tissue sections with a micrometric spatial resolution in less than one second per image with an imaging depth of ～ 200 μm. Moreover, with an appropriate set of filters, we can acquire simultaneously the SRB and the second harmonic generation (SHG) signals of collagen fibers. Here, we report various applications of this new staining method on different arterial rings. The layers of the arterial wall, as well as, the elastic lamellae are observed and are numbered. With the addition of a nuclear stain such as the Hoechst 33342, a more accurate morphological study of the arterial walls can be accomplished. Finally, an intravital observation of the saphenous artery morphology is presented.
机译：自九十年代初以来，多光子显微镜已成为研究体内或厚离体切片的形态和生理参数的强大工具。为了染色感兴趣的结构，已经开发了许多染料，并且已经表征了现有染料的双光子特性（截面，激发和发射光谱）。最近，我们的团队展示了磺基罗丹明B（SRB）的新特性。这种染料具有特异性结合弹性纤维的能力。利用弹性蛋白的内荧光特性观察弹性蛋白已被广泛描述，但需要长时间的曝光延迟，最长可达10s，并且成像深度限于约50μm。借助多光子显微镜和SRB，可以在活体动物或厚组织切片中直接观察到弹性纤维，并且具有微米级的空间分辨率，每幅图像的成像时间小于200微米，每幅图像的显示时间不到一秒钟。此外，使用一组适当的滤波器，我们可以同时获取胶原纤维的SRB和二次谐波生成（SHG）信号。在这里，我们报告了这种新的染色方法在不同动脉环上的各种应用。观察并编号动脉壁各层以及弹性薄片。通过添加诸如Hoechst 33342的核染色剂，可以完成对动脉壁的更精确的形态学研究。最后，提出了隐性大动脉形态的活体观察。

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