A series of flexible design adaptations to the Nikon E-C1 and E-C2 confocal microscope systems for UV, multiphoton and FLIM imaging
A series of flexible design adaptations to the Nikon E-C1 and E-C2 confocal microscope systems for UV, multiphoton and FLIM imaging
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DOI：
10.1111/jmi.12218
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发表时间：
2015-04-01
影响因子：
2
通讯作者：
Parker, Anthony W.
中科院分区：
文献类型：
--
作者：
Botchway, Stanley W.;Scherer, Kathrin M.;Parker, Anthony W.
多光子显微镜广泛应用于生命科学中，使用低分子量和高分子量标记的外源荧光，其激发和发射光谱在可见光和近红外区域。对于在紫外区域中具有激发光谱的内在和外在荧光团的成像，用单色或双色激光的多光子激发避免了对紫外透射激发光学器件的需要，并且在样品中的光学穿透和降低的光毒性方面具有优势。在典型的倒置共焦显微镜中，激发和检测300 nm左右及以下的紫外发射更困难，并且需要使用昂贵的石英光学器件，包括物镜。在本技术说明中，我们描述了一个商业共聚焦显微镜（尼康，日本E-C1或E-C2）的适应与钛蓝宝石和OPO激光源的多功能使用，并增加了第二个检测通道，使紫外荧光检测和增加检测灵敏度在一个典型的荧光寿命成像显微镜实验。使用此设置的一些实验的结果说明了所产生的能力。
Multiphoton microscopy is widely employed in the life sciences using extrinsic fluorescence of low- and high-molecular weight labels with excitation and emission spectra in the visible and near infrared regions. For imaging of intrinsic and extrinsic fluorophores with excitation spectra in the ultraviolet region, multiphoton excitation with one- or two-colour lasers avoids the need for ultraviolet-transmitting excitation optics and has advantages in terms of optical penetration in the sample and reduced phototoxicity. Excitation and detection of ultraviolet emission around 300 nm and below in a typical inverted confocal microscope is more difficult and requires the use of expensive quartz optics including the objective. In this technical note we describe the adaptation of a commercial confocal microscope (Nikon, Japan E-C1 or E-C2) for versatile use with Ti-sapphire and OPO laser sources and the addition of a second detection channel that enables detection of ultraviolet fluorescence and increases detection sensitivity in a typical fluorescence lifetime imaging microscopy experiment. Results from some experiments with this setup illustrate the resulting capabilities.