Highly specific label-free molecular imaging with spectrally tailored excitation stimulated Raman scattering (STE-SRS) microscopy.
Highly specific label-free molecular imaging with spectrally tailored excitation stimulated Raman scattering (STE-SRS) microscopy.
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DOI：
10.1038/nphoton.2010.294
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发表时间：
2011
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影响因子：
35
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中科院分区：
文献类型：
--
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最近，受激拉曼散射（SRS）显微镜使无标记显微镜具有化学对比和高采集速度，达到视频速率成为可能。虽然SRS成像具有极高的灵敏度，但原始窄带实现的光谱特异性有限，使得区分具有重叠拉曼波段的化学物质变得困难。在这里，我们提出了一种高度特异性的成像方法，允许在存在干扰物质的情况下，基于其振动谱的定制多重激发，对特定化学物质进行映射。这是通过在高频（bbb1mhz）对宽带泵浦光束进行频谱调制来实现的，允许以高灵敏度检测窄带斯托克斯光束的受激拉曼增益信号。利用该方案，我们演示了在存在脂质的情况下胆固醇的定量，以及活秀丽隐杆线虫中蛋白质、硬脂酸和油酸的实时三维光谱成像。
Label-free microscopy with chemical contrast and high acquisition speed up to video-rate has recently been made possible by stimulated Raman scattering (SRS) microscopy. While SRS imaging offers superb sensitivity, the spectral specificity of the original narrowband implementation is limited, making distinguishing chemical species with overlapping Raman bands difficult. Here we present a highly specific imaging method that allows mapping of a particular chemical species in the presence of interfering species based on tailored multiplex excitation of its vibrational spectrum. This is done by spectral modulation of a broadband pump beam at a high-frequency (>1MHz), allowing detection of the stimulated Raman gain signal of the narrowband Stokes beam with high sensitivity. Using the scheme, we demonstrate quantification of cholesterol in the presence of lipids, and real-time three-dimensional spectral imaging of protein, stearic acid and oleic acid in live C.elegans.
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