Multimodal spectral focusing CARS and SFG microscopy with a tailored coherent continuum from a microstructured fiber
Multimodal spectral focusing CARS and SFG microscopy with a tailored coherent continuum from a microstructured fiber
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DOI：
10.1007/s00340-020-7406-6
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发表时间：
2020-04
期刊：
影响因子：
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通讯作者：
K. P. Herdzik;K. Bourdakos;P. Johnson;Adam Lister;Aleksandra P. Pitera;Chun-yu Guo;P. Horák;
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文献类型：
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作者：
K. P. Herdzik;K. Bourdakos;P. Johnson;Adam Lister;Aleksandra P. Pitera;Chun-yu Guo;P. Horák;
我们报告了一种技术新颖的生物成像显微镜系统，该系统基于100 fs钛：蓝宝石（Ti:Sa）激光泵浦相干连续体，来自定制的9厘米长全正常色散（ANDi）光纤，实现了(a)光谱聚焦相干反斯托克斯拉曼散射（SF-CARS）（跨越900-3200 cm−1）和(b)和频率生成（SFG）的并发图像对比。这两种模式都有效地激发了与生物样品兼容的显微镜焦点的功率水平。此外，使用连续体，图像被记录在反向散射（epi-detection）几何中，而不需要昂贵的、计算机控制的空间光调制器（SLM），清楚地显示了所获得的强信号水平。与单独使用任何一种技术成像相比，多种模式的图像对比提供了更多的化学和结构信息。数值模拟支持了SC产生的最佳光纤长度和SF-CARS中实现高光谱分辨率的这些进展，通过在泵浦和Stokes脉冲之间进行仔细的群延迟色散匹配，只需在Stokes光束中插入廉价的短玻璃块序列。我们展示了通过多种方式同时记录的小鼠组织的生物图像：CARS，双光子自动荧光（TPaF）和二次谐波/和频率生成（SHG/SFG）。总体而言，我们的方法在背散射（epi-）检测配置中提供了最佳性能，适用于厚样品，降低了复杂性和成本。将这种简单的光纤附加到已经广泛用于TPF显微镜的激光器上，可以扩展大量现有显微镜实验室的能力。
We report a technologically novel microscopy system for bioimaging based on a 100 fs titanium:sapphire (Ti:Sa) laser pumped coherent continuum from a tailored, 9-cm long, all normal dispersion (ANDi) fiber, enabling concurrent image contrast with (a) spectral focusing coherent anti-Stokes Raman scattering (SF-CARS) (spanning 900–3200 cm−1) and (b) sum frequency generation (SFG). Both modalities were efficiently excited with power levels at the microscope focus compatible with biological samples. Moreover, using the continuum, images were recorded in the back-scattering (epi-detection) geometry, without the necessity for an expensive, computer-controlled, spatial light modulator (SLM), clearly demonstrating the strong signal levels achieved. Image contrast from the multiple modalities provided greater chemical and structural insights than imaging with any single technique in isolation. Numerical simulations supported these developments in regard to both the optimum fiber length for SC generation and the achievement of high spectral resolution in SF-CARS via careful group delay dispersion matching across the pump and Stokes pulses using just an inexpensive sequence of short glass blocks inserted into the Stokes beam. We show bio-images of mouse tissue recorded concurrently via label/stain-free contrast from multiple modalities: CARS, two-photon auto-fluorescence (TPaF) and second harmonic/sum frequency generation (SHG/SFG). Overall, our approach delivers optimum performance in back-scattered (epi-) detection configuration, suited for thick samples, at reduced complexity and cost. The addition of this simple fiber add-on to lasers already widely used for TPF microscopy can thus extend the capabilities of a significant number of existing microscopy laboratories.