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Photo-Induced Cell Damage Analysis for Single- and Multifocus Coherent Anti-Stokes Raman Scattering Microscopy

机译：单次和多焦点相干抗斯托克斯拉曼散射显微镜的光诱导的电池损伤分析

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摘要

In this study, we investigated photo-induced damage to living cells during single- and multifocus excitations for coherent anti-Stokes Raman scattering (CARS) imaging. A near-infrared pulsed laser (709 nm) was used to induce cell damage. We compared the photo-induced cell damage in the single- and the multifocus excitation schemes with the condition to obtain the same CARS signal in the same frame rate. For the evaluation of cell viability, we employed 4′,6-diamidino-2-phenylindole (DAPI) fluorophores that predominantly stained the damaged cells. One- and two-photon fluorescence of DAPI fluorophores were, respectively, excited by an ultraviolet light source and the same near-infrared light source and were monitored to evaluate the cell viability during near-infrared pulsed laser irradiation. We found lower uptake of DAPI fluorophores into HeLa cells during the multifocus excitation compared with the single-focus excitation scheme in both the one- and the two-photon fluorescence examinations. This indicates a reduction of photo-induced cell damage in the multifocus excitation. Our findings suggested that the multifocus excitation scheme is expected to be suitable for CARS microscopy in terms of minimal invasiveness.
机译：在这项研究中，我们在单一和多焦点激发过程中调查了对活细胞的皮肤造成的损伤，用于连贯的反斯托克斯拉曼散射（汽车）成像。近红外脉冲激光器（709nm）用于诱导细胞损伤。我们将光诱导的电池损坏与多焦点激励方案中的光电诱导的电池损坏进行了比较，条件以相同的帧速率获得相同的汽车信号。为了评价细胞活力，我们使用的4'，6-二脒基-2-苯基吲哚（DAPI）荧光团，其主要染色受损细胞。 DAPI荧光团的单光子和两光子荧光分别由紫外光源和相同的近红外光源激发，并监测在近红外脉冲激光照射期间评估细胞活力。与双光子荧光检查中的单聚焦激发方案相比，我们发现将DAPI荧光团的吸收降低到HeLa细胞中。这表明多焦点激励中的光诱导的电池损伤降低。我们的研究结果表明，多焦励磁方案预计在最小的侵袭性方面适用于汽车显微镜。

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