Couture Charles-André, Bancelin Stéphane, Van der Kolk Jarno, Popov Konstantin, Rivard Maxime, Légaré Katherine, Martel Gabrielle, Richard Hélène, Brown Cameron, Laverty Sheila, Ramunno Lora, Légaré François
Institut National de la Recherche Scientifique, Centre Énergie Matériaux Télécommunications, Varennes, Quebec, Canada.
Department of Physics, University of Ottawa, Ottawa, Ontario, Canada.
Biophys J. 2015 Dec 15;109(12):2501-2510. doi: 10.1016/j.bpj.2015.10.040.
In this work, we report the implementation of interferometric second harmonic generation (SHG) microscopy with femtosecond pulses. As a proof of concept, we imaged the phase distribution of SHG signal from the complex collagen architecture of juvenile equine growth cartilage. The results are analyzed in respect to numerical simulations to extract the relative orientation of collagen fibrils within the tissue. Our results reveal large domains of constant phase together with regions of quasi-random phase, which are correlated to respectively high- and low-intensity regions in the standard SHG images. A comparison with polarization-resolved SHG highlights the crucial role of relative fibril polarity in determining the SHG signal intensity. Indeed, it appears that even a well-organized noncentrosymmetric structure emits low SHG signal intensity if it has no predominant local polarity. This work illustrates how the complex architecture of noncentrosymmetric scatterers at the nanoscale governs the coherent building of SHG signal within the focal volume and is a key advance toward a complete understanding of the structural origin of SHG signals from tissues.
在这项工作中，我们报告了飞秒脉冲干涉二次谐波产生（SHG）显微镜的实现。作为概念验证，我们对幼年马生长软骨复杂胶原结构的SHG信号相位分布进行了成像。结合数值模拟对结果进行分析，以提取组织内胶原纤维的相对取向。我们的结果揭示了恒定相位的大区域以及准随机相位区域，它们分别与标准SHG图像中的高强度和低强度区域相关。与偏振分辨SHG的比较突出了相对纤维极性在确定SHG信号强度方面的关键作用。实际上，似乎即使是组织良好的非中心对称结构，如果没有主要的局部极性，也会发出低SHG信号强度。这项工作说明了纳米级非中心对称散射体的复杂结构如何控制焦体积内SHG信号的相干构建，是朝着全面理解组织SHG信号的结构起源迈出的关键一步。
