Membrane imaging by second-harmonic generation microscopy
Membrane imaging by second-harmonic generation microscopy
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DOI：
10.1364/josab.17.001685
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发表时间：
2000-10-01
影响因子：
1.9
通讯作者：
Mertz, J
中科院分区：
文献类型：
--
作者：
Moreaux, L;Sandre, O;Mertz, J
我们提出了一个详细的分析与苯乙烯基染料标记的生物膜的二次谐波辐射的产生。特别是，我们认为高数值孔径限制适合高分辨率显微镜，其中激发光束是紧紧集中在从一侧到膜表面上。在此限制下，有助于二次谐波产生（SHG）的有效表面积仅取决于光束焦点的紧密度，并且SHG辐射通过相位匹配被限制在两个明确定义的离轴波瓣中。我们推导出表达式的SHG辐射功率，角分布，和偏振依赖的情况下，理想或非理想的分子排列在膜和单轴性的分子超极化。我们定义了一个类似于双光子激发荧光（TPEF）中使用的SHG横截面，以允许直接比较两种成像方式。最后，我们证实了我们的结果与实验的基础上激发的苯乙烯基染料在巨大的单层囊泡与锁模钛：蓝宝石激光。(C)2000年美国光学学会。
We present a detailed analysis of the generation of second-harmonic radiation from biological membranes labeled with a styryl dye. In particular, we consider the high-numerical-aperture limit appropriate to highresolution microscopy in which an excitation beam is tightly focused from the side onto a membrane surface. In this limit the active surface area that contributes to second-harmonic generation (SHG) depends only on the tightness of the beam focus and the SHG radiation is confined by phase matching into two well-defined off-axis lobes. We derive expressions for the SHG radiation power, angular distribution, and polarization dependence in the cases of ideal or nonideal molecular alignment in the membrane and uniaxiality of the molecular hyperpolarizability. We define an SHG cross section similar to that used in two-photon-excited fluorescence (TPEF) to permit direct comparison of the two imaging modalities. Finally, we corroborate our results with experiments based on the excitation of a styryl dye in giant unilamellar vesicles with a mode-locked Ti:sapphire laser.(C) 2000 Optical Society of America.