Galiani Silvia, Harke Benjamin, Vicidomini Giuseppe, Lignani Gabriele, Benfenati Fabio, Diaspro Alberto, Bianchini Paolo
Nanophysics, Istituto Italiano di Tecnologia, Genoa, Italy.
Opt Express. 2012 Mar 26;20(7):7362-74. doi: 10.1364/OE.20.007362.
In stimulated emission depletion (STED) microscopy, the spatial resolution scales as the inverse square root of the STED beam's intensity. However, to fully exploit the maximum effective resolution achievable for a given STED beam's intensity, several experimental precautions have to be considered. We focus our attention on the temporal alignment between the excitation and STED pulses and the polarization state of the STED beam. We present a simple theoretical framework that help to explain their influence on the performance of a STED microscope and we validate the results by imaging calibration and biological samples with a custom made STED architecture based on a supercontinuum laser source. We also highlight the advantages of using time gating detection in terms of temporal alignment.
在受激发射损耗（STED）显微镜中，空间分辨率与STED光束强度的平方根成反比。然而，要充分利用给定STED光束强度下可实现的最大有效分辨率，必须考虑几个实验注意事项。我们将注意力集中在激发脉冲与STED脉冲之间的时间对准以及STED光束的偏振态上。我们提出了一个简单的理论框架，有助于解释它们对STED显微镜性能的影响，并通过使用基于超连续谱激光源的定制STED架构对校准图像和生物样品进行成像来验证结果。我们还强调了在时间对准方面使用时间选通检测的优势。
