Key Laboratory of Optoelectronic Devices and Systems of Ministry of Education and Guangdong Province, College of Optoelectronic Engineering, Shenzhen University, Shenzhen, China.
College of Physics and Energy, Shenzhen University, Shenzhen, China.
J Biophotonics. 2018 Sep;11(9):e201800071. doi: 10.1002/jbio.201800071. Epub 2018 Jun 8.
With tunable excitation light, multiphoton microscopy is widely used for imaging biological structures at subcellular resolution. Axial chromatic dispersion, present in virtually every transmissive optical system including the multiphoton microscope, leads to focal (and the resultant image) plane separation. Here, we experimentally demonstrate a technique to measure the axial chromatic dispersion in a multiphoton microscope, using simultaneous 2-color third-harmonic generation imaging excited by a 2-color soliton source with tunable wavelength separation. Our technique is self-referenced, eliminating potential measurement error when 1-color tunable excitation light is used which necessitates reciprocating motion of the mechanical translation stage. Using this technique, we demonstrate measured axial chromatic dispersion with 2 different objective lenses in a multiphoton microscope. Further measurement in a biological sample also indicates that this axial chromatic dispersion, in combination with 2-color imaging, may open up opportunity for simultaneous imaging of 2 different axial planes.
利用可调谐激发光，多光子显微镜被广泛用于亚细胞分辨率的生物结构成像。轴向色散几乎存在于包括多光子显微镜在内的每一个透射光学系统中，导致焦点（和相应的图像）平面分离。在这里，我们通过使用具有可调波长分离的双色孤子源同时激发双色三次谐波成像，实验性地演示了一种测量多光子显微镜中轴向色散的技术。我们的技术是自参考的，消除了使用 1 种可调谐激发光时的潜在测量误差，这种误差需要机械平移台的往复运动。使用该技术，我们在多光子显微镜中展示了使用 2 个不同物镜测量的轴向色散。在生物样本中的进一步测量还表明，这种轴向色散与双色成像相结合，可能为同时成像 2 个不同轴向平面开辟机会。
