Laboratory for Advanced Medical Photonics, Cancer Imaging Department, British Columbia Cancer Research Centre and Photomedicine Institute, University of British Columbia & Vancouver Coastal Health Research Institute, Vancouver, BC, Canada.
Skin Res Technol. 2009 Nov;15(4):418-26. doi: 10.1111/j.1600-0846.2009.00381.x.
BACKGROUND/PURPOSES: Understanding the two-photon excitation spectral characteristics and microscopic morphology of cutaneous collagen and elastic tissue components is important for applying multiphoton microscopy (MPM) in basic skin biology research and for clinical diagnosis.
We developed a system for two-photon excitation spectral measurements at various excitation wavelengths. The microscopic morphology was studied using a commercial multiphoton microscope.
We obtained two-photon excitation fluorescence (TPEF) excitation-emission matrices (EEM), for the first time, of purified collagen and elastin samples, as well as in situ collagen and elastic fibers within excised human dermis. The EEM of the dermis was found to be similar to that of elastin. The excitation spectra for second harmonic generation (SHG) from purified collagen and excised dermis were also studied and were found to have similar spectral shapes.
This study, using the EEM spectroscopic approach, confirmed a previous imaging study inference that in the dermis, TPEF predominantly originates from elastic fibers, while SHG originates solely from collagen fibers. The EEM data and SHG excitation spectra obtained in this study can be used to guide the selection of excitation wavelengths for MPM applications in basic skin biology research and for clinical diagnosis.
背景/目的：了解皮肤胶原和弹性组织成分的双光子激发光谱特性和微观形态对于将多光子显微镜（MPM）应用于基础皮肤生物学研究和临床诊断非常重要。
我们开发了一种用于在不同激发波长下进行双光子激发光谱测量的系统。使用商业多光子显微镜研究微观形态。
我们首次获得了纯化的胶原和弹性蛋白样品以及离体人真皮内胶原和弹性纤维的双光子激发荧光（TPEF）激发-发射矩阵（EEM）。真皮的 EEM 与弹性蛋白相似。还研究了从纯化胶原和离体真皮获得的二次谐波产生（SHG）的激发光谱，发现它们具有相似的光谱形状。
本研究使用 EEM 光谱方法证实了先前的成像研究推断，即在真皮中，TPEF 主要来源于弹性纤维，而 SHG 仅来源于胶原纤维。本研究获得的 EEM 数据和 SHG 激发光谱可用于指导 MPM 在基础皮肤生物学研究和临床诊断中的应用选择激发波长。
