Influence of optical properties on two-photon fluorescence imaging in turbid samples
Influence of optical properties on two-photon fluorescence imaging in turbid samples
复制标题
DOI：
10.1364/ao.39.001194
复制
发表时间：
2000-03-01
期刊：
影响因子：
1.9
通讯作者：
Tromberg, BJ
中科院分区：
文献类型：
--
作者：
Dunn, AK;Wallace, VP;Tromberg, BJ
建立了一个数值模型，模拟了组织光学特性、物镜数值孔径（N. A.）和仪器性能对混浊样品的双光子激发荧光成像。模型数据进行了比较与测量的荧光微球在一个组织状散射幻影。我们的结果表明，测量的双光子激发信号随焦深的增加呈指数衰减。总衰减常数是激发和发射波长处的吸收和散射参数的函数。双光子荧光的产生被证明是独立的散射各向异性，g，除了g > 0.95。这个疏证收集到的最大信号随深度而变化，尽管直到焦平面大于进入样品的两个散射平均自由程才能看到这种效果。总体而言，测量和模型结果表明，在双光子显微镜的分辨率是完全依赖于提供足够的弹道光子密度的焦点体积的能力。因此，我们表明，在双光子显微镜的横向分辨率是在很大程度上不受组织的光学性质的范围内通常遇到软组织，虽然最大成像深度是强烈依赖于吸收和散射系数，散射各向异性，和客观的NA。(C)2000年美国光学学会。
A numerical model was developed to simulate the effects of tissue optical properties, objective numerical aperture (N.A.), and instrument performance on two-photon-excited fluorescence imaging of turbid samples. Model data are compared with measurements of fluorescent microspheres in a tissuelike scattering phantom. Our results show that the measured two-photon-excited signal decays exponentially with increasing focal depth. The overall decay constant is a function of absorption and scattering parameters at both excitation and emission wavelengths. The generation of two-photon fluorescence is shown to be independent of the scattering anisotropy, g, except for g > 0.95. The N.A. for which the maximum signal is collected varies with depth, although this effect is not seen until the focal plane is greater than two scattering mean free paths into the sample. Overall, measurements and model results indicate that resolution in two-photon microscopy is dependent solely on the ability to deliver sufficient ballistic photon density to the focal volume. As a result we show that lateral resolution in two-photon microscopy is largely unaffected by tissue optical properties in the range typically encountered in soft tissues, although the maximum imaging depth is strongly dependent on absorption and scattering coefficients, scattering anisotropy, and objective N.A.. (C) 2000 Optical Society of America.