心肌膜电位的光学标测技术及实验研究

基于电压敏感染料膜电位光学标测的原理,建立了一套光标测系统,主要包括LED激发光源、以及由滤光片、CCD、A/D采集卡及计算机构成的荧光信号采集、处理部分.目前,整套系统已可自动完成图像采集、实时显示,以及部分后处理等工作,已获得兔心室组织细胞膜动作电位.另外,在建立系统的同时,研究了光学标测实验方法,对离体兔心脏急性缺血的主要电生理特征进行了标测实验,并结合仿真研究的结论说明了缺血后有效不应期时空异质性的存在以及组织中电兴奋传导速度的减慢是缺血导致折返性心律失常的主要机制.     

电压敏感染料di-4-ANEPPS在灌注液中的光谱特性

研究了电压敏感染料di-4-ANEPPS在溶液中的吸收光谱和荧光光谱特性.结果表明,染料的吸收峰在480 nm附近,加入钙阻断剂BDM不会影响染料的吸收峰位.染料在去离子水和新鲜台氏液中的荧光发射光谱峰位近似,而在灌注后的台氏液中染料的荧光峰位有蓝移现象.这些研究结果为心脏光学标测系统的设计提供了理论依据.     

电压敏感染料di-4-ANEPPS的光谱研究

实验研究了电压敏感染料di-4-ANEPPS在家兔心肌组织中的吸收光谱和荧光光谱特性。结果表明,含染料组织的光吸收普遍大于对照组,在450～550 nm波段吸收谱差异更明显;染料在心室组织中的最大吸收峰为(479.75±0.44) nm。通过测量含染料心脏不同部位的荧光光谱,首次发现心室组织、心房组织和主动脉的最大荧光峰位有一定差异,其相对荧光强度则与染料的分布浓度有关。根据三维和二维荧光光谱分别确定了含染料心房和心室组织的最佳荧光激发波长和荧光测定波长。利用心房和心室组织的静息电位差,在不同波长激发光下测量了染料的荧光光谱移动,确定了光标测量实验的最佳激发光和相应荧光检测波长范围。这些研究结果为心脏光学标测系统的设计提供了理论依据。     

罗丹明101染料的光谱特性研究

观测了罗丹明101染料在甲醇和酸性甲醇溶液巾的稳态吸收、稳态荧光和时间分辨荧光光谱,得到了吸收与荧光光谱的特征信息以及荧光寿命;通过拉曼光谱、红外光谱和密度泛函理论计算,对罗丹明101染料分子的振动模式进行了指认.研究结果全面系统地表征了罗丹明101染料的光谱特征以及分子结构和振动信息,为罗丹明101染料在染料敏化太阳能电池和生物荧光标记等方面的应用研究提供了依据.     

基于荧光强度比值法可用于现场测量的低成本聚合物光纤温度传感器

基于荧光强度比值法,设计了一种使用两种荧光染料的光纤温度传感器.实验中,罗丹明B和罗丹明110分别为对温度敏感和对温度不敏感的荧光传感物质,利用聚合物光纤来传导激发光及接收荧光.由于两种染料的荧光谱峰相距60 nm,因此容易将二者对应的荧光谱分开.通过确定能代表两种染料的最优荧光光谱范围,获得具有良好线性度的温度-荧光强度标定曲线.实验研究了不同浓度的荧光染料对标定曲线的影响,当染料浓度为0.3 g/L时,可获得0.28℃的最小均方误差及0.0128/℃的灵敏度.此外,该传感器还具备一定的抗光源扰动和抗荧光染料漂白的能力.     

基于荧光强度比值法可用于现场测量的低成本聚合物光纤温度传感器（英文）

基于荧光强度比值法,设计了一种使用两种荧光染料的光纤温度传感器.实验中,罗丹明B和罗丹明110分别为对温度敏感和对温度不敏感的荧光传感物质,利用聚合物光纤来传导激发光及接收荧光.由于两种染料的荧光谱峰相距60nm,因此容易将二者对应的荧光谱分开.通过确定能代表两种染料的最优荧光光谱范围,获得具有良好线性度的温度-荧光强度标定曲线.实验研究了不同浓度的荧光染料对标定曲线的影响,当染料浓度为0.3g/L时,可获得0.28℃的最小均方误差及0.0128/℃的灵敏度.此外,该传感器还具备一定的抗光源扰动和抗荧光染料漂白的能力.     

基于矩阵分析的STR荧光解谱技术

用多种荧光标记物进行STR检测时,由于荧光光谱的谱带展宽特性,各个荧光光谱之间有重叠部分,如何实现将相互重叠的各个波长上的能量有效利用起来,对提高荧光利用效率至关重要。本文给出了一种基于矩阵分析的数据处理方法,该方法在进行荧光探测前首先要对荧光谱进行光谱校正,即要得到每种所用标记染料受激发射荧光的光谱分布,根据光谱分布建立染料组合荧光信号矩阵,然后对矩阵进行归一,利用归一后的矩阵对所探测的荧光谱进行解谱,从而得到所期望的荧光谱图。理论分析和实验结果表明,该方法可以有效的利用各个波长的荧光光谱能量,并实现对不同荧光重叠谱的有效光谱解谱。     

利用电压敏感荧光染料测定贝体中的麻痹性贝毒

麻痹性贝毒(paralytic shellfish poisoning, PSP)是分布最广、危害最大的赤潮毒素,可阻断钠离子通道从而抑制动作电位的形成导致动物体中毒。以膀胱癌移行细胞T24为实验材料,采用电压敏感荧光染料bis-oxonol,根据PSP毒素GTX2,3对藜芦定诱导细胞去极化的抑制作用,建立了一种麻痹性贝毒电压敏感荧光染料检测法。实验结果显示,在2～100 ng·mL-1范围内,PSP毒素GTX2,3可显著改变培养体系的荧光强度,GTX浓度与荧光强度之间存在很好的线性关系。市售贝类PSP毒素检测结果表明,此方法测定结果与小鼠法基本一致,但灵敏度更高,说明根据GTX浓度与荧光强度之间的线性关系可实现对样品中PSP毒素的快速检测。利用电压敏感荧光染料测定贝体中的麻痹性贝毒是一种非常有潜力的贝毒快速筛查方法。     

具有三种颜色的白癜风皮肤的荧光光谱

将荧光光谱方法应用于具有三种颜色的白癜风皮肤特性的研究,在对荧光测量以及皮肤组织病理检查的基础上,建立了三色白癜风皮肤的光学模型,分析了皮肤黑色素的含量与分布,并进行了蒙特卡罗计算,其结果与实测结果基本一致。研究表明:具有三种颜色的白癜风皮肤可作为一个较好的皮肤黑色素含量与分布的模型;荧光光谱法可以探测到皮肤较深层(真皮层)的黑色素分布或其他组织光学特性;所建立的白癜风皮肤的光学模型及建模方法,所采用的光谱比率法等可以在皮肤光学的其他研究中应用。     

阴阳离子菁染料J-聚集体的荧光光谱研究

采用荧光光谱技术,研究了两种阴阳离子菁染料及对应的阴离子和阳离子菁染料在溴碘化银立方体颗粒上形成的J-聚集体的荧光光谱特性,并与其在溴碘化银立方体上的光谱增感率进行了实验对比。结果表明：阴阳离子菁染料的荧光光谱峰值明显低于阴离子、阳离子菁染料,说明其吸收光子后,大部分电子转移到溴碘化银表面。因此具有较高的光谱增感效率。     

Direct measurement of the voltage sensitivity of second-harmonic generation from a membrane dye in patch-clamped cells

We report what is to our knowledge the first optical imaging of voltage-clamped cells by second-harmonic generation. For the membrane-staining styryl dye di-4-ANEPPS, we determined the sensitivity of second-harmonic generation to be 18%/100 mV at an excitation wavelength of 850 ns. This sensitivity is significantly better than the optimal 10%/100 mV under fluorescence and further establishes the importance of second-harmonic generation for the functional imaging of membrane potential in living cells.     

An investigation into the role of the optical detection set-up in the recording of cardiac optical mapping signals: A Monte Carlo simulation study

Photon scattering is known to distort the fluorescence signals recorded from optically mapped cardiac tissue. However, the contribution of the parameters which define the optical detection set-up has not been assessed. In this study, Monte Carlo (MC) simulations of photon scattering within ventricular tissue are combined with a detailed model of a tandem-lens optical detection apparatus to characterise (i) the spatial origin upon emission of photons recorded in voltage-sensitive fluorescence measurements of cardiac electrical activity (using the fluorescent dye di-4-ANEPPS) and how this affects signal distortion, and (ii) the role the detector characteristics could play in modulating signal distortion during uniform illumination and photon emission from tissue depth. Results show that, for the particular excitation/emission wavelengths considered (488 nm and 669 nm, respectively), the dimensions of the scattering volume during uniform illumination extend around 3 times further in the surface recording plane than in depth. As a result, fluorescence recordings during electrical propagation are more distorted when transmembrane potential levels differ predominantly in the surface plane than in depth. In addition, MC simulation results show that the spatial accuracy of the fluorescence signal is significantly limited due to photon scattering, with only a small fraction of the recorded signal intensity originating from tissue beneath the pixel (approximately 11% for a 0.25×0.25 mm pixel). Increasing pixel size increases this fraction, however, it also results in an increase in the scattering volume dimensions, thus reducing the spatial resolution of the optical system, and increasing signal distortion. MC simulations also demonstrate that photon scattering in cardiac tissue limits the ability of optical detection system tuning in accurately locating fluorescent emission from depth. Specifically, our results prove that the focal plane depth that yields maximum signal intensity provides an underestimation of the emission depth. In conclusion, our study demonstrates the potential of MC simulations of photon scattering in guiding the design of optical mapping set-ups to optimise performance under diverse experimental conditions.     

Hybrid modeling of electrical and optical behavior in the heart

Optical mapping of transmembrane potential using voltage-sensitive dyes has revolutionized cardiac electrophysiology by enabling the visualization of electrical excitation waves in the heart. However, the interpretation of the optical mapping data is complicated by the fact that the optical signal arises not just from the surface, but also from some depth into the heart wall. Here, we review modeling efforts, in which the diffusion of photons is incorporated into the computer simulations of cardiac electrical activity (“hybrid” modeling), with the goal of improving our understanding of optical signals. We discuss the major accomplishments of hybrid modeling which include: (i) the explanation of the optical action potential upstroke morphology and prediction of its dependence on the subsurface wave front angle, (ii) the unexpectedly low magnitudes of optically recorded surface potentials during electrical shocks, and (iii) the “depolarization” of the core of the spiral wave and odd dual-humped optical action potentials during reentrant activation. We critically examine current optical mapping techniques and controversies in our understanding of electroporation during defibrillation. Finally, we provide a brief overview of recent theoretical studies aimed at extending optical mapping techniques for imaging intramural excitation to include transillumination imaging of scroll wave filaments and depth-resolved optical tomographic methods.     

Single-beam Z-scan measurement of the third-order optical nonlinearities of triarylmethane dyes

Solid-state dye-doped polymers are an attractive alternative to the conventional liquid-dye solutions. The search for new materials which have potential application in optoelectronic devices has lead us to probe the organic dyes. The solid state being a better medium when compared to a liquid medium has lead to the incorporation of dyes in the polymer matrix. The study of nonlinear characteristics of dyes in polymeric media is essential for developing such potential application devices. In this paper, the third-order nonlinear optical properties of three dyes from the Triarylmethane family were measured in 1-Butanol and in dye-doped polymer films by the Z-scan technique using a cw diode-pumped Nd:YAG laser at 532 nm. The Z-scan technique has also been used to present the observation of a nonlinear refractive index resulting from the photochromism of one of the dyes. These materials exhibit a large negative optical nonlinearity resulting due to the thermal effect. The relative contributions from the nonlinear absorption (NLA) and nonlinear refraction (NLR) are dependent on the chemical structure and linear absorption of the dyes. The dyes exhibited a nonlinear refractive coefficient n ₂, a nonlinear absorption coefficient β, and susceptibility x⁽³⁾ on the order of 10^?8 cm²/W, 10^?4 cm/W, and 10^?6 esu, respectively, in both liquid and solid media. The results show that these dyes have potential applications in nonlinear optics.     

空间遥感测绘光学系统研究综述

将遥感技术应用到测绘当中是现代地质测绘技术的发展趋势,随着光学载荷分辨率的不断提高,遥感测绘已经成为社会发展和国民经济发展的重要保障。光学载荷决定了测绘空间遥感器的分辨率、测绘精度、卫星平台体积与重量,是遥感器的核心部分。本文对高成像质量透射光学系统、同轴三反光学系统、离轴三反系统等常用的空间遥感测绘光学系统的结构形式和光学性能分别进行了介绍,并对处于研发阶段的新型空间反射光学系统的结构形式和光学性能进行了展望。分析认为,根据不同的应用环境和技术指标,合理选用不同种类的遥感测绘光学系统,可以最大程度利用平台资源,满足遥感测绘需求。     

Action potential-induced fluorescence changes resolved with an optical fiber carrying excitation light

With the use of a single, implantable, optical fiber, to excite fluorescence and detect changes from voltage-sensitive dyes, transmembrane potential changes were measured without the need for a clear line-of-sight path between the excitation light, the tissue, and the detector. In a previous study, we were required to use signal averaging and could detect only cardiac action potentials from frog. In the present study we improved this system so that unaveraged cardiac action potentials were resolved with high fidelity, and action potentials from single nerve axons were detected. Endeavors to optimize the signal-to-noise ratio resulted in the selection of a larger core fiber with a rounded tip, styryl dyes, and filters based upon fluorescence spectra of the dyes when bound to membrane (rather than in solution). The frog gave signals nearly comparable in magnitude and signal-to-noise ratio to those seen with systems that use a fluorescence microscope. Action potential-induced signals could be detected in single lobster axons with the intracellular injection of a dye. The improvement in the signal-to-noise ratio allowed the use of a reduced-intensity excitation illumination which produced less bleaching of the dye.     

Membrane Dipole Potential as Measured by Ratiometric 3-Hydroxyflavone Fluorescence Probes: Accounting for Hydration Effects

We previously applied the electrochromic modulation of excited-state intramolecular proton-transfer (ESIPT) reaction for the design of novel 3-hydroxyflavone (3-HF) derivatives as fluorescent probes for measuring the dipole potential, Ψ_D, in lipid bilayers (Klymchenko et al., Proc. Natl. Acad. Sci. USA, 2003, 100, 11219). In the present work, this method was revisited to take into account the influence of the bilayer hydration on the emission ratiometric response of 3-HF probes. For this reason, it was necessary to deconvolute the whole fluorescence spectra into three bands corresponding to the non H-bonded forms, normal N^* and tautomer T^* forms, both participating to the ESIPT reaction, and to the H-bonded H–N^* form, excluded from this reaction. This allowed us to determine the pure N^*/T^* intensity ratio, without any contribution from the H–N^* form emission depending essentially on the bilayer hydration. This new approach allowed us to confirm the correlation we obtained between the response of 3-HF probes on dipole potential modifications and the corresponding response of the reference fluorescent probe di-8-ANEPPS, thus further confirming the potency of 3-HF probes as excellent emission ratiometric probes to measure dipole potential in lipid membranes.     

Artifacts,assumptions, and ambiguity: Pitfalls in comparing experimental results to numerical simulations when studying electrical stimulation of the heart

Insidious experimental artifacts and invalid theoretical assumptions complicate the comparison of numerical predictions and observed data. Such difficulties are particularly troublesome when studying electrical stimulation of the heart. During unipolar stimulation of cardiac tissue, the artifacts include nonlinearity of membrane dyes, optical signals blocked by the stimulating electrode, averaging of optical signals with depth, lateral averaging of optical signals, limitations of the current source, and the use of excitation-contraction uncouplers. The assumptions involve electroporation, membrane models, electrode size, the perfusing bath, incorrect model parameters, the applicability of a continuum model, and tissue damage. Comparisons of theory and experiment during far-field stimulation are limited by many of these same factors, plus artifacts from plunge and epicardial recording electrodes and assumptions about the fiber angle at an insulating boundary. These pitfalls must be overcome in order to understand quantitatively how the heart responds to an electrical stimulus. (c) 2002 American Institute of Physics.