离焦系统下的单分子三维取向研究

提出了一种新思路来确定单分子的三维取向,在全内反射荧光显微成像的基础上,在显微镜的油镜上加一个水层,水和油的折射率不同造成光程差,产生离焦,利用全内反射产生隐失波激发了单分子的纵向分量,并在探测器前增加了一个分光镜,把光路分成偏振方向相互垂直的两束光,使得探测器上的光场分布包含单分子三维取向的信息。推导了隐失场激发的偶极矩光场在探测器上的光强分布表达式,通过分析偏振方向相互垂直的两束光在探测器上的不同的强度分布,来判断单分子偶极矩的三维取向。     

全内反射荧光显微术应用于单分子荧光的纵向成像

利用这种显微术中激发场的纵向强度呈指数衰减的分布特性，测得的荧光强 度将强烈依赖渗透深度（强度衰减到1/e时的距离），从而快速直接的确定单分子荧光 团间的纵向间隔、每个荧光团的纵向绝对位置和荧光团的半径大小，即实现荧光分子三维分 布的重构.在整个重构的过程中，只需要改变一次入射角的大小，即只需探测两幅荧光分子 的全内反射成像. 关键词： 全内反射荧光显微术 纵向重构     

隐失驻波全内反射荧光显微术的数值计算

从理论上详细计算了在全内反射的条件下，两束入射光产生的隐失干涉场(即隐失驻波)的强度分布，并分析了其不同于传统传播波干涉场的特点。同时使用数值模拟证明了利用隐失干涉场，即隐失驻波的激发方式，可以提高系统的分辨力，在横向实现超经典分辨的荧光成像。具体的分析表明，两束光以相等的角度入射，同时振幅相等，偏振态相同，所形成干涉条纹的反衬度最高，此时成像系统的有效点扩展函数最优化；入射介质1的折射率越大，隐失干涉场的空间周期越短(空间频率越高)，其对应的调制点扩展函数中心瓣的半峰全宽越小，可能分辨更小的物体，但同时旁瓣的强度也增强，最终成像的分辨力受两者的共同制约。     

全内反射荧光显微术

全内反射荧光显微技术是当今世界上最具前途的新型生物光学显微技术之一，可以用来实现对单个荧光分子的直接探测。它利用全内反射产生的隐失波照明样品，使照明区域限定在样品表面的一薄层范围内，因此具有其它光学成像技术无法比拟的高的信噪比和对比度。近上来，已被生物物理学家们广泛应用于单分子的荧光成像中。本文系统的介绍了全内反射荧光显微技术的原理。国内外的发展和现状及其在生物学上的应用，并对其未来做了展望。     

全内反射荧光显微术

全内反射荧光显微技术是当今世界上最具前途的新型生物光学显微技术之一 ,可以用来实现对单个荧光分子的直接探测。它利用全内反射产生的隐失波照明样品 ,使照明区域限定在样品表面的一薄层范围内 ,因此具有其它光学成像技术无法比拟的高的信噪比和对比度。近年来 ,已被生物物理学家们广泛应用于单分子的荧光成像中。本文系统的介绍了全内反射荧光显微技术的原理、国内外的发展和现状及其在生物学上的应用 ,并对其未来做了展望。     

全内反射构型下分子在十六烷-水界面取向角度的二次谐波测量

测量了在全内反射条件及非全内反射条件下,两种探针分子在十六烷-水界面上的取向角度.通过详细的偏振分析,发现探针分子在两种构型下的取向角度测量结果一致.表明全内反射条件下,取向角度的测量和偏振分析同样是准确且可行的.     

用偏振荧光光谱表征乙醇-水团簇分子的取向行为

采用波长为236nm紫外光激励乙醇水溶液获得荧光光谱并对其偏振特性进行了研究.实验中分别改变入射光的偏振度，在300—400nm波段内进行荧光光谱、水平和垂直偏振荧光光谱检测.实验结果表明，当采用线偏振光照射乙醇水溶液时，其发射的荧光具有一定的偏振性，并计算了其偏振度.经理论分析得到了描述荧光偏振态的四个斯托克斯矢量，进而对该荧光的偏振状态进行了描述，通过对乙醇-水溶液的米勒矩阵的计算，对溶液中分子的取向特性进行了分析.本结果能对乙醇-水团簇分子的结构提供理论参考. 关键词： 荧光光谱 偏振光谱 斯托克斯矢量 米勒矩阵     

活细胞内的单分子荧光成像方法

文章介绍近年来新发展的几种重要的活细胞内单分子荧光成像方法,如转盘式共聚焦显微术、全内反射荧光显微术、荧光共振能量转移技术等。通过介绍它们的原理、特点和在活细胞内单分子行为研究中的应用实例,展示了这些新方法在生命科学领域广阔的应用前景。     

牛奶水溶液的偏振三维荧光光谱研究

测得由伊利、蒙牛公司生产的纯牛奶、高钙奶、高钙低脂奶的偏振三维荧光光谱,发现在激发偏振片和发射偏振片取向平行时,偏振荧光强度高于取向垂直时的值。通过一个简单的模型估算了各种牛奶样品等效荧光团的激发偶极子和发射偶极子之间的夹角,均小于40°,且高钙低脂奶对应荧光团激发偶极子和发射偶极子之间的夹角略小于纯牛奶和高钙奶。实验还发现,在激发偏振片和发射偏振片取向平行时,高钙低脂奶的偏振荧光强度明显高于纯牛奶和高钙奶,与理论计算相符。     

异丙醇-水配合液的荧光偏振和时间分辨特性

研究了紫外光照射下异丙醇-水配合液的偏振荧光光谱,以及不同荧光峰处光子强度随时间的衰变过程,计算了偏振度并讨论了其偏振特性,测试了不同峰位对应的荧光寿命并分析了其荧光发射特性.结果表明,异丙醇-水配合液在紫外光激励下发射的荧光为具有确定分子取向的部分偏振光,偏振度和各向异性度分别为0.542和0.441.在波长为220...     

Controlling fluorescence emission with split‐ring‐resonator‐based plasmonic metasurfaces

Metasurfaces, which consist of resonant metamaterial elements in the form of two‐dimensional thin planar structures, retain great capabilities in manipulating electromagnetic wave and potential applications in modifying interaction with fluorescent molecules. The metasurfaces with magnetic responses are favorable to weakening fluorescence quenching while less investigated in controlling fluorescence. In this paper, we demonstrate control over fluorescence emission by engineering the magnetic and electric modes in plasmonic metasurfaces consisting of 45‐nm‐thick gold split‐ring‐resonators (SRRs). The fluorescence emission exhibits an enhancement factor of ∼18 and is predominantly x‐polarized with assistance of the magnetic mode excited by oblique incidence with an x‐polarized electric field. The magnetic and electric modes excited by oblique incidence with a y‐polarized electric field contribute to the rotation of emission polarization with respect to the incident polarization. The results demonstrate manipulating the interaction of fluorescent emitters with different resonant modes of the SRR‐based metasurface at the nanoscale by the polarization of incident light, providing potential applications of metasurfaces in a wide variety of areas, including optical nanosources, fluorescence spectroscopy and compact biosensors.

     

Study of the frequency distribution of the fluorescent light induced by monochromatic radiation

The frequency distribution of the fluorescent light induced by monochromatic dye laser radiation was investigated. To exclude the influence of the Doppler width a strongly collimated atomic beam was used. The spectrum was measured by means of a piezoelectrically tunable spherical Fabry Perot. The interaction region between the laser light and the atomic beam was placed into the center of the interferometer. Thus the observed fluorescence spectrum was considerably more intense than in the case where the interferometer is used separately from the beam. The fluorescence spectrum was observed for different directions of polarization of the incident laser beam. In the case of weak excitation the spectrum consists of a sharp component essentially due to elastically scattered light. At high intensities a structure of three components is observed which is in agreement with theoretical predictions when circularly polarized light is used for excitation.     

Study of the frequency distribution of the fluorescent light induced by monochromatic radiation

The frequency distribution of the fluorescent light induced by monochromatic dye laser radiation was investigated. To exclude the influence of the Doppler width a strongly collimated atomic beam was used. The spectrum was measured by means of a piezoelectrically tunable spherical Fabry Perot. The interaction region between the laser light and the atomic beam was placed into the center of the interferometer. Thus the observed fluorescence spectrum was considerably more intense than in the case where the interferometer is used separately from the beam. The fluorescence spectrum was observed for different directions of polarization of the incident laser beam. In the case of weak excitation the spectrum consists of a sharp component essentially due to elastically scattered light. At high intensities a structure of three components is observed which is in agreement with theoretical predictions when circularly polarized light is used for excitation.     

Improving the fluorescence polarization method to evaluate the orientation of fluorescent systems adsorbed in ordered layered materials

The fluorescence polarization method, recently developed for the evaluation of the preferential orientation of fluorescent dyes adsorbed in layered materials [F. López Arbeloa, V. Martínez Martínez, J. Photochem. Photobiol. A: Chem. 181 (2006) 44], is readapted to improve its application. Fluorescence polarization was previously obtained by recording the emission intensity for two orthogonal orientations of the emission polarizer (i.e., the horizontal and vertical polarized light) after excitation with vertical or horizontal polarized light. In the method proposed in this work, samples are excited with unpolarized light, reducing the polarization effect of the excitation light scattering at those emission wavelengths close to the excitation wavelength. Moreover, the present method decreases the effect of the orientation of other non-fluorescent species present in the system, which are active in the excitation process. Consequently, the new method is more simple, precise and sensitive. It is applied to evaluate the orientation of rhodamine 6G dye adsorbed in ordered laponite clay films with low and moderated dye loadings.     

Polarization-induced control of two-photon excited fluorescence in a chiral polybinaphthyl

The fluorescence behavior of a chiral polybinaphthyl excited with 100 fs 800 nm laser pulses was investigated in tetrahydrofuran solution. The peak fluorescence intensity versus the input irradiance was measured to meet a square dependence, giving evidence for two-photon excited fluorescence (TPF). The variations of the TPF intensity were found to be strongly modulated by the different polarized incident lights and tightly depend on the linearly polarized component of the incident light. Furthermore, combining with the characteristics of chiral molecules, the two-photon polarization ratio was studied to reveal the symmetry of the involved excited states.     

Whispering-gallery modes in glass microspheres: optimization of pumping in a modified confocal microscope

Whispering-gallery modes (WGMs) on Nd3+-doped glass microspheres with a radius of ～15?μm were measured in a modified confocal microscope, where a dual spatial resolution in both excitation and detection zones was possible. As an alternative to the standard excitation mechanism by an evanescent wave, we used an efficient pumping/detecting scheme, focusing a laser in the microsphere and exciting the Nd3+ ions, whose fluorescent emission produces the WGMs. We have also measured the generated WGMs by changing the detection zone, where higher amplitude resonances were found when exciting in the center and detecting at the edge of the microsphere.     

激光光源-在线荧光光谱检测中的光强度比较研究

基于激光光源和氙灯光源在线荧光光谱法测定罗丹明B、维生素B₂、荧光素和异硫氰酸荧光素含量比较研究激光光源产生的荧光强度和氙灯光源产生的荧光强度。罗丹明B、维生素B₂、荧光素和异硫氰酸荧光素浓度均为10 μg·mL-1,积分时间100 ms,测定3次得其平均值。在线荧光光谱法最大吸收波长分别为580,450,488和510 nm;最大发射波长依次为594,530,525和524 nm。紫外-可见分光光度法测得其最大吸收波长为557,441,481和490 nm;荧光分光光度法测得其最大发射波长为586,520,519和520 nm。通过测定药物,发现激光光源产生的荧光光强度较强于氙灯光源产生的荧光光强度,原因不仅跟光源有关,而且与药物分子的共轭体系大小、共轭大π键的共平面性及其刚性程度、分子母体上取代基的种类有关,分子所处的外界环境如温度、溶剂、溶液酸碱度、激发光的照射等因素也会影响荧光效率。激光光源和氙灯光源产生的荧光光强度大小顺序为罗丹明B>荧光素>异硫氰酸荧光素>维生素B₂。激光光源在线荧光光谱法在一定程度上填补了在线荧光光谱仪在食品、药品痕量检测方面应用的空白。     

Optimization of evanescent wave imaging for the visualization of protein adsorption layers

We have optimized the settings of evanescent wave imaging for the visualization of a protein adsorption layer.The enhancement of the evanescent wave at the interface brought by the incident angle,the polarized state of light beam as well as a gold layer is considered.In order to improve the image contrast of a protein monolayer in experiments,we have optimized three factors- the incident angle,the polarization of light beam,and the thickness of an introduced thin gold layer with a theoretical simulation.     

Intensification and location of relativistic laser pulses against pointing fluctuations with micro-tube plasma lenses

We analyze, via an off-axis incident model and three-dimensional particle-in-cell simulations, the influence of beam pointing fluctuations (BPFs) on the propagation properties of relativistic laser pulses in micro-tubes. It is observed that the in-tube laser intensity can be further amplified in the BPFs-induced off-axis incident case. But the intensification factor exhibits strong polarization-dependence. When the laser pulse is linearly polarized in the off-axis incident plane (p-polarization), more electrons may be dragged out from about a half of the tube inner surface in each across section than in the on-axis incident case, enhancing the effects of relativistic nonlinearity and channel focusing. The area for generating more electrons is reduced by one half in the s-polarization case, resulting in less efficient light intensification. The BPFs-induced off-axis incident also leads to pulse shortening. Moreover, light confinement in the tube core is evident and the laser pulse tends to be coaxial with the tube.