Hydrophobic optical elements for near-field optical analysis (NOA) in liquid environment--a preliminary study.

Near-field Scanning Optical Microscopy (NSOM) in liquid environment is expected to allow time resolved morphological mappings on cellular surfaces on the nanoscale level. Near-field Optical Analysis (NOA) via NSOM exploits the energy transfer from the tip of an optical element (tip diameter > or = 20nm), oscillating within the range of the characteristic length of the energy transfer ( approximately 10nm) in the near-field of the surface to be analysed. In NOA, a molecular assembly is monitored by visible light with a resolution far below the wavelength of visible light. Actually, NOA is successfully applied in mapping local optical contrasts, for instance in photonic crystals with dielectric periodicities on the nanoscale. NSOM could in principle be performed in two different modes: tapping mode, with tip-oscillations perpendicular, or shear force mode, with tip-oscillations parallel to the substrate. Both basic modes have specific advantages and disadvantages. In biological systems (e.g. in cell cultures), where scanning in liquids is prevalent, elongated optical elements non-invasively operated in the shear force modus could have some specific advantages when compared to contact modus systems. While tapping mode NSOM provides satisfactory nanoscale images even on solid surfaces covered with millimetres of liquids, the performance of shear force mode NSOM is presently largely confined to operations on dry samples. This is due to the inability of conventional shear force mode NSOM systems to provide sharp topographic images of sample surfaces substantially covered with liquids. By equipping a conventional NSOM system with hydrophobic optical elements, shear force mode based topographic images could be obtained on biological samples in dry as well as in aqueous environment, and with resolutions on the nanoscale level.     

超分辨率成像荧光探针材料应用进展

为了进一步认知复杂环境中的细胞生物学过程,研究人员发展了各种各样的生物成像技术。在这些技术中,生物荧光成像因简单的成像条件以及对生物样品的相容性而得到了广泛的发展。然而,传统的荧光成像技术受到了光学衍射极限的限制,无法分辨低于200 nm的空间结构,阻碍了对亚细胞结构的生物学过程研究。超分辨荧光显微镜技术突破了传统光学衍射对成像分辨率的限制,能够获取纳米尺度的细胞动态过程。除了对传统的宽场荧光显微镜框架的改进及升级改造之外,目前典型的超分辨成像显微镜技术通常依赖于荧光探针材料的光物理性质。常用的荧光探针材料包括荧光蛋白、有机荧光分子和纳米荧光材料等。本文介绍了几种主流的超分辨荧光显微成像技术并总结了已经成功应用到超分辨生物荧光成像中的荧光探针材料的应用进展。     

Full-field imaging-based instantaneous hyperspectral absolute refractive index measurement

Multispectral refractometers typically measure refractive index (RI) at discrete monochromatic wavelengths via a serial process. We report on the demonstration of a white light full-field imaging-based refractometer capable of instantaneous multispectral measurement of absolute RI of clear liquid-gel samples across the entire visible light spectrum. The broad optical bandwidth refractometer is capable of hyperspectral measurement of RI in the range 1.30-1.70 between 400 and 700 nm with a maximum error of 0.0036 units (0.24% of actual) at 414 nm for an η=1.50 sample. We present system design and calibration method details as well as results from a system validation sample.     

Two-color far-field fluorescence nanoscopy based on photoswitchable emitters

We demonstrate two-color far-field fluorescence microscopy with nanoscale spatial resolution based on the photoswitching of individual fluorescent markers. By enabling, recording, and disabling the emission of the reversibly switchable fluorescent protein rsFastLime and of the organic fluorophore cyanine5, we recorded two-color nanoscale images inside whole cells. The position of individual emitters was determined with a typical accuracy of 20 nm, which largely constitutes the lateral resolution of the system. Photoswitching in two-color colocalization experiments represents a major step towards the application of far-field fluorescence nanoscopy to the study of (biological) samples on the macromolecular level. PACS 33.50.Dq; 42.30.-d; 87.64.-t     

Phase-retrieval in hard x-ray diffraction imaging: Can we quantitatively reconstruct a “real” transmission function?

X-ray phase-retrieval algorithms are widely exploited in contemporary hard x-ray diffraction techniques to image at the nanoscale, less than 10-20 nm. Often reconstruction of the sample shape (image) suffices for the purpose of experiment. Identification of specimen composition requires a quantitative profiling of the complex refractive index. This Letter shows that, although such quantitative analysis is possible in many cases, there is a lower limit to variations in optical density, which can be quantitatively reconstructed using the common phase-retrieval methods.     

有孔探针近场拉曼光谱和成像技术进展

基于有孔探针SNOM的近场拉曼光谱和成像技术的出现使得拉曼光谱的分辨率突破了光学衍射极限,从而提供了一个有力的工具对样品亚波长尺度之下的化学信息进行表征。文章讨论了探针性质对实现近场拉曼光谱的影响,并全面地介绍了有孔探针近场拉曼光谱发展十余年来在纳米尺度化学分辨成像、液-液界面性质研究、微观层面解释SERS增强机理、图像化反映SERS热点分布等诸多领域的研究进展。     

FABRICATION AND APPLICATION OF NEAR-FIELD OPTICAL FIBRE PROBE

In this paper, the fabrication of a large cone angle near-field optical fibre probe, using the two-step chemical etching method and bent probe, is introduced, and the controlling parameters of the coated Cr－Al film at the probe tip are presented. The scanning electron microscopy images display that the tip diameter of the uncoated large cone angle fibre probe obtained is less than 50nm, the cone angle over 90°, and the diameter of light aperture at the coated probe tip is less than 100nm. The measured results of the optical transmission efficiency for various probe tips show that the uncoated straight optical fibre probe, film-coated straight probe and film-coated bent probe are 3×10^-1, 2×10^-3, and 1×10^-4 times that of the flat fibre probe, respectively. In addition, the force images and near-field optical images of a standard sample are acquired using a large cone angle and film-coated bent probe.     

Ultrathin side-viewing needle probe for optical coherence tomography

We present the smallest reported side-viewing needle probe for optical coherence tomography (OCT). Design, fabrication, optical characterization, and initial application of a 30-gauge (outer diameter 0.31 mm) needle probe are demonstrated. Extreme miniaturization is achieved by using a simple all-fiber probe design incorporating an angle-polished and reflection-coated fiber-tip beam deflector. When inserted into biological tissue, aqueous interstitial fluids reduce the probe's inherent astigmatism ratio to 1.8, resulting in a working distance of 300 μm and a depth-of-field of 550 μm with beam diameters below 30 μm. The needle probe was interfaced with an 840 nm spectral-domain OCT system and the measured sensitivity was shown to be only 7 dB lower than that of a comparable galvo-scanning sample arm configuration. 3D OCT images of lamb lungs were acquired over a depth range of ~600 μm, showing individual alveoli and bronchioles.     

Low voltage high-resolution SEM (LVHRSEM) for biological structural and molecular analysis

High-resolution scanning electron microscopy (HRSEM) is being used increasingly to gain new insights into three-dimensional organization of biological structure, macromolecular complexes and interactions of cellular components as well as isolated cell organelles. Modern scanning electron microscopes (SEMs) combined with adequate sample preparation can now provide resolution comparable with that achieved using transmission electron microscopes (TEMs) down to 2-5 nm for biological material. The versatility of the instrument and new sample preparation techniques have allowed detailed analysis of chromosomes, cytoskeletal components, virus and other biological material that has not been possible with TEM. The present review addresses resolution and specific specimen preparations for HRSEM, and highlights the importance of specimen preparation and choice of methods to achieve optimal results for proteins, macromolecular complexes and subcellular structures using low voltage HRSEM (LVHRSEM).     

新型光敏剂叶绿酸f对人膀胱癌T24细胞的光动力杀伤作用

将自主研发的新型光敏剂叶绿酸f与T24细胞共同孵育后,以650nm激光照射,通过MTT法检测光动力处理后T24细胞的生长抑制率,流式细胞术检测凋亡情况,同时利用激光共聚焦显微镜488nm和405nm双波长激发,应用线粒体探针,进行叶绿酸f的亚细胞定位。结果显示叶绿酸f为浓度2.5,5,10μg/mL对T24细胞生长抑制率在光能量密度4J/cm2下为17.68%,49.35%和84.42%,在1J/cm2时为4.34%,37.42%和78.38%;流式细胞术显示T24细胞光动力处理后,凋亡率为(45.23±1.2)%,显著高于对照组;激光共聚焦显微镜显示504nm激发叶绿酸f发出的红光与488nm激发线粒体探针产生的绿光分布基本一致。实验证明叶绿酸f对T24细胞杀伤效果明显,主要是通过作用于线粒体,诱导细胞凋亡来完成的。     

Unveiling facet effects in metallic nanoparticles to design an efficient plasmonic nanostructure

We investigated the role of nanoparticle surface morphology (facets) in estimating the plasmonic properties of nanoparticle-on-a-mirror design or NPOM in the presence of a thicker (20 nm) dielectric layer. Nanoparticle surface morphology differences ranging from smoother surface to multi-facets in the form of a sphere (NSOM), cube (NCOM), and singe bottom faceted sphere (SBF-NSOM) shapes have been employed. Three significant optical properties were observed. Better longer wavelength near-field and far-field resonance spectral positions from NCOM are achieved. Near-field enhancement extracted from SBF-NSOM outperformed NCOM by more than ∼ two times. Plasmonic gap mode enhancement is absent for NSOM in the presence of a larger dielectric layer. The availability of plasmonic gap modes in NCOM and SBF-NSOM, even at a 20 nm thick dielectric layer, is highly beneficial for various plasmonic applications. These differences in optical properties are understood by the role of NP facets in influencing the plasmonic cavity region.     

Influence of cavity geometry towards plasmonic gap tolerance and respective near-field in nanoparticle-on-mirror

In this work, we emphasize the importance of cavity geometry along with nanoparticle shape and plasmonic nanogap (based on a nanoparticle on a metallic film (NPOM) design) which plays significant role in understanding the complex plasmonic mode characteristics involving nanoparticle and gap mode resonances. The cross-section imprint of planar cavity on metallic film plays decisive role in near field enhancement properties at similar NP size and plasmonic nanogap conditions for spherical and cubical NPOM systems. By mimicking the NPOM structure to metal-insulator-metal design, we understand the resonant emission differences for the respective plasmonic modes. Influence of dominant and weaker gap mode resonances resulted in an interesting optical behavior (fluctuations in near field enhancement strength) in NP mode in case of cubical nanostructures. By such extensive investigation and interpretation of sub-wavelength complex plasmonic mode characteristics, various practical applications in plasmonics field can be accomplished.     

动态化学腐蚀法制备大锥角近场光纤探针

基于液压传递原理,设计出一种用动态化学腐蚀法制备大锥角近场光纤探针的装置.实验结果表明,在传统的化学腐蚀法制备光纤探针的过程中,通过控制腐蚀液液面的上升速度,可以有效地控制探针针尖的形状以及锥角的大小.在此基础上,还论述了分步控制光纤与腐蚀液液面的相对位移的方法在大锥角光纤探针的制备中所具有的独特优越性.利用所制备的大锥角近场光纤探针在扫描近场光学显微镜上对直径为200nm的小球进行探测,其力学像证实该探针具有较高的形貌分辨率(约为50nm). 关键词： 动态化学腐蚀法 大锥角 近场光纤探针     

Prism-coupled multimode waveguide refractometer

A new refractometer has been developed based on changes in the effective refractive index (RI) of the highest-order TE (or TM) mode in a prism-coupled multimode planar waveguide induced by interaction between an evanescent field and a liquid sample. The waveguide was a 100-mu;m -thick quartz plate fixed on a poly(methyl methacrylate) support containing a flow cell. A pair of prism couplers contacted the quartz plate in the flow-cell region. Such an optical sensor can detect the RI of liquid in a wide range by monitoring the resonant angle of the highest-order mode that changes order number with changes in the sample's RI. When a highest-order mode corresponding to a given RI range is used as the sensor probe, a slight RI change in this range can be detected by measurement of the output light intensity. With this method the sensor was demonstrated to have a resolution of 3x10(-5) for the RI of an aqueous solution. Combining this result with theoretical calculation indicates that the sensor can detect a 0.5-nm-thick monolayer adsorbed from an aqueous solution. Therefore, the sensor is suitable for real-time detection of biomolecular interactions.     

A high-sensitivity photonic crystal fiber (PCF) based on the surface plasmon resonance (SPR) biosensor for detection of density alteration in non-physiological cells (DANCE)

A highly sensitive photonic crystal fiber based on the surface plasmon resonance (PCF-SPR) biosensor for the detection of the density alteration in non-physiological cells (DANCE) is described. Human acute leukemia cells are determined by the discontinuous sucrose gradient centrifugation (DSGC) in which the cells are separated into several bands. The separated cells with different intracellular densities and refractive indexes (RI) ranging from 1.3342 to 1.3344 are distinguished in situ by means of the differential transmission spectrum. The biosensor shows a maximum amplitude sensitivity of 2000?nm/RIU and resolution as high as 5?×?10^?5?RIU. According to the wavelength interrogation method, a maximum spectral sensitivity of 9000?nm/RIU in the sensing range between 1.33 and 1.53 is achieved, corresponding to a resolution as high as 1.11?×?10^?5?RIU for the biosensor. The proposed PCF-SPR biosensor has promising application in biological and biochemical detection.     

扫描探针声显微镜轻敲模式中相位像的反差机理

根据扫描探针声显微镜(SPAM)轻敲工作模式中探针作周期振动的特点,以及在探针与试样相接触过程中激振力和悬臂探针自由振动的阻尼力很小的假设下,解析求解了探针与试样的碰撞运动方程,得到了最大压痕深度和碰撞时间与探针半径、等效杨氏模量以及界面吸附能等之间的关系式,较直观地说明了SPAM中轻敲模式的相位像反差机理:信号的相位与试样微区的性质、探针振幅、设置点以及试样形貌等有关。并定量预计了纳米金刚石像中的相位差值72.59°,与实验测量平均值73.2°±8.2°一致。同时,合理地解释了实验得到的光学薄膜试样相位像的反差。这些表明SPAM轻敲模式的相位成像是一种纳米分辨率测量材料表面物理性质的成像技术。      

Anisotropic fibers with elliptical deformed core

Picosecond pump and probe experiments were carried out on fully oriented transpolyacetylene. Exciting the sample at 527 nm, photoinduced bleaching and absorption were observed at 527 nm and 1054 nm, respectively. The measurements give an upper limit of 2 ps for the relaxation time of the photogenerated 0.5 eV hot carriers to the band edges. The evaluation of the measured optical processes strongly suggests the conclusion that photoinduced structural distortions of the polyacetylene chain result in observable transient decrease of the interband absorption. This may be ascribed to the reduction of the density of electronic states and to that of the interband transition matrix element of the momentum operator.     

Sensitivity investigation of the different direction vibrations of a SNOM probe

In this article, we analyze the sensitivity of lateral, axial, and torsional vibration of a scanning near-field optical microscope (SNOM) probe. An exact solution for the title problem is obtained using the theory of beam and torsional vibrations. Sensitivities are obtained for a wide range of the intervening physical parameters. This paper discusses the relationship between the sensitivity of a SNOM probe and its geometry. The analysis is presented for a probe in three different cases lateral vibration, axial vibration, and torsional vibration, for a fixed-free configuration. To derive the mathematical relation between the probe and sample (by utilizing springs and damping to simulate different environments) through the process of theory analysis for different environments. Thus a universal touchless SNOM probe sensitivity controller is developed for appropriate control and adjustment of the probe sensitivity suitable under different environments. Such controller helps the SNOM probe achieve the nanoscale resolution through proper control and adjustment probe sensitivity in accordance to changes to different environments and reduce the chance of being damaged during the scan, which in the end further advances the sample inspection technique.     

Near-field optical microscope with a multiheight scanning imaging mode

A near-field scanning optical microscope has been developed to yield optical images with various gap distances between the probe and the sample surface. The microscope uses an apertureless metallic probe, the position of which is controlled by regulation of the tunneling-electron current from the probe to the sample and by computer-generated bias voltage. Experimental results of near-field optical imaging with the developed microscope at different gap distances are shown. Thirteen images at gap distances of 0 to 500nm demonstrate that the near-field image depends strongly on the gap distance. The imaging characteristics of a near-field imaging system are shown with the spatial-frequency spectra of images. Future investigation of the developed microscope is also discussed.     

Finite-Difference-Time-Domain Analysis for Electro-Magnetic Field Distribution on Near-Field Optical Recording Probe Head

In recent years there has been a strong trend and movement towards digitalism and high memory capacity in the multi-media field including digital movies. It is expected that new kinds of high density data storage to match this environment will be developed. The optical disk system using near-field optics and technology is considered to be a strong candidate for ultra-high density optical memory. In order to realize a new optical recording system we have focused on the writing head with high optical efficiency and resolution. We evaluated the properties of pyramidal probe heads using the FDTD (finite difference time domain) method and obtained simulation results in order to find the best structure and design for new near-field optical probes. From these results, we understood that a novel probe structure having metal layers on just 3 side planes (scoop type) shows the best performance in optical power density and beam size. We believe that optical enhancement and beam confinement arise from the effect of surface plasmon on a metal layer and the beam blocking of metal, respectively. Using this scoop-type probe having a tip size of 55 nm, we can get a very small spot size of 33 #x00D7; 60 nm² in FWHM (full with half maximum), which allows us to realize the recording density of 300 Gb/in².