Scanning Near-field Optical Microscopy Images Research Articles

Heterogeneous Molecular Distribution in Supported Multicomponent Lipid Bilayers

Membrane domains contribute important structural and functional attributes to biological membranes. We describe the heterogeneous nanoscale distribution of lipid molecules within microscale membrane domains in multicomponent lipid bilayers composed of dipalmitoylphosphatidylcholine (DPPC), dilauroylphosphatidylcholine (DLPC), and cholesterol (chol). The lipids were labeled with the fluorescent lipid analogues Bodipy-PC and DiI-C20:0 to identify the distribution of individual membrane components. We used a near-field scanning optical microscope (NSOM) at room temperature to identify the nanoscale structures in the membrane. Simultaneous multicolor NSOM imaging at the emission maxima of the fluorescent analogues revealed a patchy distribution of Bodipy-PC and DiI-C20:0 indicative of phase separations in the bilayer. In a cholesterol-free system (DPPC/DLPC = 1:1), NSOM images proved that the two phosphatidylcholine molecules can coexist in domains at the micrometer level but form nanoscopic patches within the domains; DPPC occurs at the edge of the domains, whereas DLPC is present throughout the domains. In the presence of cholesterol (DPPC/DLPC = 7:3, chol = 18.9%), the two lipid molecules were more miscible but incomplete phase separations also occurred. The average domain sizes were 140-200 nm, well below the resolution capabilities of diffraction-limited light microscopy techniques; the domains were unresolvable by confocal microscopy. Our high-resolution NSOM studies of membrane domain behavior provide a better understanding of complex membrane phase phenomena in multicomponent biological membranes.

Scanning Near-Field Optical Microscope Study of Ag Nanoprotrusions Fabricated by Nano-oxidation with Atomic Force Microscope

We investigate light transmission of Ag nano protrusions with a scanning near-field optical microscope (SNOM) to study the surface plasmon (SP) in metallic films. A Ag nanoprotrusion whose height is ∼20 nm is fabricated by the deposition of Ag on the Ti nano-protrusion template which is made by nano-oxidation with an atomic force microscope (AFM). It is found that through several scans, the tip is coated with Ag from the sample, and this modified tip shows apparent undulation patterns in SNOM imaging, while a fresh tip does not. The undulation pattern in the SNOM image is observed only along the direction of polarization light close to the expected undulation period. This result indicates that local launching of SP is effective with an Ag-coated tip and the SP launching characteristic is consistent with the so-called cos 2Φ SP distribution. By finite domain time difference (FDTD) simulations, it is confirmed that light transmission is enhanced in the Ag-coated tip compared to the fresh tip. Our results imply that local illumination with a noble-metal-coated cantilever probe tip is desirable for obtaining more detailed features in SNOM images involved with SP. Our results also indicate that a nano protrusion fabricated by AFM-based nano-oxidation can be applied to an optical element using surface plasmon.

Optimizing the near field around silver tips.

Owing to their promise of obtaining optical as well as topographic information in nanometer scale, apertureless near-field scanning optical microscopy (NSOM) and apertureless near-field scanning optical spectroscopy have drawn much attention recently. However, NSOM is still not a mature technique. A proper understanding of and the ability to tune the near field around the tip end is critically important in NSOM instrumentation and in NSOM image interpretation. On the basis of reflection geometry, we systematically studied the effects of a number of parameters pertinent in the application of apertureless NSOM, e.g., polarization, incident angle, wavelength of the incident laser, tip material, and tip length, by using the generalized field propagator technique. Our results show that all the above parameters have a significant influence on near-field enhancement and that care must be taken in the design of the experiment in order to maximize the near field. In addition to apertureless NSOM and spectroscopy, apertureless near-field lithography can benefit from these simulation results.

Exploring the Photorefractive Effect in Polymer-Dispersed Liquid Crystals Using Near-Field Scanning Optical Microscopy

Static and dynamic near-field scanning optical microscopy (NSOM) methods are used to study physical phenomena associated with photorefractivity in dye-doped polymer dispersed liquid crystals (PDLCs). Micrometer-sized liquid crystal droplets doped with perylene and N,N'-dioctyl-1,4,5,8-naphthalene diimide (NDI) dyes, which serve as the photoexcitable electron donor and acceptor, respectively, are studied. The droplets are encapsulated within a thin poly(vinyl alcohol) film. Simultaneously recorded topographic, fluorescence, and birefringence NSOM images show the films to be morphologically complex, containing ellipsoidal and collapsed ellipsoidal droplets. The presence of the collapsed droplets suggests the dissolved dyes may be surface active. Nevertheless, the NSOM images prove the vast majority of dye is contained within the well-defined liquid crystal droplets. Data on the local ion migration and liquid crystal reorientation dynamics is obtained by dynamic NSOM imaging methods. A sinusoidally modulated electric field is applied across the sample, between the aluminized NSOM probe and the electrically conductive film substrate. Lockin detection of the modulated optical signal resulting from field-induced liquid crystal reorientation is used to obtain amplitude and phase images of the sample. Comparison of the images recorded before, during, and after photogeneration of ions shows spatial variations in the relative importance of ion generation and migration on the liquid crystal orientation state. Computer simulations of the field-dependent ion migration dynamics and the liquid crystal response are used to better understand the data obtained.

Chemically Resolved Imaging of Biological Cells and Thin Films by Infrared Scanning Near-Field Optical Microscopy

The infrared (IR) absorption of a biological system can potentially report on fundamentally important microchemical properties. For example, molecular IR profiles are known to change during increases in metabolic flux, protein phosphorylation, or proteolytic cleavage. However, practical implementation of intracellular IR imaging has been problematic because the diffraction limit of conventional infrared microscopy results in low spatial resolution. We have overcome this limitation by using an IR spectroscopic version of scanning near-field optical microscopy (SNOM), in conjunction with a tunable free-electron laser source. The results presented here clearly reveal different chemical constituents in thin films and biological cells. The space distribution of specific chemical species was obtained by taking SNOM images at IR wavelengths (λ) corresponding to stretch absorption bands of common biochemical bonds, such as the amide bond. In our SNOM implementation, this chemical sensitivity is combined with a lateral resolution of 0.1μm (≈λ/70), well below the diffraction limit of standard infrared microscopy. The potential applications of this approach touch virtually every aspect of the life sciences and medical research, as well as problems in materials science, chemistry, physics, and environmental research.

Analysis of the interferometric effect of the background light in apertureless scanning near-field optical microscopy

We investigate in detail the interferometric nature of the signal delivered by an apertureless scanning near-field optical microscope (SNOM). This nature is first brought to the fore by near-field images of an integrated waveguide. The detection process of an evanescent wave generated by total internal reflection is then studied by both lateral near-field scans and signal detection as a function of the tip-to-sample distance. This study permits interpretation of fringe patterns appearing in apertureless SNOM images and provides important information about the nature of the signal. In particular, both experimental data and simple calculations show that, because of interference with background light coming from the sample, the detected signal can describe the complex field amplitude, or the field intensity, or a subtle mix of both, depending on the tip environment and the tip position.

Fully time-resolved near-field scanning optical microscopy fluorescence imaging

Time-correlated single photon counting has been coupled with near-field scanning optical microscopy (NSOM) to record complete fluorescence lifetime decays at each pixel in an NSOM image. The resulting three-dimensional data sets can be binned in the time dimension to create images of photons at particular time delays or images of the fluorescence lifetime. Alternatively, regions of interest identified in the topography and fluorescence images can be used to bin the data in the spatial dimensions resulting in high signal to noise fluorescence decays of particular regions of the sample. The technique has been demonstrated on films of poly(vinylalcohol), doped with the fluorescent dye, cascade blue (CB). The CB segregates into small circular regions of high concentration within the films during the drying process. The lifetime imaging shows that the spots have slightly faster excited state decays due to quenching of the luminescence as a result of the higher concentration. The technique is also used to image the fluorescence lifetime of an annealed film of poly(dihexylfluorene). The samples show high contrast in the total intensity fluorescence image, but the lifetime image reveals the sample to be extremely uniform.

AFM and SNOM characterization of carboxylic acid terminated silicon and silicon nitride surfaces

Silicon and silicon nitride surfaces have been successfully terminated with carboxylic acid monolayers and investigated by atomic force microscopy (AFM) and scanning near-field optical microscopy (SNOM). On clean Si surface, AFM showed topographical variations of 0.3–0.4 nm while for the clean Si 3N 4 surface the corrugation was around 3–4 nm. After material deposition, the corrugation increased in both samples with a value in topography of 1–2 nm for Si and 5–6 nm for Si 3N 4. The space distribution of specific chemical species was obtained by taking SNOM reflectivity at several infrared wavelengths corresponding to stretch absorption bands of the material. The SNOM images showed a constant contribution in the local reflectance, suggesting that the two surfaces were uniformly covered.

Characterization of Domain Ordering in Polymer and Dendrimer Thin Films Using Photoluminescence and Third Harmonic Generation (THG) Near-field Scanning Optical Microscopy (NSOM)

We have utilized the high spatial resolution of near-field scanning optical microscopy (NSOM) to characterize nanoscopic electronic inhomogeneity in as-cast thin films of a light-harvesting dendrimer consisting of coumarin-343 (core) and coumarin-2 (peripheral) chromophores and in thermally annealed thin films of the semiconductive polymer poly(2-methoxy-5-(2'-ethylhexyloxy)-1,4-phenylene vinylene) (MEH-PPV). Using photoluminescence (PL) and third harmonic generation (THG) NSOM techniques, we have observed nanoscopic domains in these films that exhibit increased delocalization in the excited (using PL) and ground states (using THG). In addition, we have developed a procedure for examining NSOM images via calculation of radial distribution functions (RDFs). RDF analysis of the PL and THG NSOM images indicates that the domains exhibit correlated structure in annealed MEH-PPV films while the light-harvesting dendrimeric material does not. The existence or absence of such nanoscopic amorphous structure can be understood in terms of the molecular structure of each material.

Near-field scanning optical microscopy imaging of multimode interference.

By use of a near-field scanning optical microscope (NSOM) in collection mode, the intensity distribution along a 2 x 2 multimode interference coupler was directly imaged as a function of wavelength. Although calculations can predict the general trend of wavelength dependence and the approximate positions of multiple images in the coupler, the accuracy is poor because of uncertainties in the waveguide width. We show that direct imaging using a NSOM bypasses calculational uncertainties and proves to be a powerful technique for studying these waveguide devices.

Near-field imaging of surface plasmon on gold nano-dots fabricated by scanning probe lithography.

We obtained scanning near-field optical microscopy images to study the excitation of surface plasmons on metallic dots fabricated using scanning probe lithography. Gold nano-dots were fabricated by applying electric voltages to conducting probes installed in an atomic force microscope using the mechanism of field-induced diffusion and nano-oxidation plus Au-coating. High spatial resolution of scanning near-field optical microscopy revealed a 'bifold' pattern of surface plasmon mode on fabricated Au dots in the polarization direction of incident light. We found that scanning near-field optical microscopy imaging combined with scanning probe lithography is able to provide a systematic study of surface plasmon excitation on nano-metallic structures.

Photonic nanopatterns of gold nanostructures indicate the excitation of surface plasmon modes of a wavelength of 50–100 nm by scanning near‐field optical microscopy

Scanning near-field optical microscopy images of metal nanostructures taken with the tetrahedral tip (T-tip) show a distribution of dark and bright spots at distances in the order of 25-50 nm. The images are interpreted as photonic nanopatterns defined as calculated scanning near-field optical microscopy images using a dipole serving as a light-emitting scanning near-field optical microscopy probe. Changing from a positive to a negative value of the dielectric function of a sample leads to the partition of one spot into several spots in the photonic nanopatterns, indicating the excitation of surface plasmons of a wavelength in the order of 50-100 nm in metal nanostructures.

Apertureless scanning near-field optical microscopy for ion exchange channel waveguide characterization.

We report the characterization of an integrated Ag+/Na+ ion exchange waveguide realized in a silicate glass substrate using apertureless scanning near-field optical microscopy. Our experimental set-up is based on the combination of a commercial atomic force microscope with an optical confocal detection system. Thanks to this system, the topography and evanescent optical field at the waveguide top surface are mapped simultaneously. Also, the process of apertureless scanning near-field optical microscopy image formation is analysed. In particular, fringe patterns appearing in the image reveal the intrinsic interferometric nature of the collected signal, due to interference between the field scattered by the tip end and background fields related to guide losses. We give a quantitative interpretation of these fringes. Evanescent intensity mapping on the sample surface allowed us to extract physical waveguide parameters. In particular, it shows an unambiguous multimode beat along the waveguide propagation axis. Furthermore, we show that analysis of this intensity profile reveals back-reflection effects from the waveguide exit facet. The resulting standing waves pattern allows us to evaluate the eigenmode propagation constants.

SNOM imaging of very fine pits formed by EB lithography for ultrahigh density optical recording

The possibility to optically image the very fine pit structure formed by electron beam (EB) writing has been researched using scanning near-field optical microscopy (SNOM). Fine test pit samples were formed in an electron resist (ZEP520) with a minimum pit size of 30 nm×160 nm. From experimental results using the pits, a conventional reflection SNOM could not image fine pit structures with a size of less than 100 nm. The technique was improved by coating the metal film on the optical probe and adopting an optical depolarization technique in the SNOM optics efficiently to detect near-field light reflected from the sample surface. We demonstrated that very fine pits with a minimum size of 30 nm were imaged and discussed that reflection type depolarization SNOM has a potential to achieve an ultrahigh density optical reading with 1.2 Tb/in. 2, which is limited by EB fabrication of the pits.

Labelling quality and chromosome morphology after low temperature FISH analysed by scanning far-field and near-field optical microscopy.

A non-enzymatic, low temperature fluorescence in situ hybridization (LTFISH) procedure was applied to metaphase spreads and interphase cell nuclei. In this context 'low temperature' means that the denaturation procedure of the chromosomal target DNA usually applied by heat treatment and chaotropic agents such as formamide was completely omitted so that the complete hybridization reaction took place at 37 degrees C. For LTFISH, the DNA probe had to be single-stranded, which was achieved by means of separate thermal denaturation of the DNA probe only. The DNA probe pUC1.77 was used for all LTFISH experiments. The labelling quality (number of binding sites, relative background intensity, relative intensity of major and minor binding sites) was analysed by confocal laser scanning microscopy (CLSM). An optimum in specificity and signal quality was obtained for 15 h hybridization time. For this hybridization condition of LTFISH, the chromosomal morphology was analysed by scanning near-field optical microscopy (SNOM). The results were compared with the morphology of chromosomes after (a) labelling of all centromeres using the same chemical treatment in the FISH procedure but with the application of target denaturation, and (b) labelling of all centromeres using a standard FISH protocol including thermal denaturation of the DNA probe and the chromosomal target. Depending on the FISH-procedure applied, SNOM images show substantial differences in the chromosome morphology. After LTFISH the chromosome morphology appeared to be much better preserved than after standard FISH. In contrast, the application of the LTFISH chemical treatment accompanied by heat denaturation had a very destructive influence on chromosomal morphology. The results indicate that, at least for certain DNA probes, specific chromosome labelling can be obtained without the usually applied heat and chemical denaturation of the DNA target, resulting in an apparently well preserved chromatin morphology as visualized by SNOM. LTFISH may be therefore a useful labelling technique whenever the chromosomal morphology had to be preserved after specific labelling of DNA regions. Binding mechanisms of single-stranded DNA probes to double-stranded DNA targets are discussed.

Near-field imaging of light propagation in photonic crystal waveguides: Explicit role of Bloch harmonics

We employ a collection scanning near-field optical microscope (SNOM) to image the propagation of light at telecommunication wavelengths along straight and bent regions of silicon-on-insulator photonic crystal waveguides (PCWs) formed by removing a single row of holes in the triangular 410-nm-period lattice along the $\ensuremath{\Gamma}M$ direction of the irreducible Brillouin zone. We obtain high quality SNOM images of PCWs excited in the wavelength range of 1520--1570 nm, which indicate good PCW mode confinement and low propagation loss. Using averaged cross sections of the intensity distributions before and after PCW bends, bend loss is evaluated and found to noticeably increase with the increase of the light wavelength from \ensuremath{\sim}1 dB at 1520 nm to \ensuremath{\sim}6 dB at 1570 nm. We analyze light intensity variations along PCWs measured with the SNOM at different distances from the sample surface. Considering the interference between a quasihomogeneous background field and Bloch harmonics of the PCW mode, we account for spatial frequency spectra of the intensity variations and determine the propagation constant of the PCW mode at 1520 nm. The possibilities and limitations of SNOM imaging for the characterization of PCWs are discussed.

Intermolecular coupling in nanometric domains of light-harvesting dendrimer films studied by photoluminescence near-field scanning optical microscopy (PL NSOM).

Near-field scanning optical microscopy (NSOM) has been used to investigate the photophysical characteristics of first- to fourth-generation (G1 to G4) light-harvesting dendrimer thin films containing coumarin-343 and coumarin-2 as the core and peripheral chromophores, respectively. Thin film photoluminescence (PL) spectra exhibit a significant red shift in the lower generations (G1, G2, and G3) as compared to their respective solution PL spectra, implying the formation of excimers. Spatially resolved PL NSOM images exhibit pronounced nanoscopic domains in G1, which become more homogeneous in higher generations due to site-isolation of the core chromophore. G4 exhibits complete site-isolation for these light-harvesting dendrimer films.

Rewritable organic films for near-field recording

Photochromic thin films were prepared for near-field recording. Acetyl substituted diarylethene were synthesized from 1,2-bis(2-methylbenzo[b]thiophene-3-yl)hexafluorocyclopentene in one step. Transparent and homogeneous thin films were coated on a substrate by vacuum deposition method. A colorless vacuum deposited diarylethene film turned to deep red hue upon exposure to a UV light. Near-field scanning optical microscopy (NSOM) was used to characterize nanoscale color change of the films. NSOM images showed distinct recording mark by 514 nm laser with mark speed of 30 ms. The records were completely erased upon excitation with a UV light, and rewritable with visible light (514 nm) on a UV activated colored film.

The Imaging of Small Domains of J-Aggregated Dye Molecules by Scanning Near-Field Optical Microscopy

The concept of high resolution imaging of a sample on a metal substrate by scanning near field optical microscopy (SNOM) with the tetrahedral tip is used for imaging of domains of oriented dye molecules on a metal substrate at a resolution in the order of 10 nm. The metal substrate is important for obtaining a high resolution in the SNOM images and at the same time it is expected to provide a mechanism for an increased contrast in SNOM imaging by scanning probe enhanced elastic scattering of molecules in the gap between a metal tip and a metal surface.

Imaging of photonic nanopatterns by scanning near-field optical microscopy

We define photonic nanopatterns of a sample as images recorded by scanning near-field optical microscopy with a locally excited electric dipole as a probe. This photonic nanopattern can be calculated by use of the Green’s dyadic technique. Here, we show that scanning near-field optical microscopy images of well-defined gold triangles taken with the tetrahedral tip as a probe show a close similarity to the photonic nanopattern of this nanostructure with an electric dipole at a distance of 15 nm to the sample and tilted 45° with respect to the scanning plane.