Keiko Tawa

Optical microscopic observation of fluorescence enhanced by grating-coupled surface plasmon resonance

On the substrate carrying a sub-wavelength grating covered with a thin metal layer, a fluorescent dye-labeled cell was observed by fluorescence microscope. The fluorescence intensity was more than 20 times greater than that on an optically flat glass substrate. Such a great fluorescence enhancement from labeled cells bound to the grating substrate was due to the excitation by grating coupled surface plasmon resonance. The application of a grating substrate to two-dimensional detection and fluorescence microscopy appears to offer a promising method of taking highly sensitive fluorescence images.

Sensitive Detection of a Tumor Marker, α-Fetoprotein, with a Sandwich Assay on a Plasmonic Chip

Two types of plasmonic silver- and gold-coated grating biosensor chips (plasmonic chip) were applied in the detection of α-fetoprotein (AFP) with a sandwich imunoassay and surface plasmon field-enhanced fluorescence. On the plasmonic chip, unlabeled marker in the sandwich immunoassay was first quantitatively detected over a wide range between 10(-12) and 10(-8) g/mL. The affinity constants between AFP and anti-AFP antibody, which were obtained by fitting the experimental data to the Langmuir isotherm adsorption curve, were 1 × 10(8) g(-1) mL regardless of the kind of metal in the plasmonic chips. Although the fluorescence intensity on the silver plasmonic chip was 5 times larger than that on the gold plasmonic chip, the limit of detection (LOD) was on the order of 10(-11) g/mL and not improved with a silver plasmonic chip. Herein, we used a new setup that generated less dispersions of both the fluorescence intensity for nonspecific adsorption and the background (optical blank) signal and improved the LOD of AFP to 4 pg/mL (55 fM) with the silver plasmonic chip. With the highly sensitive detection in the sandwich immunoassay, the development of a plasmonic chip for clinical diagnosis by a blood test is promising.

Application of 300× Enhanced Fluorescence on a Plasmonic Chip Modified with a Bispecific Antibody to a Sensitive Immunosensor

The grating substrate covered with a metal layer, a plasmonic chip, and a bispecific antibody can play a key role in the sensitive detection of a marker protein with an immunosensor, because of the provision of an enhanced fluorescence signal and the preparation of a sensor surface densely modified with capture antibody, respectively. In this study, one of the tumor markers, a soluble epidermal growth factor receptor (sEGFR), was selected as the target to be detected. The ZnO- and silver-coated plasmonic chip with precise regularity and the appropriate duty ratio in the periodic structure further enhanced the fluorescence intensity. As for sensor surface modification with capture antibody, a bispecific antibody (anti-sEGFR and anti-ZnO antibody), the concentrated bispecific antibody solution was found to nonlinearly form a surface densely immobilized with antibody, because the binding process of a bispecific antibody to the ZnO surface can be a competitive process with adsorption of phosphate. As a result, the interface on the plasmonic chip provided a 300× enhanced fluorescence signal compared with that on a ZnO-coated glass slide, and therefore sEGFR was found to be quantitatively detected in a wide concentration range from 10 nM to 700 fM on our plasmonic surface.

FEMTOSECOND OPTICAL KERR STUDY OF HEAVY-ATOM EFFECTS ON THE THIRD-ORDER OPTICAL NON-LINEARITY OF THIOPHENE HOMOLOGUES : ELECTRONIC HYPERPOLARIZABILITY OF TELLUROPHENE

Abstract Second hyperpolarizability γ of tellurophene (C 4 H 4 Te), a tellurium-containing homologue of thiophene, was measured by optically heterodyne-detected optical Kerr effect (OHD–OKE) experiments. We found that the γ of tellurophene was +6.3×10 −36 esu and was larger than twice of that for selenophene and almost three times that for thiophene. The large γ for tellurophene as the homologue was discussed in terms of the symmetry species of the two lowest excited states and their transition energies from the ground state under the 3-state approximation. Low-lying excited A 1 state is presented as the plausible cause of the large γ .

Synthesis and Nonlinear Properties of Poly[1,4‐bis(4‐methylpyridinium)butadiyne triflate)]

1,4-Bis(4-methylpyridinium)butadiyne triflate (1), which was prepared from the reaction of 1,4-bis(4-pyridyl)butadiyne with methyl trifluoromethanesulfonate, was found to be piled up along the c axis with two triflates as counter anions. The UV irradiation and thermal treatment of 1 gave rise to 1,4-topochemical polymerization to yield poly[1,4-bis(4-methylpyridinium) butadiyne triflate)]. The third-order nonlinear optical susceptibility (X (3) ) of the polymer was determined by the degenerate four wave mixing (DFWM) method to exhibit the remarkable high value of 2.2 x 10 -10 esu.

Duty ratio-dependent fluorescence enhancement through surface plasmon resonance in Ag-coated gratings

One-dimensional gratings with different duty ratios were designed and implemented for enhanced fluorescence detection and imaging. Verified by finite difference time domain simulations, our results showed that the enhancement strongly depended on the duty ratio of the land width to pitch of the grating structure. The maximum enhancement factor was achieved when the duty ratio was equal to 0.50 in our trapezoidal gratings with pitch=400 nm and depth=20 nm. Such a facile grating mold will exert a considerable influence on microarray biosensors and fluorescence microscopy.

Synthesis and nonlinear optical properties of 1,3- and 1,4-disubstituted type of poly(phenyleneethynylene)s containing electron-donor and acceptor group

The third-order nonlinear optical properties of 1,3-and 1,4-disituted poly (phenyleneethynylene)s containing electron-donor and accpetor groups were investigated in terms of the local polirization effect. The electron-donor and acceptor groups being set apart in the benzene ring adjacent to the triple bond in 1,4-disusbstituted poly (phenyleneethynylene)s was found to increase the X (3) value, together with the shift of the maximum absorption to longer wavelengths, while no significant effect was observed in the X (3) value for 1,3-disubstituted poly (phenyleneethynylene)s having both electron-donor and acceptor groups present in a benzene ring, in accordance with the fact that the maximum absorption remains at shorter wavelength.

In situ sensitive fluorescence imaging of neurons cultured on a plasmonic dish using fluorescence microscopy.

A plasmonic dish was fabricated as a novel cell-culture dish for in situ sensitive imaging applications, in which the cover glass of a glass-bottomed dish was replaced by a grating substrate coated with a film of silver. Neuronal cells were successfully cultured over a period of more than 2 weeks in the plasmonic dish. The fluorescence images of their cells including dendrites were simply observed in situ using a conventional fluorescence microscope. The fluorescence from neuronal cells growing along the dish surface was enhanced using the surface plasmon resonance field. Under an epi-fluorescence microscope and employing a donut-type pinhole, the fluorescence intensity of the neuron dendrites was found to be enhanced efficiently by an order of magnitude compared with that using a conventional glass-bottomed dish. In a transmitted-light fluorescence microscope, the surface-selective fluorescence image of a fine dendrite growing along the dish surface was observed; therefore, the spatial resolution was improved compared with the epi-fluorescence image of the identical dendrite.

Sensitive detection of interleukin-6 on a plasmonic chip by grating-coupled surface-plasmon-field-enhanced fluorescence imaging

A plasmonic chip, which is a grating substrate coated with metal films, was fabricated and applied to grating-coupled surface-plasmon-field-enhanced fluorescence imaging (GC-SPFI). With the enhanced fluorescence excited by the surface plasmon field on the plasmonic chip, interleukin-6 (IL-6), a marker protein of lifestyle-related diseases, was measured using a sandwich assay system. IL-6 was quantitatively determined to be up to 2 pg/mL by GC-SPFI. The detection sensitivity for IL-6 was superior to that with an enzyme-linked immunosorbent assay, and GC-SPFI was shown to be a valid method for immunosensing.

Photoinduced Anisotropy in a Polymer Doped with Azo Dyes in the Photostationary State Studied by Polarized FT-IR Spectroscopy

Photoinduced optical anisotropy in poly(methyl methacrylate) (PMMA) films doped with Disperse Orange 3 (DO3, NO2–C6H4–N=N–C6H4–NH2) was investigated by polarized FT-IR spectroscopy. Symmetric (NOs2) and antisymmetric (NOas2) stretching modes of the NO2 group and the C–N stretching mode of C–NH2(C–N) in the infrared absorption spectra of DO3 were found to be useful in order to evaluate the degree of orientation of DO3 molecules in the polymer matrix. Infrared dichroism was observed in the NOs2, NOas2, and C–N bands under the stationary irradiation by visible light. The orientation factors, K(n) Zf (f = x, y, z, n = trans, cis), determined from the observed dichroism of each isomer in the photostationary state indicate that the p-NO2–C6H4 group is less mobile than the p-NH2–C6H4 group in PMMA during trans-cis-trans isomerization. This dynamic behavior of the DO3 molecules can be interpreted in terms of restriction of rotation within the free volume of PMMA. This study shows that polarized FT-IR spectroscopy is one of the most powerful methods available for the analysis of the physical mechanisms of photoinduced anisotropy.