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This PDF file contains the front matter associated with SPIE
Proceedings Volume 7577, including the Title Page, Copyright
information, Table of Contents, and the Conference Committee listing.
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Within this contribution we convincingly demonstrate that the enhancement of the intrinsically weak Raman signals
through an interaction between an analyte molecule and enhanced electromagnetic fields in the vicinity of metallic
nanostructured surfaces is an extremely potent tool in bioanalytical science because such a SERS approach comprises
high sensitivity with molecular specificity. In particular innovative approaches to realize reproducible plasmonic
nanostructures i.e. SERS substrates like e.g. lithographically produced nanostructured gold surfaces or the defined
deposition of silver nanoparticles through an enzymatic reaction are introduced.
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Surface-enhanced Raman scattering (SERS) on gold nanohole and nanodisk arrays with precisely controlled size,
spacing and shapes of rod, circle, and triangle fabricated via focused ion beam was investigated by 632- and 785-nm
lasers. Results demonstrate that nanodisks and nanoholes exhibit strong SERS signals at 632-nm excitation, but only
triangular nanoholes present SERS signals at 785-nm excitation. The normal incidence transmission spectrum of
nanoholes shows the extension of near-infrared wavelength, but not for nanodisks. The broadband wavelength of
plasmon resonance and space confined by triangular nanoholes suggest promise for being a functional component in
biosensing, Raman spectroscopy, and photonic devices.
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The overall goal of this research is to develop a new point-of-care system for early detection and
characterization of cardiac markers to aid in diagnosis of acute coronary syndrome. The envisioned final technology
platform incorporates functionalized gold colloidal nanoparticles trapped at the entrance to a nanofluidic device
providing a robust means for analyte detection at trace levels using surface enhanced Raman spectroscopy (SERS).
To discriminate a specific biomarker, we designed an assay format analogous to a competitive ELISA. Notably, the
biomarker would be captured by an antibody and in turn displace a peptide fragment, containing the binding epitope
of the antibody labeled with a Raman reporter molecule that would not interfere with blood serum proteins. To
demonstrate the feasibility of this approach, we used C-reactive protein (CRP) as a surrogate biomarker. We
functionalized agarose beads with anti-CRP that were placed outside the nanochannel, then added either
Rhodamine-6-G (R6G) labeled-CRP and gold (as a surrogate of a sample without analyte present), or R6G labeled
CRP, gold, and unlabeled CRP (as a surrogate of a sample with analyte present). Analyzing the spectra we see an
increase in peak intensity in the presence of analyte at characteristic peaks for R6G specifically, 1284 and1567 cm-
1. Further, our results illustrate the reproducibility of the Raman spectra collected for R6G-labeled CRP in the
nanochannel. Overall, we believe that this method will provide the advantage of sensitivity and narrow line widths
characteristic of SERS as well as the specificity toward the biomarker of interest.
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The plasmonic nature of discontinuous thin films with micro-patterned structures such as triangles and hole arrays
present distinct optical properties with a Kretschmann surface plasmon resonance (SPR) instrument. Au microstructures
were prepared with a modified nanospheres lithography (NSL) method using 3.2 μm spheres which gives 1.8 μm
triangles and hole arrays with hole diameter ranging from 2.5 to 0.5 μm. The sensitivity to refractive index in thin film is
increased by up to 45% with using microhole arrays instead of continuous film. A transition in the microstructure aspect
from triangles to hole arrays with large hole diameter affects the spectral aspect of the SPR active band. Triangles
present a characteristic broad transmission maximum band while in hole arrays, a broad and weak absorption band first
appears for large holes, which sharpens and increases in intensity as the hole diameter decreases. Moreover, the SPR
penetration depth is tuned between 230 and 30 nm as the microstructure aspect shifts from continuous film, to small hole
arrays and to isolated triangles. Thus, these new plasmonic properties were observed in microhole arrays excited in
Kretschmann SPR configuration, which are spectrally similar to continuous film. These can significantly improve the
existing SPR sensing methods.
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The objective of this study was to develop rapid, inexpensive, and easily applied in vivo phenotyping strategies for
characterizing drug-metabolizing phenotypes with reference to the cytochrome P450 (CYP) enzymes in biological
fluids. Therefore, the accurate detection of low concentration of theophylline, which can be used as a probe for
cytochrome P450 (CYP450) enzymes (e.g. CYP1A2) activity, could benefit drug-metabolizing studies. In this study, a
portable, specific, and sensitive functionalized surface plasmon resonance (SPR) sensor using polyacrylamide
molecularly imprinted polymers (MIPs) as the highly specific selector is developed for the detection of low
concentration theophylline in the presence of other confounding components, such as, caffeine which has a very similar
chemical structure.
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Circular polarization interferometry configuration was used to develop surface plasmon based instrument, which
had two light beams with p- and s- polarization states individually within the common path. We used evanescent field to
determine the concentration of the biological sample via varying incident angles enabled phase interrogation. The
instrument named "OBMorph" includes a light source, an easy to use incident angle varying scheme based on a parabolic
and a spherical mirrors, and prism coupled sample stages. To increase the metrology sensitivity, which depends on
precisely control the angular resolution, a precision step-motor coupled with a parabolic mirror were used to control the
incident angle accurately. By using fault tolerance algorithm, the imperfect adjustment of circular polarization
interferometer was eliminated to obtain a perfect Lissajous curve needed for circular polarization interferometry. The
instrument developed was shown to have resolution as high as 4.92×10-6 RIU. The effect that refractive index of ITO thin
film changes with respect to externally applied voltage was also adopted by coating an ITO thin film onto biochips so as
to shift the surface plasmon resonance angle for larger phase interrogation ranges. We successfully measured CRP and
anti-CRP specific interaction in 0.75 μg/ml ~ 400 μg/ml ranges. In addition, the concentrations of tuberculosis inhibitor -
DHFR and compound Mg2P4O7 that can interact with CYP450 were also quantified successfully. The OBMorph was
shown to have potential applications in areas such as flat panel displays, optical coating, and drug delivery, to name a
few.
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Complementary metal oxide semiconductor (CMOS) cameras that can measure the phase and amplitude of periodically
modulated optical signals have been developed. These allow parallel lock-in imaging at up to 256 x 256 pixels resolution
without the need for slow and costly mechanical scanning. In conjunction with a differential surface plasmon resonance
(dSPR) system, spatially resolved SPR imaging has been achieved with sensitivities of better than 10 microRIU per pixel
per second. Results demonstrating the performance of modulated light cameras for dSPR imaging and high resolution
SPR microscopy are presented and discussed.
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Nanoapertures milled in opaque metallic films offer a simple and robust photonic tool to significantly enhance
the fluorescence of single molecules. We provide a detailed physical characterization of this phenomenon
for apertures milled in gold and aluminum, and discuss its application to biophotonics. For the first time,
the most general figures are provided to predict the awaited enhancement factors for almost every kind of
fluorescent molecule. This knowledge is essential to discuss the ability to detect low-quantum yield species.
We also report the first demonstration of single metal nanoapertures to perform DNA hybridization sensing,
and measure similar enhancement factors as for experiments on diffusing molecules.
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Most of the applications of fluorescence require the use of labeled drugs and labeled biomolecules. Due to the
need of labeling biomolecules with extrinsic fluorophores, there is a rapidly growing interest in methods which provide
label-free detection (LFD). Proteins are highly fluorescent, which is due primarily to tryptophan residues. However,
since most proteins contain tryptophan, this emission is not specific for proteins of interest in a biological sample. This is
one of the reasons of not utilizing intrinsic tryptophan emission from proteins to detect specific proteins. Here, we
present the intrinsic fluorescence for several proteins bound to the silver or aluminum metal nanostructured surfaces. We
demonstrate the metal enhanced fluorescence (MEF) of proteins with different numbers of tryptophan residues. Large
increases in fluorescence intensity and decreases in lifetime provide the means of direct detection of bound protein
without separation from the unbound. We present specific detection of individual types of proteins and measure the
binding kinetics of proteins such as IgG and streptavidin. Additionally, specific detection of IgG and streptavidin has
been accomplished in the presence of large concentrations of other proteins in sample solutions. These results will allow
design of surface-based assays with biorecognitive layer that specifically bind the protein of interest and thus enhance its
intrinsic fluorescence. The present study demonstrates the occurrence of MEF in the UV region and thus opens new
possibilities to study tryptophan-containing proteins without labeling with longer wavelength fluorophores and provides
an approach to label-free detection of biomolecules.
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A metallic nanoparticle can act as an optical antenna, enhancing the fluorescence of nearby fluorescent molecules. We
investigate the effects of a silica spacer layer on this fluorescence enhancement. Numerical simulations using the Finite-
Difference Time Domain (FDTD) method is performed to find the optimum size for the gold nanosphere core as well as
the optimum thickness of the silica shell. We present the dependence of radiative decay rate and quantum yield of the
fluorescent molecule with respect to varying silica layer thicknesses.
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We consider the possibility of using aluminum nanostructures for enhancing the intrinsic emission of biomolecules. We
used the finite-difference time-domain (FDTD) method to calculate the effects of aluminum nanoparticles on nearby
fluorophores that emit in the ultra-violet (UV). We find that the radiated power of UV fluorophores is significantly
increased when they are in close proximity to aluminum nanostructures. We show that there will be increased localized
excitation near aluminum particles at wavelengths used to excite intrinsic biomolecule emission. We also examine the
effect of excited-state fluorophores on the near-field around the nanoparticles. Finally we present experimental evidence
showing that a thin film of amino acids and nucleotides display enhanced emission when in close proximity to aluminum
nanostructured surfaces. Our results suggest that biomolecules can be detected and identified using aluminum
nanostructures that enhance their intrinsic emission. We hope this study will ignite interest in the broader scientific
community to take advantage of the plasmonic properties of aluminum and the potential benefits of its interaction with
biomolecules to generate momentum towards implementing fluorescence-based bioassays using their intrinsic emission.
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The study was performed on 16 CBA-line female mice with transplanted cervical cancer. 0.2 ml of gold
nanoparticle solution with a concentration of 109 particles/ml were injected into the animals intravenously. The particles
were 200-250 nm in size; the plasmon-resonance related extinction maximum was at the wavelength of 850-950 nm.
Accumulation of the nanoparticles into tumor node was visualized by the method of optical coherence tomography
(OCT). When the accumulation of nanoparticles in the tumor was maximal, hyperthermia was accomplished using the
LSP-AZOR laser setup generating cw radiation at 810 nm. The duration of exposition was 20 min. The therapeutical
effect was assessed by the rate of tumor growth inhibition (TGI, %).
Determining the instant when nanoparticle concentration in tumor tissue reaches its maximum enables more
efficient laser impact. The use of nanoparticles decreases laser irradiation power and ensures local action.
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This work describes the development and fabrication of a novel nanofluidic flow-through sensing chip that utilizes a
plasmonic resonator to excite fluorescent tags with sub-wavelength resolution. We cover the design of the microfluidic
chip and simulation of the plasmonic resonator using Finite Difference Time Domain (FDTD) software. The fabrication
methods are presented, with testing procedures and preliminary results.
This research is aimed at improving the resolution limits of the Direct Linear Analysis (DLA) technique developed by
US Genomics [1]. In DLA, intercalating dyes which tag a specific 8 base-pair sequence are inserted in a DNA sample.
This sample is pumped though a nano-fluidic channel, where it is stretched into a linear geometry and interrogated with
light which excites the fluorescent tags. The resulting sequence of optical pulses produces a characteristic "fingerprint"
of the sample which uniquely identifies any sample of DNA. Plasmonic confinement of light to a 100 nm wide metallic
nano-stripe enables resolution of a higher tag density compared to free space optics. Prototype devices have been
fabricated and are being tested with fluorophore solutions and tagged DNA. Preliminary results show evanescent
coupling to the plasmonic resonator is occurring with 0.1 micron resolution, however light scattering limits the S/N of
the detector. Two methods to reduce scattered light are presented: index matching and curved waveguides.
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We recently demonstrated that liposome-supported plasmon resonant gold nanoshells are degradable into
components of a size compatible with renal clearance, potentially enabling their use as multifunctional agents in
applications in nanomedicine, including imaging, diagnostics, therapy, and drug delivery (Troutman et al., Adv.
Mater. 2008, 20, 2604-2608). When illuminated with laser light at the wavelength matching their plasmon resonance
band, gold-coated liposomes rapidly release their encapsulated substances, which can include therapeutic and
diagnostic agents. The present research demonstrates that release of encapsulated agents from gold-coated liposomes
can be spectrally controlled by varying the location of the plasmon resonance band; this spectral tuning is
accomplished by varying the concentration of gold deposited on the surface of liposomes. Furthermore, the amount
of laser energy required for release is qualitatively explained using the concept of thermal confinement (Jacques,
Appl. Opt. 1993, 32(3), 2447-2454). Overlapping thermal confinement zones can be avoided by minimizing the laser
pulse width, resulting in lower energy requirements for liposomal content release and less global heating of the
sample. Control of heating is especially important in drug delivery applications, where it enables spatial and spectral
control of delivery and prevents thermal damage to tissue.
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Polarized light scattering is utilized to measure the optical anisotropy and the aspect ratio for nearly spherical colloidal
gold nanoparticles as well as to observe their rotational dynamics which were detected as fluctuations in the time-trace of
the scattering polarization. A comparison between the measured distributions of maximum anisotropy and aspect ratio
with those distributions calculated based on the TEM images analysis are found to show an excellent agreement
confirming the validity of our approach. Our method has the advantage to be simple, easy to implement, and can yield
access to different projections of the particle due to its rotational diffusion. The range of aspect ratios for the sample
being studied is 1 ~ 1.3 and another results for rods with aspect ratio of 2.4 is discussed. Gold nanoparticles have a good
biocompatibility where the possibility of conjugation to a variety of biomolecules and antibodies make them suitable for
optical imaging and optical probes which can be used for nanoscale orientaional sensing to monitor orientations and
rotations of biomolecules during their functional task.
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Surface Plasmon Resonnance (SPR) techniques have been mostly set-up as angular reflectivity
interrogation mode using quasi-monochromatic light or as spectral reflectivity interrogation mode at one given
wavelength, providing information about variation of effective optical thickness ▵n.e above the metal surface. In this
communication we present a dual mode sensor working both in angular and spectral interrogation modes. A white light
illuminates the sensor surface and the reflectivity spectra in TE and TM polarization are measured with a spectrometer.
By changing the angular coupling conditions, a complete reflectivity surface R(θ, λ) can be measured. The 2D
reflectivity decrease valley is affected by both the real and imaginary part of the optical index of the dielectric medium as
well as their spectral dispersion. With such experimental data set, it is possible to back calculate the dispersion of the
complex refractive index of the dielectric layer. This is demonstrated using a turquoise dye doped solution. According to
the Kramers-Kronig relations, the imaginary part of the refractive index for an absorbing medium is proportional to the
absorption while the real part presents a large dispersion around the absorption wavelength. The reflectivity surface R(θ, λ) was measured from 500 nm to 750 nm over about 8° angular range. The whole complex refractive optical index of the
doped solution, absorbing around 630 nm, was reconstructed from the SPR reflectivity experimental data, using a
homemade program based on an extended Rouard method to fit the experimental angular plasmon data for each
wavelength. These results show that the classical SPR technique can be extended to acquire precise spectral information
about biomolecular interactions occurring on the metallic layer.
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In this paper, we present experimental and numerical analysis on Extraordinary Optical Transmission (EOT) through
various nano-hole arrays in a thick metal film within the visible and near infra-red spectrum of light. Large nano-hole
arrays with different spacing between adjacent holes in the square lattice arrangement were fabricated using Electron
Beam Lithography (EBL). Optical transmission properties (wavelength, peak, and spectral bandwidth of transmission
resonances) of the fabricated nano-hole arrays were characterized and validated by numerical analysis based on Finite
Difference Time Domain (FDTD). Finally, the dependencies and discrepancies between EOT properties of various nanohole
arrays were analyzed.
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Nanoparticles such as gold and silver with plasmonic resonances in the near-infrared (NIR) optical region,
where soft tissue is the most transparent, are of great interest in biomedical applications. A major roadblock
in translation of inorganic nanoparticles to clinical practice for systemic targeting of disease is their nonbiodegradable
nature. In addition, gold nanoparticles that absorb in the NIR are typically greater than 50 nm,
which is above the threshold size of 5.5 nm required for effective excretion from the body. Here we describe
a new class of biodegradable gold nanoparticles with plasmon resonances in the NIR region. The synthesis is
based on controlled assembly of very small (less than 5 nm) primary gold particles into nanoclusters with
sub-100 nm overall diameter and an intense NIR absorbance. The assembly is mediated by biodegradable
polymers, polyethylene glycol (PEG) and polylactic acid (PLA) copolymer, and small capping ligands on the
constituent nanoparticles. Nanoclusters deaggregate into sub-5nm primary gold particles upon biodegradation
of the polymer binder in live cells over one week, as shown by dark-field reflectance and hyperspectral
imaging.
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Metal nanostructures are known to amplify the spontaneous emission of fluorescent molecules by resonant coupling to external electromagnetic fields. We have used spectroscopy to characterize the structural properties of emodin molecules, a natural anthraquinone dye, and bovine serum albumin, the most abundant protein in plasma, in the presence of silver nanoparticles. Aggregation of emodin at pH=10 and pH=6 gives rise to SERS and MEF effects in silver colloid. We have obtained MEF spectra at acidic pH=2.9 using two different silver nanostructures. We have also studied the change in the secondary structure of bovine serum albumin adsorbed on metal nanoparticles surface. Circular dichroism, fluorescence emission and fluorescence lifetime measurements indicate an increase in the alfa-helical content of the protein and a change in the environment of the tryptophan residues that bury in the interior of the biomolecule. This variation on the secondary structure could have further influence in the binding of the drug to form transport and regulatory complexes.
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We describe the synthesis of novel hybrid nanoparticles composed of a gold nanorods core and a bovine serum albuminconjugated
porous silica shell. The pores of the porous silica shell are loaded with the cytotoxic surfactant cetrimonium
bromide. Throughout the various steps of the synthesis, these nanoparticles exhibit excellent light extinction in a near
infrared window around 700 - 900 nm, and good colloidal stability in aqueous environment. On replacement of the
bovine serum albumin and cetrimonium bromide with functional macromolecules (e.g. antibodies) and appropriate drugs,
this architecture may find application in a variety of biomedical applications including optical and photoacoustic
diagnoses and imaging, photothermal and photoacoustic therapies and controlled drug release.
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We provide novel insight into the photothermal stability of different solutions of gold nanorods embedded in a semi-solid
PVA environment, which may produce different effects with respect e.g. to the aqueous colloidal suspension. We
investigate gold nanorods with different average dimensions and surface modifications (cetrimonium bromide (CTAB)
and porous silica (p-silica)). Under near-infrared laser pulses between 20 ms and 20 s, the effective photothermal
stability is higher in large than in small nanoparticles and in CTAB-protected than in p-silica-modified nanoparticles. At
least up to ~200 °C there occurs only partial deformation of the gold nanorods with negligible formation of stable
byproducts. We understand these results as the interplay of different factors, including the protection exerted from the
semi-solid environment and the aggregation of the nanoparticles.
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A novel plasmonic Raman sensor using periodic nano-hole and, potentially, nanofocusing arrays is investigated
numerically and experimentally. The effect of structural parameters (such as periodicity of the structure, hole
dimensions, etc.) is determined and investigated. The analysed structures are fabricated in thin gold films by means of
focused ion beam lithography. Optical characteristics of the fabricated arrays are determined experimentally and
compared with the theoretical predictions. Experimental field enhancements are determined and also compared with the
theoretical predictions.
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In this study, we explore the relation between near-field and far-field characteristics of plasmon-enhanced total internal
reflection fluorescence (TIRF) imaging. It was found that a significant disparity exists between near-field intensity
maximum of evanescent fields and reflectance minimum in the far-field. The disparity tends to be larger when plasmons
are localized by surface nanostructures. Experimental data with nanogratings and nanoislands as well as theoretical
results are presented. The disparity can be considered to optimize plasmon-enhanced TIRF microscopy.
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