Diffraction patterns observed in surface plasmon resonance imaging (SPRI) microscopy measurements of single gold nanorods (AuNRs) exhibit a complex behavior at wavelengths near the longitudinal plasmonic resonance band. SPRI microscopy measurements at 814 nm from AuNRs in three samples with resonance extinction maxima at 670, 816, and 980 nm reveal a variety of diffraction patterns with central peaks that are either positive, negative, or biphasic. A unitless ratio parameter MR (-1 ≤ MR ≤ 1) is created to describe the distribution of diffraction patterns. A purely negative (MR = -1) central peak is observed for 30%, 57%, and 98% of the diffraction patterns in the 670, 816, and 980 nm samples, respectively. These results along with a theoretical modeling of the diffraction patterns with an anisotropic complex scattering coefficient suggests that this behavior only occurs for AuNRs when the laser wavelength used in SPRI experiments is shorter than the AuNR plasmonic resonance maxima, that is, in the anomalous dispersion region.

N-Isopropylacrylamide-based hydrogel nanoparticles (HNPs) that incorporate 30 mer single-stranded DNA (ssDNA) oligonucleotides were synthesized and characterized with single-nanoparticle surface plasmon resonance imaging (SPRI) microscopy, dynamic light scattering, transmission electron microscopy, and fluorescence measurements. The synthesized HNPs had an averaged diameter of 230 nm and exhibited a large (5-10x) increase in the average single-nanoparticle SPRI refractive index (Delta% R-NP), as compared with HNPs without DNA. A combination of SPRI and fluorescence measurements was used to measure the uptake of approximately 20 000 complementary ssDNA into each HNP, resulting in a Langmuir isotherm adsorption coefficient of 4.89 X 10(8) M-1. Single-nanoparticle SPRI measurements also showed that approximately 35% of the incorporated ssDNA was accessible for both hybridization uptake and enzymatic hydrolysis using exonuclease I. We attribute the presence of an inactive ssDNA population in the nanoparticle to a combination of acrylamide Michael addition reactions with adenine, cytosine, and guanine nucleotides, as well as the possible formation of self-complementary secondary structures in the polymerized ssDNA.

We investigated sequence-specific and simultaneous microRNA (miRNA) detections by surface plasmon resonance (SPR) imaging measurements on SPR chips possessing an Au spot array modified with probe DNAs based on a miRNA-detection-selective SPR signal amplification method. MiRNAs were detected with the detection limit of the attomole level by SPR imaging measurements for different miRNA concentrations on a single chip. SPR signals were enhanced based on a combination process of sequence-specific hybridization of the miRNA to the probe DNAs, extension reaction of polyadenine (poly(A)) tails by poly(A) polymerase, binding of a ternary complex of T30-biotin/horseradish peroxidase (HRP)-biotin/streptavidin to the poly(A) tails, and the oxidation reaction of tetramethylbenzidine (TMB) on the HRP by providing a blue precipitate on the surface. This process sequence-specifically and dramatically amplified the SPR signals. This is a simple, cost-effective, and feasible signal amplification method based on the organic compound TMB instead of metal nanoparticles.

To quantitatively characterize the optical coupling between ZnO (a high refractive index but nonabsorptive material) and an electrochromic material (either tungsten oxide (WO3) or poly(3,4-ethylenedioxythiophene) (PEDOT)), electrodiffractive and electrochromic properties were investigated from a novel two-component composite nanostructured electrodeposited grating that incorporated both ZnO and either WO3 or PEDOT. The sequential electrodeposition of these two materials through a photopatterned photoresist layer onto a fluorine-doped tin oxide (FTO)-coated glass substrate created a grating structure that exhibited optical diffraction that could be modulated electrochemically. SEM images of the electrochromic gratings revealed that the ZnO electrodeposition created a high surface area nanostructured thin film. The nanostructured ZnO is nonabsorbing, but its high refractive index and large surface area redirect light into the electrochromic grating and create a 4-fold enhancement in the grating's electrodiffractive response.

We describe the development and performance of a new type of optical sensor suitable for registering the binding/dissociation of nanoscopic particles near a gold sensing surface. The method shares similarities with surface plasmon resonance microscopy but uses a completely different optical signature for reading out binding events. This new optical read-out mechanism, which we call confined optical field enhanced fluorescence emission (Cofefe), uses pulsed surface plasmon polariton fields at the gold/liquid interface that give rise to confined optical fields upon binding of the target particle to the gold surface. The confined near-fields are sufficient to induce two-photon absorption in the gold sensor surface near the binding site. Subsequent radiative recombination of the electron-hole pairs in the gold produces fluorescence emission, which can be captured by a camera in the far-field. Bound nanoparticles show up as bright confined spots against a dark background on the camera. We show that the Cofefe sensor is capable of detecting gold and silicon nanoparticles, as well as polymer nanospheres and sub-μm lipid droplets in a label-free manner with average illumination powers of less than 10 μW/μm2.

Novel broadband ultra-antireflective surfaces were created via the electrodeposition of a nanostructured zinc oxide thin film onto conductive, light absorbing periodic nanocone arrays. Nanocone arrays of (i) fluorinated ethylene propylene (FEP) coated with a 50 nm plasmonic gold thin film and (ii) the electroactive polymer poly(3,4-ethylenedioxythiophene) (PEDOT) exhibited a very low broadband reflectivity of less than 0.1% from 475 to 800 nm at a wide range of incident angles after the electrodeposition of a nanostructured ZnO thin film onto the surface. SEM images reveal the formation of ZnO nanoflowers and nanorods on both nanocone array surfaces; these additional ZnO nanostructures enhance the coupling of the incident visible light into the absorptive gold or PEDOT nanocones to significantly reduce the reflectivity of these surfaces. The ZnO-coated nanocone array surfaces also exhibited an enhanced photoreactivity for the oxidative degradation of methylene blue, suggesting their potential to be used as a self-cleaning antireflective surface.

Near-infrared surface plasmon resonance imaging (SPRI) microscopy is used to detect and characterize the adsorption of single polymeric and protein nanoparticles (PPNPs) onto chemically modified gold thin films in real time. The single-nanoparticle SPRI responses, Δ%RNP, from several hundred adsorbed nanoparticles are collected in a single SPRI adsorption measurement. Analysis of Δ%RNP frequency distribution histograms is used to provide information on the size, material content, and interparticle interactions of the PPNPs. Examples include the measurement of log-normal Δ%RNP distributions for mixtures of polystyrene nanoparticles, the quantitation of bioaffinity uptake into and aggregation of porous NIPAm-based (N-isopropylacrylamide) hydrogel nanoparticles specifically engineered to bind peptides and proteins, and the characterization of the negative single-nanoparticle SPRI response and log-normal Δ%RNP distributions obtained for three different types of genetically encoded gas-filled protein nanostructures derived from bacteria.

Ordered nanocone arrays of the electroactive polymer poly(3,4-ethylenedioxythiophene) (PEDOT) were fabricated by the simultaneous oxygen plasma etching of an electrodeposited PEDOT thin film coated with a hexagonally closed packed polystyrene bead monolayer. PEDOT nanocone arrays with an intercone spacing of 200 nm and an average nanocone height of 350 nm exhibited a low broadband reflectivity of <1.5% from 550 to 800 nm. Electrochemical modulation of the oxidation state of the PEDOT nanocone array film was used to change both its ex situ absorption spectrum (electrochromism) and reflection spectrum (electroreflectivity). The sign of the PEDOT nanocone array electroreflectivity was opposite to that observed from unmodified PEDOT thin films; this significant difference is attributed to the unique optical behavior of nanostructured surfaces with an interfacial layer that contains a graded mix of air and highly absorptive nanocones. The combined electrochromic and electroreflective behavior of the antireflective PEDOT nanocone array films should find promising applications in solar energy cells, sensors and other optical devices.

The specific binding and uptake of protein molecules to individual hydrogel nanoparticles is measured with real-time single-nanoparticle surface plasmon resonance imaging (SPRI) microscopy. Nanoparticles that adsorb onto chemically modified gold thin films interact with traveling surface plasmon polaritons and create individual point diffraction patterns in the SPRI microscopy differential reflectivity images. The intensity of each point diffraction pattern depends on the integrated refractive index of the nanoparticle; an increase in this single nanoparticle point diffraction intensity (Delta%R-NP) is observed for nanoparticles that bind proteins. SPRI adsorption measurements can be used to measure an average increase in Delta%R-NP that can be correlated with bulk dynamic light scattering measurements. Moreover, the distribution of Delta%R-NP values observed for individual nanoparticles can be used to learn more about the nature of the protein-nanoparticle interaction. As a first example, the binding of the lectin Concanavalin A to 180 nm N-Isopropylacrylamide hydrogel nanoparticles that incorporate a small percentage of mannose sugar monomer units is characterized.

A two-step templated, ribosomal biosynthesis/printing method for the fabrication of protein microarrays for surface plasmon resonance imaging (SPRI) measurements is demonstrated. In the first step, a sixteen component microarray of proteins is created in microwells by cell free on chip protein synthesis; each microwell contains both an in vitro transcription and translation (IVTT) solution and 350 femtomoles of a specific DNA template sequence that together are used to create approximately 40 picomoles of a specific hexahistidine-tagged protein. In the second step, the protein microwell array is used to contact print one or more protein microarrays onto nitrilotriacetic acid (NTA)-functionalized gold thin film SPRI chips for real-time SPRI surface bioaffinity adsorption measurements. Even though each microwell array element only contains approximately 40 picomoles of protein, the concentration is sufficiently high for the efficient bioaffinity adsorption and capture of the approximately 100 femtomoles of hexahistidine-tagged protein required to create each SPRI microarray element. As a first example, the protein biosynthesis process is verified with fluorescence imaging measurements of a microwell array containing His-tagged green fluorescent protein (GFP), yellow fluorescent protein (YFP) and mCherry (RFP), and then the fidelity of SPRI chips printed from this protein microwell array is ascertained by measuring the real-time adsorption of various antibodies specific to these three structurally related proteins. This greatly simplified two-step synthesis/printing fabrication methodology eliminates most of the handling, purification and processing steps normally required in the synthesis of multiple protein probes, and enables the rapid fabrication of SPRI protein microarrays from DNA templates for the study of protein-protein bioaffinity interactions.

Liposomes are small vesicles that can be used in various targeting applications as carrier vehicles. In this paper, we show that real-time surface plasmon resonance imaging microscopy (SPRI microscopy) can be used to detect diffraction patterns of these singular vesicles in water phase at room temperature and without any additives. The diffraction pattern intensities, related to the particle size, are shown to follow the log-normal distribution in a cumulative distribution function (CDF) that is very well in accordance with the normal size distribution of liposomes prepared with the extrusion method. In addition, this distribution is further analyzed to determine the number of gold nanoparticle (GNP) encapsulated liposomes in a set of liposomal adsorption events. Thus, we obtain the encapsulation efficiency and present a method to study the intrinsic properties of liposomes and other soft nanomaterials.