Single molecule tracking on supported membranes with arrays of optical nanoantennas

Coupling of the localized surface plasmons between two closely apposed gold nanoparticles (nanoantenna) can cause strong enhancements of fluorescence or Raman signal intensity from molecules in the plasmonic "hot-spot". Harnessing these properties for practical applications is challenging due to the need to fabricate gold particle arrays with well-defined nanometer spacing and a means of delivering functional molecules to the hot-spot. We report fabrication of billions of plasmon-coupled nanostructures on a single substrate by a combination of colloid lithography and plasma processing. Controlled spacing of the nanoantenna gaps is achieved by taking advantage of the fact that polystyrene particles melt together at their contact point during plasma processing. The resulting polymer thread shadows a gap of well-defined spacing between each pair of gold triangles in the final array. Confocal surface-enhanced Raman spectroscopy imaging confirms the array is functionally uniform. Furthermore, a fully intact supported membrane can be formed on the intervening substrate by vesicle fusion. Trajectories of freely diffusing individual proteins are traced as they sequentially pass through, and are enhanced by, multiple gaps. The nanoantenna array thus enables enhanced observation of a fluid membrane system without static entrapment of the molecules.     

Micropatterned fluid lipid bilayer arrays created using a continuous flow microspotter

We have developed a new method for creating micropatterned lipid bilayer arrays (MLBAs) using a 3D microfluidic system. An array of fluid lipid membranes was patterned onto a glass substrate using a Continuous Flow Microspotter. Fluorescence microscopy experiments were used to verify the formation of a bilayer structure on the glass substrate. Fluorescence recovery after photobleaching experiments demonstrated the bilayers' fluidity was maintained while being individually corralled on the substrate. The reproducibility of bilayer formation within an array was demonstrated by the linear response of membrane fluorescence versus mol % rhodamine functionalized lipids incorporated into the vesicles prior to fusion to the surface. The highly customizable nature of the MLBAs was demonstrated utilizing three different fluorescently labeled lipids to generate a multiple component lipid array. Finally, the cholera toxin B/ganglioside GM 1, antidinitrophenyl (DNP) antibody/DNP, and NeutrAvidin/biotin protein-ligand systems were used to model multiple protein-ligand binding on the MLBAs. The multicomponent patterned bilayers were functionalized with GM 1, DNP, and biotin lipids, and binding curves was generated by recording surface fluorescence versus increasing concentration of membrane bound ligands.     

Metallic photonic crystals based on solution-processible gold nanoparticles

We demonstrate the fabrication of metallic photonic crystals, in the form of a periodic array of gold nanowires on a waveguide, by spin-coating a colloidal gold suspension onto a photoresist mask and subsequent annealing. The photoresist mask with a period below 500 nm is manufactured by interference lithography on an indium tin oxide (ITO) glass substrate, where the ITO layer has a thickness around 210 nm and acts as the waveguide. The width of the nanowires can be controlled from 100 to 300 nm by changing the duty cycle of the mask. During evaporation of solvent, the gold nanoparticles are drawn to the grooves of the grating with apparently complete dewetting off the photoresist for channels less than 2 microm in width, which therefore form nanowires after the annealing process. Strong coupling between the waveguide mode and the plasmon resonance of the nanowires, which is dependent on the polarization and incidence angle of the light wave, is demonstrated by optical extinction measurements. Continuity of the nanowires is confirmed by conductivity properties. Simplicity, high processing speed, and low cost are the main advantages of this method, which may have a plethora of applications in telecommunication, all-optical switching, sensors, and semiconductor devices.     

Manipulation of extinction spectra of P3HT/PMMA medium arrays on silicon substrate containing self-assembled gold nanoparticles

In this study, we report a simple novel approach to modulate the extinction spectra of P3HT/PMMA by manipulating the medium arrays on a substrate that is coated with self-assembled gold nanoparticles. The 20 nm gold nanoparticles were synthesized and then self-assembled on the APTMS/silicon substrate surface by immersing the substrate into the gold colloid suspension. A high-resolution P3HT/PMMA photoluminescent electron beam resist was used to fabricate various square hole arrays on the substrate containing gold nanoparticles. The P3HT/PMMA medium composition causes the blue shifts in the extinction peaks of up to 40.6 nm by decreasing the period from 500 nm to 200 nm for P3HT/PMMA square hole arrays with a diameter of 100 nm. The magnitude of blue shift is directly proportional to the product of the changes of medium refractive index and the array structure factor. These peak shifts and intensity of extinction spectra for various P3HT/PMMA medium arrays are well described by the finite-difference time-domain (FDTD) simulation results. Since this simple cost-effective technique can tune the extinction spectrum of medium and adding the gold nanoparticles can give more functionalities for sensing applications, such as surface-enhanced Raman scattering (SERS), that provides good opportunities for the design and fabrication of new optoelectronic devices and sensors.     

Wafer-scale fabrication of nanofluidic arrays and networks using nanoimprint lithography and lithographically patterned nanowire electrodeposition gold nanowire masters

Wafer scale (cm(2)) arrays and networks of nanochannels were created in polydimethylsiloxane (PDMS) from a surface pattern of electrodeposited gold nanowires in a master-replica process and characterized with scanning electron microscopy (SEM), atomic force microscopy (AFM), and fluorescence imaging measurements. Patterns of gold nanowires with cross-sectional dimensions as small as 50 nm in height and 100 nm in width were prepared on silica substrates using the process of lithographically patterned nanowire electrodeposition (LPNE). These nanowire patterns were then employed as masters for the fabrication of inverse replica nanochannels in a special formulation of PDMS. SEM and AFM measurements verified a linear correlation between the widths and heights of the nanowires and nanochannels over a range of 50 to 500 nm. The PDMS replica was then oxygen plasma-bonded to a glass substrate in order to create a linear array of nanofluidic channels (up to 1 mm in length) filled with solutions of either fluorescent dye or 20 nm diameter fluorescent polymer nanoparticles. Nanochannel continuity and a 99% fill success rate was determined from the fluorescence imaging measurements, and the electrophoretic injection of both dye and nanoparticles in the nanochannel arrays was also demonstrated. Employing a double LPNE fabrication method, this master-replica process was also used to create a large two-dimensional network of crossed nanofluidic channels.     

Physicochemical Properties and Route of Systemic Delivery Control the In Vivo Dynamics and Breakdown of Radiolabeled Gold Nanostars

The in vivo dynamics of nanoparticles requires a mechanistic understanding of multiple factors. Here, for the first time, the surprising breakdown of functionalized gold nanostars (F-AuNSs) conjugated with antibodies and ⁶⁴Cu radiolabels in vivo and in artificial lysosomal fluid ex vivo, is shown. The short-term biodistribution of F-AuNSs is driven by the route of systemic delivery (intravenous vs intraperitoneal) and long-term fate is controlled by the tissue type in vivo. In vitro studies including endocytosis pathways, intracellular trafficking, and opsonization, are combined with in vivo studies integrating a milieu of spectroscopy and microcopy techniques that show F-AuNSs dynamics is driven by their physicochemical properties and route of delivery. F-AuNSs break down into sub-20 nm broken nanoparticles as early as 7 days postinjection. Martini coarse-grained simulations are performed to support the in vivo findings. Simulations suggest that shape, size, and charge of the broken nanoparticles, and composition of the lipid membrane depicting various tissues govern the interaction of the nanoparticles with the membrane, and the rate of translocation across the membrane to ultimately enable tissue clearance. The fundamental study addresses critical gaps in the knowledge regarding the fate of nanoparticles in vivo that remain a bottleneck in their clinical translation.     

Effect of near-field coupling on far-field inelastic scattering imaging of gold nanoparticles

The optical near-field enhancement induced by coupling between noble nanoparticles and the substrate has been studied by a far-field imaging method. The longitudinal mode of the incident laser is revealed to contribute to the coupling. The far-field images of individual gold nanoparticles exhibit a peanut-shaped pattern; these were constructed by the intensity of inelastically scattered light. The coupling between gold nanoparticles and the silicon substrate leads to the patterned image. By tuning the separation between the gold nanoparticles and substrate using SiO(2) layers of different thickness, the coupling efficiency decreases with the thickness of the SiO(2) layer.     

苯乙烯-马来酸酐共聚物/金纳米微粒的合成

以氯金酸为原料,DMF(N,N-二甲基甲酰胺)为溶剂及还原剂,苯乙烯-马来酸酐共聚物为大分子稳定剂,合成了金纳米微粒。通过紫外-可见吸收光谱、透射电子显微镜等方法对纳米金样品进行了表征。结果表明:所得到的金纳米微粒可以在520 nm~530 nm范围内产生明显的纳米金所具有的特征等离子共振吸收峰,金纳米微粒的尺寸在3 nm~5nm且具有较窄的分布,证明苯乙烯-马来酸酐共聚物可以对金纳米微粒表面产生较好的修饰作用,从而为制备纳米金材料提供了一种新的途径。     

Characterizing the Lateral Friction of Nanoparticles on On‐Chip Integrated Black Lipid Membranes

The use of nanoparticles (NPs) in biomedical applications creates a need for appropriate model systems to systematically investigate NP–membrane interactions under well‐defined conditions. Black lipid membranes (BLMs) are free‐floating membranes with defined composition that are ideally suited for characterizing NP–membrane interactions free of any potential perturbation through a supporting substrate. Herein, arrays of microfabricated BLMs are integrated into a chip‐based platform that is compatible with high‐speed optical NP tracking. This system is used to investigate the lateral diffusion of 40 nm gold spheres tethered to biotinylated lipids through antibody‐functionalized ligands (single‐stranded DNA or polyethylene glycol). Although the NPs show an almost free and ergodic diffusion, their lateral motion is subject to substantial drag at the membrane surface, which leads to systematically smaller diffusion coefficients than those obtained for lipids in the membrane through fluorescence recovery after photobleaching. The lateral mobility of the NPs is influenced by the chemical composition and salt concentration at the NP‐membrane interface, but is independent of the ligand density in the membrane. Together with the observation that nanoprisms, which have a larger relative contact area with the membrane than spherical NPs, show an even slower diffusion, these findings indicate that the lateral mobility of NPs tethered in close vicinity to a membrane is significantly reduced by the friction at the NP‐membrane interface.     

Direct writing of large-area plasmonic photonic crystals using single-shot interference ablation

We report direct writing of metallic photonic crystals (MPCs) through a single-shot exposure of a thin film of colloidal gold nanoparticles to the interference pattern of a single UV laser pulse before a subsequent annealing process. This is defined as interference ablation, where the colloidal gold nanoparticles illuminated by the bright interference fringes are removed instantly within a timescale of about 6 ns, which is actually the pulse length of the UV laser, whereas the gold nanoparticles located within the dark interference fringes remain on the substrate and form grating structures. This kind of ablation has been proven to have a high spatial resolution and thus enables successful fabrication of waveguided MPC structures with the optical response in the visible spectral range. The subsequent annealing process transforms the grating structures consisting of ligand-covered gold nanoparticles into plasmonic MPCs. The annealing temperature is optimized to a range from 250 to 300?°C to produce MPCs of gold nanowires with a period of 300 nm and an effective area of 5 mm in diameter. If the sample of the spin-coated gold nanoparticles is rotated by 90° after the first exposure, true two-dimensional plasmonic MPCs are produced through a second exposure to the interference pattern. Strong plasmonic resonance and its coupling with the photonic modes of the waveguided MPCs verifies the success of this new fabrication technique. This is the simplest and most efficient technique so far for the construction of large-area MPC devices, which enables true mass fabrication of plasmonic devices with high reproducibility and high success rate.     

Corona Generation and Deposition of Metal Nanoparticles on Conductive Surfaces and Their Effects on the Substrate Surface Texture and Chemistry

A corona discharge ion bombardment technique was used successfully to generate gold particles of submicron diameters. In a negative corona discharge, the glow region contains electrons, negative ions, and positive ions. Positive ions collided with the negative corona tip electrode, causing it to sputter and emit fine particles of the electrode material. These nanoparticles were deposited on grounded metal substrates or thin mica sheets supported by grounded metal substrates. Scanning electron microscopy (SEM) and atomic force microscopy (AFM) were used to study the size distribution and deposition pattern of the metal nanoparticles. The diameter of these nanoparticles was dependent upon the material of the electrode and ranged from 20 to 450 nm for gold and from 15 to 240 nm for tungsten. The nanoparticles were deposited on aluminum, mica, and carbon steel test panels for different amounts of time. The electrochemical response of the carbon steel panels exposed to aerated salt solution was measured by direct current (DC) polarization technique before and after the gold nanoparticles were deposited. This technique was employed to determine the changes in the surface chemistry because of the presence of gold nanoparticles, and it proved to be a sensible method for detecting the presence of fine layers of nanoparticles on the metallic substrate. The presence of the gold nanoparticles increased the electrochemical potential E_corr from -0.640 V to -0.211 V, compared with the value for a noncorrosive surface, like that of pure gold, which is 0 V.     

Tunable subradiant lattice plasmons by out-of-plane dipolar interactions

Plasmonic nanostructures concentrate optical fields into nanoscale volumes, which is useful for plasmonic nanolasers, surface enhanced Raman spectroscopy and white-light generation. However, the short lifetimes of the emissive plasmons correspond to a rapid depletion of the plasmon energy, preventing further enhancement of local optical fields. Dark (subradiant) plasmons have longer lifetimes, but their resonant wavelengths cannot be tuned over a broad wavelength range without changing the overall geometry of the nanostructures. Also, fabrication of the nanostructures cannot be readily scaled because their complex shapes have subwavelength dimensions. Here, we report a new type of subradiant plasmon with a narrow (～5 nm) resonant linewidth that can be easily tuned by changing the height of large (>100 nm) gold nanoparticles arranged in a two-dimensional array. At resonance, strong coupling between out-of-plane nanoparticle dipolar moments suppresses radiative decay, trapping light in the plane of the array and strongly localizing optical fields on each nanoparticle. This new mechanism can open up applications for subradiant plasmons because height-controlled nanoparticle arrays can be manufactured over wafer-scale areas on a variety of substrates.     

Synthesis of gold nanoclusters: a fluorescent marker for water-soluble TiO2 nanotubes

The first example of a water-soluble wrapped titania nanotube (TNT) decorated with fluorescent gold nanoparticles has been prepared. Gold nanoparticles ～ 1.6 nm in diameter were grown on the TiO(2) nanotubes using a thiolactic acid linker to control the size. The gold clusters emit at 660 nm in water and were imaged using confocal microscopy. The gold decorated TNTs were suspended in water by wrapping the nanotubes with poly-L-arginine.     

Engineering Functional Membrane–Membrane Interfaces by InterSpy

Engineering synthetic interfaces between membranes has potential applications in designing non-native cellular communication pathways and creating synthetic tissues. Here, InterSpy is introduced as a synthetic biology tool consisting of a heterodimeric protein engineered to form and maintain membrane–membrane interfaces between apposing synthetic as well as cell membranes through the SpyTag/SpyCatcher interaction. The inclusion of split fluorescent protein fragments in InterSpy allows tracking of the formation of a membrane–membrane interface and reconstitution of functional fluorescent protein in the space between apposing membranes. First, InterSpy is demonstrated by testing split protein designs using a mammalian cell-free expression (CFE) system. By utilizing co-translational helix insertion, cell-free synthesized InterSpy fragments are incorporated into the membrane of liposomes and supported lipid bilayers with the desired topology. Functional reconstitution of split fluorescent protein between the membranes is strictly dependent on SpyTag/SpyCatcher. Finally, InterSpy is demonstrated in mammalian cells by detecting fluorescence reconstitution of split protein at the membrane–membrane interface between two cells each expressing a component of InterSpy. InterSpy demonstrates the power of CFE systems in the functional reconstitution of synthetic membrane interfaces via proximity-inducing proteins. This technology may also prove useful where cell-cell contacts and communication are recreated in a controlled manner using minimal components.     

Liquid-crystal micropolarizer array for polarization-difference imaging

Fabrication and applications are discussed for a visible-wavelength micropolarizer array consisting of a linear polarizer and a micropatterned liquid-crystal (LC) cell. LC alignment direction is controlled by means of depositing an optically transparent gold film at an oblique angle and coating the surface with an alkanethiol self-assembled monolayer. Microdomains of two perpendicular LC alignment directions are created by photolithography and etching of the gold layer, rotating the substrate 90 deg, and depositing a second oblique gold layer in the etched areas. The resulting array is used for polarization-difference imaging (PDI), a technique that enhances image contrast in the presence of scattering. Images obtained with the array require more processing than do conventional PDI images, but this method eliminates the need for an electronically activated LC filter and is especially suited to systems whose filters are closely integrated with optical sensor arrays.     

Detection and identification of proteins using nanoparticle-fluorescent polymer 'chemical nose' sensors

A sensor array containing six non-covalent gold nanoparticle-fluorescent polymer conjugates has been created to detect, identify and quantify protein targets. The polymer fluorescence is quenched by gold nanoparticles; the presence of proteins disrupts the nanoparticle-polymer interaction, producing distinct fluorescence response patterns. These patterns are highly repeatable and are characteristic for individual proteins at nanomolar concentrations, and can be quantitatively differentiated by linear discriminant analysis (LDA). Based on a training matrix generated at protein concentrations of an identical ultraviolet absorbance at 280 nm (A280 = 0.005), LDA, combined with ultraviolet measurements, has been successfully used to identify 52 unknown protein samples (seven different proteins) with an accuracy of 94.2%. This work demonstrates the construction of novel nanomaterial-based protein detector arrays with potential applications in medical diagnostics.     

Characterization of different sequences of DNA on si substrate by atomic force microscopy and gold nanoparticle labeling

In this paper, different sequences of single-strand DNA modified on Si substrate were studied taking advantages of the high resolution of atomic force microscopy (AFM) and signal enhancement of gold nanoparticles. Two sequences of single-strand DNA, as a model, were immobilized on Si substrate and hybridized with their sequence-complementary DNA molecules modified respectively with two sizes of gold nanoparticles. The surface of Si substrate was characterized through detecting the size and coverage of gold nanoparticles by AFM. Results demonstrated that different sizes of gold nanoparticles represented different sequences of DNA immobilized on the substrate. Density and distribution of DNA on Si substrate can be investigated by AFM imaging using gold nanoparticles as topographic markers. Compared to other sensitive methods such as fluorescence energy transfer, X-ray photoelectron, and radiolabeling experiments, this approach is advantageous in terms of high spatial resolution in sub-micrometer scale. This new method will be beneficial in the characterization of DNA immobilized on chip surfaces.     

Immobilization of gold nanoparticles on solid supports utilizing DNA hybridization

Self-organization of colloidal metal nanoparticles into micro- and nanostructured assemblies is currently of tremendous interest promising to find new size- and structure-dependent physical properties. Owing to its unique recognition capabilities and physicochemical stability, DNA can be used as a molecular linker for gold nanoparticles and is a promising construction material for their precise spatial positioning. Due to the enormous specificity of nucleic acid hybridization, the site-specific immobilization of DNA-functionalized gold colloids (1–40 nm) to solid supports, previously functionalized with a complementary DNA array, allows the fabrication of novel nanostructured surface architectures. Scanning force microscopy (SFM), used to characterize the intermediate steps of the DNA-directed immobilization (DDI) on a gold substrate, provides initial insight into the specificity and efficiency of this technique.     

Assemblies of carbon nanotubes and unencapsulated sub-10-nm gold nanoparticles

The development of assemblies consisting of unencapsulated, sub-10-nm gold particles attached to individual carbon nanotubes (CNTs) with diameters of 2 nm is described. The assemblies are formed on the surface of a porous anodic alumina (PAA) template on which the CNTs (single- or double-walled) are grown by plasma-enhanced chemical vapor deposition. The Au nanoparticles are formed through an indirect evaporation technique using a silicon nitride membrane mask, and diffuse along the PAA surface into the regions containing CNTs. The nanoparticles bind relatively strongly to the CNTs, as indicated by observations of nanoparticles that are suspended over pores or that move along with the CNTs. This approach may provide a new method to functionalize CNTs for chemical or biological sensing and fundamental studies of nanoscale contacts to CNTs.     

Interaction of nanoparticles with lipid membrane

A nanoscale range of surface feature curvatures where lipid membranes lose integrity and form pores has been found experimentally. The pores were experimentally observed in the l-alpha-dimyristoyl phosphatidylcholine membrane around 1.2-22 nm polar nanoparticles deposited on mica surface. Lipid bilayer envelops or closely follows surface features with the curvatures outside of that region. This finding provides essential information for the understanding of nanoparticle-lipid membrane interaction, cytotoxicity, preparation of biomolecular templates and supported lipid membranes on rough and patterned surfaces.