Surin Hong

Sensitive and colorimetric detection of the structural evolution of superoxide dismutase with gold nanoparticles.

The detection and characterization of protein aggregates are critical in terms of advanced diagnostic applications and investigations of protein stability. A variety of analytical methods (e.g., circular dichroism, size exclusion chromatography, and fluorescence microscopy) have been used in this regard, but they are limited in the trace detection of the structural evolution of protein aggregation. Here we report the gold nanoparticle (AuNP)-based highly sensitive and colorimetric detection of the temporal evolution of superoxide dismutase (SOD1) aggregates implicated in the pathology of amyotrophic lateral sclerosis (ALS). For the temporal discrimination of SOD1 aggregation, AuNPs were conjugated with SOD1 monomers (SOD1-AuNPs). Upon exposure of the probes (SOD1-AuNPs) with SOD1 aggregates, significant changes in both surface plasmon resonance spectra and concomitant colors were observed which are attributed to the formation of probe aggregates of variable sizes onto the SOD1 aggregates.

Inhibition of excision-repair of ultraviolet damage in human cells by exposure to methyl methanesulfonate.

Unscheduled DNA synthesis and excision of pyrimidine dimers in human cells exposed to ultraviolet let were inhibited by exposure to methyl methanesulfonate (MMS, 1-2 mM), but repair of MMS damage was not inhibited by UV light. Because the pathways for excision of pyrimidine dimers and alkylation damage have previously been shown to be different, this observation implies a direct effect of alkylation on repair enzymes. We estimate that if inhibition is due to protein alkylation, the UV repair system must present an extremely large target to alkylation and may involve a complex of protein subunits in the order of 1 million daltons such that 1 or more alkylations occur per complex at the concentrations used. These results also indicate that the method of exposing cells to 2 DNA-damaging agents to determine whether they are repaired by common or different pathways can be quite unreliable because of other effects on the repair systems themselves.

Picomolar selective detection of mercuric ion (Hg2 + ) using a functionalized single plasmonic gold nanoparticle

A highly sensitive method for the selective detection and quantification of mercuric ions (Hg(2+)) using single plasmonic gold nanoparticle (GNP)-based dark-field microspectroscopy (DFMS) is demonstrated. The method is based on the scattering property of a single GNP that is functionalized with thiolated molecules, which is altered when analytes bind to the functionalized GNP. The spectral resolution of the system is 0.26 nm and a linear response to Hg(2+) was found in the dynamic range of 100 pM-10 microM. The method permits Hg(2+) to be detected at the picomolar level, which is a remarkable reduction in the detection limit, considering the currently proscribed Environmental Protection Agency regulation level (10 nM, or 2 ppb) and the detection limits of other optical methods for detecting Hg(2+) (recently approx. 1-10 nM). In addition, Hg(2+) can be sensitively detected in the presence of Cd(2+), Pb(2+), Cu(2+), Zn(2+) and Ni(2+), which do not interfere with the analysis. Based on the findings reported herein, it is likely that single-nanoparticle-based metal ion sensing can be extended to the development of other chemo- and biosensors for the direct detection of specific targets in an intracellular environment as well as in environmental monitoring.

Effect of end group modification of DNA-functionalized gold nanoparticles on cellular uptake in HepG2 cells.

Understanding the dynamics of the cellular uptake of nanoparticles in human derived (cancer) cells is crucial to the rational design of functional nanoprobes that can be used for the targeting and delivery of drugs. This study reports on the cellular uptake of gold nanoparticles (GNPs) that were functionalized with different oligonucleotide derivatives using HepG2 cancer cells as a model system. DNA oligomers, in which the end group was modified (NH3, PO3, OH, CH3, and SH groups) were introduced onto the GNP surface. Then, quantitative and qualitative analyses using each DNA-GNP complex were carried out via dark-field scattering microscopy and ICP-MS measurements. Visualization of microscopic images of single cells indicated that the uptake of DNA-GNPs was highly dependent on the type of functionality of the end group in the DNA-GNP complex; the functionality of CH3, and SH resulted in less cellular uptake than that for modifications with NH3, PO3, OH for the same incubation time. This result was reinforced by ICP-MS quantitative analysis. These results were also strongly supported by the events of a DNA-GNP/protein corona; the different association and dissociation rates of proteins around the GNPs was dependent on the functionality of the end group in the DNA-GNP complex, providing further evidence for the conclusion that the components on the surface of nanoparticles directly affected cellular uptake. The findings reported herein provide a basis for the understanding of the fate of GNP-based delivery and provide important insights into the rational design of nanoprobes for the effective treatment of various diseases.

Real-time analysis and direct observations of different superoxide dismutase (SOD1) molecules bindings to aggregates in temporal evolution step

The misfolding and intracellular aggregation of Cu-Zn superoxide dismutase (SOD1) is pathologically key feature of amyotrophic lateral sclerosis (ALS). Although details of the mechanisms continue to be unclear, there are key steps in the possible pathway to the development of ALS. This study focuses on interactions between different SOD1 molecules (A4V apo/holo, and WT apo/holo) and homogeneous aggregates in the temporal evolution step, and a determination of whether any of the SOD1 molecules are reactive to the aggregates with the extent of binding, as determined by surface plasmon resonance (SPR) measurements. Using a kinetic binding model, the association constant of A4V apo was found to be three times larger than that for the WT apo species. Differences in the extent of the interactions were also simultaneously measured and visualized by means of SPR imaging techniques. The SPR-based approach suggests direct correlation between SPR signal and the extent of molecular binding, which can identify the significant contributors to the formation of macroaggregates of SOD1 in the temporal evolution step.

Sensitive and molecular size-selective detection of proteins using a chip-based and heteroliganded gold nanoisland by localized surface plasmon resonance spectroscopy.

A highly sensitive and molecular size-selective method for the detection of proteins using heteroliganded gold nanoislands and localized surface plasmon resonance (LSPR) is described. Two different heteroligands with different chain lengths (3-mercaptopionicacid and decanethiol) were used in fabricating nanoholes for the size-dependent separation of a protein in comparison with its aggregate. Their ratios on gold nanoisland were optimized for the sensitive detection of superoxide dismutase (SOD1). This protein has been implicated in the pathology of amyotrophic lateral sclerosis (ALS). Upon exposure of the optimized gold nanoisland to a solution of SOD1 and aggregates thereof, changes in the LSPR spectra were observed which are attributed to the size-selective and covalent chemical binding of SOD1 to the nanoholes. With a lower detection limit of 1.0 ng/ml, the method can be used to selectively detect SOD1 in the presence of aggregates at the molecular level.

The sensitive, anion-selective detection of arsenate with poly(allylamine hydrochloride) by single particle plasmon-based spectroscopy.

The use of single gold nanoparticle plasmon-based spectroscopy for the sensitive, anion-selective detection of arsenate is described. The method is based on the selective formation of electrostatic complexes between arsenate and poly(allylamine hydrochloride) (PAH) and changes in the single particle plasmon in Rayleigh scattering profiles. PAH, when modified with gold nanoparticles, binds arsenate via its amine-functionalities. The scattering properties of the resulting selectively formed complexes are altered, leading to significant changes in the surface plasmon resonance wavelength. The limit of detection of the method was determined to be 10 nM, which is ca. 13 times more sensitive than U.S. EPA regulation levels. The response is essentially linear in the concentration range of 50-300 nM. The method also shows good selectivity for arsenate in the presence of other environmentally relevant anions, including H(2)PO(4)(-), SO(4)(2-), NO(3)(-), and Cl(-).

Construction of pcAFM module to measure photoconductance with a nanoscale spatial resolution.

A photoconductive atomic force microscopy (pcAFM) module was designed and the performance was tested. This module consisted of three units: the conductive mirror-plate, the steering mirror and the laser source. The module with a laser irradiation unit was equipped to a conventional conducting probe atomic force microscopy (CP-AFM) instrument to measure photoconductance in a nanoscale resolution. As a proof-of-concept experiment, the photoconductance of aggregated fullerene on indium tin oxide (ITO) substrate was measured with this module. The electrical signals (currents) of aggregated fullerene under the conditions of laser on/off at about -10 V sample bias voltage were -100 to -160 nA and 0 to -20 nA, respectively. Results indicated that the pcAFM with this module allowed one to observe photoinduced changes of electrical properties in nanodevices with nanoscale spatial resolution.

Mesoporous silica thin films as a spatially extended probe of interfacial electric fields for amplified signal transduction in surface plasmon resonance spectroscopy

A new simpler concept about the signal amplification of surface plasmon resonance (SPR) that is based on the utilization of mesoporous silica thin films is demonstrated. As compared to monolayer based coatings, mesoporous silica thin films of approximately 200 nm extend the interaction arena away from the metal, thus permitting the integration of the change in optical contrast at different distances from the sensor surface.

A combined top-down/bottom-up approach to structuring multi-sensing zones on a thin film and the application to SPR sensors.

The development of a thin film with well-defined metallic micro/nanostructures, diverse surface functionalities, and superior electronic/optical properties has been a great challenge to researchers seeking an efficient method for the detection of various analytes in chemical and biological sensing applications. Herein, we report a facile and effective approach to the fabrication of an ordered gold island pattern on a glass substrate with contrasted chemical functionalities, which can provide spatially separated sensing zones for multi-detection. In the proposed method, the combination between the micro/nano-imprint lithography and sequential self-assembly approaches exhibited synergistic effects that allowed well-defined structuring and easy surface functionalization in separated sensing zones. Via imprint lithography, the uniform gold islands/glass structure was successfully fabricated from a readily available gold-coated glass film. In addition, a sequential self-assembling strategy and specific chemical-substrate interactions, such as thiol-gold and silane-glass, enabled the surfaces of gold islands and exposed portions of the glass substrate with contrasting chemical functionalities-SH-functionalized gold islands and NH2-functionalized glass substrate. A proof-of-concept experiment for the multi-detection of heavy metal ions (Hg(2+) and Cu(2+)) in an aqueous media was also successfully conducted using the dual-functionalized gold islands/glass structure and surface plasmon resonance measurements. The SH groups on the gold islands and the NH2 groups on the glass substrate functioned as spatially separated and selective receptors for Hg(2+) and Cu(2+) ions, respectively. Therefore, both the detection and quantification of Hg(2+) and Cu(2+) ions could be achieved using a single sensing substrate.