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ABSTRACT: Rapid and ultrasensitive immunoassays were developed by using biofunctional Fe3O4/ZnO/Au nanorices as Raman probes. Taking advantage of the superparamagnetic property of the nanorices, the labeled proteins can rapidly be separated and purified with a commercial permanent magnet. The unsusceptible multiphonon resonant Raman scattering of the nanorices provided a characteristic spectroscopic fingerprint function, which allowed an accurate detection of the analyte. High specificity and selectivity of the assay were demonstrated. It was found that the diffusion barriers and the boundary layer effects had a great influence on the detection limit. Manipulation of the nanorice probes using an external magnetic field can enhance the assay sensitivity by several orders of magnitude, and reduce the detection time from 1 h to 3 min. This magnetic-field-assisted rapid and ultrasensitive immunoassay based on the resonant Raman scatting of semiconductor shows significant value for potential applications in biomedicine, food safety, and environmental defence.
Biosensors & bioelectronics 10/2010; 26(2):918-22. · 5.43 Impact Factor
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ABSTRACT: Up to now, experimental limitations have prevented researchers from achieving the molecular-level understanding for the initial steps of the enzymatic hydrolysis of cellulose, where cellulase breaks down the crystal structure on the surface region of cellulose and exposes cellulose chains for the subsequent hydrolysis by cellulase. Because one of these non-hydrolytic enzymatic steps could be the rate-limiting step for the entire enzymatic hydrolysis of crystalline cellulose by cellulase, being able to analyze and understand these steps is instrumental in uncovering novel leads for improving the efficiency of cellulase. In this communication, we report an innovative application of the microcantilever technique for a real-time assessment of the morphological change of cellulose induced by a treatment of sodium chloride. This sensitive nanomechanical approach to define changes in surface structure of cellulose has the potential to permit a real-time assessment of the effect of the non-hydrolytic activities of cellulase on cellulose and thereby to provide a comprehensive understanding of the initial steps of the enzymatic hydrolysis of cellulose.
Biotechnology and Bioengineering 09/2010; 107(1):190-4. · 3.95 Impact Factor
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ABSTRACT: We describe a method to correct the errors induced by viscous drag on the cantilever in macromolecular manipulation experiments using the atomic force microscope. The cantilever experiences a viscous drag force in these experiments because of its motion relative to the surrounding liquid. This viscous force superimposes onto the force generated by the macromolecule under study, causing ambiguity in the experimental data. To remove this artifact, we analyzed the motions of the cantilever and the liquid in macromolecular manipulation experiments, and developed a novel model to treat the viscous drag on the cantilever as the superposition of the viscous force on a static cantilever in a moving liquid and that on a bending cantilever in a static liquid. The viscous force was measured under both conditions and the results were used to correct the viscous drag induced errors from the experimental data. The method will be useful for many other cantilever based techniques, especially when high viscosity and high cantilever speed are involved.
The Review of scientific instruments 06/2010; 81(6):063703. · 1.52 Impact Factor
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ABSTRACT: Single molecule methods are becoming routine biophysical techniques for studying biological macromolecules. In mechanical unfolding of proteins, an externally applied force is used to induce the unfolding of individual protein molecules. Such experiments have revealed novel information that has significantly enhanced our understanding of the function and folding mechanisms of several types of proteins. To obtain information on the unfolding kinetics and the free energy landscape of the protein molecule from mechanical unfolding data, a Monte Carlo simulation based on a simple two-state kinetic model is often used. In this paper, we provide a detailed description of the procedure to perform such simulations and discuss the approximations and assumptions involved. We show that the appearance of the force versus extension curves from mechanical unfolding of proteins is affected by a variety of experimental parameters, such as the length of the protein polymer and the force constant of the cantilever. We also analyze the errors associated with different methods of data pooling and present a quantitative measure of how well the simulation results fit experimental data. These findings will be helpful in experimental design, artifact identification, and data analysis for single molecule studies of various proteins using the mechanical unfolding method.
International journal of biological macromolecules 12/2009; 46(2):159-66. · 2.37 Impact Factor
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ABSTRACT: The eight-residue alanine oligopeptide Ac-A(4)KA(2)Y-NH(2) (AKY8) was found to form amyloid-like fibrils upon incubation at room temperature in acidified aqueous solution at peptide concentrations >10 mM. The fibril solution exhibits an enhanced vibrational circular dichroism (VCD) couplet in the amide I' band region that is nearly 2 orders of magnitude larger than typical polypeptide/protein signals in this region. The UV-CD spectrum of the fibril solution shows CD in the region associated with the tyrosine side chain absorption. A similar peptide, Ac-A(4)KA(2)-NH(2) (AK7), which lacks a terminal tyrosine residue, does not aggregate. These results suggest a pivotal role for the C-terminal tyrosine residue in stabilizing the aggregation state of this peptide. It is speculated that interactions between the lysine and tyrosine side chains of consecutive strands in an antiparallel arrangement (e.g., cation-pi interactions) are responsible for the stabilization of the resulting fibrils. These results offer considerations and insight regarding the de novo design of self-assembling oligopeptides for biomedical and biotechnological applications and highlight the usefulness of VCD as a tool for probing amyloid fibril formation.
Journal of the American Chemical Society 12/2009; 131(51):18218-9. · 9.91 Impact Factor
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ABSTRACT: A new method of protein detection was explored on the resonant Raman scattering signal of ZnO nanoparticles. A probe for the target protein was constructed by binding the ZnO/Au nanoparticles to secondary protein by eletrostatic interaction. The detection of proteins was achieved by an antibody-based sandwich assay. A first antibody, which could be specifically recognized by target protein, was attached to a solid silicon surface. The ZnO/Au protein probe could specifically recognize and bind to the complex of the target protein and first antibody. This method on the resonant Raman scattering signal of ZnO nanoparticles showed good selectivity and sensitivity for the target protein.
The Journal of Physical Chemistry B 02/2009; 113(5):1468-72. · 3.70 Impact Factor
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ABSTRACT: Macromolecular crowding, a common phenomenon in the cellular environments, can significantly affect the thermodynamic and kinetic properties of proteins. A single-molecule method based on atomic force microscopy (AFM) was used to investigate the effects of macromolecular crowding on the forces required to unfold individual protein molecules. It was found that the mechanical stability of ubiquitin molecules was enhanced by macromolecular crowding from added dextran molecules. The average unfolding force increased from 210 pN in the absence of dextran to 234 pN in the presence of 300 g/L dextran at a pulling speed of 0.25 microm/sec. A theoretical model, accounting for the effects of macromolecular crowding on the native and transition states of the protein molecule by applying the scaled-particle theory, was used to quantitatively explain the crowding-induced increase in the unfolding force. The experimental results and interpretation presented could have wide implications for the many proteins that experience mechanical stresses and perform mechanical functions in the crowded environment of the cell.
Protein Science 10/2008; 17(12):2156-66. · 2.80 Impact Factor
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ABSTRACT: A novel method for identifying DNA microarrays based on ZnO/Au nanocomposites functionalized with thiol-oligonucleotide as probes is descried here. DNA labeled with ZnO/Au nanocomposites has a strong Raman signal even without silver acting as a surface-enhanced Raman scattering promoter. X-ray photoelectron spectra confirmed the formation of a three-component sandwich assay, i.e., constituted DNA and ZnO/Au nanocomposites. The resonance multiple-phonon Raman signal of the ZnO/Au nanocomposites as a spectroscopic fingerprint is used to detect a target sequence of oligonucleotide. This method exhibits extraordinary sensitivity and the detection limit is at least 1 fM.
The Journal of Physical Chemistry B 06/2008; 112(20):6484-9. · 3.70 Impact Factor
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ABSTRACT: The synthesis and characterization of novel electroactive periodic mesoporous organosilica (PMO) are reported. The silsesquioxane precursor, N,N'-bis(4'-(3-triethoxysilylpropylureido)phenyl)-1,4-quinonene-diimine (TSUPQD), was prepared from the emeraldine base of amino-capped aniline trimer (EBAT) using a one-step coupling reaction and was used as an organic silicon source in the co-condensation with tetraethyl orthosilicate (TEOS) in proper ratios. By means of a hydrothermal sol-gel approach with the cationic surfactant cetyltrimethyl-ammonium bromide (CTAB) as the structure-directing template and acetone as the co-solvent for the dissolution of TSUPQD, a series of novel MCM-41 type siliceous materials (TSU-PMOs) were successfully prepared under mild alkaline conditions. The resultant mesoporous organosilica were characterized by Fourier transform infrared (FT-IR) spectroscopy, thermogravimetry, X-ray diffraction, nitrogen sorption, and transmission electron microscopy (TEM) and showed that this series of TSU-PMOs exhibited hexagonally patterned mesostructures with pore diameters of 2.1-2.8 nm. Although the structural regularity and pore parameters gradually deteriorated with increasing loading of organic bridges, the electrochemical behavior of TSU-PMOs monitored by cyclic voltammetry demonstrated greater electroactivities for samples with higher concentration of the incorporated TSU units.
Chemistry 02/2008; 14(9):2909-17. · 5.93 Impact Factor
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ABSTRACT: In this paper we demonstrate the manipulation of single and multiple micro-particles using a bulk acoustic wave (BAW) piezoelectric actuator. Particle motion is achieved by a combination of inertial and acoustic forces, exerted by a piezoelectric actuator, onto particles placed on its surface. This manipulation technique is capable of controlled manipulation of single and multiple micro-particles. The particles can be moved as far as ten times their diameters. The forces generated by the actuator are characterized using a finite element method (FEM) simulation and a theoretical study of the interfacial forces. A comparative study of the forces generated by the actuator and the particle-surface interfacial forces measured using atomic force microscopy (AFM) is then performed to show the feasibility of this technique. Experimental and theoretical results show good agreement indicating that this technique can prove to be a versatile tool for the controlled non-destructive manipulation of single as well as multiple micro-particles in the fields of biosensors, tissue engineering, bio-chips, microfabrication and MEMS devices.
Ultrasonics Symposium, 2007. IEEE; 11/2007
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ABSTRACT: The polymer decoration technique has been widely used to study the chain folding behavior of polymer single crystals. In this article, we demonstrate that this method can be successfully adopted to pattern a variety of polymers on carbon nanotubes (CNTs). The resulting structure is a two-dimensional nanohybrid shish kebab (2D NHSK), wherein the CNT forms the shish and the polymer crystals form the kebabs. 2D NHSKs consisting of CNTs and polymers such as polyethylene, nylon 66, polyvinylidene fluoride and poly(L-lysine) have been achieved. Transmission electron microscopy and atomic force microscopy were used to study the nanoscale morphology of these hybrid materials. Relatively periodic decoration of polymers on both single-walled and multi-walled CNTs was observed. It is envisaged that this unique method offers a facile means to achieve patterned CNTs for nanodevice applications.
Langmuir 08/2007; 23(16):8522-5. · 4.19 Impact Factor
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ABSTRACT: Reactive oxygen species, such as hydroxyl or superoxide radicals, can be generated by exogenous agents as well as from normal cellular metabolism. Those radicals are known to induce various lesions in DNA, including strand breaks and base modifications. These lesions have been implicated in a variety of diseases such as cancer, arteriosclerosis, arthritis, neurodegenerative disorders and others. To assess these oxidative DNA damages and to evaluate the effects of the antioxidant N-acetyl-L-cysteine (NAC), atomic force microscopy (AFM) was used to image DNA molecules exposed to hydroxyl radicals generated via Fenton chemistry. AFM images showed that the circular DNA molecules became linear after incubation with hydroxyl radicals, indicating the development of double-strand breaks. The occurrence of the double-strand breaks was found to depend on the concentration of the hydroxyl radicals and the duration of the reaction. Under the conditions of the experiments, NAC was found to exacerbate the free radical-induced DNA damage.
FEBS Letters 08/2006; 580(17):4136-42. · 3.54 Impact Factor
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ABSTRACT: The design and implementation of a temperature control device for the atomic force microscope (AFM) are described. This device is based on a thermoelectric module which can be used for both heating and cooling the samples in the AFM liquid chamber within the range of 5–50 °C. A thermocouple is inserted in the liquid chamber to measure the sample temperature. A commercial thermoelectric temperature controller is used to keep the temperature constant during a measurement, which can be stabilized within 0.1 °C. To dissipate the heat generated by the thermoelectric module during cooling, a water cooled heat sink is used. Using this device, imaging and mechanical unfolding experiments were carried out at different temperatures. The results show that the temperature control device does not significantly reduce the imaging capacities of the AFM, and that the force-induced unfolding of individual protein molecules can be readily observed at different temperatures. Temperature dependent single molecule measurements can broaden the applications of AFM and reveal new insights into the macromolecular structures and processes.
Review of Scientific Instruments 06/2006; 77(6):063701-063701-5. · 1.37 Impact Factor
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ABSTRACT: Periodic patterning on one-dimensional (1D) carbon nanotubes (CNTs) is of great interest from both scientific and technological points of view. In this letter, we report using a facile physical vapor deposition method to achieve periodic polyethylene (PE) oligomer patterning on individual CNTs. Upon heating under vacuum, PE degraded into oligomers and crystallized into rod-shaped single crystals. These PE rods periodically decorate on CNTs with their long axes perpendicular to the CNT axes. The formation mechanism was attributed to "soft epitaxy" growth of PE oligomer crystals on CNTs. Both SWNTs and MWNTs were decorated successfully with PE rods. The intermediate state of this hybrid structure, MWNTs absorbed with a thin layer of PE, was captured successfully by depositing PE vapor on MWNTs detached from the solid substrate, and was observed using high-resolution transmission electron microscopy. Furthermore, this hybrid structure formation depends critically on CNT surface chemistry: alkane-modification of the MWNT surface prohibited the PE single-crystal growth on the CNTs. We anticipate that this work could open a gateway for creating complex CNT-based nanoarchitectures for nanodevice applications.
Nano Letters 06/2006; 6(5):1007-12. · 13.20 Impact Factor
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ABSTRACT: Interactions of microparticles and nanoparticles with various surfaces under different ambient conditions are of great importance to many applications such as micro-electrical-mechanical systems (MEMS), biomedical and chemical engineering, and have attracted broad interests in the last decade. However, there are only limited numbers of techniques currently available for interfacial particle analysis. In this paper, we propose to use a piezoelectric quartz sensor in a thickness shear mode (TSM) to characterize the interfacial interaction of a single microparticle with a surface. A theoretical model was developed based on the analysis of the various interaction forces, including Van der Waals (VDW) force, capillary force and electrostatic force, etc. The dependence of the particle-surface interaction forces on the particle size was demonstrated. Experimental results from a 5MHz TSM sensor have shown an average frequency shift of 0.5 Hz for a 40 mum stainless steel sphere, which indicated that a coupling strength between a sphere and a surface is about 1.3times10<sup>4</sup> N/m. An atomic force microscope (AFM) was used to provide a complementary tool to help study the interfacial interactions. Adhesion forces between particles and surfaces were measured and favorably compared with the simulation results. It is shown that a TSM sensor is capable of real-time analyzing an interaction of a single microparticle with a surface and an AFM is helpful to identify the effect of each individual interaction force. A combined TSM-AFM technique may create a novel measurement platform for characterization of nanoparticles and their interactions with surfaces
Frequency Control Symposium and Exposition, 2005. Proceedings of the 2005 IEEE International; 09/2005
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ABSTRACT: Nanoscale electrochemical probes with dimensions as small as 50 nm in diameter and 12 micrometers in length have been fabricated using electron beam deposition on the apex of conventional micron size electrodes. The electroactive region is limited to the extreme tip of the nanoprobe by coating with insulating polymer and re-opening of the coating at the extreme tip. We show that our nanoelectrodes are capable of recording neuronal signals from neurons of the mouse hippocampal slice preparation. It is conceivable that nanoprobes can be potentially valuable for the direct measurement of electrical signals at a single presynaptic terminal or postsynaptic site.
Journal of Biomedical Nanotechnology 08/2005; 1(3):336-340. · 4.22 Impact Factor
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ABSTRACT: Single-molecule manipulation techniques have enabled the characterization of the unfolding and refolding process of individual protein molecules, using mechanical forces to initiate the unfolding transition. Experimental and computational results following this approach have shed new light on the mechanisms of the mechanical functions of proteins involved in several cellular processes, as well as revealed new information on the protein folding/unfolding free-energy landscapes. To investigate how protein molecules of different folds respond to a stretching force, and to elucidate the effects of solution conditions on the mechanical stability of a protein, we synthesized polymers of the protein ubiquitin and characterized the force-induced unfolding and refolding of individual ubiquitin molecules using an atomic-force-microscope-based single-molecule manipulation technique. The ubiquitin molecule was highly resistant to a stretching force, and the mechanical unfolding process was reversible. A model calculation based on the hydrogen-bonding pattern in the native structure was performed to explain the origin of this high mechanical stability. Furthermore, pH effects were studied and it was found that the forces required to unfold the protein remained constant within a pH range around the neutral value, and forces decreased as the solution pH was lowered to more acidic values.
Biophysical Journal 01/2005; 87(6):3995-4006. · 3.65 Impact Factor
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ABSTRACT: Macromolecular crowding exists in all living cells and affects protein
folding, rates of diffusion, and amyloid formation. Although the
biophysical theory of macromolecular crowding by thermodynamics and
statistical thermodynamics approaches has been well developed, the
effect of crowding on protein stability and kinetics has not received
sufficient attention experimentally. We have carried out experiments to
characterize the effects of macromolecular crowding on the mechanical
force-induced unfolding and refolding of individual protein molecules of
ubiquitin. To facilitate the mechanical unfolding experiments, polymers
of ubiquitin molecules were synthesized. Using the atomic force
microscope, we determined that, as the concentration of the crowding
agents (dextran) increases from zero to 300 g/l, the average unfolding
force of ubiquitin increase from 228 pN to 296 pN with a pulling speed
of 1000 nm/s and in neutral pH. This result suggests that the unfolding
rate of ubiquitin is reduced due to molecular crowding.
02/2004; -1:9004.
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ABSTRACT: Pathways of unfolding a protein depend in principle on the perturbation-whether it is temperature, denaturant, or even forced extension. Widely-shared, helical-bundle spectrin repeats are known to melt at temperatures as low as 40-45 degrees C and are also known to unfold via multiple pathways as single molecules in atomic force microscopy. Given the varied roles of spectrin family proteins in cell deformability, we sought to determine the coupled effects of temperature on forced unfolding. Bimodal distributions of unfolding intervals are seen at all temperatures for the four-repeat beta(1-4) spectrin-an alpha-actinin homolog. The major unfolding length corresponds to unfolding of a single repeat, and a minor peak at twice the length corresponds to tandem repeats. Increasing temperature shows fewer tandem events but has no effect on unfolding intervals. As T approaches T(m), however, mean unfolding forces in atomic force microscopy also decrease; and circular dichroism studies demonstrate a nearly proportional decrease of helical content in solution. The results imply a thermal softening of a helical linker between repeats which otherwise propagates a helix-to-coil transition to adjacent repeats. In sum, structural changes with temperature correlate with both single-molecule unfolding forces and shifts in unfolding pathways.
Biophysical Journal 12/2003; 85(5):3286-93. · 3.65 Impact Factor
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ABSTRACT: Interactions of microparticles and nanoparticles with various surfaces under different ambient conditions are of great importance to many applications such as microelectrical- mechanical systems (MEMS), biomedical and chemical engineering, and have attracted broad interests in the last decade. However, there are only limited numbers of techniques currently available for interfacial particle analysis. In this paper, we propose to use a piezoelectric quartz sensor in a thickness shear mode (TSM) to characterize the interfacial interaction of a single microparticle with a surface. A theoretical model was developed based on the analysis of the various interaction forces, including Van der Waals (VDW) force, capillary force and electrostatic force, etc. The dependence of the particle-surface interaction forces on the particle size was demonstrated. Experimental results from a 5MHz TSM sensor have shown an average frequency shift of 0.5 Hz for a 40 μm stainless steel sphere, which indicated that a coupling strength between a sphere and a surface is about 1.3×104 N/m. An atomic force microscope (AFM) was used to provide a complementary tool to help study the interfacial interactions. Adhesion forces between particles and surfaces were measured and favorably compared with the simulation results. It is shown that a TSM sensor is capable of real-time analyzing an interaction of a single microparticle with a surface and an AFM is helpful to identify the effect of each individual interaction force. A combined TSM–AFM technique may create a novel measurement platform for characterization of nanoparticles and their interactions with surfaces.
