Article

Biofouling Removal and Protein Detection Using a Hypersonic Resonator

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Abstract

Nonspecific binding (NSB) is a general issue for surface based biosensors. Various approaches have been developed to prevent or remove the NSBs. However, these approaches either increased the background signals of the sensors or limited to specific transducers interface. In this work, we developed a hydrodynamic approach to selectively remove the NSBs using a microfabricated hypersonic resonator with 2.5 gigahertz (GHz) resonant frequency. The high frequency device facilitates to generate multiple controlled micro-vortices which then create cleaning forces at the solid-liquid interfaces. The competitive adhesive and cleaning forces have been investigated using the finite element method (FEM) simulation, identifying the feasibility of the vortices induced NSB removal. NSB proteins have been selectively removed experimentally both on the surface of the resonator and on other substrates which contact the vortices. Thus, the developed hydrodynamic approach is believed to be a simple and versatile tool for NSB removal and compatible to many sensor systems. The unique feature of the hypersonic resonator is that it can be used as a gravimetric sensor as well, thus a combined NSB removal and protein detection dual functional biosensor system is developed.

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... The shear force occurs due to the friction between the nanoparticle and the aqueous fluid flow. It enables the preferential selection of strongly bound specific proteins and thus enhances target signals [22][23][24][25]. Most microfluidic devices for protein capture have focused on the nanoparticles at the constant flow rate. ...
... Cadmium selenide tetrapods (CdSe TPs) were prepared as probe nanoparticles for protein capture. Rotating CdSe TPs by the vortex flow can generate a shear force on the surface of nanoparticles and enhance the target protein captures [23] and removal of non-specific proteins [35]. In this experiment, we applied the vortex flow on CdSe TPs to simultaneously capture and remove proteins. ...
... In contrast, the PL intensities of anti-mouse IgG antibody and BSA that form non-specific binding are reduced by 35% (Figure 1b, middle) and 45% (Figure 1b, bottom) after the vortex flow. Subsequently, the shear force was increased to remove protein [23] by controlling the rotation rate, which was controlled to evaluate the effects on protein removal, as shown in Figure 1c. The PL intensity of anti-goat IgG antibody is slightly reduced with respect to rotation rates. ...
Article
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Non-specific binding (NSB) is one of the important issues in biosensing performance. Herein, we designed a strategy for removing non-specific binding including anti-mouse IgG antibody and bovine serum albumin (BSA) by utilizing anisotropic cadmium selenide tetrapods (CdSe TPs) in a vortex flow. The shear force on the tetrapod nanoparticles was increased by controlling the rotation rate of the vortex flow from 0 rpm to 1000 rpm. As a result, photoluminescence (PL) signals of fluorescein (FITC)-conjugated protein, anti-mouse IgG antibody-FITC and bovine serum albumin (BSA)-FITC, were reduced by 35% and 45%, respectively, indicating that NSB can be removed under vortex flow. In particular, simultaneous NSB removal and protein capture can be achieved even with mixture solutions of target antibodies and anti-mouse IgG antibodies by applying cyclic mode vortex flow on anisotropic CdSe TPs. These results demonstrate successfully that NSB can be diminished by rotating CdSe TPs to generate shear force under vortex flow. This study opens up new research protocols for utilization of anisotropic nanoparticles under vortex flow, which increases the feasibility of protein capture and non-specific proteins removal for biosensors.
... - [90] Magnetic actuator ---Removed 85% of bovine serum albumin (BSA) after 5 min actuation. ...
... (b) Using a hypersonic resonator, non-specific bound proteins were selectively removed. Adopted from [90]. (c) Proteins adsorbed to the actuators and surrounding tubing surfaces was removed during magnetic actuation. ...
... Their results suggested that the most effective protein removal happened when the generated acoustic wavelengths were close to the protein radius. In the work by Pan et al., non-specific bound proteins were removed by micro-vortexes induced by the microfabricated hypersonic resonator with 2.5 GHz resonant frequency [90]. They also showed the non-specific protein removal did not affect the specific antibody-antigen binding (Figure 2b), meaning such approach will not compromise the performance of the potential biosensors (Table 2). ...
Article
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The growing trend for personalized medicine calls for more reliable implantable biosensors that are capable of continuously monitoring target analytes for extended periods (i.e., >30 d). While promising biosensors for various applications are constantly being developed in the laboratories across the world, many struggle to maintain reliable functionality in complex in vivo environments over time. In this review, we explore the impact of various biotic and abiotic failure modes on the reliability of implantable biosensors. We discuss various design considerations for the development of chronically reliable implantable biosensors with a specific focus on strategies to combat biofouling, which is a fundamental challenge for many implantable devices. Briefly, we introduce the process of the foreign body response and compare the in vitro and the in vivo performances of state-of-the-art implantable biosensors. We then discuss the latest development in material science to minimize and delay biofouling including the usage of various hydrophilic, biomimetic, drug-eluting, zwitterionic, and other smart polymer materials. We also explore a number of active anti-biofouling approaches including stimuli-responsive materials and mechanical actuation. Finally, we conclude this topical review with a discussion on future research opportunities towards more reliable implantable biosensors.
... However, biomolecular areas of surface-based sensors often come into contact with complex mixtures of proteins and other molecules during their use [7]. Most surfaces are particularly prone to non-specific and irreversible adsorption of proteins, known as non-specific adsorption (NSA) [7][8][9][10]. NSA happens when a molecule adsorbs to a sensor's surface, resulting in Figure 1. ...
... This phenomenon can result in four different types of NSA: (1) molecules adsorbed on vacant spaces (2) molecules adsorbed on non-immunological sites (3) molecules adsorbed on immunological sites, allowing access to antigens and (4) molecules adsorbed on immunological sites. Non-specific adsorption leads to elevated background signals that cannot be discriminated from specific binding [9]. These false-positive signals affect the dynamic range [25], limit of detection [2,9], reproducibility, selectivity, and sensitivity [9]. ...
... Non-specific adsorption leads to elevated background signals that cannot be discriminated from specific binding [9]. These false-positive signals affect the dynamic range [25], limit of detection [2,9], reproducibility, selectivity, and sensitivity [9]. The reduction of NSA is crucial in the development of biosensors, especially for point-of-care clinical diagnostics [5]. ...
Article
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Non-specific adsorption (NSA) is a persistent problem that negatively affects biosensors, decreasing sensitivity, specificity, and reproducibility. Passive and active removal methods exist to remedy this issue, by coating the surface or generating surface forces to shear away weakly adhered biomolecules, respectively. However, many surface coatings are not compatible or effective for sensing, and thus active removal methods have been developed to combat this phenomenon. This review aims to provide an overview of methods of NSA reduction in biosensing, focusing on the shift from passive methods to active methods in the past decade. Attention is focused on protein NSA, due to their common use in biosensing for biomarker diagnostics. To our knowledge, this is the first review to comprehensively discuss active NSA removal methods. Lastly, the challenges and future perspectives of NSA reduction in biosensing are discussed.
... In our previous research, it was confusing that the proteins behaved with a much stronger dynamic response monitored by optical microscopy when motivated by the SMR device in fluid [18]. Consequently, the dynamic behavior of the bare SMR device without proteins was characterized using Garcia-Sanchez's method [15]. ...
... In the same year, San Paulo et al. successively utilized atomic force microscopy in contact mode and dynamic force microscopy to characterize the mechanical vibrations of RF resonators including FBAR [17] in air. In our previous research, it was confusing that the proteins behaved with a much stronger dynamic response monitored by optical microscopy when motivated by the SMR device in fluid [18]. Consequently, the dynamic behavior of the bare SMR device without proteins was characterized using Garcia-Sanchez's method [15]. ...
... The much stronger hydrodynamic motion of water hinders the contact of the tip and the SMR surface. As observed previously, the strong dynamic behavior of the proteins in fluid is reasonable to be considered the excitation of vibrating water [18]. Figure 5a shows the "step-type" topography of the static and vibrating SMR in fluid obtained from bottom to top. ...
Article
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The very small vibration of a solidly-mounted resonator (SMR) in fluid may trigger a relatively large motion of the covering fluid, which was implied by our protein-related experimental results. Therefore, a series of experimental methods for characterizing the mechanical longitudinal vibration of the SMR and the corresponding out-of-plane dynamic response of the fluid above the SMR surface is described in this paper. A SMR device with theoretical resonance frequency of 2.5 GHz was driven by an amplitude-modulated (AM) signal, in which the amplitude is modulated by a signal of the second resonance frequency of the atomic force microscope (AFM) cantilever. A lock-in amplifier is used to demodulate the vibration response of the AFM cantilever, which is proportional to the amplitude of the sample vibration in contact mode and tapping mode. The amplitude-frequency curve of the SMR surface is obtained in contact mode with a relatively stronger interaction force between the AFM tip and the SMR surface. The amplitude-frequency curve of the motion of the liquid above the SMR device and the peak amplitude of the fluid at different distances above the SMR surface are measured in tapping mode with a relatively weak interaction force between the AFM tip and the fluid sample.
... Because the free acoustic field is nonuniform and the scale of the medium is so large in comparison with the GHz traveling BAW-based AST, this momentum flux is a typical Eckart streaming, which pushes the fluid along the propagation direction of the acoustic wave. 39 As a consequence, the constitutive equation of a Newtonian fluid giving the relationship between stress, shear strain, normal strain and pressure is ...
... More detailed information about the fabrication process can be found in our previous paper. 39 ...
Article
Contactless acoustic manipulation of micro/nanoscale particles has attracted considerable attention owing to its near independence of the physical and chemical properties of the targets, making it universally applicable to almost all biological systems. Thin-film bulk acoustic wave (BAW) resonators operating at gigahertz (GHz) frequencies have been demonstrated to generate localized high-speed microvortices through acoustic streaming effects. Benefitting from the strong drag forces of the high-speed vortices, BAW-enabled GHz acoustic streaming tweezers (AST) have been applied to the trapping and enrichment of particles ranging in size from micrometers to less than 100 nm. However, the behavior of particles in such 3D microvortex systems is still largely unknown. In this work, the particle behavior (trapping, enrichment, and separation) in GHz AST is studied by theoretical analyses, 3D simulations, and microparticle tracking experiments. It is found that the particle motion in the vortices is determined mainly by the balance between the acoustic streaming drag force and the acoustic radiation force. This work can provide basic design principles for AST-based lab-on-a-chip systems for a variety of applications.
... Such resonators have recently been reported by us to generate high-speed (> m s À1 ) acoustic streaming with strong forces (> nn), which has been applied to enhance the solution mixing in microfluidic chips [39] and to remove nonspecific binding at solid-liquid interfaces. [40] Since acoustic streaming can exert mechanical forces on cells that are immobilized at the solid-liquid interface, [41] we envisaged that vesicles, which are soft and hollow structures, would also be affected and could experience ...
... Such resonators have recently been reported by us to generate high-speed (> m s À1 ) acoustic streaming with strong forces (> nn), which has been applied to enhance the solution mixing in microfluidic chips [39] and to remove nonspecific binding at solid-liquid interfaces. [40] Since acoustic streaming can exert mechanical forces on cells that are immobilized at the solid-liquid interface, [41] we envisaged that vesicles, which are soft and hollow structures, would also be affected and could experience mechanical deformation under such acoustic stimulation. We hypothesized that owing to the fluidic nature of the lipid membranes, mechanical deformation of the vesicles might induce transient pores in the membrane, which would change the membrane permeability and facilitate materials exchange between the interior and exterior of the vesicles (Scheme 1). ...
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Controllable exchange of molecules between the interior and the external environment of vesicles is critical in drug delivery and micro/nano‐reactors. While many approaches exist to trigger release from vesicles, controlled loading remains a challenge. Here, we show that gigahertz acoustic streaming generated by a nanoelectromechanical resonator can control the loading and release of cargo into/from vesicles. Polymer‐shelled vesicles showed loading and release of molecules both in solution and on a solid substrate. We observed deformation of individual giant unilamellar vesicles and propose that the shear stress generated by gigahertz acoustic streaming induces the formation of transient nanopores in the vesicle membranes. The size of these pores was estimated to be on the order of 100 nm by loading nanoparticles of different sizes into the vesicles. Forming such pores with gigahertz acoustic streaming provides a non‐invasive method to control materials exchange across membranes of different types of vesicles. This method could allow site‐specific release of therapeutics and controlled loading into cells, as well as tunable microreactors.
... Due to the piezoelectric effect, the radio frequency (RF) source applied to the SMR device can generate high-frequency longitudinal acoustic waves. When the piezoelectric material is immersed in solution, the high-frequency acoustic waves will trigger the acoustic streaming effects due to acoustic power leakage during the propagation of the acoustic waves into liquid [29][30][31]. High-frequency acoustic waves will contribute to the streaming flow strength due to the great energy attenuation coefficient after their transmission into liquid. Moreover, longitudinal waves can provide more effective energy coupling into the liquid than the shear mode acoustic waves to induce stronger turbulent flow [31]. ...
... High-frequency acoustic waves will contribute to the streaming flow strength due to the great energy attenuation coefficient after their transmission into liquid. Moreover, longitudinal waves can provide more effective energy coupling into the liquid than the shear mode acoustic waves to induce stronger turbulent flow [31]. To be exact, SMR devices have both features, namely high frequency and longitudinal acoustic waves, which can increase the acoustic streaming. ...
Article
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Low-abundance biomolecule detection is very crucial in many biological and medical applications. In this paper, we present a novel electrolyte-gated graphene field-effect transistor (EGFET) biosensor consisting of acoustic tweezers to increase the sensitivity. The acoustic tweezers are based on a high-frequency bulk acoustic resonator with thousands of MHz, which has excellent ability to concentrate nanoparticles. The operating principle of the acoustic tweezers to concentrate biomolecules is analyzed and verified by experiments. After the actuation of acoustic tweezers for 10 min, the IgG molecules are accumulated onto the graphene. The sensitivities of the EGFET biosensor with accumulation and without accumulation are compared. As a result, the sensitivity of the graphene-based biosensor is remarkably increased using SMR as the biomolecule concentrator. Since the device has advantages such as miniaturized size, low reagent consumption, high sensitivity, and rapid detection, we expect it to be readily applied to many biological and medical applications.
... In this acoustical disruption process that occurs by coupling with the acoustic waves at a sub-micron length scale, some transient nanopores are generated to allow for the ion transfer across the SLB. Since hypersound can form strong turbulent flow inside fluids, 50 the ion transfer can also be facilitated by the accompanying acoustic streaming effects. Due to the flexibility of the lipid membrane and the molecular diffusion of the lipid molecules therein, the membrane defects can then be healed in a re-assembly process by turning off the hypersound. ...
... It is likely that more and/or longer lasting transient nanopores were generated with hypersound of higher power, and this high acoustic intensity also enhanced the ion transfer by accelerating the velocity of acoustic streaming. 50 The magnitude of the current through the transient pores was plotted as a function of input power, which shows a response to hypersound that levels off at high powers (Fig. 3b). ...
Article
Hypersound (ultrasound of gigahertz (GHz) frequency) has been recently introduced as a new type of membrane-disruption method for cells, vesicles and supported lipid bilayers (SLBs), with the potential to improve the efficiency of drug and gene delivery for biomedical applications. Here, we fabricated an integrated microchip, composed of a nano-electromechanical system (NEMS) resonator and a gold electrode as the extended gate of a field effect transistor (EGFET), to study the effects of hypersonic poration on an SLB in real time. The current recordings revealed that hypersound enabled ion conduction through the SLB by inducing transient nanopores in the membrane, which act as the equivalent of ion channels and show gating behavior. The mechanism of pore formation was studied by cyclic voltammetry (CV), atomic force microscopy (AFM) and laser scanning microscopy (LSM), which support the causality between hypersound-triggered deformation and the reversible membrane disruption of the SLB. This finding contributes to the development of an approach to reversibly control membrane permeability by hypersound.
... The fabrication process of the hypersonic device-Solid Mount Resonator (SMR) was performed according to a published procedure [36]. The experimental set-up was very similar to the published one [25,36]. ...
... The fabrication process of the hypersonic device-Solid Mount Resonator (SMR) was performed according to a published procedure [36]. The experimental set-up was very similar to the published one [25,36]. ...
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6′/3′-Sialylated N-acetyllactosamine (6′/3′-SLN) is important for discrimination of the source (human or avian) of influenza virus strains. Biotinylated oligosaccharides have been widely used for analysis and quick detection. The development of efficient strategies to synthesize biotin-tagged 6′/3′-SLN have become necessary. Effective mixing is essential for enzymatic solid-phase oligosaccharide synthesis (SPOS). In the current study, newly developed technology ultrahigh-frequency-acoustofluidics (UHFA), which can provide a powerful source for efficient microfluidic mixing, solid-phase oligosaccharide synthesis and one-pot multienzyme (OPME) system, were used to develop a new strategy for oligosaccharide synthesis. Firstly, biotinylated N-acetylglucosamine was designed and chemically synthesized through traditional approaches. Secondly, biotinylated 6′- and 3′-sialyl-N-acetylglucosamines were prepared in solution through two sequential OPME modules in with a yield of ~95%. Thirdly, 6′-SLN was also prepared through UHFA-based enzymatic solid-phase synthesis on magnetic beads with a yield of 64.4%. The current strategy would be potentially used for synthesis of functional oligosaccharides.
... To minimize these problems, different strategies can be applied, which are classified into passive and active approaches [19]. Active approaches are based on removing already-adsorbed fouling elements, and can be achieved using temperature-responsive materials [20], acoustic waves [21], or mechanical actuation [22]. By comparison, passive approaches are related to the prevention of incoming fouling. ...
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Analytical Parameters of a Novel Glucose Biosensor Based on Grafted PFM as a Covalent Immobilization Technique. Abstract: Bioanalytical methods, in particular electrochemical biosensors, are increasingly used in different industrial sectors due to their simplicity, low cost, and fast response. However, to be able to reliably use this type of device, it is necessary to undertake in-depth evaluation of their fundamental analytical parameters. In this work, analytical parameters of an amperometric biosensor based on covalent immobilization of glucose oxidase (GOx) were evaluated. GOx was immobilized using plasma-grafted pentafluorophenyl methacrylate (pgPFM) as an anchor onto a tailored HEMA-co-EGDA hydrogel that coats a titanium dioxide nanotubes array (TiO 2 NTAs). Finally, chitosan was used to protect the enzyme molecules. The biosensor offered outstanding analytical parameters: repeatability (RSD = 1.7%), reproducibility (RSD = 1.3%), accuracy (deviation = 4.8%), and robustness (RSD = 2.4%). In addition, the Ti/TiO 2 NTAs/ppHEMA-co-EGDA/pgPFM/GOx/Chitosan biosensor showed good long-term stability; after 20 days, it retained 89% of its initial sensitivity. Finally, glucose concentrations of different food samples were measured and compared using an official standard method (HPLC). Deviation was lower than 10% in all measured samples. Therefore, the developed biosensor can be considered to be a reliable analytical tool for quantification measurements.
... So far, ultrahigh frequency acoustofluidics has rarely been studied owing to the lack of such high-frequencya coustic devices.Inthis work, we report amethod for controlling both loading and release of materials into and from vesicles without damaging their structures using the gigahertz (GHz) acoustic streaming generated by at hin film-based nanoelectromechanical (NEMS) resonator.S uch resonators have recently been reported by us to generate high-speed (> ms À1 ) acoustic streaming with strong forces (> nn), which has been applied to enhance the solution mixing in microfluidic chips [39] and to remove nonspecific binding at solid-liquid interfaces. [40] Since acoustic streaming can exert mechanical forces on cells that are immobilized at the solid-liquid interface, [41] we envisaged that vesicles,w hich are soft and hollow structures,w ould also be affected and could experience mechanical deformation under such acoustic stimulation. We hypothesized that owing to the fluidic nature of the lipid membranes,m echanical deformation of the vesicles might induce transient pores in the membrane,which would change the membrane permeability and facilitate materials exchange between the interior and exterior of the vesicles (Scheme 1). ...
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Full-text available
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... Protein adsorption is also used to make biocompatible surfaces and artificial scaffolds [11]. However, in marine and biomedical implants, the adsorption of protein on substrates [12] or nanostructured materials [13] is considered as an undesirable effect [14], as it results in bacterial adhesion and biofouling formation on the materials that reduces the functioning of material (e.g. heat transfer efficiencies, reduced flow rates, bacterial influenced corrosion etc.) [15]. ...
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Here we report the adsorption behavior of protein at an oil-water interface, that is covered with polyelectrolytes (PE) and the interaction forces involved. The weak and strong PE's used are poly(acrylic acid), PAA and poly(diallyldimethylammonium chloride), PDDA, respectively. The protein and oil used are bovine serum albumin (BSA) and octane droplets in water, containing superparamagnetic magnetite nanoparticles, respectively. The intermolecular forces, hydrodynamic diameter and electrophoretic mobility measurements are used to study the adsorption behavior of protein. The results showed an enhanced displacement of the adsorbed PDDA and PAA with increasing concentrations of BSA. In the absence of BSA, the emulsion droplets covered with PDDA and PAA exhibited a long range repulsive interaction. The addition of BSA into PDDA and PAA stabilized emulsion resulted in a decrease in the onset of interaction of repulsive force profile. The reversal of the sign and the change in the magnitude of zeta potential indicates the displacement of PDDA by BSA molecules at the oil-water (O/W) interface, which was in agreement with the findings of Satulovsky et.al. [Proc. Natl. Acad. Sci. 97 (2000) 9037–9041] that polymers on hydrophobic surfaces are not effective in preventing protein adsorption. These results indicate the displacement of macromolecule of very large molecular weight and preferential adsorption of small macromolecules at the O/W interfaces, irrespective of the nature of adsorbed PE's (strong or weak, anionic or cationic with high or low charge density). These finding provides new insights into adsorption of proteins at the oil-water interfaces and may have important implication in tailoring industrial formulations with an extended shelf life and in increasing biocompatibility of implants.
... Protein adsorption is also used to make biocompatible surfaces and artificial scaffolds [11]. However, in marine and biomedical implants, the adsorption of protein on substrates [12] or nanostructured materials [13] is considered as an undesirable effect [14], as it results in bacterial adhesion and biofouling formation on the materials that reduces the functioning of material (e.g. heat transfer efficiencies, reduced flow rates, bacterial influenced corrosion etc.) [15]. ...
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Hydrodynamic approaches are important for biomedical diagnostics, chemical analysis, and a broad range of industrial applications. Size-based separation and sorting is an important tool for these applications. We report the integration of hypersound technology with patterned protein films to provide efficient sorting of microparticles based on particle charge and size. We employed a hypersonic resonator for the acoustic streaming of the fluidic system to generate microvortices that exert drag forces on the objects on the surface that are dictated by their radius of curvature. We demonstrate a size-based sorting of anionic silica particles using protein patterns and gradients fabricated using attractive cationic and repulsive anionic proteins.
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The development of rapid and efficient tools to modulate neurons is vital for the treatment of nervous system diseases. Here, a novel non-invasive neurite outgrowth modulation method based on a controllable acoustic streaming effect induced by an electromechanical gigahertz resonator microchip is reported. The results demonstrate that the gigahertz acoustic streaming can induce cell structure changes within a 10 min period of stimulation, which promotes a high proportion of neurite bearing cells and encourages longer neurite outgrowth. Specifically, the resonator stimulation not only promotes outgrowth of neurites, but also can be combined with chemical mediated methods to accelerate the direct entry of nerve growth factor (NGF) into cells, resulting in higher modulation efficacy. Owing to shear stress caused by the acoustic streaming effect, the resonator microchip mediates stress fiber formation and induces the neuron-like phenotype of PC12 cells. We suggest that this method may potentially be applied to precise single-cell modulation, as well as in the development of non-invasive and rapid disease treatment strategies.
Conference Paper
In this study, an acoustofluidic based wireless micropump for drug delivery was proposed and fabricated. The key actuator of this micropump is a small gigahertz piezoelectric resonator, which could induce strong fluidic streaming at low applied power. This acoustofluidic micropump has stable and accurate dosage resolution (7.0 μL), and sufficient flow rate (1.34 mL/min). The miniaturized size and wireless controlled operation prove it as a portable drug delivery system.Clinical Relevance- The acoustofluidic based micropump could apply for the drug administration in a safe, effective and stable form. It has potential to integrate with miniaturized sensors and electronic circuit to form portable drug delivery systems, realizing smart on-demand drug delivery.
Thesis
Randomly oriented nanowire networks (nanonets) exhibit interesting properties, arising from the intrinsic properties of the individual nanostructures, but also offer remarkable new properties related to the nanonet structure, namely a better reproducibility, and high fault tolerance and flexibility. In this context, this project investigated the possibility of using a 2D assembly of randomly oriented Si nanowires (Si nanonet), as building blocks for the fabrication and development of two different types of biosensors: Si nanonet-based FET biosensors for the electrical detection of DNA hybridization, and Si nanonet-based electrochemical biosensors for thrombin detection.For this purpose, the development of each type of sensor was divided into three parts: (i) fabrication of the Si nanonet-based device, (ii) biofunctionalization of the device allowing the grafting of the bioreceptor (TBA-15 aptamer), and (iii) detection of the target molecule.Regarding the Si nanonet-based FET biosensors, the devices were fabricated following a process previously developed at LMGP. Here, the work focused on the implementation and the optimization of a biofunctionalization protocol, enabling the proper immobilization of the bioreceptor on the surface of the Si nanonet-based FETs. Then, the ability to electrically detect the DNA hybridization in dry environment was studied. Concerning the electrochemical biosensors, it was developed for the first time a process allowing the fabrication and biofunctionalization of a 2D Si nanonet-based biosensor, enabling thrombin detection in nM concentrations, indicating either the risk or the presence of thrombosis.These promising results open the door to future projects implying the use of 2D Si nanonet-based devices in the field of biodetection.
Article
Surface-enhanced Raman spectroscopy (SERS) has become a growing ultrasensitive analytical technique to quantify toxic molecules in foodstuffs. Monitoring the levels of chemical contaminants not only ensures food security but also offers a guideline on the production, processing, and risk analysis of consumer’s health protection. The objective of this study was to point out the possible challenges associated with the detection of mycotoxins in foodstuffs. Herein, we have discussed briefly as to selectivity, accuracy, precision, robustness, ruggedness, non-specific adsorption (NSA), cross-reactivity (for both label-free and the target analyte capture approaches like the application of antibody, aptamer, molecularly imprinted polymer (MIP), linear polymer affinity agents and/or specific surface-modified nanomaterials) and their potential solution.
Article
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We present an acoustic microfluidic mixing approach via acousto-mechanically induced micro-vortices sustained by localized ultrahigh frequency (UHF) acoustic fields. A micro-fabricated solid-mounted thin-film piezoelectric resonator (SMR) with a frequency of 1.54 GHz has been integrated into microfluidic systems. Experimental and simulation results show that UHF-SMR triggers strong acoustic field gradients to produce efficient and highly localized acoustic streaming vortices, providing a powerful source for microfluidic mixing. Homogeneous mixing with 87% mixing efficiency at a Peclet number of 35520 within 1 ms has been achieved. The proposed strategy shows a great potential for microfluidic mixing and enhanced molecule transportation in minimized analytical systems.
Article
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Despite increasing demand in the manipulation of nanoscale objects for next generation biological and industrial processes, there is a lack of methods for reliable separation, concentration and purification of nanoscale objects. Acoustic methods have proven their utility in contactless manipulation of microscale objects mainly relying on the acoustic radiation effect, though the influence of acoustic streaming has typically prevented manipulation at smaller length scales. In this work, however, we explicitly take advantage of the strong acoustic streaming in the vicinity of a highly focused, high frequency surface acoustic wave (SAW) beam emanating from a series of focused 6 μm substrate wavelength interdigital transducers patterned on a piezoelectric lithium niobate substrate and actuated with a 633 MHz sinusoidal signal. This streaming field serves to focus fluid streamlines such that incoming particles interact with the acoustic field similarly regardless of their initial starting positions, and results in particle displacements that would not be possible with a travelling acoustic wave force alone. This streaming-induced manipulation of nanoscale particles is maximized with the formation of micro-vortices that extend the width of the microfluidic channel even with the imposition of a lateral flow, occurring when the streaming-induced flow velocities are an order of magnitude larger than the lateral one. We make use of this acoustic streaming to demonstrate the continuous and differential focusing of 100 nm, 300 nm and 500 nm particles.
Article
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Controlled drug release has a high priority for the development of modern medicine and biochemistry. To develop a versatile method for controlled release, a miniaturized acoustic gigahertz (GHz) resonator is designed and fabricated which can transfer electric supply to mechanical vibrations. By contacting with liquid, the GHz resonator directly excites streaming flows and induces physical shear stress to tear the multilayered polyelectrolyte (PET) thin films. Due to the ultra-high working frequency, the shear stress is greatly intensified, which results in a controlled disassembling of the PET thin films. This technique is demonstrated as an effective method to trigger and control the drug release. Both theory analysis and controlled release experiments prove the thin film destruction and the drug release..
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In this paper, we have modeled and analyzed affinities and kinetics of volatile organic compounds (VOCs) adsorption (and desorption) on various surface chemical groups using multiple self-assembled monolayers (SAMs) functionalized film bulk acoustic resonator (FBAR) array. The high-frequency and micro-scale resonator provides improved sensitivity in the detections of VOCs at trace levels. With the study of affinities and kinetics, three concentration-independent intrinsic parameters (monolayer adsorption capacity, adsorption energy constant and desorption rate) of gas-surface interactions are obtained to contribute to a multi-parameter fingerprint library of VOC analytes. Effects of functional group's properties on gas-surface interactions are also discussed. The proposed sensor array with concentration-independent fingerprint library shows potential as a portable electronic nose (e-nose) system for VOCs discrimination and gas-sensitive materials selections.
Article
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Affinity biosensors use biorecognition elements and transducers to convert a biochemical event into a recordable signal. They provides the molecule binding information, which includes the dynamics of biomolecular association and dissociation, and the equilibrium association constant. Complementary metal oxide semiconductor-compatible silicon (Si) nanowires configured as a field-effect transistor (NW FET) have shown significant advantages for real-time, label-free and highly sensitive detection of a wide range of biomolecules. Most research has focused on reducing the detection limit of Si-NW FETs but has provided less information about the real binding parameters of the biomolecular interactions. Recently, Si-NW FETs have been demonstrated as affinity biosensors to quantify biomolecular binding affinities and kinetics. They open new applications for NW FETs in the nanomedicine field and will bring such sensor technology a step closer to commercial point-of-care applications. This article summarizes the recent advances in bioaffinity measurement using Si-NW FETs, with an emphasis on the different approaches used to address the issues of sensor calibration, regeneration, binding kinetic measurements, limit of detection, sensor surface modification, biomolecule charge screening, reference electrode integration and nonspecific molecular binding.
Article
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In acoustofluidic manipulation and sorting devices, Rayleigh streaming flows are typically found in addition to the acoustic radiation forces. However, experimental work from various groups has described acoustic streaming that occurs in planar devices in a plane parallel to the transducer face. This is typically a four-quadrant streaming pattern with the circulation parallel to the transducer. Understanding its origins is essential for creating designs that limit or control this phenomenon. The cause of this kind of streaming pattern has not been previously explained as it is different from the well-known classical streaming patterns such as Rayleigh streaming and Eckart streaming, whose circulation planes are generally perpendicular to the face of the acoustic transducer. In order to gain insight into these patterns we present a numerical method based on Nyborg's limiting velocity boundary condition that includes terms ignored in the Rayleigh analysis, and verify its predictions against experimental PIV results in a simple device. The results show that the modelled particle trajectories match those found experimentally. Analysis of the dominant terms in the driving equations shows that the origin of this kind of streaming pattern is related to the circulation of the acoustic intensity.
Article
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Monitoring the binding affinities and kinetics of protein interactions is important in clinical diagnostics and drug development because such information is used to identify new therapeutic candidates. Surface plasmon resonance is at present the standard method used for such analysis, but this is limited by low sensitivity and low-throughput analysis. Here, we show that silicon nanowire field-effect transistors can be used as biosensors to measure protein-ligand binding affinities and kinetics with sensitivities down to femtomolar concentrations. Based on this sensing mechanism, we develop an analytical model to calibrate the sensor response and quantify the molecular binding affinities of two representative protein-ligand binding pairs. The rate constant of the association and dissociation of the protein-ligand pair is determined by monitoring the reaction kinetics, demonstrating that silicon nanowire field-effect transistors can be readily used as high-throughput biosensors to quantify protein interactions.
Article
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Mechanical interactions are fundamental to biology. Mechanical forces of chemical origin determine motility and adhesion on the cellular scale, and govern transport and affinity on the molecular scale. Biological sensing in the mechanical domain provides unique opportunities to measure forces, displacements and mass changes from cellular and subcellular processes. Nanomechanical systems are particularly well matched in size with molecular interactions, and provide a basis for biological probes with single-molecule sensitivity. Here we review micro- and nanoscale biosensors, with a particular focus on fast mechanical biosensing in fluid by mass- and force-based methods, and the challenges presented by non-specific interactions. We explain the general issues that will be critical to the success of any type of next-generation mechanical biosensor, such as the need to improve intrinsic device performance, fabrication reproducibility and system integration. We also discuss the need for a greater understanding of analyte-sensor interactions on the nanoscale and of stochastic processes in the sensing environment.
Article
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Despite its fundamental role in the dynamics of compressible fluids, bulk viscosity has received little experimental attention and there remains a paucity of measured data. Acoustic spectroscopy provides a robust and accurate approach to measuring this parameter. Working from the Navier-Stokes model of a compressible fluid one can show that the bulk viscosity makes a significant and measurable contribution to the frequency-squared acoustic attenuation. Here we employ this methodology to determine the bulk viscosity of Millipore water over a temperature range of 7 to 50 degrees Celsius. The measured attenuation spectra are consistent with the theoretical predictions, while the bulk viscosity of water is found to be approximately three times larger than its shear counterpart, reinforcing its significance in acoustic propagation. Moreover, our results demonstrate that this technique can be readily and generally applied to fluids to accurately determine their temperature dependent bulk viscosities.
Article
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The ability to detect single protein molecules in blood could accelerate the discovery and use of more sensitive diagnostic biomarkers. To detect low-abundance proteins in blood, we captured them on microscopic beads decorated with specific antibodies and then labeled the immunocomplexes (one or zero labeled target protein molecules per bead) with an enzymatic reporter capable of generating a fluorescent product. After isolating the beads in 50-fl reaction chambers designed to hold only a single bead, we used fluorescence imaging to detect single protein molecules. Our single-molecule enzyme-linked immunosorbent assay (digital ELISA) approach detected as few as approximately 10-20 enzyme-labeled complexes in 100 microl of sample (approximately 10(-19) M) and routinely allowed detection of clinically relevant proteins in serum at concentrations (<10(-15) M) much lower than conventional ELISA. Digital ELISA detected prostate-specific antigen (PSA) in sera from patients who had undergone radical prostatectomy at concentrations as low as 14 fg/ml (0.4 fM).
Article
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Protein biochips are at the heart of many medical and bioanalytical applications. Increasing interest has been focused on surface activation and subsequent functionalization strategies for immobilizing these biomolecules. Different approaches using covalent and noncovalent chemistry are reviewed; particular emphasis is placed on the chemical specificity of protein attachment and on retention of protein function. Strategies for creating protein patterns (as opposed to protein arrays) are also outlined. An outlook on promising and challenging future directions for protein biochip research and applications is also offered.
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We describe highly sensitive, label-free, multiplexed electrical detection of cancer markers using silicon-nanowire field-effect devices in which distinct nanowires and surface receptors are incorporated into arrays. Protein markers were routinely detected at femtomolar concentrations with high selectivity, and simultaneous incorporation of control nanowires enabled discrimination against false positives. Nanowire arrays allowed highly selective and sensitive multiplexed detection of prostate specific antigen (PSA), PSA-alpha1-antichymotrypsin, carcinoembryonic antigen and mucin-1, including detection to at least 0.9 pg/ml in undiluted serum samples. In addition, nucleic acid receptors enabled real-time assays of the binding, activity and small-molecule inhibition of telomerase using unamplified extracts from as few as ten tumor cells. The capability for multiplexed real-time monitoring of protein markers and telomerase activity with high sensitivity and selectivity in clinically relevant samples opens up substantial possibilities for diagnosis and treatment of cancer and other complex diseases.
Article
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Nonspecific binding is a universal problem that reduces bioassay sensitivity and specificity. We demonstrate that ultrasonic waves, generated by 5-MHz quartz crystal resonators, accelerate nonspecifically bound protein desorption from sensing and nonsensing areas of micropatterned protein arrays, controllably and nondestructively cleaning the micropatterns. Nonsensing area fluorescent intensity values dropped by more than 85% and sensing area fluorescent intensity dropped 77% due to nonspecific binding removal at an input power of 14 W. After patterning, antibody films were many layers thick with nonspecifically bound protein, and aggregates obscured patterns. Quartz crystal resonators removed excess antibody layers and aggregates leaving highly uniform films, as evidenced by smaller spatial variations in fluorescent intensity and atomic force microscope surface roughness values. Fluorescent intensity values obtained after 14-W QCR operation were more repeatable and uniform.
Article
The advancement in micro/nanotechnologies has been continuously providing possibilities for inventing novel biochemical sensors. However, variations in the transducer type can cause different sensing results due to the differences in their mechanisms of analyzing biomolecular interactions. In this work, we focused on the comparative analysis of static and non-static assays for molecular interactions using on-chip integrated extended-gate field effect transistor (EGFET) as a static sensing interface and solidly mounted resonator (SMR) as a non-static sensing interface. Analysis of polyelectrolytes (PETs) surface assembly and antigen-antibody interaction using the two types of biochemical sensors presented consistent and complementary sets of information. Meanwhile, due to the difference in their operating mechanisms, variations on the detection efficiency, kinetics and thermodynamics were observed. Our results highlighted the critical dependence of signal detection on biochemical sensors’ operating mechanisms and provided a valuable guidance for static and non-static assays for biomolecular detections.
Article
Concentration and separation of particles and biological specimens is a fundamental function of micro/nanofluidic systems. Acoustic streaming is an effective and biocompatible way to create rapid microscale fluid motion and induce particle capture, though the >100 MHz frequencies required to directly generate acoustic body forces on the microscale have traditionally been difficult to generate and localize in a way that is amenable to efficient generation of streaming. Moreover, acoustic, hydrodynamic and electrical forces as typically applied have difficulty manipulating specimens in the sub-micron regime. In this work, we introduce highly focused travelling surface acoustic waves (SAW) at high frequencies between 193-636 MHz for efficient and highly localized production of acoustic streaming vortices on microfluidic length scales. Concentration occurs via a novel mechanism, whereby the combined acoustic radiation and streaming field results in size-selective aggregation in fluid streamlines in the vicinity of a high-amplitude acoustic beam, as opposed to previous acoustic radiation induced particle concentration where objects typically migrate towards minimum pressure locations. Though the acoustic streaming is induced by a travelling wave, we are able to manipulate particles an order of magnitude smaller than possible using the travelling wave force alone. We experimentally and theoretically examine the range of particle sizes that can be captured in fluid streamlines using this technique, with rapid particle concentration demonstrated down to 300 nm diameters. We also demonstrate that locations of trapping and concentration are size-dependent, which is attributed to the combined effects of the acoustic streaming and acoustic forces.
Article
A new surface functionalization scheme for nano-Bio field effect transistors (FETs) using biocompatible polyelectrolyte thin films (PET) is developed. PET assemblies on Si nanowires (Si-NWs) are driven by electrostatic interactions between the positively charged polymer backbone and negatively charged Si/SiO2 surface. Such assemblies can be directly coated from PET aqueous solutions and result in a uniform nanoscale thin film, which is more stable compared to the conventional amine silanization. Short oligo-ethylene glycol chains are grafted on the PETs to prevent nonspecific protein binding. Moreover, the reactive groups of the polymer chains can be further functionalized to other chemical groups in specific stoichiometry for biomolecules detection. Therefore, it opens a new strategy to precisely control the functional group densities on various biosensor surfaces at the molecular level. In addition, such assemblies of the polymers together with the bound analytes can be removed with the pH stimulation resulting in regeneration of a bare sensor surface without compromising the integrity and performance of the Si-NWs. Thus, it is believed that the developed PET coating and sensing systems on Si-NW FETs represent a versatile, promising approach for regenerative biosensors which can be applied to other biosensors and will benefit real device applications, enhancing sensor lifetime, reliability, and repeatability.
Article
We report a simple method to remove nonspecifically adsorbed species from sensor surface and also improve the detection sensitivity of the sensor using tuneable alternating current (ac) electrohydrodynamics (ac-EHD) forces. These forces generated within few nanometers of an electrode surface (i.e., double layer) engender fluid flow within a serpentine channel containing a long array of the asymmetric electrode pairs, and can easily be tuned externally by changing the frequency and amplitude of the ac-EHD field. Under the optimized experimental conditions, we achieved a 3.5-fold reduction in nonspecific adsorption of non-target proteins with a 1000-fold enhancement in detection sensitivity of the device for the analysis of human epidermal growth factor receptor 2 (HER2) protein spiked in serum. This approach can be applicable in diverse fields including biosensors, cellular and molecular separation systems and biomedical applications to remove/reduce nonspecific adsorption of molecular and cellular species.
Article
Theories for calculating steady streaming associated with sound fields are reviewed, comparing the methods and approximations of various authors. Two illustrative problems are worked out, both for rectilinear flow due to irrotational sound fields. The first deals with a single attenuated plane wave traveling down a tube, as in Cady&apos;s quartz wind experiments. In the second, a pair of crossed plane waves is treated, giving rise to a quite different kind of streaming. In obtaining solutions, attention is given to boundary conditions; here, gradients of She excess static pressure, another second‐order quantity, come into consideration. Significantly, streaming speeds depend critically upon α, the attenuation constant, where α may be due to any common cause, such as heat conduction, scattering, thermal relaxation, etc. From these results it appears that streaming measurements cannot be used to distinguish between absorption mechanisms. Numerical values are given for a few cases; high flow speeds may be expected in a bubbly medium.
Article
Biosensors typically operate in liquid media for detection of biomarkers and suffer from fouling resulting from nonspecific binding of protein molecules to the device surface. In the current work, using a coupled field finite element fluid-structure interaction simulation, we have identified that fluid motion induced by high intensity sound waves, such as those propagating in these sensors, can lead to the efficient removal of the nonspecifically bound proteins thereby eliminating sensor fouling. We present a computational analysis of the acoustic-streaming phenomenon induced biofouling elimination by surface acoustic-waves (SAWs) propagating on a lithium niobate piezoelectric crystal. The transient solutions generated from the developed coupled field fluid solid interaction model are utilized to predict trends in acoustic-streaming induced forces for varying design parameters such as voltage intensity, device frequency, fluid viscosity, and density. We utilize these model predictions to compute the various interaction forces involved and thereby identify the possible mechanisms for removal of nonspecifically-bound proteins. For the range of sensor operating conditions simulated, our study indicates that the SAW motion acts as a body force to overcome the adhesive forces of the fouling proteins to the device surface whereas the acoustic-streaming induced hydrodynamic forces prevent their reattachment. The streaming velocity fields computed using the finite element models in conjunction with the proposed particle removal mechanism were used to identify the optimum conditions that lead to improved removal efficiency. We show that it is possible to tune operational parameters such as device frequency and input voltage to achieve effective elimination of biofouling proteins in typical biosensing media. Our simulation results agree well with previously reported experimental observations. The findings of this work have significant implications in designing reusable, selective, and highly sensitive biosensors.
Article
This paper demonstrates that surface plasmon resonance (SPR) spectroscopy can be used to measure the nonspecific adsorption of proteins to self-assembled monolayers (SAMs) of alkanethiolates on gold in situ and in realtime. Mixed SAMs comprising hexa(ethylene glycol) and methyl groups that have values of (chi Me) < 0.5 resisted the adsorption of four test proteins: RNase A, lysozyme, fibrinogen, and pyruvate kinase. These four proteins adsorbed irreversibly to surfaces having values of (chi Me) > 0.5: the amount of adsorbed protein correlated with (chi Me). The initial rate for adsorption of fibrinogen to a methyl-terminated SAM ((chi Me) = 1.0) followed first-order kinetics. The combination of SAMs and SPR described here is particularly well suited for investigations of the interactions of proteins with structurally well-defined organic surfaces.
Article
Acoustic streaming is a stationary fluid motion induced by intense ultrasound. This paper describes the mechanism of generation of acoustic streaming, theoretically and experimentally. A qualitative explanation in relation to acoustic radiation pressure is followed by a detailed discussion based on the formulation of the phenomenon. The spatial non-uniformity of the sound field and energy dissipation due to viscosity are requisite for the generation of streaming. In the development of streaming, two kinds of nonlinearity are important: one is fluid-dynamic nonlinearity, which accounts for effects of inertial force compared to viscous force in fluid motion, and the other is acoustic nonlinearity, which determines the driving force of streaming. © 1998 Scripta Technica, Electron Comm Jpn Pt 3, 81(10): 1–8, 1998
Article
This article reviews acoustic microfluidics: the use of acoustic fields, principally ultrasonics, for application in microfluidics. Although acoustics is a classical field, its promising, and indeed perplexing, capabilities in powerfully manipulating both fluids and particles within those fluids on the microscale to nanoscale has revived interest in it. The bewildering state of the literature and ample jargon from decades of research is reorganized and presented in the context of models derived from first principles. This hopefully will make the area accessible for researchers with experience in materials science, fluid mechanics, or dynamics. The abundance of interesting phenomena arising from nonlinear interactions in ultrasound that easily appear at these small scales is considered, especially in surface acoustic wave devices that are simple to fabricate with planar lithography techniques common in microfluidics, along with the many applications in microfluidics and nanofluidics that appear through the literature.
Article
Piezoelectric microelectromechanical systems (MEMS) resonant sensors, known for their excellent mass resolution, have been studied for many applications, including DNA hybridization, protein-ligand interactions, and immunosensor development. They have also been explored for detecting antigens, organic gas, toxic ions, and explosives. Most piezoelectric MEMS resonant sensors are acoustic sensors (with specific coating layers) that enable selective and label-free detection of biological events in real time. These label-free technologies have recently garnered significant attention for their sensitive and quantitative multi-parameter analysis of biological systems. Since piezoelectric MEMS resonant sensors do more than transform analyte mass or thickness into an electrical signal (e.g., frequency and impedance), special attention must be paid to their potential beyond microweighing, such as measuring elastic and viscous properties, and several types of sensors currently under development operate at different resonant modes (i.e., thickness extensional mode, thickness shear mode, lateral extensional mode, flexural mode, etc.). In this review, we provide an overview of recent developments in micromachined resonant sensors and activities relating to biochemical interfaces for acoustic sensors.
Article
A label-free biosensor for Escherichia coli (E. coli) ORN 178 based on faradaic electrochemical impedance spectroscopy (EIS) was developed. α-Mannoside or β-galactoside was immobilized on a gold disk electrode using a self-assembled monolayer (SAM) via a spacer terminated in a thiol functionality. Impedance measurements (Nyquist plot) showed shifts due to the binding of E. coli ORN 178, which is specific for α-mannoside. No significant change in impedance was observed for E. coli ORN 208, which does not bind to α-mannoside. With increasing concentrations of E. coli ORN 178, electron-transfer resistance (R(et)) increases before the sensor is saturated. After the Nyquist plot of E. coli/mixed SAM/gold electrode was modeled, a linear relationship between normalized R(et) and the logarithmic value of E. coli concentrations was found in a range of bacterial concentration from 10(2) to 10(3) CFU/mL. The combination of robust carbohydrate ligands with EIS provides a label-free, sensitive, specific, user-friendly, robust, and portable biosensing system that could potentially be used in a point-of-care or continuous environmental monitoring setting.
Article
All transducers used in biological sensing suffer from fouling resulting from nonspecific binding of protein molecules to the device surface. The acoustic-streaming phenomenon, which results from the fluid motion induced by high-intensity sound waves, can be used to remove these nonspecifically bound proteins to allow more accurate determinations and reuse of these devices. We present a computational and experimental study of the acoustic-streaming phenomenon induced by surface acoustic waves.A coupled-field fluid-structure interaction (FSI) model of a surface-acoustic-wave (SAW) device based on a micrometer-sized piezoelectric substrate (YZ-LiNbO3) in contact with a liquid loading was developed to study the surface-acoustic-wave interaction with fluid loading. The fluid domain was modeled using the Navier-Stokes equation; the arbitrary Lagrangian-Eulerian approach was employed to handle the mesh distortions arising from the motion of the solid substrate. The fluid-solid coupling was established by maintaining stress and displacement continuity at the fluid-structure interface. A transient analysis was carried out by applying a time-varying voltage to the transmitter interdigital transducer (IDT) fingers. Simulation results predict strong coupling of ultrasonic surface waves on the piezoelectric substrate with the thin liquid layer causing wave mode conversion from Rayleigh to leaky SAWs, which leads to acoustic streaming. The transient solutions generated from the FSI model were utilized to predict trends in acoustic-streaming velocity for varying design parameters such as voltage intensity, device frequency, fluid viscosity, and density. The induced streaming velocities typically vary from 1 mum/s to 1 cm/s, with the exact values dictated by the device operating conditions as well as fluid properties. Additionally, the model predictions were utilized to compute the various interaction forces involved and thereby identify the possible mechanisms for removal of nonspecifically bound proteins. Our study indicates that the SAW body force overcomes the adhesive forces of the fouling proteins to the device surface and the fluid-induced drag and lift forces prevent their reattachment. The streaming velocity fields computed using the finite-element model in conjunction with the proposed mechanism were used to identify the conditions leading to improved removal efficiency. Predictions of the model are in good agreement with those of simple analytical theories as well as the experimentally observed trends of nonspecific protein removal in typical SAW biosensing operations.
Article
The efficiency of a pre-absorbed bovine serum albumin (BSA) layer in blocking the non-specific adsorption of different proteins on hydrophobic and hydrophilic surfaces was evaluated qualitatively and quantitatively using infrared reflection spectroscopy supported by spectral simulations. A BSA layer with a surface coverage of 35% of a close-packed monolayer exhibited a blocking efficiency of 90-100% on a hydrophobic and 68-100% on a hydrophilic surface, with respect to the non-specific adsorption of concanavalin A (Con A), immunoglobulin G (IgG), and staphylococcal protein A (SpA). This BSA layer was produced using a solution concentration of 1 mg/mL and 30 min incubation time. BSA layers that were adsorbed at conditions commonly employed for blocking (a 12 h incubation time and a solution concentration of 10 mg/mL) exhibited a blocking activity that involved competitive adsorption-desorption. This activity resulted from the formation of BSA-phosphate surface complexes, which correlated with the conformation of adsorbed BSA molecules that was favourable for blocking. The importance of optimisation of the adsorbed BSA layer for different surfaces and proteins to achieve efficient blocking was addressed in this study.
Article
SPR imaging (SPRi) is at the forefront of optical label-free and real-time detection. It offers the possibility of monitoring hundreds of biological interactions simultaneously and from the binding profiles, allows the estimation of the kinetic parameters of the interactions between the immobilised probes and the ligands in solution. We review the current state of development of SPRi technology and its application including commercially available SPRi instruments. Attention is also given to surface chemistries for biochip functionalisation and suitable approaches to improve sensitivity.
Article
Tumor suppressor p53 is an attractive cancer therapeutic target because it can be functionally activated to eradicate tumors. Direct gene alterations in p53 or interaction between p53 and MDM2 proteins are two alternative mechanisms for the inactivation of p53 function. Designing small molecules to block the MDM2-p53 interaction and reactivate the p53 function is a promising therapeutic strategy for the treatment of cancers retaining wild-type p53. This review will highlight recent advances in the design and development of small-molecule inhibitors of the MDM2-p53 interaction as new cancer therapies. A number of these small-molecule inhibitors, such as analogs of MI-219 and Nutlin-3, have progressed to advanced preclinical development or early phase clinical trials.
Article
The current technology available for patterning proteins is reviewed. Examples of two-dimensional protein patterning using conventional photolithographic methods, photochemistry, and self-assembled monolayers are given. Consideration is also given to some major issues affecting protein patterning, including non-specific binding, protein pattern uniformity, and measurement techniques. S-layer nanopatterning and three-dimensional biochip patterning are discussed in possible future directions. In addition, a discussion of the impact of protein patterning technology on the field of biomaterials is discussed.
Article
A DNA piezoelectric sensor has been developed for the detection of genetically modified organisms (GMOs). Single stranded DNA (ssDNA) probes were immobilised on the sensor surface of a quartz crystal microbalance (QCM) device and the hybridisation between the immobilised probe and the target complementary sequence in solution was monitored. The probe sequences were internal to the sequence of the 35S promoter (P) and Nos terminator (T), which are inserted sequences in the genome of GMOs regulating the transgene expression. Two different probe immobilisation procedures were applied: (a) a thiol-dextran procedure and (b) a thiol-derivatised probe and blocking thiol procedure. The system has been optimised using synthetic oligonucleotides, which were then applied to samples of plasmidic and genomic DNA isolated from the pBI121 plasmid, certified reference materials (CRM), and real samples amplified by the polymerase chain reaction (PCR). The analytical parameters of the sensor have been investigated (sensitivity, reproducibility, lifetime etc.). The results obtained showed that both immobilisation procedures enabled sensitive and specific detection of GMOs, providing a useful tool for screening analysis in food samples.
Article
Dense, ultrathin networks of isocyanate terminated star-shaped poly(ethylene oxide) (PEO) molecules, cross-linked at their chain ends via urea groups, were shown to be extremely resistant to unspecific adsorption of proteins while at the same time suitable for easy biocompatible modification. Application by spin coating offers a simple procedure for the preparation of minimally interacting surfaces that are functionalized by suitable linker groups to immobilize proteins in their native conformations. These coatings form a versatile basis for biofunctional and biomimetic surfaces. We have demonstrated their advantageous properties by using single-molecule fluorescence microscopy to study immobilized proteins under destabilizing conditions. Biotinylated ribonuclease H (RNase H) was labeled with a fluorescence resonance energy transfer (FRET) pair of fluorescent dyes and attached to the surface by a biotin-streptavidin linkage. FRET analysis demonstrated completely reversible denaturation/renaturation behavior upon exposure of the surface-immobilized proteins to 6 M guanidinium chloride (GdmCl) followed by washing in buffer. A comparison with bovine serum albumin (BSA) coated surfaces and linear PEO brush surfaces yielded superior performance in terms of chemical stability, inertness and noninteracting nature of the star-polymer derived films.
Article
The success of microarrays, such as DNA chips, for biosample screening with minimal sample usage has led to a variety of technologies for assays on glass slides. Unfortunately, for small molecules, such as carbohydrates, these methods usually rely on covalent bond formation, which requires unique functional handles and multiple chemical steps. A new simpler concept in microarray formation is based on noncovalent fluorous-based interactions. A fluorous tail is designed not only to aid in saccharide purification but also to allow direct formation of carbohydrate microarrays on fluorous-derivatized glass slides for biological screening with lectins, such as concanavalin A. The noncovalent interactions in the fluorous-based array are even strong enough to withstand the detergents used in assays with the Erythrina crystagalli lectin. Additionally, the utility of benzyl carbonate protecting groups on fucose building blocks for the formation of alpha-linkages is demonstrated.
Article
A facile, self-assembly approach to the fabrication of a robust, mesoporous, biocompatible polymeric network for the spatial organization of proteins is described. Surface-deposited poly(styrene) (PS) beads that assemble into a two-dimensional (2-D) hexagonal array are used to template cross-linked poly(vinyl alcohol) (PVA), yielding an inverse opal structure. The porous, water insoluble network is used to entrain a model, soluble protein, green fluorescent protein (GFP). The polymeric network is characterized by atomic force microscopy (AFM) and optical microscopy, and the spatial localization of the incorporated GFP is determined by fluorescence microscopy. The results demonstrate that this system may constitute a versatile platform for the lateral organization of biomolecules.
Article
A novel anti-fouling mechanism based on the combined effects of electric field and shear stress is reported. A lead zirconate titanate (PZT) composite is used to generate an electric field and an acoustic streaming shear stress that increase nanomolecule desorption. In vitro characterization showed that (1) 58+/-5.5% and 39+/-5.2% of adsorbed bovine serum albumin (BSA) proteins can be effectively removed from fired silver and titanium coated PZT plate, respectively; and (2) 43+/-9.7% of the anti-mouse immunoglobulin G (IgG) can be effectively removed from a fired silver coated PZT plate. Theoretical calculations on protein-surface interactions (van der Waals (VDW), electrostatic, and hydrophobic) and shear stress describe the mechanism for protein desorption from model surfaces. We have shown that the applied electric potential is the major contributor in reducing the adhesive force between protein and surface, and the desorbed protein is taken away by acoustic streaming shear stress. We strongly believe that the present method offers the possibility of minimizing nanomolecule adsorption without further surface treatment.
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Testing for prostate-specific antigen (PSA) has profoundly affected the diagnosis and treatment of prostate cancer. PSA testing has enabled physicians to detect prostate tumours while they are still small, low-grade and localized. This very ability has, however, created controversy over whether we are now diagnosing and treating insignificant cancers. PSA testing has also transformed the monitoring of treatment response and detection of disease recurrence. Much current research is directed at establishing the most appropriate uses of PSA testing and at developing methods to improve on the conventional PSA test.
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This study examined six different polymer and self-assembled monolayer (SAM) surface modifications for their interactions with human serum and plasma. It was demonstrated that zwitterionic polymer surfaces are viable alternatives to more traditional surfaces based on poly(ethylene glycol) (PEG) as nonfouling surfaces. All polymer surfaces were formed using atom transfer radical polymerization (ATRP) and they showed an increased resistance to nonspecific protein adsorption compared to SAMs. This improvement is due to an increase in the surface packing density of nonfouling groups on the surface, as well as a steric repulsion from the flexible polymer brush surfaces. The zwitterionic polymer surface based on carboxybetaine methacrylate (CBMA) also incorporates functional groups for protein immobilization in the nonfouling background, making it a strong candidate for many applications such as in diagnostics and drug delivery.
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Nonspecific binding of proteins is an ongoing problem that dramatically reduces the sensitivity and selectivity of biosensors. We demonstrate that ultrasonic waves generated by surface acoustic wave (SAW) devices remove nonspecifically bound proteins from the sensing and nonsensing regions of the microarrays. We demonstrate our approach for controllably and nondestructively cleaning the microarray interface. In this work, SAWs were generated using 128 YX lithium niobate, chosen for its high coupling coefficient and efficient power transfer to mechanical motion. These waves propagating along the surface were coupled into specifically bound and nonspecifically bound proteins on a patterned surface of 40 mum feature size. Fluorescence intensity was used to quantify cleaning efficacy of the microarrays. Our results have shown that excess protein layers and aggregates are removed leaving highly uniform films as evidenced by fluorescence intensity profiles. Selected antigen-receptor interactions remained bound during the acoustic cleaning process when subjected to 11.25 mW of power and retained their efficacy for subsequent antigen capture. Results demonstrate near-complete fluorescence signal recovery for both the sensing and nonsensing regions of the microarrays. Of significance is that our approach can be integrated into existing array technologies where sensing and nonsensing regions are extensively fouled. We believe that this technology will be pivotal in the development and advancement of microsensors and their biological applications.
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In this paper, theories of particle removal by high-frequency ultrasonic waves are discussed and tested against recent experimental data. First, the principal adhesion forces such as van der Waals forces are briefly reviewed and the typical uncertainties in their size in particle-surface systems are assessed. The different ultrasound-induced forces-linear forces such as added mass, drag, lift, and Basset forces and nonlinear ones due to radiation pressure, and drag exerted by acoustic streaming-are discussed and their magnitudes are evaluated for typical cleaning operations. It is shown that high-frequency ultrasound can clean particles most effectively in media with properties like water because: (1) the wavelength can be made comparable to the particle radius to promote effective sound-particle interaction; (2) the viscous boundary layer is thin, minimizing particle "hide-out;" and (3) both the added mass and radiation pressure forces exceed typical adhesion forces at high frequencies. Based on these analyses, possible mechanisms of particle removal are discussed and interpreted in terms of experimental observations of particle cleaning.< >