Gravimetric Biosensors Research Articles

Microfluidic QCM enables ultrahigh Q-factor: a new paradigm for in-liquid gravimetric sensing

Acoustic gravimetric biosensors attract attention due to their simplicity, robustness, and low cost. However, a prevailing challenge in these sensors is dissipation which manifests in a low quality factor (Q-factor), which limits their sensitivity and accuracy. To mitigate dissipation of acoustic sensors in liquid environments we introduce an innovative approach in which we combine microfluidic channels with gravimetric sensors. To implement this novel paradigm we chose the quartz crystal microbalance (QCM) as our model system, owing to its wide applicability in biosensing and the relevance of its operating principles to other types of acoustic sensors. We postulate that the crucial determinant for enhancing performance lies in the ratio between the width of the microfluidic channels and the wavelength of the pressure wave generated by the oscillating channel side walls driven by the QCM. Our hypothesis is supported by finite element analysis (FEA) and dimensional studies, which revealed two key factors that affect device performance: (1) the ratio of the channel width to the pressure wavelength (W/λp\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$W/{\\lambda }_{{\\rm {p}}}$$\\end{document}) and (2) the ratio of the channel height to the shear evanescent wavelength (H/λs\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$H/{\\lambda }_{{\\rm {s}}}$$\\end{document}). To validate our hypothesis, we fabricated a microfluidic QCM (µ-QCM) and demonstrated a remarkable 10-fold improvement in its dissipation when compared to conventional QCM. The novel microfluidic approach offers several additional advantages, such as direct data interpretation, reduced volume requirement for sample liquids, and simplified temperature control, augmenting the sensor’s overall performance. By fostering increased sensitivity, accuracy, and ease of operation, our novel paradigm unlocks new possibilities for advancing gravimetric technologies, potentially for biosensing applications.

Acoustic radiation-free surface phononic crystal resonator for in-liquid low-noise gravimetric detection

Acoustic wave resonators are promising candidates for gravimetric biosensing. However, they generally suffer from strong acoustic radiation in liquid, which limits their quality factor and increases their frequency noise. This article presents an acoustic radiation-free gravimetric biosensor based on a locally resonant surface phononic crystal (SPC) consisting of periodic high aspect ratio electrodes to address the above issue. The acoustic wave generated in the SPC is slower than the sound wave in water, hence it prevents acoustic propagation in the fluid and results in energy confinement near the electrode surface. This energy confinement results in a significant quality factor improvement and reduces frequency noise. The proposed SPC resonator is numerically studied by finite element analysis and experimentally implemented by an electroplating-based fabrication process. Experimental results show that the SPC resonator exhibits an in-liquid quality factor 15 times higher than a conventional Rayleigh wave resonator at a similar operating frequency. The proposed radiation suppression method using SPC can also be applied in other types of acoustic wave resonators. Thus, this method can serve as a general technique for boosting the in-liquid quality factor and sensing performance of many acoustic biosensors.

Direct growth of few-layer graphene on AlN-based resonators for high-sensitivity gravimetric biosensors.

We present the successful growth of few-layer graphene on top of AlN-based solidly mounted resonators (SMR) using a low-temperature chemical vapour deposition (CVD) process assisted by Ni catalysts, and its effective bio-functionalization with antibodies. The SMRs are manufactured on top of fully insulating AlN/SiO2 acoustic mirrors able to withstand the temperatures reached during the CVD growth of graphene (up to 650 °C). The active AlN films, purposely grown with the c-axis tilted, effectively excite shear modes displaying excellent in-liquid performance, with electromechanical coupling and quality factors of around 3% and 150, respectively, which barely vary after graphene integration. Raman spectra reveal that the as-grown graphene is composed of less than five weakly coupled layers with a low density of defects. Two functionalization protocols of the graphene are proposed. The first one, based on a covalent binding approach, starts with a low-damage O2 plasma treatment that introduces a controlled density of defects in graphene, including carboxylic groups. After that, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride/N-hydroxysuccinimide (EDC/NHS) chemistry is used to covalently bind streptavidin molecules to the surface of the sensors. The second functionalization protocol is based on the non-covalent bonding of streptavidin on hydrophobic graphene surfaces. The two protocols end with the effective bonding of biotinylated anti-IgG antibodies to the streptavidin, which leaves the surface of the devices ready for possible IgG detection.

Gravimetric biosensor based on a 1.3 GHz AlN shear-mode solidly mounted resonator

We investigate the performance of solidly mounted resonators based on Ir/tilted-AlN/Ir piezoelectric stacks as biosensors. These films are deposited by varying the pressure, the cathode power and the temperature of a two-step process based on depositing (00·2)-tilted AlN active layers over an (10·3)-oriented AlN seed layer. To minimize the influence of the temperature coefficient of frequency on the stability of the biosensor, we use insulating acoustic mirrors made of layers of SiO2 and amorphous TaOx with non-λ/4 thicknesses, which enables to reduce the TCF to −14ppm/°C. The mass loading of the resonators with SiO2 thin films results in a sensitivity of 1800kHz/pg·cm2. Surface functionalization consists on the binding of silane groups on plasma oxidized SiO2 surfaces. After a glutaraldehyde link, streptavidin is bonded to the surface to receive biotinylated receptors for several species. We test thrombin-binding aptamer (TBA29 against thrombin, and IgG antibody against immunoglobulin). The sensors response to species of different molecular weight like TBA-29 (9.75kDa) or IgG antibody (150kDa) is monitored. Finally, we assess the response of the biosensors to different thrombin concentrations (ranging from 4nM to 270nM) on surfaces functionalized with the TBA29 aptamer.

Direct Comparison of the Sensitivity of QCMs and AlN-based TFRs Biosensors

We present the direct comparison of the performance of two gravimetric biosensors based on acoustic resonators, a quartz crystal microbalance and a high frequency AlN-based bulk acoustic wave film solidly mounted resonator (SMR). Both sensors are functionalized with streptavidin to detect the response to TBA29 aptamer biotin modified and different concentrations of thrombin. Experimental results reveal that both sensors succeed in detecting the targeted species, although SMRs show significantly greater sensitivity and a lower limit of detection.

Bacteria Detection with High-frequency Gravimetric Biosensors Based on AlN Thin Film Resonators

Gravimetric sensors based on shear-mode resonators are suitable for in-liquid detection of biological species because their quality factors barely decrease during in-liquid operation. However, we have found that in the particular case of large ligands, such as bacteria, the transmission of the surface movement to them appears to be more efficient when movement takes place normal to the surface (longitudinal modes) instead of to parallel to it (shear modes). In this work, we succeeded in detecting bacteria with AlN-based bulk acoustic wave solidly mounted resonators operating in longitudinal modes at 2 GHz that we were unable to detect with shear modes.

Effects of biologically compatible buffers on the electrical response of gravimetric sensors operating at GHz frequencies

Gravimetric biosensors based on piezoelectric resonators have their active area just atop an electrical contact. Sometimes both contacts are located on the same surface (coplanar electrodes). The use of bio-buffers with high ionic content makes often electrical isolation of the electrodes from the liquid mandatory. This restricts the use of surfaces for functionalization and worsens the response of the resonator. In this paper we demonstrate that, if operating frequencies are high enough, there is little effect of the liquid ionic conductivity on the performance of the resonators with both electrodes in contact with the latter. We test the response of 1.32GHz AlN shear mode resonators in NaCl solutions of different concentrations to show the effects of the ionic conductivity on their performance. The conclusion is that, even the presence of a parallel conductance due to the ionic conductivity into the liquid, the shear mode resonance is not significantly disturbed. To prove the viability of non-isolated AlN resonators as biosensors, we present measurements of thrombin detection using devices with aptamer-functionalized iridium electrode surfaces.

On-chip temperature-compensated Love mode surface acoustic wave device for gravimetric sensing

Love mode surface acoustic wave (SAW) sensors have been recognized as one of the most sensitive devices for gravimetric sensors in liquid environments such as bio sensors. Device operation is based upon measuring changes in the transmitted (S21) frequency and phase of the first-order Love wave resonance associated with the device upon on attachment of mass. However, temperature variations also cause a change in the first order S21 parameters. In this work, shallow grooved reflectors and a “dotted” single phase unidirectional interdigitated transducer (D-SPUDT) have been added to the basic SAW structure, which promote unidirectional Love wave propagation from the device's input interdigitated transducers. Not only does this enhance the first-order S21 signal but also it allows propagation of a third-order Love wave. The attenuation coefficient of the third-order wave is sufficiently great that, whilst there is a clear reflected S11 signal, the third-order wave does not propagate into the gravimetric sensing area of the device. As a result, whilst the third-order S11 signal is affected by temperature changes, it is unaffected by mass attachment in the sensing area. It is shown that this signal can be used to remove temperature effects from the first-order S21 signal in real time. This allows gravimetric sensing to take place in an environment without the need for any other temperature measurement or temperature control; this is a particular requirement of gravimetric biosensors.

Silaffin Peptides as a Novel Signal Enhancer for Gravimetric Biosensors

Application of biomimetic silica formation to gravimetric biosensors has been conducted for the first time. As a model system, silaffin peptides fused with green fluorescent protein (GFP) were immobilized on a gold quartz crystal resonator for quartz crystal microbalances using a self-assembled monolayer. When a solution of silicic acid was supplied, silica particles were successfully deposited on the Au surface, resulting in a significant change in resonance frequency (i.e., signal enhancement) with the silaffin-GFP. However, frequency was not altered when bare GFP was used as a control. The novel peptide enhancer is advantageous because it can be readily and quantitatively conjugated with sensing proteins using recombinant DNA technology. As a proof of concept, this study shows that the silaffin domains can be employed as a novel and efficient biomolecular signal enhancer for gravimetric biosensors.

AlN-based BAW resonators with CNT electrodes for gravimetric biosensing

Solidly mounted resonators (SMRs) with a top carbon nanotubes (CNTs) surface coating that doubles as an electrode and as a sensing layer have been fabricated. The influence of the CNTs on the frequency response of the resonators was studied by direct comparison to identical devices with a top metallic electrode. It was found that the CNTs introduced significantly less mass load on the resonators and these devices exhibited a greater quality factor, Q (>2000, compared to ∼1000 for devices with metal electrodes), which increases the gravimetric sensitivity of the devices by allowing the tracking of smaller frequency shifts. Protein solutions with different concentrations were loaded on the top of the resonators and their responses to mass-load from physically adsorbed coatings were investigated. Results show that resonators using CNTs as the top electrode exhibited a higher frequency change for a given load (∼0.25 MHz cm 2 ng −1) compared to that of a metal thin film electrode (∼0.14 MHz cm 2 ng −1), due to the lower mass of the CNT electrodes and their higher active surface area compared to that of a thin film metal electrode. It is therefore concluded that the use of CNT electrodes on resonators for their use as gravimetric biosensors is a significant improvement over metallic electrodes that are normally employed.

Deposition and characterisation of ultralow-stress ZnO thin films for application in FBAR-based gravimetric biosensors

Zinc oxide (ZnO) thin films were deposited at high rates ( > 50 nm min –1 ) using a unique technique known as high target utilisation sputtering (HiTUS). The films obtained possess good crystallographic orientation, low surface roughness, very low stress and excellent piezoelectric properties. We have utilised the films to develop highly sensitive biosensors based on thickness longitudinal mode (TLM) thin film bulk acoustic resonators (FBARs). The FBARs have the fundamental TLM at a frequency near 1.5 GHz and quality factor Q higher than 1,000, which is one of the largest values ever reported for ZnO-based FBARs. Bovine Serum Albumin (BSA) solutions with different concentrations were placed on the top of different sets of identical FBARs and their responses to mass-loading from physically adsorbed protein coatings were investigated. These resonators demonstrated a high sensitivity and thus have a great potential as gravimetric sensors for biomedical applications.

Modelling of a Piezoelectric Polymer Film System for Biosensing Applications

Most piezoelectric gravimetric biosensors are based on quartz crystal microbalances or surface acoustic wave devices. In this paper we describe a polymer film system, made of piezoelectric polyvinylidene difluoride (PVDF) and the Immobilon-P membrane (porous PVDF), for biosensing applications. In operation a film is accommodated in a flow cell and connected to an electronic circuit, constituting an oscillatory resonant device; the output signal is its resonance frequency. This device successfully detected the binding between bovine IgG and bovine anti-IgG. Work on the modelling of this film system is presented. Two different approaches are being considered: the finite element method (FEM) and the ABCD matrices. Results comparing theoretical and experimental data are presented.

Nanoscale Labels: Nanoparticles and Liposomes in The Development of High-Performance Biosensors

Technology for the detection of biological species has generated considerable interest in a variety of fields including healthcare, defense, food and environmental monitoring. In a biosensor, labeled specific binding partners are used to emit a detectable signal. Owing to their unique properties, nanomaterials have been proposed as a novel label category and have led to the development of new assays and new transduction mechanisms. In this article, the role of three major types of nanoscale labels (metallic, semiconductor and liposome nanoparticles) in the development of a new generation of optical, electrochemical or gravimetric biosensors will be presented. The underlying transduction principles will be briefly explained and assay strategies relying on the use of these 'nanolabels' will be described. The contribution to increased assay performance and sensitivity will be highlighted. Approaches towards simple, cost efficient and sensitive assays are essential to meet the demands of a growing number of applications.

A new PZT piezoelectric sensor for gravimetric applications using the resonance-frequency detection

Piezoelectric sensors made of quartz, Pb(Zr,Ti)O 3, Pb(La,Zr,Ti)O 3, or Pb(Zr,Ti,Sn)O y have rapidly been developed recently because of the potential applications in devices such as biosensors, accelerometers, pressure sensors, and ultrasonic transducers. However, the development of devices with reduced size but with improved sensitivity is highly important. With this idea, the study aims to develop a lead zironate titanate (PZT) chip as a novel gravimetric biosensor using the resonance feature in biomolecule detection. The PZT thin film characterized by the X-ray diffractometer (XRD) has a perovskite structure. The fabricated PZT sensor was designed with 250 μm × 250 μm-sensing areas of electrodes, determined by the measured impedance at 98.5 MHz frequency. The morphology and cross-section of scanning electron microscope (SEM) images show that the thickness of the spin-on PZT layer was about 500 nm and the crystal grain size was less than 50 nm. Moreover, an evaluation circuit consisted of a resonance circuit and a frequency-to-voltage circuit was established to detect bovine serum albumin (BSA) protein. The PZT sensor, combined with evaluation circuit, showed a high sensitivity of 62.8 Hz cm 2/ng. The detection sensitivity was enhanced by 300 folds than that of traditional quartz crystal microbalance (QCM).

Piezoquartz biosensors for the analysis of environmental objects, foodstuff and for clinical diagnostic

It is reviewed opportunities and practical application features of gravimetric biosensors for determination of high-and low-molecular biologically active substances and microorganisms in environments, food stuffs and clinical diagnostics. Characteristics of piezosensors based on different nature receptor molecules (an antibodies, antigens, hapten-protein conjugates etc.) are given. It’s reflected the modern view on the wide range of questions, connected with their function in liquids. The special attention is directed towards immunosensors, the most perspective for quantification of trace concentrations in complex matrixes. Possible immunoassay formats in liquid (direct and competitive detection) are given. The questions of formation and regeneration of sensitive layer on the base of biomolecules of different nature are discussed.

Development of a Biosensor Based on a Piezoelectric Film

Most gravimetric biosensors use thin piezoelectric quartz crystals (quartz crystal microbalance, QCM), either as resonating crystals, or as bulk/surface acoustic wave (SAW) devices. This paper illustrates a related technique where a polymer film system made of two different types of the polymer polyvinylidene difluoride (PVDF) are used: one has piezoelectric properties and the other has a high protein binding capacity. Acoustic waves are launched in this polymer film to produce an oscillatory resonant device. This device is being tested in protein detection. Current work and preliminary results are presented.

Gravimetric biosensors based on acoustic waves in thin polymer films

Most gravimetric biosensors use thin piezoelectric quartz crystals, either as resonating crystals (quartz crystal microbalance, QCM), or as bulk/surface acoustic wave (SAW) devices. In the majority of these the mass response is inversely proportional to the crystal thickness which, at a limit of about 150 microns, gives inadequate sensitivity. A new system is described in which acoustic waves are launched in very thin (10 microns) tensioned polymer films to produce an oscillatory device. A theoretical equation for this system is almost identical to the well-known Sauerbrey equation used in the QCM method. Because the polymer films are so thin, a 30-fold increase in sensitivity is predicted and verified by adding known surface masses. Temperature sensitivity is a problem so a separate control sensor and careful temperature regulation are necessary. Preliminary results showing the real time binding of protein (IgG), a step towards immunosensor development, and the use of mass enhancing particles are presented. Inexpensive materials are used so disposable gravimetric biosensors may become feasible.