Magnetoelastic Biosensor Research Articles

Performance of Optimized Phage-Based Magnetoelastic Biosensors for Salmonella Typhimurium Detection on Tomatoes

An optimized, phage-based magnetoelastic (ME) biosensor method was performed to detect Salmonella Typhimurium contaminates on the surface of tomatoes and to determine the detection limit. After serial inoculations of S. Typhimurium on the surface of the tomatoes, the S. Typhimurium on the surfaces was analyzed using an optimized ME biosensor method. Detection limit and sensitivity of the ME biosensor for detection of S. Typhimurium on tomatoes were determined. The resonant frequency shifts of the ME measurement sensors increased linearly with an increase in the concentration of S. Typhimurium. Contrastingly, the control sensors showed relatively constant and low magnitude frequency shifts. Detection limit and sensitivity of the ME biosensor method was determined to be (1.78 {plus minus} 0.17) log CFU/cm2 and 922.7 Hz.

Signal Optimization for Salmonella Typhimurium Detection on Food Surfaces Using Phage-Based Biosensors

This paper presents the direct detection of Salmonella typhimurium on egg shells using a phage-based magnetoelastic (ME) biosensor. The ME biosensor consists of an ME resonator as the sensor platform and E2 phage as the bio-recognition element. The bio-recognition element is genetically engineered to bind specifically with Salmonella typhimurium. The ME biosensor, a wireless sensor, vibrates with a characteristic resonant frequency under an externally applied magnetic field. Multiple sensors can easily be remotely monitored. Multiple measurement and control sensors were placed on egg shells contaminated by Salmonella typhimurium solutions with different known concentrations. The resonant frequency of sensors before and after the exposure to the spiked egg shells was measured. The frequency shift of the measurement sensors was significantly different than the control sensors indicating Salmonella contamination. Scanning electron microscopy was used to confirm binding of Salmonella to the sensor surface and the resulting frequency shift results.

The analytical comparison of phage-based magnetoelastic biosensor with TaqMan-based quantitative PCR method to detect Salmonella Typhimurium on cantaloupes

A phage-based magnetoelastic (ME) biosensor method for the detection of Salmonella Typhimurium was compared with a TaqMan-based quantitative real-time PCR (qPCR) method on cantaloupes. The selectivity of a qPCR primer set was investigated and different DNA extraction methods were compared. The limits of detection (LOD) of both detection methods were determined by serial inoculation of S. Typhimurium suspensions on cantaloupe surfaces. The repeatability of both detection methods was determined over three-day periods. Among 74 bacteria tested, only S. Typhimurium exhibited positive amplification, and other bacterial strains did not show any positive amplification. The DNeasy tissue kit showed the best performance for DNA purification due to the significantly superior threshold cycle (Ct) values, compared with either direct applications of qPCR without any extraction procedure or after employing a simple boiling method for extraction. The LOD and repeatability of qPCR and ME biosensor methods for S. Typhimurium detection were determined to be 1.35 ± 0.07 and 2.47 ± 0.50 log CFU/2 mm2 surface of cantaloupe, and 2.41 and 6.28%, respectively. The results demonstrate that the ME biosensor method is one of promising and practical on-site detection methods for S. Typhimurium on fresh produce.

Kinetic Analysis of Glucose with Wireless Magnetoelastic Biosensor

Kinetic Analysis of Glucose with Wireless Magnetoelastic Biosensor

Effects of Surface Morphologies of Fresh Produce on the Performance of Phage-Based Magnetoelastic Biosensors

This study investigated the effects of different surface morphologies on the performance of phage-based magnetoelastic (ME) biosensors for Salmonella Typhimurium detection on selected fresh produce. The ME biosensor is composed of an ME resonator platform and filamentous E2 phage for the specific recognition of S. Typhimurium. After SEM observation of the surface morphologies of tomatoes and spinach (adaxial/abaxial), S. Typhimurium was inoculated serially onto the surfaces to examine the attachment and distribution of bacterial cells on the surfaces. Finally, the linearity and detection limit of the ME biosensors detecting S. Typhimurium on food surfaces were tested. The morphologies of tomato and spinach surfaces were obviously different due to the existence of stomata and the differences in roughness and curvature. However, there were no obvious differences in attachment and distribution of bacterial cells on the surfaces. The resonant frequency shifts of measurement sensors increased linearly with an increase in the concentration of S. Typhimurium in contrast to relatively constant and negligible shifts of control sensors. Detection limit of the ME biosensor method was determined to be 1.87 log CFU/cm2 for the tomatoes, 1.72 log CFU/cm2 for the adaxial surface of spinach, and 2.16 log CFU/cm2 for the abaxial surface of spinach.

Evaluation of phage-based magnetoelastic biosensors for direct detection of Salmonella Typhimurium on spinach leaves

Abstract For a phage-based magnetoelastic (ME) biosensor method to become accepted for evaluating foodborne contamination, the method of use must be objectively compared, evaluated and validated against more widely accepted standard methods. In this study, a ME biosensor method was evaluated by comparison with TaqMan-based quantitative PCR (qPCR) for the detection of Salmonella Typhimurium on spinach leaves. Limit of detection (LOD) was determined after serial inoculation of S. Typhimurium on spinach leaves. The LOD for the ME biosensor was 2.17 and 1.94 log CFU/spinach for adaxial and abaxial surface, respectively. The LOD for qPCR was 1.37 log CFU/spinach. The repeatability of both methods was measured over a three day period by inoculation of S. Typhimurium on spinach leaves. The repeatability of the ME biosensor method was determined to be 6.38%, competitive with qPCR at 1.92%. For the direct comparison of both methods, 3 log CFU/spinach of S. Typhimurium was grown on two groups of 25 spinach leaves for 24 h and the number of S. Typhimurium was quantified by both methods. After confirmation of the growth, S. Typhimurium was positively detected and the quantified numbers were 5.79 ± 0.88 and 6.11 ± 0.26 log CFU/spinach for the ME biosensor and the qPCR method, respectively. This study demonstrated that the ME biosensor method was competitive and promising as an on-site and in-field detection method for the detection of pathogens.

Amorphous metallic glass biosensors

Amorphous metallic glasses possess a unique combination of magnetostriction and extremely soft magnetic properties making them ideal candidates for developing high performance biosensors. This paper presents the results of an investigation of novel free-standing magnetoelastic (ME) biosensors based on Fe–B amorphous metallic glasses. The principle of operation of wireless ME biosensors and the advantages of metallic glasses for their construction are discussed. The materials with the highest possible elastic–magnetic energy conversion efficiency are favored for sensor applications. Due to the unique properties of amorphous metallic glasses, acoustic wave sensors based on amorphous ME alloys with mild magnetostriction properties show a very high elastic–magnetic energy conversion efficiency. In this study, ME biosensor resonators 4 × 100 × 500 μm in size were manufactured by dual beam sputtering and non-traditional microelectronic fabrication techniques. E2 phage, genetically engineered to bind with Salmonella typhimurium, was immobilized on the sensor surfaces as a bio-recognition element. The detection of Salmonella in liquid using these phage-based ME biosensors was demonstrated. The ME biosensor exhibited high sensitivity and a detection limit better than 50 CFU/mL.

Rapid and sensitive detection of Salmonella Typhimurium on eggshells by using wireless biosensors.

This article presents rapid, sensitive, direct detection of Salmonella Typhimurium on eggshells by using wireless magnetoelastic (ME) biosensors. The biosensor consists of a freestanding, strip-shaped ME resonator as the signal transducer and the E2 phage as the biomolecular recognition element that selectively binds with Salmonella Typhimurium. This ME biosensor is a type of mass-sensitive biosensor that can be wirelessly actuated into mechanical resonance by an externally applied timevarying magnetic field. When the biosensor binds with Salmonella Typhimurium, the mass of the sensor increases, resulting in a decrease in the sensor's resonant frequency. Multiple E2 phage-coated biosensors (measurement sensors) were placed on eggshells spiked with Salmonella Typhimurium of various concentrations (1.6 to 1.6 × 10(7) CFU/cm(2)). Control sensors without phage were also used to compensate for environmental effects and nonspecific binding. After 20 min in a humidity-controlled chamber (95%) to allow binding of the bacteria to the sensors to occur, the resonant frequency of the sensors was wirelessly measured and compared with their initial resonant frequency. The resonant frequency change of the measurement sensors was found to be statistically different from that of the control sensors down to 1.6 × 10(2) CFU/cm(2), the detection limit for this work. In addition, scanning electron microscopy imaging verified that the measured resonant frequency changes were directly related to the number of bound cells on the sensor surface. The total assay time of the presented methodology was approximately 30 min, facilitating rapid detection of Salmonella Typhimurium without any preceding sampling procedures.

The effect of incubation time for Salmonella Typhimurium binding to phage-based magnetoelastic biosensors

This study presents an investigation of the effect of incubation time on the binding of Salmonella Typhimurium (Salmonella) with E2 phage-based magnetoelastic (ME) biosensors. After inoculation of Salmonella on a tomato surface, the attachment and existence of Salmonella on the tomato surface was observed using scanning electron microscopy (SEM). Measurement sensors with immobilized E2 phage and control sensors devoid of phage were placed on the inoculated tomato surface and incubated for various times. The extent of binding of Salmonella to the ME biosensors was determined by measuring the resonant frequency shift of the sensors and confirmed by SEM. The attachment of Salmonella on the tomato surface increased as the inoculation concentration increased. Significant increases in the frequency shifts for the measurement sensors were observed between 15 and 30min (P<0.001), exhibiting 3439±185 and 5312±248Hz, respectively. Further incubation times did not increase Salmonella binding with E2 phage whereas longer incubation times increased non-specific attachment from residual nutrients on the tomato surface. In contrast, there was either no or negligible binding of Salmonella on the control sensors. This study will contribute to the development of a practical protocol for the ME biosensor method.

Comparison of Phage-Based Magnetoelastic Biosensors with Taqman- Based Quantitative Real-Time PCR for the Detection of Salmonella typhimurium Directly Grown on Tomato Surfaces

A phage-based magnetoelastic (ME) biosensor method was compared with a TaqMan-based quantitative real- time PCR (Q-PCR) method for the detection of Salmonella typhimurium on tomato surfaces. This ME biosensor method utilizes magnetoelastic resonators coated with E2 filamentous phage to bind with and measure the concentration of S. typhimurium . In this study, standard curves, correlations, and limits of detection (LOD) for the ME biosensor and Q-PCR methods were determined by inoculating tomato surfaces with S. typhimurium suspensions in concentrations ranging from 1 to 8 log CFU/tomato. The LOD for the ME biosensor method and Q-PCR were 3 and 2 log CFU/tomato, respectively. In a direct comparison of the detection methods, S. typhimurium suspensions (3 log CFU/tomato) were inoculated on 65 tomato surfaces, then incubated at 37°C and 100% RH for 24 h. After 24 h, S. typhimurium was positively detected by both methods and the quantified concentrations were nearly the same, (6.35 ± 2.03) and (6.34 ± 0.17) log CFU/tomato respectively for the ME biosensor method and the Q-PCR method, which were significantly greater than the concentration determined by the BGS-plate count method (5.33 ± 0.21). Scanning electron microscopy (SEM) was used to confirm the growth of S. typhimurium on the tomato surfaces and the binding of S. typhimurium on the measurement sensors. This study demonstrated that the ME biosensor method was robust and competitive with Q-PCR for S. typhimurium detection on fresh produce.

Blocking Agent Optimization for Nonspecific Binding on Phage Based Magnetoelastic Biosensors

Magnetoelastic biosensors have been studied for the past few years for use in detecting bacteria for food safety. These magnetoelastic biosensors are composed of a bio-molecular recognition element and transducer platform (magnetoelastic resonator). Phages (bio-molecular recognition element) have been developed to provide very specific binding with Salmonella typhimurium bacteria. However, there is some binding (non-specific binding) of bacteria to the gold areas not covered with phage on both the measurement and control sensors. In those areas, a blocking agent is necessary to reduce the non-specific binding. The purpose of this study is to compare three of the often used blocking agents, BSA, milk and casein in different concentrations to determine the agent and the optimum concentration for blocking. To accomplish this, a set of measurement and control sensors were prepared using identical procedures except the control sensors were not immobilized with phage. The measurement and control sensors were then coated with a blocking agent of concentrations ranging from 0.1 to 5 mg/mL. SEM observations show that coating the magnetoelastic biosensors with BSA at a concentration of 1mg/mL yields the largest amount of specific binding of bacteria cells while also maintaining the largest ratio of specific to non-specific binding. This binding characteristic is a result of the physical and electrical structure of the blocking proteins. To verify that the blocking solutions made good contact with the sensor's surface, contact angle measurements were used to evaluate the wettability of the surface. Changes in the resonant frequency of the biosensors were found to agree with the results from SEM observations.

Detection of Salmonella typhimurium Grown Directly on Tomato Surface Using Phage-Based Magnetoelastic Biosensors

A phage-based magnetoelastic (ME) biosensor method is being developed for on-site pathogen detection in fresh fruits and vegetables. Salmonella typhimurium was directly grown with minimal nutrients on tomato surfaces in order to mimic natural environmental conditions. S. typhimurium was inoculated on the surface of fresh tomatoes, and ME biosensors were used to detect the bacteria. The populations of S. typhimurium after 24-h incubation time at 37°C and 100% relative humidity were 6.1 and 7.8 logCFU/cm2 after starting with initial inoculations of 3.0 and 5.0 logCFU/cm2, respectively. After evident growth, measurement sensors with E2 phage immobilized on their surfaces and control sensors devoid of E2 phage were placed on the inoculated tomato surfaces and the resonant frequency shifts were measured. As the population of S. typhimurium increased, the resonant frequency shifts of the measurement sensors significantly increased (P < 0.01), exhibiting 6,680 ± 665 and 9,384 ± 457 Hz for the populations of 6.1 and 7.8 logCFU/cm2, respectively. SEM images confirmed that the measurement sensor resonant frequency shifts were due to S. typhimurium binding with E2 phage. This study demonstrated that the ME biosensor method could detect S. typhimurium grown directly on tomato surfaces with limited sample preparation procedures. Therefore, the ME biosensor could be applied as a cost and time effective, relatively simple, practically suitable on-site detection method for S. typhimurium in fresh produce.

The effect of incubation temperature on the binding of Salmonella typhimurium to phage-based magnetoelastic biosensors

The objective of this study is to evaluate the effect of incubation temperature on the binding of Salmonella typhimurium with E2 phage magnetoelastic (ME) biosensors. S. typhimurium was inoculated on tomato surfaces and the attachment of S. typhimurium was observed using scanning electron microscopy (SEM). Measurement biosensors, immobilized with E2 phage, and control sensors devoid of E2 phage were placed on the inoculated tomato surface and incubated at different temperatures for a fixed time in order to determine the effect of incubation temperature on the binding. After incubation, the resonant frequencies of both measurement and control sensors were measured using a network analyzer. The binding of S. typhimurium with E2 phage on both types of sensors was confirmed by SEM analysis. The SEM images of the inoculated tomato surfaces confirmed that S. typhimurium attached across the surface, however, its distribution was not uniform. After placement and incubation of the ME biosensors on the spiked tomato surfaces, the resonant frequencies of the ME biosensors were measured. The resonant frequency shifts in the measurement sensors increased as temperature was increased up to 35 °C, while the resonant frequency shifts of the control sensors were small and relatively constant across the entire range of incubation temperatures. There were no significant differences between incubation temperatures of 20, 25, 30, 35, 40, 45, and 50 °C in measurement sensors ( P > 0.05). SEM images of the measurement sensors confirmed that increasing resonant frequency shifts were caused by increasing numbers of bound S. typhimurium cells to the measurement sensors. This is contrasted with none or negligible binding of S. typhimurium cells on the control sensors. Overall, the optimum incubation temperature was chosen as 30 °C. This result will contribute to the development of a practical ME biosensor protocol for in situ detection of S. typhimurium on fresh produce .

Prediction of the Mass Sensitivity of Phage-Coated Magnetoelastic Biosensors for the Detection of Single Pathogenic Bacteria

ABSTRACTFreestanding, strip-shaped magnetoelastic (ME) biosensors are a class of wireless, mass-based biosensors that are being developed for the real-time detection of pathogenic bacteria for food safety and bio-security. The mass sensitivity of these biosensors operating in longitudinal-vibration modes is known to be largely dependent on the position of masses attached to the sensor surfaces. Hence, considering this dependence is crucial to the detection of low-concentration target pathogens, including single pathogenic bacteria, because their local attachment may cause varying sensor responses. In a worst case scenario, the resultant sensor responses (i.e., mass-induced resonance frequency changes of the sensor) may be too small to be detected despite the attachment of the target pathogenic masses. To address the issue, phage-coated ME biosensors (magnetostrictive strips (4 mm × 0.8 mm × 30 μm) coated with a phage probe specifically binding streptavidin protein) with localized masses (streptavidin-coated polystyrene beads) were fabricated, and mass-position-dependence of the sensor’s sensitivity under the fundamental-mode vibration was experimentally measured. In addition, three-dimensional finite element (FE) modal analysis was performed using the CalculiX software to simulate the phenomena. The experimental and theoretical results show close agreement: (1) the mass sensitivity was low when the mass was positioned in the middle of the sensor’s longest dimension and (2) a much higher mass sensitivity was, by contrast, obtained for the equivalent masses placed at both ends of the strip-shaped sensor. Furthermore, FE models were constructed for differently sized, phage-coated ME biosensors (100 – 500 μm in length with different widths and thicknesses) loaded with a single bacterial mass (2 μm × 0.4 μm × 0.4 μm, 1.05 g/cm3) at varying longitudinal positions. The mass sensitivity was found to be approximated by a mass-position-dependent Boltzmann function whose amplitude is inversely proportional to the length squared, width, and thickness of the sensor.

Biosensing technologies for Mycobacterium tuberculosis detection: status and new developments.

Biosensing technologies promise to improve Mycobacterium tuberculosis (M. tuberculosis) detection and management in clinical diagnosis, food analysis, bioprocess, and environmental monitoring. A variety of portable, rapid, and sensitive biosensors with immediate “on-the-spot” interpretation have been developed for M. tuberculosis detection based on different biological elements recognition systems and basic signal transducer principles. Here, we present a synopsis of current developments of biosensing technologies for M. tuberculosis detection, which are classified on the basis of basic signal transducer principles, including piezoelectric quartz crystal biosensors, electrochemical biosensors, and magnetoelastic biosensors. Special attention is paid to the methods for improving the framework and analytical parameters of the biosensors, including sensitivity and analysis time as well as automation of analysis procedures. Challenges and perspectives of biosensing technologies development for M. tuberculosis detection are also discussed in the final part of this paper.

Effects of surface functionalization on the surface phage coverage and the subsequent performance of phage-immobilized magnetoelastic biosensors

One of the important applications for which phage-immobilized magnetoelastic (ME) biosensors are being developed is the wireless, on-site detection of pathogenic bacteria for food safety and bio-security. Until now, such biosensors have been constructed by immobilizing a landscape phage probe on gold-coated ME resonators via physical adsorption. Although the physical adsorption method is simple, the immobilization stability and surface coverage of phage probes on differently functionalized sensor surfaces need to be evaluated as a potential way to enhance the detection capabilities of the biosensors. As a model study, a filamentous fd-tet phage that specifically binds streptavidin was adsorbed on either bare or surface-functionalized gold-coated ME resonators. The surface functionalization was performed through the formation of three self-assembled monolayers with a different terminator, based on the sulfur-gold chemistry: AC (activated carboxy-terminated), ALD (aldehyde-terminated), and MT (methyl-terminated). The results, obtained by atomic force microscopy, showed that surface functionalization has a large effect on the surface phage coverage (46.8%, 49.4%, 4.2%, and 5.2% for bare, AC-, ALD-, and MT-functionalized resonators, respectively). In addition, a direct correlation of the observed surface phage coverage with the quantity of subsequently captured streptavidin-coated microbeads was found by scanning electron microscopy and by resonance frequency measurements of the biosensors. The differences in surface phage coverage on the differently functionalized surfaces may then be used to pattern the phage probe layer onto desired parts of the sensor surface to enhance the detection capabilities of ME biosensors.

Detection of Salmonella Typhimurium on Fresh Food Produce Using Phage-Based Magnetoelastic Biosensors

Salmonella Typhimurium on fresh tomato surface using phage-based magnetoelastic (ME) biosensors. The ME biosensors are composed of a ME resonator platform that is coated with filamentous E2 phage (genetically engineered to bind with Salmonella). Actuated and detected through magnetic field, the ME biosensors are wireless sensors. In this study, tomato surfaces were spiked with a known number of Salmonella cells and then allowed to dry in air. The detection was conducted by directly placing ME biosensors on the spiked surface. Measurements of the resonance frequencies of both control and measurement ME biosensors were made prior to the placement and after 30 minutes of the placement. Shifts in the resonance frequency of the measurement biosensors were observed while control sensors showed negligible change. The specific binding of Salmonella to the biosensors was verified by Scanning Electron Microscopy (SEM). The results indicate that this methodology may provide real-time pathogen detection without traditional sampling process.

Effects of Surface Phage Coverage on the Performance of Wireless Phage-Immobilized Magnetoelastic Biosensors

One of the important applications for which wireless phage-immobilized magnetoelastic (ME) biosensors are being developed is the real-time detection of pathogenic bacteria for bio-security and food safety. Until now, such ME biosensors have been manufactured by immobilizing a phage probe on gold-coated ME sensor platforms via physical adsorption. Although the physical adsorption method is simple, the surface coverage as well as immobilization stability of phage probes must be evaluated as a potential way to enhance the performance of the ME biosensors. In addition, recent studies have shown that chemical immobilization of phage probes potentially improves the performance of phage-based bio/chemical sensors. Hence, we here present performance comparisons between phage-immobilized ME biosensors manufactured by different phage immobilization methods: physical adsorption and covalent attachment. The biosensor's performance was found to be directly related to the surface phage coverage.

A Multiple Magnetoelastic Sensor System for Detection of Salmonella Typhimurium Using Pulse Method

Magnetoelastic sensors show a resonance frequency decrease once target pathogens are attached to the sensors and thus add a small mass to the sensors. These sensors are fast, sensitive, and have higher mass sensitivity with smaller sized sensors. However, signals from smaller sensors are weaker and have more noise due to manufacturing defects. In this paper, we present a pulse biosensor system that is able to avoid bias magnetic field and allows simultaneous detection using multiple sensors. With multi-sensors, the effect of a manufacturing defect is decreased and we get the benefit of averaging for more accurate and reliable results. Stability tests show that the variance of frequency detection is less than 122 ppm of its resonance frequency. Real detection of pathogen is demonstrated using Salmonella typhimurium and the results show that multiple magnetoelastic biosensors are able to detect S. typhimurium simultaneously and offer good sensitivity and reliability.

Direct detection of Salmonella typhimurium on fresh produce using phage-based magnetoelastic biosensors

Current bacterial detection methods require the collection of samples followed by preparation and analysis in the laboratory, both time and labour consuming steps. More importantly, because of cost, only a limited number of samples can be taken and analyzed. This paper presents the results of an investigation to directly detect Salmonella typhimurium on fresh tomato surfaces using phage-based magnetoelastic (ME) biosensors. The biosensor is composed of a ME resonator platform coated with filamentous E2 phage, engineered to bind with S. typhimurium. The ME biosensors are wireless sensors, whose resonance oscillation and resonance frequency are actuated and detected through magnetic fields. The sensors used in this study were 0.028 mm × 0.2 mm × 1 mm in size. In this study, the tomato surface was spiked with S. typhimurium suspensions with concentrations ranging from 5 × 10 1 to 5 × 10 8 CFU/ml and then allowed to dry in air. The detection was conducted by directly placing ME measurement biosensors and control sensors on the spiked surface for 30 min in a humid environment. The control sensors were identical to the measurement biosensors, but without phage. Both measurement and control sensors were blocked with BSA to reduce non-specific binding. The resonance frequencies of both measurement and control sensors were measured prior to and after the placement of the sensors on the tomato. Shifts in the resonance frequency of the measurement biosensors were observed, while the control sensors showed negligible change. Scanning electron microscopy (SEM) was used to verify the specific binding of S. typhimurium to the biosensor. Results of multiple biosensor detection and corresponding analyzes showed statistically different responses between the measurement and control sensors for tomatoes spiked with S. typhimurium suspensions with concentrations of 5 × 10 2 CFU/ml and greater. This study demonstrates the direct detection of food-borne bacteria on fresh produce.