Michael L. Johnson

Sequential detection of Salmonella typhimurium and Bacillus anthracis spores using magnetoelastic biosensors.

Multiple phage-based magnetoelastic (ME) biosensors were simultaneously monitored for the detection of different biological pathogens that were sequentially introduced to the measurement system. The biosensors were formed by immobilizing phage and 1mg/ml BSA (blocking agent) onto the magnetoelastic resonators surface. The detection system included a reference sensor as a control, an E2 phage-coated sensor specific to S. typhimurium, and a JRB7 phage-coated sensor specific to B. anthracis spores. The sensors were free standing during the test, being held in place by a magnetic field. Upon sequential exposure to single pathogenic solutions, only the biosensor coated with the corresponding specific phage responded. As the cells/spores were captured by the specific phage-coated sensor, the mass of the sensor increased, resulting in a decrease in the sensors resonance frequency. Additionally, non-specific binding was effectively eliminated by BSA blocking and was verified by the reference sensor, which showed no frequency shift. Scanning electron microscopy was used to visually verify the interaction of each biosensor with its target analyte. The results demonstrate that multiple magnetoelastic sensors may be simultaneously monitored to detect specifically targeted pathogenic species with good selectivity. This research is the first stage of an ongoing effort to simultaneously detect the presence of multiple pathogens in a complex analyte.

Micro-fabricated wireless biosensors for the detection of S. typhimurium in liquids

Food borne illnesses from the ingestion of S. typhimurium have been of primary concern due to their common occurrence in food products of daily consumption. In this paper, micron size, magnetoelastic (ME) biosensors for the detection of S. typhimurium were fabricated and tested in liquid solutions containing known concentrations of S. typhimurium cells. The biosensors are comprised of a ME sensor platform and immobilized bio-molecular recognition element (E2 phage) that has been engineered to bind the S. typhimurium multi-valently. The micron size ME sensor platforms were manufactured using microelectronics fabrication techniques. Phage was engineered at Auburn University and immobilized onto all surfaces of the sensor. The ME biosensor oscillates with a characteristic resonance frequency when subjected to a time varying magnetic field. Binding between the phage and bacteria, adds mass to the sensor that causes a shift in the sensors resonance frequency. Sensors with the dimension of 500x100x4 μm were exposed to S. typhimurium with increasing known concentrations ranging from 5 x101 to 5 x 107 cfu/ml. The ME biosensor exhibited high sensitivity and a detection limit better than 50 cfu/ml.

Multiple Phage-Based Magnetoelastic Biosensors System for the Detection of Salmonella typhimurium and Bacillus anthracis Spores

This paper presents a multiple magnetoelastic (ME) biosensor system for in-situ detection of S. typhimurium and B. anthracis spores in a flowing bacterial/spore suspension (5 x 10 1 - 5 x 10 8 cfu/ml). The ME biosensor was formed by immobilizing filamentous phage (specific to each detection target) on the ME platforms. An alternating magnetic field was used to resonate the ME biosensor to determine its resonance frequency. When cells/spores are bound to a ME biosensor surface, the additional mass of the cells/spores causes a decrease in the resonance frequency of the biosensor. The detection system was composed of a control sensor, an E2 phage-based biosensor (specific to S. typhimurium ) and a JRB7 phage-based biosensor (specific to B. anthracis spores). The frequency response curves of the ME biosensors as a function of exposure time were then measured and the detection limit of the ME biosensor was observed to be 5 x 10 3 cfu/ml. The results show that phage-based ME biosensors can detect multiple pathogens simultaneously and offer good performance, including good sensitivity and rapid detection.

Magnetoelastic Material as a Biosensor for the Detection of Salmonella Typhimurium

ABSTRACT Magnetoelastic materials are amorphous, ferromagnetic alloys that usually include a combination of iron, nickel, molybdenum and boron. Magnetoelastic biosensors are mass sensitive devices comprised of a magnetoelastic material that serves as the transducer and bacteriophage as the bio-recognition element. By applying a time varying magnetic field, the magnetoelastic sensor thin films can be made to oscillate, with the fundamental resonant frequency of oscillations depends on the physical dimensions and properties of the material. The change in the resonance frequency of these mass based sensors can be used to evaluate the amount of analyte attached on the sensor surface. Filamentous bacteriophage specific to S. typhimurium was used as a bio-recognition element in order to ensure specific and selective binding of bacteria onto the sensor surface. The sensitivity of magnetoelastic materials is known to be dependent on the physical dimensions of the material. An increase in sensitivity from 159Hz/decade for a 2mm sensor to 770Hz/decade for a 1mm sensor and 1100Hz/decade for a 500micron sensor was observed. The sensors were characterized by scanning electron microscopy (SEM) analysis assayed biosensors to provide visual verification of frequency responses and an insight into the characteristics of the distribution of phage on the sensor surface. The magnetoelastic sensors immobilized with filamentous phage are suitable for specific and selective detection of target analyte in different media. Certain modifications to the measurement circuit resulted in better signal to noise ratios for sensors with smaller dimensions (L

Characterization of phage-coupled magnetoelastic micro-particles for the detection of Bacillus anthracis Sterne spores

A microfabricated magnetoelastic sensor immobilized with specific phage for the detection of Bacillus anthracis spores is presented. This micro-sensor can detect response signal remotely through magnetic field, enabling the in-situ measurement of target pathogens in aqueous environment. The sensors surface was modified by selected phage which provides the binding ability to B. anthracis spores. When exposed to B. anthracis spore solution, the resonant frequency of the sensor decreased due to the accumulated spore attachment on the surface. Sensors with dimensions of 200 times 40 times 4 mum were employed in the tests of increasing concentrations (102 to 108 cfu/ml) of B. anthracis spores. A detection limit of 102 cfu/ml, and a sensitivity of 13.1 kHz/decade, were observed. Scanning electron microscopy (SEM) photographs demonstrated binding of target analytes to biosensors compared to controls.

Detection of S. Typhimirium and Bacillus Anthracis Spores in a Flow System Using ME Biosensors by Optimizing Phage Chemistry

This paper presents the results of a study that investigates the optimization of phage chemistry during the fabrication of magnetoelastic (ME) biosensors for the detection of Salmonella typhimurium or Bacillus anthracis spores. The bundling characteristics of the phage filaments limit the ability of the biosensor to bind bacterial cells/spores. Experiments were performed to determine the proper phage concentration for the prevention of bundling in aqueous environments. Based on the transmission electron microscopy (TEM) and scanning electron microscopy (SEM) results, which verify the structure of phage under different concentrations and binding numbers of target species to the sensor surface, we found that phage concentrations of 1011 vir/ml exhibit the best sensor performance in terms of binding sensitivity. Additionally, the sensors immobilized with phage under this condition were tested in a flowing liquid system using S. typhimurium and B. anthracis spores suspensions in concentrations ranging from 5 times101 to 5 times 108 cfu/ml, separately. As cells/spores are bound to a ME biosensor surface, the additional mass of the spores causes a decrease in the resonance frequency of the sensor. The frequency response curves of the ME biosensors as a function of exposure time were then measured, and the detection limit of the ME biosensor was determined to be 5 times 103 cfu/ml.