Simultaneous Detection of Salmonella Typhimurium and Bacillus Anthracis Spores Using Remote Phage-Based Magnetoelastic Microbiosensors
Type of Degreedissertation
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This dissertation discusses the results of a study to develop a free standing, magnetoelastic (ME) biosensor for the detection of S. typhimurium and B. anthracis spores. This affinity-based biosensor is composed of ME resonator platforms immobilized with filamentous phage as the bio-recognition element. Two different phages, E2 phage, genetically engineered to bind with S. typhimurium, and JRB7 phage, genetically engineered to bind with B. anthracis spores, were used to form the biosensors. A time-varying magnetic field is used to oscillate the magnetoelastic resonator at its characteristic resonance frequency. Upon contact with a target pathogen, the specific phage binds the pathogen to the biosensor, increasing the mass of the sensor. This mass change causes a corresponding decrease in the resonance frequency. The effect of annealing and sputtering a gold layer onto the surface was studied, and annealing between 200-250 °C found to greatly improve both the corrosion resistance and the Q factor of the signal spectrum. The effect of different salt and phage concentrations on the binding affinity of ME biosensors was also investigated and 420 mM NaCl at a phage concentration of 5 × 1011 vir/ml found to produce the optimal distribution of coated phages onto the sensor surface, consequently promoting better binding of spores to the biosensor’s surface. When sensors immobilized with JRB7 iii phage under this optimum condition were exposed to B. anthracis spores in different concentrations (5 × 101 - 5 × 108 cfu/ml) in a flowing system, the binding sensitivity of the ME biosensor was 226 Hz/decade. The multi-sensors detection system was exposed to different conditions of analyte including: S. typhimurium / B. anthracis spores (5 × 101-5 × 108 cfu/ml) in water, S. typhimurium (5 × 101-5 × 108 cfu/ml) in a mixture of E. coli O157:H7 (5 × 107 cfu/ml) and B. anthracis spores (5 × 101-5 ×108 cfu/ml) in a mixture of B. cereus (5 × 107 cfu/ml), separately. The sensitivity of both measurement biosensors (length ≈ 2 mm) was about 170-280 Hz/decade, with an average binding valence of about 3 and a dissociation constant Kd of about 250 cfu/ml under different testing analytes.