Optical and impedance based sensors for chemical/biological and foodborne pathogens applications
Abstract
[EMBARGOED UNTIL 5/1/2025] An impedance based microfluidic biosensor for rapid simultaneous detection of different Salmonella serogroups in food product was investigated. Foodborne pathogens contamination in the United States alone has caused millions of infections which are dangerous and deadly sometimes. Salmonella is one of the public foodborne bacterial pathogens and can result in severe illness such as diarrhea, abdominal cramps, and fever. The economic impact due to food recall and foodborne illnesses is $7 billion per year. The food industry is seeking new pathogen detection techniques that can perform the diagnosis rapidly while maintaining high accuracy by eliminating the enrichment step. To resolve these challenges, a highly sensitive microfluidic impedance-based biosensor was fabricated, in a horizontal microchannel, for the rapid and simultaneous detection of three Salmonella serogroups or other pathogen types while the second biosensor was fabricated in a couple of parallel fluidic microchannels for detection of two pathogen type. The first design has the following innovation. (1) a focusing region uses a ramp down vertical electrode pair along with tilted finger pairs (45o) with a ramp down channel that generates positive dielectrophoresis (p-DEP) forces to concentrate the Salmonella pathogens into the center of the microchannel and direct them toward the sensing regions that have a narrower microchannel. This design achieved highly concentrated samples. (2) Three bacteria sensing regions, each consists of interdigitated electrode array (IDE array) with 10-finger pairs. Microfabrication techniques such as photolithography, sputter-deposition, wet etching and surface plasma treatment were utilized to fabricate the sensor. The three sensing electrodes were pre-functionalized by delivery specific anti-Salmonella antibody serotypes B, D and E, each for one electrode through independent antibody inlets without causing cross-contamination. All the antibodies were pre-linked with crosslinker. The impedance was then measured. The bacterial samples were spiked with Salmonella type B and injected via the sample inlet into the focusing region. An AC signal, i.e., 6 V peak-to-peak at 6 MHz from an AC power supply was applied across the focusing electrode pair to generate a non-uniform E-field gradient and p-DEP forces to move the Salmonella cells/ microbeads toward the center of the channel where the E-field intensity/gradient is high. The Salmonella cells kept moving toward the sensing region where they bonded with the antibody selectively. The antibody antigen binding process has caused the impedance to change, which was measured by an impedance analyzer. The change of impedance represents the presence/absence of bacterial cells. The detection limit of the sensor was found to be 8 Cells/ml, which was confirmed using bacterial culture and colony counting. For all concentrations, impedance decreased as function of frequency. The device cannot quantify the number of detected bacterial cells, but it informs the presence or absence of bacterial cells. The impedance value could indicate a range for the bacterial concentration. The overall antigen detection time was [less than] 1 hour. The second sensor had a simplified design, i.e., the sensor starts with a single microchannel and splits into two microchannel. Each channel includes a region that consists of 50 pairs of IDE arrays for bacteria sensing. The sensor was fabricated using similar microfabrication techniques. Two types of Salmonella antibodies (type B and D) were mixed separately with the cross-linker (Sulfo-LC-SPDP) to functionalize the electrode surface, one for each sensing region and without causing cross contamination. The Salmonella samples were spiked with Salmonella type B and introduced into the biosensor via the sample inlet towards the detecting regions to bind with the antibody. The antibody antigen binding caused the change in the impedance, i.e., measured by an impedance analyzer. A low-cost chemical sensor based on a fiber optic platform was investigated. The sensor consists of a thin film metal patterned with holes on a fiber tip, used for extraordinary optical transmission (EOT). The sensing technique is based on the fact that change of the refractive index at the interface of the metal layer to the surrounding environment producing a wavelength shift in resonance wavelength. This project investigated the integration of EOT with single mode optical fiber. In addition, the microsphere lithography (MPL) process was used to pattern nano hole arrays in Al thin film and thus creating refractive index (RI) sensor on the cleaved tips of optical fiber was also studied. The resonant wavelength of metal elements on the surface is dependent on the local dielectric environment and allows the refractive index of an analyte to be resolved spectrally. The reflection spectra of the tested fibers were recorded using optical signal analyzer. The sensor resonant wavelength can be controlled by changing the hole diameter, periodicity, and the thickness. The sensor was validated in multiple mediums with different RI. The result demonstrated that the wavelength was shifted when the medium was changed. The sensor was tested using several glucose concentrations. The limit of detection (LoD) was 10 mg/ml with a sensitivity of 613 nm/RIU. To increase the sensor sensitivity, we have used self-assembled microspheres monolayer with multiple UV illumination jets to pattern multiple holes groups. This included three-hole group and four-hole group patterns. The sensors were then tested with glucose at various concentrations in water with various refractive indices, which result in shifting the resonant wavelength of the nano holes arrays, represented as sensitivity. The testing results show that three-hole group and four- hole group have the sensitivity of 906 nm/RIU and 675 nm/RIU, respectively. The sensitivity of these new metasurface coated fiber's has increased by 40 percent compared to our single hole array. In this research, MPL was also used as an alternative fabrication method to Focus Ion Beam (FIB) milling and other expensive nanofabrication techniques for performing photolithography for creating nanodisks on the endface of the fiber optic to create a Surface Enhanced Raman Spectroscopy (SERS) sensor. The SERS sensor consists of an array of periodic disks patterned on the endface of a multi-mode fiber optic of core diameter of 62.5 [m. The fabricated disks are made of gold because this material can efficiently reflect Raman signals with high intensities. MPL with off-exposure technique was also exploited to create 3-disks and 4-disks groups which would increase the total number of disks pattern in the core area of the fiber endface, decreasing the periodicity between fabricated disks. The sensor was tested and validated in different concentrations of Rhodamine 6G (R6G) dye solutions. A transformative low-cost surface-enhanced Raman scattering (SERS) sensor was designed and fabricated on the side of a multimode optical fiber core with large surface area. The optical fiber (diameter 100 [mu]m) was first integrated into 3-dimensional printed structure, especially designed for this purpose, where the fiber was manually polished with various surface areas. Microsphere Photolithography (MPL) technique was utilized to pattern periodic arrays of metallic nano disks with multiple diameters, on the side polished optical fiber. This technique provides a versatile, low-cost fabrication method to produce highly periodic hexagonal nanodisks, which in this study can significantly help increase the SERS signal. The sensor was validated by testing the relatively low Rhodamine 6G (R6G) solution concentration. A comparable SERS signal was found on the patterned side polished fiber. The detection limit of R6G was found to be near 10-9 mol/L.
Degree
Ph. D.