In this proposal, we report the design of two bioanalytical sensors: (1) a fluorescence optrode-based optical fiber sensor and (2) a carbon-based optically transparent electrode. The first provides a means for spatially organizing microorganisms to monitor their social behavior in response to different chemical environments. The electrode sensor enables the detection of signaling molecules that are responsible for, or a consequence of, sociomicrobial behavior. Upon their completion, these sensors will be coupled with a micro three-dimensional printing (µ3DP) technique. These sensors will provide key insights to understanding cellular communication and behavior in organized in vitro microenvironments that closely mimic in vivo settings (Scheme
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To carry out these electrochemical measurements in biological environments involving cells, we created an electrochemical well from a biocompatible material, PDMS. Teflon is composed of polytetrafluoroethylene chains with fluorine atoms containing a partial negative charge, which causes cell death. Consequently, we created a PDMS-based electrochemical well via a two-step soft lithography casting process.33 This process is illustrated in Scheme 3, where the Teflon-based electrochemical well serves as a template over which PDMS pre-polymer is cast. In the second step, the casting process is repeated where the PDMS negative serves as the new template to create the PDMS-based electrochemical …show more content…
In addition, light from fluorescent bacteria can be collected through the optical fiber. Bacteria within the 3D structures express green fluorescent proteins that are activated by QS signals in the environment to which the optical fiber probe is introduced. Consequently, we present our progress in constructing an optrode-based optical fiber biosensor (Scheme 5). This optrode sensor will spatially organize microorganisms to monitor their social behavior in response to different chemical environments. Scheme 5 illustrates the components of our custom-built bioanalytical sensor. A 532 nm solid-state laser is used as the excitation source to closely match the excitation wavelength of mCherry, a green fluorescent protein derivative. The laser beam is coupled into a 0.22 NA objective to match the refractive index with the cone of acceptance of the optical fiber. The input laser will excite bacterial green fluorescent proteins within the microstructures fabricated on the tip of the optical fiber. The collected fluorescence will be filtered through a dichroic filter (cut-off wavelength ~570 nm) before entering a photomultiplier tube (PMT). The PMT is an extremely sensitive, low noise, high gain detector, capable of detecting signal photons. The signal from the PMT will be converted to a number
To carry out these electrochemical measurements in biological environments involving cells, we created an electrochemical well from a biocompatible material, PDMS. Teflon is composed of polytetrafluoroethylene chains with fluorine atoms containing a partial negative charge, which causes cell death. Consequently, we created a PDMS-based electrochemical well via a two-step soft lithography casting process.33 This process is illustrated in Scheme 3, where the Teflon-based electrochemical well serves as a template over which PDMS pre-polymer is cast. In the second step, the casting process is repeated where the PDMS negative serves as the new template to create the PDMS-based electrochemical …show more content…
In addition, light from fluorescent bacteria can be collected through the optical fiber. Bacteria within the 3D structures express green fluorescent proteins that are activated by QS signals in the environment to which the optical fiber probe is introduced. Consequently, we present our progress in constructing an optrode-based optical fiber biosensor (Scheme 5). This optrode sensor will spatially organize microorganisms to monitor their social behavior in response to different chemical environments. Scheme 5 illustrates the components of our custom-built bioanalytical sensor. A 532 nm solid-state laser is used as the excitation source to closely match the excitation wavelength of mCherry, a green fluorescent protein derivative. The laser beam is coupled into a 0.22 NA objective to match the refractive index with the cone of acceptance of the optical fiber. The input laser will excite bacterial green fluorescent proteins within the microstructures fabricated on the tip of the optical fiber. The collected fluorescence will be filtered through a dichroic filter (cut-off wavelength ~570 nm) before entering a photomultiplier tube (PMT). The PMT is an extremely sensitive, low noise, high gain detector, capable of detecting signal photons. The signal from the PMT will be converted to a number