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%0 Thesis %A Kasahara, Keitaro %T Structuring spatiotemporal oxygen environments for microbial single-cell analysis in microfluidics %I Rheinisch-Westfälische Technische Hochschule Aachen %V Dissertation %C Aachen %M RWTH-2025-06324 %P 1 Online-Ressource : Illustrationen %D 2025 %Z Veröffentlicht auf dem Publikationsserver der RWTH Aachen University %Z Dissertation, Rheinisch-Westfälische Technische Hochschule Aachen, 2025 %X Microbial single-cell analysis using microfluidics is a promising method for studying microbial growth behavior in detail under precisely controlled environments. However, little effort has been made to incorporate spatiotemporal O2 control, a critical factor influencing microbial growth and physiology. This dissertation explores various strategies for establishing straightforward O2 control, temporal O2 control in the range of seconds to minutes, and spatial O2 control in the range of micrometers. First, a comprehensive experimental platform was developed that is transferable to microbial single-cell analysis within various formats of microfluidic devices. Using a low-cost 3D-printed mini-incubator surrounding the air-permeable PDMS microfluidic chip, the O2 concentration in the microfluidic chip was controlled. The O2 sensing method using FLIM and an O2-sensitive dye was also implemented, allowing direct measurement of the O2 availability inside the fluid channels. Subsequent imaging with timelapse microscopy and deep-learning-based image analysis provided a solid platform for data analysis. Furthermore, a double-layer microfluidic chip was developed to implement spatiotemporal O2 control in microbial single-cell analysis. The newly developed microfluidic platform could reproduce O2 oscillations occurring within seconds to minutes, thus enabling time-resolved microbial growth analysis at single-cell resolution. The case studies were performed by studying the aerobic and anaerobic growth and adaptation of E. coli and C. glutamicum. The growth analysis results revealed aerobic/anaerobic specific growth and growth adaptation in response to O2 oscillations, insights that cannot be obtained using conventional cultivation setups. Lastly, several different designs of the double-layer microfluidic chip were introduced to achieve spatial O2 control in microbial single-cell analysis in the range from millimeters down to micrometers. The experimental results demonstrated the capability of spatial O2 control by diffusion. The proposed concepts and devices are expected to be used for further microbial growth characterization under spatiotemporally structured O2 microenvironments. %F PUB:(DE-HGF)11 %9 Dissertation / PhD Thesis %R 10.18154/RWTH-2025-06324 %U https://publications.rwth-aachen.de/record/1015345