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Dissertation Lucille Ray Final.pdf (3 MB)

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Live single cell fluorescence microscopy; from antibiotic resistance detection to mitochondrial dysfunction

Ray, Lucille Alexandria
http://rave.ohiolink.edu/etdc/view?acc_num=akron1597342775751888

Abstract Details

2020, Doctor of Philosophy, University of Akron, Chemistry.
Single-cell fluorescence microscopy is a powerful tool which can be used to investigate the nature of cellular responses to external stimuli. The application of single-cell fluorescence microscopy to the growing problem of antimicrobial susceptibility detection stands to improve and refine our ability to address bacterial resistance in human infections. In cases of severe bacterial infection, determining the correct antibiotic to use to combat an infection is a race against the patient’s dwindling lifespan. By combining the RedoxSensor™ Green fluorescent dye with live single-cell imaging, we have developed a method for antimicrobial susceptibility testing which can identify susceptibility to a given antibiotic within 100 minutes of treatment. We show that this method is reproducible and can identify susceptibility to several bacterial cell wall targets and bacterial cell membrane targeted antibiotics. Our methodology has considerable applicability within the sphere of rational antibiotic drug design as well in its ability to identify antibiotic efficacy as a function of time instead of antibiotic concentration. We use our method to compare the efficacy of 4 recently synthesized polyurethane antimicrobials. Our work lays the framework for expansion upon our method into microfluidics systems and use in screening candidate antimicrobial drugs. Mitochondrial morphological analysis within living eukaryotic cells represents another challenge which requires careful application of single-cell fluorescence microscopy. The cuprizone mouse treatment model for multiple sclerosis is known to generate enlarged mitochondria within oligodendrocytes, but much remains unknown about the dynamics of their formation. Cultured MO3.13 oligodendrocyte cells treated with cuprizone are shown to undergo mitochondrial enlargement within 8 hours of direct cuprizone exposure via single-cell mitochondrial size determination with MitoTracker Red fluorescent dye. Cuprizone treatment is also shown to increase oxygen consumption rapidly following oligodendrocyte exposure using porphyrin-based fluorescence lifetime oxygen consumption measurement.
Michael Konopka (Advisor)
Shriver Leah (Committee Member)
Smith Adam (Committee Member)
Paruchuri Sailaja (Committee Member)
Joy Abraham (Committee Member)
113 p.
Biochemistry; Chemistry; Microbiology
microscopy; single-cell; antibiotic resistance; fluorescence; mitochondria; cuprizone; antibacterial susceptibility testing; antimicrobial polymer

Recommended Citations

Citations

  • Ray, L. A. (2020). Live single cell fluorescence microscopy; from antibiotic resistance detection to mitochondrial dysfunction [Doctoral dissertation, University of Akron]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=akron1597342775751888

    APA Style (7th edition)

  • Ray, Lucille. Live single cell fluorescence microscopy; from antibiotic resistance detection to mitochondrial dysfunction. 2020. University of Akron, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=akron1597342775751888.

    MLA Style (8th edition)

  • Ray, Lucille. "Live single cell fluorescence microscopy; from antibiotic resistance detection to mitochondrial dysfunction." Doctoral dissertation, University of Akron, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=akron1597342775751888

    Chicago Manual of Style (17th edition)

akron1597342775751888
374
© 2020, all rights reserved.
This open access ETD is published by University of Akron and OhioLINK.