New phosphorescence based probes and techniques for the analysis of cellular oxygen and respiration

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dc.contributor.advisor Papkovsky, Dmitri B. en Kondrashina, Alina 2015-11-20T13:17:36Z 2015-11-20T13:17:36Z 2014 2014
dc.identifier.citation Kondrashina, A. 2014. New phosphorescence based probes and techniques for the analysis of cellular oxygen and respiration. PhD Thesis, University College Cork. en
dc.identifier.endpage 137
dc.description.abstract Real time monitoring of oxygenation and respiration is on the cutting edge of bioanalysis, including studies of cell metabolism, bioenergetics, mitochondrial function and drug toxicity. This thesis presents the development and evaluation of new luminescent probes and techniques for intracellular O2 sensing and imaging. A new oxygen consumption rate (OCR) platform based on the commercial microfluidic perfusion channel μ-slides compatible with extra- and intracellular O2 sensitive probes, different cell lines and measurement conditions was developed. The design of semi-closed channels allowed cell treatments, multiplexing with other assays and two-fold higher sensitivity to compare with microtiter plate. We compared three common OCR platforms: hermetically sealed quartz cuvettes for absolute OCRs, partially sealed with mineral oil 96-WPs for relative OCRs, and open 96-WPs for local cell oxygenation. Both 96-WP platforms were calibrated against absolute OCR platform with MEF cell line, phosphorescent O2 probe MitoXpress-Intra and time-resolved fluorescence reader. Found correlations allow tracing of cell respiration over time in a high throughput format with the possibility of cell stimulation and of changing measurement conditions. A new multimodal intracellular O2 probe, based on the phosphorescent reporter dye PtTFPP, fluorescent FRET donor and two-photon antennae PFO and cationic nanoparticles RL-100 was described. This probe, called MM2, possesses high brightness, photo- and chemical stability, low toxicity, efficient cell staining and high-resolution intracellular O2 imaging with 2D and 3D cell cultures in intensity, ratiometric and lifetime-based modalities with luminescence readers and FLIM microscopes. Extended range of O2 sensitive probes was designed and studied in order to optimize their spectral characteristics and intracellular targeting, using different NPs materials, delivery vectors, ratiometric pairs and IR dyes. The presented improvements provide useful tool for high sensitive monitoring and imaging of intracellular O2 in different measurement formats with wide range of physiological applications. en
dc.description.sponsorship European Commission (Marie Curie Program) en
dc.format.mimetype application/pdf en
dc.language.iso en en
dc.publisher University College Cork en
dc.rights © 2014, Alina Kondrashina. en
dc.rights.uri en
dc.subject Cellular oxygen en
dc.subject Phosphorescent nanoparticles en
dc.subject Intracellular probes en
dc.subject Cell respiration en
dc.title New phosphorescence based probes and techniques for the analysis of cellular oxygen and respiration en
dc.type Doctoral thesis en
dc.type.qualificationlevel Doctoral en
dc.type.qualificationname PhD (Science) en
dc.internal.availability Full text available en No embargo required en
dc.description.version Accepted Version
dc.contributor.funder European Commission en
dc.description.status Not peer reviewed en Biochemistry en
dc.check.type No Embargo Required
dc.check.reason No embargo required en
dc.check.opt-out No en
dc.thesis.opt-out false
dc.check.embargoformat Not applicable en
dc.internal.conferring Summer Conferring 2015

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© 2014, Alina Kondrashina. Except where otherwise noted, this item's license is described as © 2014, Alina Kondrashina.
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