Biochemistry and Cell Biology - Conference Items
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Item A new macro-imager based on Tpx3Cam optical camera for PLIM applications(Society of Photo-Optical Instrumentation Engineers (SPIE), 2020-04-01) Sen, Rajannya; Zhdanov, Alexander V.; Hirvonen, Liisa; Svihra, Peter; Andersson-Engels, Stefan; Nomerotski, Andrei; Papkovsky, Dmitri B.; Science Foundation IrelandThe recently designed Tpx3Cam camera based PLIM (Phosphorescence Lifetime IMaging) macro-imager was tested using an array of phosphorescent chemical and biological samples. A series of sensor materials prepared by incorporating the phosphorescent O2-sensitive dye, PtBP, into five polymers with different O2 permeability were imaged along with several commercial and non-commercial sensors based on PtBP and PtOEPK dyes. The PLIM images showed good lifetime contrast between the different materials, and phosphorescence lifetime values obtained were consistent with those measured by alternative methods. A panel of live tissues samples stained with PtBP based nanoparticle probe were also prepared and imaged under resting conditions and upon inhibition of respiration. The macro-imager showed promising results as a tool for PLIM of O2 in chemical and biological samples.Item In vivo imaging of flavoprotein fluorescence during hypoxia reveals the importance of direct arterial oxygen supply to cerebral cortex tissue(Springer Nature Switzerland AG, 2016-01) Chisholm, K. I.; Ida, K. K.; Davies, A. L.; Papkovsky, Dmitri B.; Singer, M.; Dyson, A.; Tachtsidis, I.; Duchen, M. R.; Smith, K. J.; Wellcome Trust; University College London; Multiple Sclerosis SocietyLive imaging of mitochondrial function is crucial to understand the important role played by these organelles in a wide range of diseases. The mitochondrial redox potential is a particularly informative measure of mitochondrial function, and can be monitored using the endogenous green fluorescence of oxidized mitochondrial flavoproteins. Here, we have observed flavoprotein fluorescence in the exposed murine cerebral cortex in vivo using confocal imaging; the mitochondrial origin of the signal was confirmed using agents known to manipulate mitochondrial redox potential. The effects of cerebral oxygenation on flavoprotein fluorescence were determined by manipulating the inspired oxygen concentration. We report that flavoprotein fluorescence is sensitive to reductions in cortical oxygenation, such that reductions in inspired oxygen resulted in loss of flavoprotein fluorescence with the exception of a preserved ‘halo’ of signal in periarterial regions. The findings are consistent with reports that arteries play an important role in supplying oxygen directly to tissue in the cerebral cortex, maintaining mitochondrial function.Item Background-free fluorescence decay time sensing and imaging of pH with highly photostable diazaoxotriangulenium dyes(American Chemical Society, 2018-12-06) Dalfen, Irene; Dmitriev, Ruslan I.; Holst, Gerhard; Klimant, Ingo; Borisov, Sergey M.; Science Foundation IrelandNovel fluorescent diazaoxatriangulenium (DAOTA) pH indicators for lifetime-based self-referenced pH sensing are reported. The DAOTA dyes were decorated with phenolic receptor groups inducing fluorescence quenching via photoinduced electron transfer mechanism. Electron-withdrawing chlorine substituents ensure response in the most relevant pH range (apparent pK'a values ~5 and 7.5 for the p,p-dichlorophenol- and the p-chlorophenol-substituted dyes, respectively). The dyes feature long fluorescence lifetime (17-20 ns), high quantum yield (~60%) and high photostability. Planar optodes are prepared upon immobilization of the dyes into polyurethane hydrogel D4. Apart from the response in the fluorescence intensity, the optodes show pH-dependent lifetime behaviour which makes them suitable for studying 2D pH distribution with help of fluorescence lifetime imaging technique. The lifetime response is particularly pronounced for the sensors with high dye concentration (0.5-1% wt. in respect to the polymer) and is attributed to efficient homo-FRET mechanism.