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Development of new methods in fluorescence microscopy

dc.contributor.advisorWalla, Peter Jomo Prof. Dr.
dc.contributor.authorLin, Chao-Chen
dc.titleDevelopment of new methods in fluorescence microscopyde
dc.contributor.refereeJahn, Reinhard Prof. Dr.
dc.description.abstractengIn recent years, advancements in single-molecule imaging techniques have enabled scientists to study in great detail the cells and relevant physiological processes, including neurons and the communication between them. Nevertheless, with proper experimental design, much can still be learned from conventional fluorescence spectroscopy. In this work I use primarily the fluorescence lifetime as an indicator of FRET (Förster resonance energy transfer), which gives information on the interaction between liposomes. The dissertation consists of two connected projects, and in each a new chemical tool is developed. In the first part, control of membrane gaps by synaptotagmin-Ca2+ measured with a novel membrane distance ruler, a set of liposomes bridged by double-stranded DNAs of various lengths serves as the molecular ruler to measure the changes in membrane distances induced by binding of Ca2+ to synaptotagmin-1 (syt-1). I showed that the distance maintained by syt-1 alone was reduced by one-third from ~7-8 nm to ~5 nm, which may explain how syt-1 functions as the fast and efficient Ca2+ trigger in promoting the zippering of SNARE proteins, which leads subsequently to membrane fusion and neurotransmitter release. The major part of this project has been published in Nat. Commun. 2014, 5, 5859 (doi: 10.1038/ncomms6859). In the second part, asymmetrically labeled liposomes as a new tool to study membrane fusion, one type of liposomes is labeled differently on the inner and outer leaflets, and the two fluorescent labels are distinguished by their differently lifetimes. Theses liposomes were used to monitor SNARE-mediated membrane fusion in microfluidic channels, and it turned out that there was no apparent delay between the merging of the two leaflets. As a final remark, further characterization and improvements of the two new tools should allow for their future applications in studying other cellular mechanisms of
dc.contributor.coRefereeSteinem, Claudia Prof. Dr.
dc.contributor.thirdRefereeHöbartner, Claudia Prof. Dr.
dc.contributor.thirdRefereeBodenschatz, Eberhard Prof. Dr.
dc.contributor.thirdRefereeEnderlein, Jörg Prof. Dr.
dc.subject.engfluorescence lifetimede
dc.subject.engSNARE proteinsde
dc.affiliation.instituteGöttinger Graduiertenschule für Neurowissenschaften, Biophysik und molekulare Biowissenschaften (GGNB)de
dc.subject.gokfullBiologie (PPN619462639)de

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