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Fluorescence Imaging with Single-Molecule Sensitivity and Fluorescence Correlation Spectroscopy of Cell-Penetrating Neuropeptides
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
2011 (English)In: Neuropeptides: methods and protocols / [ed] Merighi, A., Humana Press, 2011, 147-170 p.Chapter in book (Refereed)
Abstract [en]

Neuropeptide plasma membrane interactions in the absence of a corresponding specific receptor may result in neuropeptide translocation into the cell. Trans location across the plasma membrane may represent a previously unknown mechanism by which neuropeptides can signal information to the cell interior. We introduce here two complementary optical methods with single-molecule sensitivity, fluorescence imaging with avalanche photodiode detectors (APD imaging) and fluorescence correlation spectroscopy (FCS), and demonstrate how they may be applied for the analysis of neuropeptide ability to penetrate into live cells in real time. APD imaging enables us to visualize fluorescently labeled neuropeptide molecules at very low, physiologically relevant concentrations, whereas FCS enables us to characterize quantitatively their concentration and diffusion properties in different cellular compartments. Application of these methodologies for the analysis of the endogenous opioid peptide dynorphin A (Dyn A), a ligand for the kappa-opioid receptor (KOP), demonstrated that this neuropeptide may translocate across the plasma membrane of living cells and enter the cellular interior without binding to its cognate receptor.

Place, publisher, year, edition, pages
Humana Press, 2011. 147-170 p.
, Methods in Molecular Biology, ISSN 1064-3745 ; 789
Keyword [en]
Dynorphin A, Cell-penetrating peptides, Fluorescence correlation spectroscopy, APD imaging, single-molecule detection
National Category
Biochemistry and Molecular Biology
URN: urn:nbn:se:su:diva-94465DOI: 10.1007/978-1-61779-310-3_9ISI: 000299454600009ISBN: 978-1-61779-309-7OAI: diva2:653597
Available from: 2013-10-04 Created: 2013-10-04 Last updated: 2013-10-25Bibliographically approved

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Gräslund, Astrid
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Department of Biochemistry and Biophysics
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