title: Combining reflectometry and fluorescence microscopy: an assay for the investigation of leakage processes across lipid membranes.
abstract: The passage of solutes across a lipid membrane plays a central role in many cellular processes. However, the investigation of transport processes remains a serious challenge in pharmaceutical research, particularly the transport of uncharged cargo. While translocation reactions of ions across cell membranes is commonly measured with the patch-clamp, an equally powerful screening method for the transport of uncharged compounds is still lacking. A combined setup for reflectometric interference spectroscopy (RIfS) and fluorescence microscopy measurements is presented that allows one to investigate the passive exchange of uncharged compounds across a free-standing membrane. Pore-spanning lipid membranes were prepared by spreading giant 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) vesicles on porous anodic aluminum oxide (AAO) membranes, creating sealed attoliter-sized compartments. The time-resolved leakage of different dye molecules (pyranine and crystal violet) as well as avidin through melittin induced membrane pores and defects was investigated.
authors.affiliation: Institute of Physical Chemistry, University of Göttingen , Tammannstr. 6, Göttingen, Lower Saxony 37077, Germany.
authors.forename: Milena
authors.initials: M
authors.lastname: Stephan
authors.forename: Ingo
authors.initials: I
authors.lastname: Mey
authors.forename: Claudia
authors.initials: C
authors.lastname: Steinem
authors.forename: Andreas
authors.initials: A
authors.lastname: Janshoff
mesh_terms: D000537:Aluminum Oxide / Q000737:chemistry; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism; D008555:Melitten / Q000378:metabolism; D008856:Microscopy, Fluorescence; D010713:Phosphatidylcholines / Q000737:chemistry / Q000378:metabolism; D021381:Protein Transport; D013057:Spectrum Analysis; D053835:Unilamellar Liposomes / Q000737:chemistry / Q000378:metabolism
chemical_list: D008051:Lipid Bilayers; D010713:Phosphatidylcholines; D053835:Unilamellar Liposomes; D008555:Melitten; C017251:1,2-oleoylphosphatidylcholine; D000537:Aluminum Oxide