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  • Title: Combining reflectometry and fluorescence microscopy: an assay for the investigation of leakage processes across lipid membranes.
    Author: Stephan M, Mey I, Steinem C, Janshoff A.
    Journal: Anal Chem; 2014 Feb 04; 86(3):1366-71. PubMed ID: 24377291.
    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.
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