Osmotic Stressing, Membrane Leakage, and Fluorescence: An Introductory Biochemistry Demonstration.

A fluorescence demonstration is described that incorporates several fundamental aspects of an introductory biochemistry course. A variation of a known leakage assay is utilized to prepare vesicles containing a quenched fluorophore. The vesicles are exposed to several osmotic environments ranging from isotonic to hypotonic. The degree of vesicle swelling, partial rupture or bursting, and subsequent release and dequenching of the encapsulated fluorophore can be observed. This demonstration introduces students to fluorescence, fluorescence quenching, lipid membrane dynamics, membrane permeability, and osmotic pressure. Students are also introduced to the lipid extrusion process and are able to observe the separation of compounds via size exclusion chromatography. This demonstration has the flexibility to be expanded to laboratory experiments that allow students to investigate a variety of membrane properties.