Membrane fluidity and other properties of lipids are commonly assessed using various fluorogenic membrane probes and fluorescence polarization (FP) measurements. The basic principle is that alterations in lipid packing (e.g. temperature-dependent lipid phase transitions) change the mobility of a membrane-bound fluorophore. Membrane fluidization increases mobility of the dye and accordingly decreases fluorescence polarization values.
DPH (1,6-diphenyl-hexa-1,3,5-triene) is a fluorescent molecule excitable by UV light around 355 nm that binds to the hydrophobic region of the membrane.
The FP assay was validated with unilamellar liposomes labeled with DPH. The liposomes were exposed to increasing temperatures and FP was measured on a BMG LABTECH microplate reader. As expected, the polarization value dropped at around 37 °C, indicating the transition from a high ordered gel state to liquid phase.
The HTS-compatible method allows for the monitoring of large scale alterations in membrane fluidity (e.g. phase transitions) as well as more subtle changes in lipid dynamics.