Imaging molecular transport across and nanomaterial interaction with lipid membranes

The ability of a molecule to pass through the plasma membrane without the aid of any active cellular mechanisms is central to that molecule's pharmaceutical characteristics. Passive transport has been understood in the context of Overton's rule, which states that more lipophilic molecules cross membrane lipid bilayers more readily. Standard techniques including planar lipid membrane, liposome, and cell monolayer to observe passive transport processes are flawed and lack reproducibility.;This research describes assays based on spinning-disk confocal microscopy (SDCM) of giant unilamellar vesicles (GUVs) that allow for fluorescent molecules to be tracked as they permeate the lipid membrane. This approach allows for the temporal development of the concentration field to be directly observed. Precise membrane permeability can be determined from by fitting the data to a mathematical permeation model.;A series of molecules of increasing hydrophilicity was constructed by conjugating 4-nitrobenzo-2-oxa-1,3-diazole (NBD) with poly(ethylene-glycol) (PEG). An analytical passive transport model was devised, image intensity data was regressed to the model, and permeability was calculated. The result shows that longer chain PEG molecules which are more hydrophilic permeate more slowly. This trend is consistent with Overton's rule, though it does not seem to fit a simple partition-diffusion model of membrane transport.;Low-molecular-weight carboxylic acids have crucial effects on cellular processes. We studied the transport of carboxylic acids with different carbon chains lengths into GUVs. Fluorescein-dextran was used to trace the transport of acid. GUVs were immobilized on the surface of a poly(dimethylsiloxane) (PDMS) microchannel which allows the changing of buffer solution quickly and uniformly. The results showed that the permeabilities are consistent with octanol-water partition coefficients and demonstrate that Overton's rule applies for this class of molecules.;Synthetic lipid bilayers were used to study potentially harmful interactions between nanoparticles and biomembranes. Twenty nm polystyrene nanoparticles with cationic surfaces adhere strongly to lipid membranes. Adhesion is driven by nonspecific electrostatic interactions between the lipid phosphate groups and the nanoparticles. Nanoparticle adhesion leads to membrane morphological deformation and the formation of transient nanoscale pores. These results suggest that nanoparticle adhesion imposes surface tension on biomembranes via a steric crowding mechanism, leading to poration.