fluorescence-recovery-after-merging-droplet-to-measure-two

Fluorescence Recovery after Merging a Droplet to Measure the Two-dimensional Diffusion of a Phospholipid Monolayer

We introduce a new method to measure the lateral diffusivity of a surfactant monolayer at the fluid-fluid interface, called fluorescence recovery after ...

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At low mole fractions, cholesterol segregates into 10- to 100-nm-diameter nanodomains dispersed throughout primarily dipalmitoylphosphatidylcholine (DPPC) domains in mixed DPPC:cholesterol monolayers. The nanodomains consist of 6:1 DPPC:cholesterol "complexes" that decorate and lengthen DPPC domain boundaries, consistent with a reduced line tension, λ. The surface viscosity of the monolayer, ηs, decreases exponentially with the area fraction of the nanodomains at fixed surface pressure over the 0.1- to 10-Hz range of frequencies common to respiration. At fixed cholesterol fraction, the surface viscosity increases exponentially with surface pressure in similar ways for all cholesterol fractions. This increase can be explained with a free-area model that relates ηs to the pure DPPC monolayer compressibility and collapse pressure. The elastic modulus, G', initially decreases with cholesterol fraction, consistent with the decrease in λ expected from the line-active nanodomains, in analogy to 3D emulsions. However, increasing cholesterol further causes a sharp increase in G' between 4 and 5 mol% cholesterol owing to an evolution in the domain morphology, so that the monolayer is elastic rather than viscous over 0.1-10 Hz. Understanding the effects of small mole fractions of cholesterol should help resolve the controversial role cholesterol plays in human lung surfactants and may give clues as to how cholesterol influences raft formation in cell membranes.

Effect of cholesterol nanodomains on monolayer morphology and dynamics.

Adding small fractions of cholesterol decreases the interfacial viscosity of dipalmitoylphosphatidylcholine (DPPC) monolayers by an order of magnitude per wt %. Grazing incidence X-ray diffraction shows that cholesterol at these small fractions does not mix ideally with DPPC but rather induces nanophase separated structures of an ordered, primarily DPPC phase bordered by a line-active, disordered, mixed DPPC-cholesterol phase. We propose that the free area in the classic Cohen and Turnbull model of viscosity is inversely proportional to the number of molecules in the coherence area, or product of the two coherence lengths. Cholesterol significantly reduces the coherence area of the crystals as well as the interfacial viscosity. Using this free area collapses the surface viscosity data for all surface pressures and cholesterol fractions to a universal logarithmic relation. The extent of molecular coherence appears to be a fundamental factor in determining surface viscosity in ordered monolayers.

Influence of molecular coherence on surface viscosity.

We study electrostatic interactions of polystyrene particles at an oil/water interface controlled by a chemical reaction of carboxylate surface functional groups. By replacing the carboxyl functional groups with hydrocarbon chains using the well-known EDC (1-ethyl-3-(3-(dimethylamino)propyl)carbodiimide) coupling reaction, the surface charge density decreases while the hydrophobicity of the colloid surface increases. Direct visualization of the particle-laden interface reveals that, depending on the extent of hydrocarbon coupling, the strength of the electrostatic repulsion can be regulated: the repulsive interaction increases with the reaction, removing aggregates, but rapidly decreases if the reaction proceeds too much, forming a large aggregation. This simple reaction, thus, dramatically changes the structures of the colloidal monolayers at the oil/water interface. We conclude that such structural change is the result of change of the repulsive interactions from the oil phase, although interactions in the water phase are also changed slightly.

Surface charge regulation of carboxyl terminated polystyrene latex particles and their interactions at the oil/water interface.

We present a novel technique to measure diffusion coefficients of insoluble surfactant monolayers. We merge a surfactant-coated droplet with a fluorescently labeled planar monolayer. During the merging process, a monolayer on a droplet displaces the existing planar monolayer, leaving a dark area when viewed under a fluorescence microscope. We measure fractional intensities as the dyes recover, which allows diffusion coefficients to be computed. We validate this technique with the two most common phospholipid monolayers (DPPC and DOPC) and study the diffusion of their mixtures. The proposed technique has several advantages over the FRAP technique and is potentially capable of measuring the diffusion of any soluble/insoluble surfactant monolayers.

Fluorescence recovery after merging a surfactant-covered droplet: a novel technique to measure the diffusion of phospholipid monolayers at fluid/fluid interfaces.

Article	Year
Effect of cholesterol nanodomains on monolayer morphology and dynamics. Proceedings of the National Academy of Sciences of the United States of America\| PubMed ID: 23901107	2013
Influence of molecular coherence on surface viscosity. Langmuir : the ACS journal of surfaces and colloids\| PubMed ID: 24991992	2014
Surface charge regulation of carboxyl terminated polystyrene latex particles and their interactions at the oil/water interface. Langmuir : the ACS journal of surfaces and colloids\| PubMed ID: 25226338	2014
Fluorescence recovery after merging a surfactant-covered droplet: a novel technique to measure the diffusion of phospholipid monolayers at fluid/fluid interfaces. Langmuir : the ACS journal of surfaces and colloids\| PubMed ID: 25422050	2014

KyuHan Kim

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