1-2-dioleoylphosphatidylserine and 1-2-distearoyllecithin

For quantitative analysis, data should be obtained at a sample concentration that is within the range of linearity. We examined the effect of sample concentration on nanoparticle tracking analysis (NTA) of small extracellular vesicles (sEVs), including exosomes, by comparing NTA results of sEVs with those obtained for polystyrene nanoparticles (PSN) and liposomes, which mimic lipid composition and physicochemical properties of exosomes. Initially, NTA of PSN at different concentrations was performed and the particle sizes determined were validated by dynamic light scattering. The major peak maxima for PSN mixtures of different sizes at the higher particle numbers were similar, with some fluctuation of the minor peak maxima observed at the lower particle number, which was also observed for sEVs. Sample concentration is critical for obtaining reproducible data for liposomes and exosomes and increasing the sample concentration caused an increase in data variability because of particle interactions. The inter-day repeatability of particles sizes and concentration for sEVs were 7.47 and 4.51%, respectively. Analysis of the linearity range revealed that this was narrower for sEVs when compared with that of liposomes. Owing to the use of liposomes that mimic the lipid composition and physicochemical properties of exosomes and proteinase-treated sEVs, it was demonstrated that these different analytical results could be possibly caused by the protein corona of sEVs. Consideration of the sample concentration and linearity range is important for obtaining reproducible and reliable data of sEVs.

Topics: Exosomes; Extracellular Vesicles; HeLa Cells; Hep G2 Cells; Humans; K562 Cells; Limit of Detection; Liposomes; Nanoparticles; Particle Size; Phosphatidylcholines; Phosphatidylserines; Reproducibility of Results; Single Molecule Imaging

Zeta potential is often used to approximate a nanoparticle's surface charge, i.e., cationic, anionic, or neutral character, and has become a standard characterization technique to evaluate nanoparticle surfaces. While useful, zeta potential values provide only very general conclusions about surface charge character. Without a thorough understanding of the measurement parameters and limitations of the technique, these values can become meaningless. This case study attempts to explore the sensitivity of zeta potential measurement using specifically formulated cationic, anionic, and neutral liposomes. This study examines zeta potential dependence on pH and ionic strength, resolving power, and highlights the sensitivity of zeta potential to charged liposomes. Liposomes were prepared with cholesterol, 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), and varying amounts of 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) or 1,2-dioleoyl-sn-glycero-3-phospho-L-serine (DOPS). A strong linear relationship was noted between zeta potential values and the mole percentage of charged lipids within a liposome (e.g., cationic DOTAP or anionic DOPS). This finding could be used to formulate similar liposomes to a specific zeta potential, potentially of importance for systems sensitive to highly charged species. In addition, cationic and anionic liposomes were titrated with up to two mole percent of the neutral lipid 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000] (lipid-PEG; LP). Very small amounts of the lipid-PEG (<0.2 mol%) were found to impart stability to the DOTAP- and DOPS-containing liposomes without significantly affecting other physicochemical properties of the formulation, providing a simple approach to making stable liposomes with cationic and anionic surface charge.

Topics: Anions; Cations; Cholesterol; Fatty Acids, Monounsaturated; Liposomes; Osmolar Concentration; Phosphatidylcholines; Phosphatidylserines; Polyethylene Glycols; Quaternary Ammonium Compounds; Static Electricity; Surface Properties

The partitioning behavior of gramicidin A' was investigated in four binary phospholipid mixtures with coexisting fluid and gel phases. The ratio of the equilibrium peptide concentration in the fluid phase to that in the gel phase (i.e., the partition coefficient, Kp) was determined by analysis of the quenching of gramicidin A' tryptophanyl fluorescence by a spin-labeled phosphatidylcholine. The partition coefficient was used as a measure of the relative solubility of gramicidin A' in the four types of gel phases analyzed. The composition of the gel phase was entirely Ca(dioleoylphosphatidylserine)2 (Ca(di18:1-PS)2), or was rich in either distearoylphosphatidylcholine (di18:0-PC), dipalmitoylphosphatidylcholine (di16:0-PC), or dimyristoylphosphatidylcholine (di14:0-PC). Except in the last case, the gel phase was depleted of gramicidin A': Kp approximately 30 when the gel phase was Ca(di18:1-PS)2 or di18:0-PC-rich, Kp approximately 10 when the gel phase was di16:0-PC-rich, and Kp approximately 1 when the gel phase was di14:0-PC-rich. The hydrophobic mismatch between the length of gramicidin A' and the length of the phospholipid acyl chains in the bulk gel phase is greatest with di18:1-PS and di18:0-PC, intermediate with di16:0-PC, and least with di14:0-PC. The Kp measurements presented here are consistent with increasing solubility of gramicidin A' in the gel phase with decreasing hydrophobic mismatch.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Dimyristoylphosphatidylcholine; Gels; Gramicidin; Liposomes; Molecular Conformation; Phosphatidylcholines; Phosphatidylserines; Phospholipids; Spectrometry, Fluorescence