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Abstract Small, spherical vesicles are a widely used chassis for the formation of model protocells and investigating the beginning of compartmentalized evolution. Various methods exist for their preparation, with one of the most common approaches being gentle hydration, where thin layers of lipids are hydrated with aqueous solutions and gently agitated to form vesicles. An important benefit to gentle hydration is that the method produces vesicles without introducing any organic contaminants, such as mineral oil, into the lipid bilayer. However, compared to other methods of liposome formation, gentle hydration is much less efficient at encapsulating aqueous cargo. Improving the encapsulation efficiency of gentle hydration would be of broad use for medicine, biotechnology, and protocell research. Here, we describe a method of sequentially hydrating lipid thin films to increase encapsulation efficiency. We demonstrate that sequential gentle hydration significantly improves encapsulation of water-soluble cargo compared to the traditional method, and that this improved efficiency is dependent on buffer composition. Similarly, we also demonstrate how this method can be used to increase concentrations of oleic acid, a fatty acid commonly used in origins of life research, to improve the formation of vesicles in aqueous buffer.
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Dynamics of giant vesicle assembly from thin lipid films
Motivation Giant unilamellar vesicles (GUVs), cell-like synthetic micrometer size structures, assemble when thin lipid films are hydrated in aqueous solutions. Quantitative measurements of static yields and distribution of sizes of GUVs obtained from thin film hydration methods were recently reported. Dynamic data such as the time evolution of yields and distribution of sizes, however, is not known. Dynamic data can provide insights into the assembly pathway of GUVs and guidelines for choosing conditions to obtain populations with desired size distributions. Approach We develop the ‘stopped-time’ technique to characterize the time evolution of the distribution of sizes and molar yields of populations of free-floating GUVs. We additionally capture high resolution time-lapse images of surface-attached GUV buds on the lipid films. We systematically study the dynamics of assembly of GUVs from three widely used thin film hydration methods, PAPYRUS (Paper-Abetted amPhiphile hYdRation in aqUeous Solutions), gentle hydration, and electroformation. Findings We find that the molar yield versus time curves of GUVs demonstrate a characteristic sigmoidal shape, with an initial yield, a transient, and then a steady state plateau for all three methods. The population of GUVs showed a right-skewed distribution of diameters. The variance of the distributions increased with time. The systems reached steady state within 120 min. We rationalize the dynamics using the thermodynamically motivated budding and merging (BNM) model. These results further the understanding of lipid dynamics and provide for the first-time practical parameters to tailor the production of GUVs of specific sizes for applications.
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- PAR ID:
- 10504845
- Publisher / Repository:
- Journal of Colloid and Interface Science
- Date Published:
- Journal Name:
- Journal of Colloid and Interface Science
- Volume:
- 661
- Issue:
- C
- ISSN:
- 0021-9797
- Page Range / eLocation ID:
- 1033 to 1045
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1016/j.jcis.2024.02.022
