![]() Lipids present in the oil phase assemble along both the interfaces as the oil de-wets or is extracted to form liposomes. The single emulsion is further sheared into droplets by an outer aqueous solution (OA) to form W/O/W double emulsions. This results in the formation of water-in-oil (W/O) single emulsions whose interface is assembled with a monolayer of lipids, thanks to their amphiphilic nature. 1a) involves an inner aqueous solution (IA) that is sheared by an oil phase containing lipids (LO). Typically, the production of lipid vesicles using microfluidics (see Fig. This involves microdroplet technology to create double emulsions of water-in-oil-in-water (W/O/W) with lipids in the oil phase. Interestingly, all of the aforementioned drawbacks can be mitigated by implementing microfluidics to make lipid-based vesicles 14, 15, 16, 17, 18. In our previous work, we showed that the inverted emulsion method can be a straightforward and reliable technique to produce basic models for cells, albeit with some drawbacks such as lack of size control, low throughput, and its dependency on the density of the solutions used 9. In the past few years, our group and many other researchers have turned towards emulsion-based technologies to overcome these issues and precisely control the uniformity of the encapsulates 9, 10, 11, 12, 13. Although there are reports of encapsulating biomolecules in GUVs using swelling-based techniques, the reliability and reproducibility are low 7, 8. However, the limitations of these methods prevent high and uniform encapsulation of large and charged biomolecules. Currently, the most common methods include the well-established electroformation and spontaneous swelling to produce giant unilamellar vesicles (GUVs) 6. The latter being an essential step in the emerging and accelerating field of bottom-up synthetic biology 4, 5. ![]() A cell mimic should fulfill the basic requirements of being lipid-based, vesicular in structure, and encapsulating the desired biomolecules such as enzymes, DNA, and even smaller vesicles as artificial organelles 3. While applications of lipid vesicles in the field of health care are advancing in the form of nanometer-sized liposomes 2, their usability in understanding the evolution of cells and their various biochemical and physical pathways has hit a roadblock due to the lack of appropriate methods to form truly biomimetic cellular models, i.e., artificial cells. Lipids are not only biocompatible but are also the building blocks of life-forming vesicular structures, i.e., cells. Lipid-based vesicles have grown in popularity both in basic research and in application-oriented sciences, especially in pharmaceutics and cosmetics 1. ![]() This robust method capable of creating truly biomimetic artificial cells in high-throughput will prove valuable for bottom-up synthetic biology and the understanding of membrane function. Usability as artificial cells is demonstrated by increasing their complexity, i.e., by encapsulating plasmids, smaller liposomes, mammalian cells, and microspheres. Purity, functionality, and stability of the membranes are validated by lipid diffusion, protein incorporation, and leakage assays. The versatile design allows for the production of vesicle sizes ranging anywhere from ~10 to 130 µm with either neutral or charged lipids, and in physiological buffer conditions. Here we present a microfluidic method for producing biomimetic surfactant-free and additive-free giant unilamellar vesicles. While production is high-throughput and the lipid vesicles are mono-disperse compared to bulk methods, current technologies rely heavily on the addition of additives such as surfactants, glycerol and even ethanol. Microfluidic production of giant lipid vesicles presents a paradigm-shift in the development of artificial cells. ![]()
0 Comments
Leave a Reply. |