Giant unilamellar vesicles (GUVs) are artificial membrane models which are of special interest to researchers because of their similarity in size to eukaryotic cells. The most commonly used method for GUVs production is electroformation. However, the traditional electroformation protocol involves a step in which the organic solvent is completely evaporated, leaving behind a dry lipid film. This leads to artifactual demixing of cholesterol (Chol) in the form of anhydrous crystals. These crystals do not participate in the formation of the lipid bilayer, resulting in a decrease of Chol concentration in the bilayer compared to the initial lipid solution. We propose a novel electroformation protocol which addresses this issue by combining the rapid solvent exchange, plasma cleaning and spin-coating techniques to produce GUVs from damp lipid films in a fast and reproducible manner. We have tested the protocol efficiency using 1/1 phosphatidylcholine/Chol and 1/1/1 phosphatidylcholine/sphingomyelin/Chol lipid mixtures and managed to produce a GUV population of an average diameter around 40 µm, with many GUVs being larger than 100 µm. Additionally, compared to protocols that include the dry film step, the sizes and quality of vesicles determined from fluorescence microscopy images were similar or better, confirming the benefits of our protocol in that regard as well.

Thermodynamics is a theory based on phenomenological premises and has wide applicability in science and technology. However, it remains one of the most challenging subjects to understand and teach, which makes it an excellent candidate for research and development of teaching methods. In this research, a questionnaire was used to evaluate the current knowledge of Bachelor’s and Master’s physics students, analyzing their immediate understanding of the topic and exploring their reasoning and thought processes. The questionnaire is divided into three sections which sequentially examine high school knowledge of entropy and thermodynamics; understanding of (ir)reversible processes related to energy and entropy change; and the distinction between equilibrium and nonequilibrium states. Based on the analysis of the results, we identified difficulties in understanding and articulating and applying the learned concepts. In particular, misunderstandings of entropy changes in isothermal processes and isolated systems are observed among students at all levels. Additionally, students find it difficult to distinguish between the contributions of energy and entropy changes to a system and its environment in the processes. The difficulty in defining (non)equilibrium states is present among Bachelor’s second-year physics students. To address these challenges, we propose adjustments to the teaching approach, including discussions about entropy sources and process (ir)reversibility, incorporating more theoretical and everyday examples of various processes and (non)equilibrium states and allowing more time for student discussions.