» Articles » PMID: 36984739

Electroformation of Giant Unilamellar Vesicles from Damp Lipid Films Formed by Vesicle Fusion

Overview
Date 2023 Mar 29
PMID 36984739
Authors
Affiliations
Soon will be listed here.
Abstract

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.

Citing Articles

Electroformation of Giant Unilamellar Vesicles from Damp Films in Conditions Involving High Cholesterol Contents, Charged Lipids, and Saline Solutions.

Mardesic I, Boban Z, Raguz M Membranes (Basel). 2024; 14(10).

PMID: 39452827 PMC: 11510074. DOI: 10.3390/membranes14100215.


Electroformation of Giant Unilamellar Vesicles from Damp Lipid Films with a Focus on Vesicles with High Cholesterol Content.

Mardesic I, Boban Z, Raguz M Membranes (Basel). 2024; 14(4).

PMID: 38668107 PMC: 11051717. DOI: 10.3390/membranes14040079.

References
1.
Ong S, Chitneni M, Lee K, Ming L, Yuen K . Evaluation of Extrusion Technique for Nanosizing Liposomes. Pharmaceutics. 2016; 8(4). PMC: 5198018. DOI: 10.3390/pharmaceutics8040036. View

2.
Baykal-Caglar E, Hassan-Zadeh E, Saremi B, Huang J . Preparation of giant unilamellar vesicles from damp lipid film for better lipid compositional uniformity. Biochim Biophys Acta. 2012; 1818(11):2598-604. DOI: 10.1016/j.bbamem.2012.05.023. View

3.
Ionova I, Livshits V, Marsh D . Phase diagram of ternary cholesterol/palmitoylsphingomyelin/palmitoyloleoyl-phosphatidylcholine mixtures: spin-label EPR study of lipid-raft formation. Biophys J. 2012; 102(8):1856-65. PMC: 3328704. DOI: 10.1016/j.bpj.2012.03.043. View

4.
Bhatia T, Husen P, Brewer J, Bagatolli L, Hansen P, Ipsen J . Preparing giant unilamellar vesicles (GUVs) of complex lipid mixtures on demand: Mixing small unilamellar vesicles of compositionally heterogeneous mixtures. Biochim Biophys Acta. 2015; 1848(12):3175-80. DOI: 10.1016/j.bbamem.2015.09.020. View

5.
Schindelin J, Arganda-Carreras I, Frise E, Kaynig V, Longair M, Pietzsch T . Fiji: an open-source platform for biological-image analysis. Nat Methods. 2012; 9(7):676-82. PMC: 3855844. DOI: 10.1038/nmeth.2019. View