Effects of Electrolytes and Lipid Charge on the Productivity of Giant Unilamelar Vesicle (GUV) by Electroformation

Abstract

Sugars and related compounds are increasingly being used as potential cryo-and lyo-protective agents for preservation of rare and valuable mammalian cells and tissues, although a high performance introduction of these compounds, especially oligo-saccharide, into living cells is one of the key technology. We propose Giant Unilamelar Vesicle (GUV) -cell electrofusion as a noble large molecular delivery method, which resembles native endocytosis of vesicles. Although there is few conventional methods to produce GUV, electroformation is one of the promising methods of producing GUV. For the better understanding of mechanism of electroformation, we investigate effects of extra-vesicle electrolytes (K+ and Ca2+) and charged lipids (L-α-Phosphatidylcholine; PC) on productivity of electroformed GUV. Zeta potential of PC in various concentration of CaCl2 solutions including glucose or trehalose was measured to estimate the amount of charge on PC. The results showed that; 1) trehalose inhibited PC to be charged by Ca2+, while glucose showed no inhibition. 2) PC in KCl solution was not charged. The number density and the averaged diameter of GUVs produced in various concentration of CaCl2 or KCl solutions were measured. Regardless of ion species, the GUV-productivity by electroformation decreases as the electric conductivity of extra-vesicles increases. It might be concluded that the electric conductivity of extra-vesicles plays more important role in electroformation than the charge of lipid.