In vitro Assay to Evaluate Cation Transport of Ionophores

Ion homeostasis is a fundamental regulator of cellular processes and depends upon lipid membranes, which function as ion permeability barriers. Ionophores facilitate ion transport across cell membranes and offer a way to manipulate cellular ion composition. Here, we describe a calcein quenching assay based on large unilamellar vesicles that we used to evaluate divalent cation transport of the ionophore 4-Br-A23187. This assay can be used to study metal transport by ionophores and membrane proteins, under well-defined conditions. This protocol was validated in: Proteomics (2022), DOI: 10.1002/pmic.202200061 Graphical abstract.