Abstract
Purification of plasma membranes by two-phase partitioning is based on the separation of microsomal membranes, dependent on their surface hydrophobicity. Here we explain the purification of plasma membranes from a relatively small amount of material (7–30 g). The fluorescent probe ACMA (9-amino-6-chloro-2-metoxyacridine) accumulates inside the vesicles upon protonation. Quenching of ACMA in the solution corresponds to the H+ transport across the plasma membrane. Before running the assay, the plasma membranes are incubated with the detergent Brij-58 in order to create inside-out vesicles.
Purification of plasma membranes by two-phase partitioning is based on the separation of microsomal membranes, dependent on their surface hydrophobicity. Here we explain the purification of plasma membranes from a relatively small amount of material (7–30 g). The fluorescent probe ACMA (9-amino-6-chloro-2-metoxyacridine) accumulates inside the vesicles upon protonation. Quenching of ACMA in the solution corresponds to the H+ transport across the plasma membrane. Before running the assay, the plasma membranes are incubated with the detergent Brij-58 in order to create inside-out vesicles.
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References
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Lund, A., Fuglsang, A.T. (2012). Purification of Plant Plasma Membranes by Two-Phase Partitioning and Measurement of H+ Pumping. In: Shabala, S., Cuin, T. (eds) Plant Salt Tolerance. Methods in Molecular Biology, vol 913. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-986-0_14
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DOI: https://doi.org/10.1007/978-1-61779-986-0_14
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Publisher Name: Humana Press, Totowa, NJ
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Online ISBN: 978-1-61779-986-0
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