Biophysical Journal 63: 1336-1345 (1992)
© 1992 the Biophysical Society

This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Harrigan, P R Articles by Cullis, P R Search for Related Content PubMed PubMed Citation Articles by Harrigan, P R Articles by Cullis, P R

Determination of transmembrane pH gradients and membrane potentials in liposomes.

Department of Biochemistry, University of British Columbia, Vancouver, Canada.

ABSTRACT

Techniques for determining large transbilayer pH gradients (delta pH) and membrane potentials (delta psi) induced in response to delta pH in large unilamellar vesicle liposomal systems by measuring the transbilayer redistribution of radiolabeled compounds have been examined. For liposomes with acidic interiors, it is shown that protocols using radiolabeled methylamine in conjunction with gel filtration procedures to remove untrapped methylamine provide accurate measures of delta pH in most situations. Exceptions include gel state lipid systems, where transbilayer equilibration processes are slow, and situations where the interior buffering capacity is limited. These problems can be circumvented by incubation at elevated temperatures and by using probes with higher specific activities, respectively. Determination of delta pH in vesicles with a basic interior using weak acid probes such as radiolabeled acetate in conjunction with gel filtration was found to be less reliable, and an alternative equilibrium centrifugation protocol is described. In the case of determinations of the membrane potentials induced in response to these pH gradients, probes such as tetraphenylphosphonium and thiocyanate provide relatively accurate measures of the delta psi induced. It is shown that the maximum transmembrane pH gradient that can be stably maintained by an egg phosphatidylcholine-cholesterol 100-nm-diam large unilamellar vesicle is approximately 3.7 units, corresponding to an induced delta psi of 220 mV or transbilayer electrical field of 5 x 10(5) V/cm.

This article has been cited by other articles:

				S. A. Abraham, C. McKenzie, D. Masin, R. Ng, T. O. Harasym, L. D. Mayer, and M. B. Bally In Vitro and in Vivo Characterization of Doxorubicin and Vincristine Coencapsulated within Liposomes through Use of Transition Metal Ion Complexation and pH Gradient Loading Clin. Cancer Res., January 15, 2004; 10(2): 728 - 738. [Abstract] [Full Text] [PDF]

				M. S. Webb, N. L. Boman, D. J. Wiseman, D. Saxon, K. Sutton, K. F. Wong, P. Logan, and M. J. Hope Antibacterial Efficacy against an In Vivo Salmonella typhimurium Infection Model and Pharmacokinetics of a Liposomal Ciprofloxacin Formulation Antimicrob. Agents Chemother., January 1, 1998; 42(1): 45 - 52. [Abstract] [Full Text]