Wicking: a rapid method for manually inserting ion channels into planar lipid bilayers.

The planar lipid bilayer technique has a distinguished history in electrophysiology but is arguably the most technically difficult and time-consuming method in the field. Behind this is a lack of experimental consistency between laboratories, the challenges associated with painting unilamellar bilay...

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Autores principales:	Justin A Costa, Dac A Nguyen, Edgar Leal-Pinto, Ronald E Gordon, Basil Hanss
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2013
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/9c41735cb8ba4a64b7f2cac1758dd5da
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spelling	oai:doaj.org-article:9c41735cb8ba4a64b7f2cac1758dd5da2021-11-18T07:44:28ZWicking: a rapid method for manually inserting ion channels into planar lipid bilayers.1932-620310.1371/journal.pone.0060836https://doaj.org/article/9c41735cb8ba4a64b7f2cac1758dd5da2013-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/23717384/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203The planar lipid bilayer technique has a distinguished history in electrophysiology but is arguably the most technically difficult and time-consuming method in the field. Behind this is a lack of experimental consistency between laboratories, the challenges associated with painting unilamellar bilayers, and the reconstitution of ion channels into them. While there has be a trend towards automation of this technique, there remain many instances where manual bilayer formation and subsequent membrane protein insertion is both required and advantageous. We have developed a comprehensive method, which we have termed "wicking", that greatly simplifies many experimental aspects of the lipid bilayer system. Wicking allows one to manually insert ion channels into planar lipid bilayers in a matter of seconds, without the use of a magnetic stir bar or the addition of other chemicals to monitor or promote the fusion of proteoliposomes. We used the wicking method in conjunction with a standard membrane capacitance test and a simple method of proteoliposome preparation that generates a heterogeneous mixture of vesicle sizes. To determine the robustness of this technique, we selected two ion channels that have been well characterized in the literature: CLIC1 and α-hemolysin. When reconstituted using the wicking technique, CLIC1 showed biophysical characteristics congruent with published reports from other groups; and α-hemolysin demonstrated Type A and B events when threading single stranded DNA through the pore. We conclude that the wicking method gives the investigator a high degree of control over many aspects of the lipid bilayer system, while greatly reducing the time required for channel reconstitution.Justin A CostaDac A NguyenEdgar Leal-PintoRonald E GordonBasil HanssPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 8, Iss 5, p e60836 (2013)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Justin A Costa Dac A Nguyen Edgar Leal-Pinto Ronald E Gordon Basil Hanss Wicking: a rapid method for manually inserting ion channels into planar lipid bilayers.
description	The planar lipid bilayer technique has a distinguished history in electrophysiology but is arguably the most technically difficult and time-consuming method in the field. Behind this is a lack of experimental consistency between laboratories, the challenges associated with painting unilamellar bilayers, and the reconstitution of ion channels into them. While there has be a trend towards automation of this technique, there remain many instances where manual bilayer formation and subsequent membrane protein insertion is both required and advantageous. We have developed a comprehensive method, which we have termed "wicking", that greatly simplifies many experimental aspects of the lipid bilayer system. Wicking allows one to manually insert ion channels into planar lipid bilayers in a matter of seconds, without the use of a magnetic stir bar or the addition of other chemicals to monitor or promote the fusion of proteoliposomes. We used the wicking method in conjunction with a standard membrane capacitance test and a simple method of proteoliposome preparation that generates a heterogeneous mixture of vesicle sizes. To determine the robustness of this technique, we selected two ion channels that have been well characterized in the literature: CLIC1 and α-hemolysin. When reconstituted using the wicking technique, CLIC1 showed biophysical characteristics congruent with published reports from other groups; and α-hemolysin demonstrated Type A and B events when threading single stranded DNA through the pore. We conclude that the wicking method gives the investigator a high degree of control over many aspects of the lipid bilayer system, while greatly reducing the time required for channel reconstitution.
format	article
author	Justin A Costa Dac A Nguyen Edgar Leal-Pinto Ronald E Gordon Basil Hanss
author_facet	Justin A Costa Dac A Nguyen Edgar Leal-Pinto Ronald E Gordon Basil Hanss
author_sort	Justin A Costa
title	Wicking: a rapid method for manually inserting ion channels into planar lipid bilayers.
title_short	Wicking: a rapid method for manually inserting ion channels into planar lipid bilayers.
title_full	Wicking: a rapid method for manually inserting ion channels into planar lipid bilayers.
title_fullStr	Wicking: a rapid method for manually inserting ion channels into planar lipid bilayers.
title_full_unstemmed	Wicking: a rapid method for manually inserting ion channels into planar lipid bilayers.
title_sort	wicking: a rapid method for manually inserting ion channels into planar lipid bilayers.
publisher	Public Library of Science (PLoS)
publishDate	2013
url	https://doaj.org/article/9c41735cb8ba4a64b7f2cac1758dd5da
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Wicking: a rapid method for manually inserting ion channels into planar lipid bilayers.

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