Lipid-Protein Membrane Structure-Function Characterization using Droplet Interface Bilayers

Peter T. Podar; Ariana Adkisson-Washington; Olivia Ziemer; Victoria Sullivan; Christian Milem; Janine Hagar-Montoya; Shriya Khule; Zachary Griffith; Khalid Tantawi; Xiuhong Zhai; Dima Bolmatov; John Katsaras; Charles Patrick Collier

doi:10.3791/70628

Lipid-Protein Membrane Structure-Function Characterization using Droplet Interface Bilayers

Peter T. Podar
, Ariana Adkisson-Washington
, Olivia Ziemer
, Victoria Sullivan
, Christian Milem
, Janine Hagar-Montoya
, Shriya Khule
, Zachary Griffith
, Khalid Tantawi
, Xiuhong Zhai
, Dima Bolmatov
, John Katsaras
, Charles Patrick Collier

Research output: Contribution to journal › Article › peer-review

Abstract

Droplet interface bilayers (DIBs) offer a tunable platform for probing the electromechanical properties of lipid and lipid-peptide membranes under controlled electrical stimulation. DIBs enable both single-channel and ensemble ion conductance measurements over membrane areas orders of magnitude larger than those accessible by traditional patch clamp techniques, thereby allowing membrane-level analyses of electromechanical deformation and its influence on ion-conducting peptides. By systematically tuning membrane structure through the bulk hydrocarbon oil phase (e.g., hexadecane [C16] vs. dodecane/hexadecane [C12/C16] [25%/75%, v/v]), this bottom-up platform enables systematic variation of membrane composition and oil environment, which influence membrane viscoelasticity and structural reorganization, and thereby peptide ion conduction. Detailed procedures are provided for the assembly of gramicidin A-doped 1,2-diphytanoyl-sn-glycero-3-phosphocholine (DPhPC) membranes using different hydrocarbon oil compositions and for the application of voltage-pulse protocols that drive membranes into metastable electromechanical states. Adaptive membrane ion conduction is characterized, including short-term plasticity-like (STP-like) and long-term potentiation-and depression-like (LTP-like/LTD-like) responses in a model membrane system. More broadly, this protocol provides a robust, reproducible approach for systematically investigating composition-dependent, membrane-level electromechanical contributions to synaptic-like conductive behavior and for understanding how lipid membrane environments modulate ion channel function.

Original language	English
Article number	e70628
Journal	Journal of Visualized Experiments
Volume	2026-June
Issue number	232
DOIs	https://doi.org/10.3791/70628
State	Published - Jun 2026

Funding

C.P.C. and J.K. are supported through the Scientific User Facilities Division of the Department of Energy (DOE) Office of Science, sponsored by the Basic Energy Science (BES) Program, DOE Office of Science, under Contract No. DE-AC05-00OR22725. D.B. was supported through the National Science Foundation, Division of Molecular and Cellular Biosciences (MCB), under contract no. 2219289. The research was partly funded through the Nonequilibrium and Emergent Transients in Advanced and Soft Materials (NEAT) award, sponsored by the Laboratory Directed Research and Development Program of Oak Ridge National Laboratory, managed by UT-Battelle, LLC, for the U. S. Department of Energy. P.T.P. and C.M. were supported through the DOE Omni Technology Alliance Internship Program and the Education Collaboration at ORNL (ECO) program. P.T.P. and V.S. were supported by the Oak Ridge National Laboratory (ORNL) Research Student Internships (RSI) program. P.T.P., O.Z., and Z.G. were supported through the DOE Science Undergraduate Laboratory Internships (SULI) program. A.A. and J.H.M. were supported by a Graduate Education for Minority Students (GEM) Fellowship. Data acquisition and analysis were carried out at the Shull-Wollan Center and at the Center for Nanophase Materials Sciences, which is a DOE Office of Science User Facility.

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10.3791/70628

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Podar, P. T., Adkisson-Washington, A., Ziemer, O., Sullivan, V., Milem, C., Hagar-Montoya, J., Khule, S., Griffith, Z., Tantawi, K., Zhai, X., Bolmatov, D., Katsaras, J., & Collier, C. P. (2026). Lipid-Protein Membrane Structure-Function Characterization using Droplet Interface Bilayers. Journal of Visualized Experiments, 2026-June(232), Article e70628. https://doi.org/10.3791/70628

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abstract = "Droplet interface bilayers (DIBs) offer a tunable platform for probing the electromechanical properties of lipid and lipid-peptide membranes under controlled electrical stimulation. DIBs enable both single-channel and ensemble ion conductance measurements over membrane areas orders of magnitude larger than those accessible by traditional patch clamp techniques, thereby allowing membrane-level analyses of electromechanical deformation and its influence on ion-conducting peptides. By systematically tuning membrane structure through the bulk hydrocarbon oil phase (e.g., hexadecane [C16] vs. dodecane/hexadecane [C12/C16] [25\%/75\%, v/v]), this bottom-up platform enables systematic variation of membrane composition and oil environment, which influence membrane viscoelasticity and structural reorganization, and thereby peptide ion conduction. Detailed procedures are provided for the assembly of gramicidin A-doped 1,2-diphytanoyl-sn-glycero-3-phosphocholine (DPhPC) membranes using different hydrocarbon oil compositions and for the application of voltage-pulse protocols that drive membranes into metastable electromechanical states. Adaptive membrane ion conduction is characterized, including short-term plasticity-like (STP-like) and long-term potentiation-and depression-like (LTP-like/LTD-like) responses in a model membrane system. More broadly, this protocol provides a robust, reproducible approach for systematically investigating composition-dependent, membrane-level electromechanical contributions to synaptic-like conductive behavior and for understanding how lipid membrane environments modulate ion channel function.",

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AU - Hagar-Montoya, Janine

AU - Khule, Shriya

AU - Griffith, Zachary

AU - Tantawi, Khalid

AU - Zhai, Xiuhong

AU - Bolmatov, Dima

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AB - Droplet interface bilayers (DIBs) offer a tunable platform for probing the electromechanical properties of lipid and lipid-peptide membranes under controlled electrical stimulation. DIBs enable both single-channel and ensemble ion conductance measurements over membrane areas orders of magnitude larger than those accessible by traditional patch clamp techniques, thereby allowing membrane-level analyses of electromechanical deformation and its influence on ion-conducting peptides. By systematically tuning membrane structure through the bulk hydrocarbon oil phase (e.g., hexadecane [C16] vs. dodecane/hexadecane [C12/C16] [25%/75%, v/v]), this bottom-up platform enables systematic variation of membrane composition and oil environment, which influence membrane viscoelasticity and structural reorganization, and thereby peptide ion conduction. Detailed procedures are provided for the assembly of gramicidin A-doped 1,2-diphytanoyl-sn-glycero-3-phosphocholine (DPhPC) membranes using different hydrocarbon oil compositions and for the application of voltage-pulse protocols that drive membranes into metastable electromechanical states. Adaptive membrane ion conduction is characterized, including short-term plasticity-like (STP-like) and long-term potentiation-and depression-like (LTP-like/LTD-like) responses in a model membrane system. More broadly, this protocol provides a robust, reproducible approach for systematically investigating composition-dependent, membrane-level electromechanical contributions to synaptic-like conductive behavior and for understanding how lipid membrane environments modulate ion channel function.

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Lipid-Protein Membrane Structure-Function Characterization using Droplet Interface Bilayers

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