Controlled delivery of proteins into bilayer lipid membranes on chip

Zagnoni, Michele; Sandison, Mairi E.; Marius, Phedra; Lee, Anthony G.; Morgan, Hywel

doi:10.1039/b703818f

Cited by 64 publications

(53 citation statements)

References 48 publications

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“…This assay has been used continuously in a variety of labs for fusion/reconstruction studies (48)(49)(50)(51)(54)(55)(56)(57)(58)(59)(60). Liposome fusion to planar bilayers was measured using standard electrophysiological equipment (Bilayer Clamp Amplifier model BC-535 or BC-525C, Warner Instruments, Hamden CT).…”

Section: Fusion Assaymentioning

confidence: 99%

Drunken Membranes: Short-Chain Alcohols Alter Fusion of Liposomes to Planar Lipid Bilayers

Paxman

Hunt

Hallan

et al. 2017

Biophysical Journal

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Although the effects of ethanol on protein receptors and lipid membranes have been studied extensively, ethanol's effect on vesicles fusing to lipid bilayers is not known. To determine the effect of alcohols on fusion rates, we utilized the nystatin/ergosterol fusion assay to measure fusion of liposomes to a planar lipid bilayer (BLM). The addition of ethanol excited fusion when applied on the cis (vesicle) side, and inhibited fusion on the trans side. Other short-chain alcohols followed a similar pattern. In general, the inhibitory effect of alcohols (trans) occurs at lower doses than the excitatory (cis) effect, with a decrease of 29% in fusion rates at the legal driving limit of 0.08% (w/v) ethanol (IC 50 ¼ 0.2% v/v, 34 mM). Similar inhibitory effects were observed with methanol, propanol, and butanol, with ethanol being the most potent. Significant variability was observed with different alcohols when applied to the cis side. Ethanol and propanol enhanced fusion, butanol also enhanced fusion but was less potent, and low doses of methanol mildly inhibited fusion. The inhibition by trans addition of alcohols implies that they alter the planar membrane structure and thereby increase the activation energy required for fusion, likely through an increase in membrane fluidity. The cis data are likely a combination of the above effect and a proportionally greater lowering of the vesicle lysis tension and hydration repulsive pressure that combine to enhance fusion. Alternate hypotheses are also discussed. The inhibitory effect of ethanol on liposome-membrane fusion is large enough to provide a possible biophysical explanation of compromised neuronal behavior.

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Section: Fusion Assaymentioning

confidence: 99%

Drunken Membranes: Short-Chain Alcohols Alter Fusion of Liposomes to Planar Lipid Bilayers

Paxman

Hunt

Hallan

et al. 2017

Biophysical Journal

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“…[ 27 ] Ionophores, valinomycin, and nigericin, were added to the solution in the PDMS microfl uidic channel at a fi nal concentration of 2 µM and incubated in the fl uid cell for approximately 30 min to allow suffi cient time for incorporation into the bR containing bilayer. Afterwards, the chamber was washed with buffer.…”

Section: Proteoliposomementioning

confidence: 99%

Bioelectronic Light‐Gated Transistors with Biologically Tunable Performance

et al. 2014

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“…20 Following droplet contact, a salt gradient was established across the droplet-interface BLM initiating vesicle fusion and leading to observable KcsA electrical activity (the conductance level and open probability of the channel is similar to that observed previously). 19, 20 A network of BLMs could also be formed. The formation of two BLMs at the interfaces between three droplets is illustrated in Figs.…”

mentioning

confidence: 99%

Formation of artificial lipid bilayers using droplet dielectrophoresis

Aghdaei

Sandison

Zagnoni³

et al. 2008

Lab Chip

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We describe the formation of artificial bilayer lipid membranes (BLMs) by the controlled, electrical manipulation of aqueous droplets immersed in a lipid-alkane solution. Droplet movement was generated using dielectrophoresis on planar microelectrodes covered in a thin insulator. Droplets, surrounded by lipid monolayers, were brought into contact and spontaneously formed a BLM. The method produced BLMs suitable for single-channel recording of membrane protein activity and the technique can be extended to create programmable BLM arrays and networks.Artificial BLM techniques are of great importance in membrane protein research. Electrophysiological studies of proteins reconstituted into BLMs can generate detailed information at the single molecule level on protein activity, ligand-binding and kinetics. 1 Classical BLM techniques have been in use for over 40 years, but are not amenable to applications where high-throughput processes and reproducibility are required (e.g. drug screening and biosensing applications). 2 New labon-chip technologies are being developed which allow for ever greater integration of complex functions with precise control of fluids on the micro-scale. 3 This technology can be exploited to combine parallelization, achieved through microfluidics, with the sensitivity and selectivity of BLM approaches. Although various miniaturized devices have been reported, only a few BLM formation methods 4-7 have been developed on-chip that are reproducible, potentially automatable and suitable for singlechannel recording.An alternative technique was recently described by Holden et al.,[8][9][10][11][12] which uses an alkane-lipid solution as the bulk phase, as opposed to an aqueous electrolyte. Droplets of buffer are immersed in the organic solution and at the interface between the two phases lipid monolayers form. When two droplets are brought into contact, a BLM forms at the interface between the droplets. As well as forming single BLMs, this approach allows for the creation of BLM networks. However, the method required manual manipulation of the droplets using micromanipulators.Electrical methods for droplet manipulation exist, based on electrodynamic techniques, such as dielectrophoresis (DEP) and electro-wetting on dielectric (EWOD).

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Controlled delivery of proteins into bilayer lipid membranes on chip

Cited by 64 publications

References 48 publications

Drunken Membranes: Short-Chain Alcohols Alter Fusion of Liposomes to Planar Lipid Bilayers

Drunken Membranes: Short-Chain Alcohols Alter Fusion of Liposomes to Planar Lipid Bilayers

Bioelectronic Light‐Gated Transistors with Biologically Tunable Performance

Formation of artificial lipid bilayers using droplet dielectrophoresis

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