Whole Cell Patch Clamp Recording Performed on a Planar Glass Chip

Fertig, Niels; Blick, Robert H.; Behrends, Jan C.

doi:10.1016/s0006-3495(02)75646-4

Cited by 335 publications

(227 citation statements)

References 41 publications

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“…Other nonelastomer devices have been used to perform whole-cell recording of mammalian cells by employing a similar planar design with vertical, lithographically defined pores in silicon and other substrates (4,11,20,21). Although integration of planar patch designs with microfluidics is a stated goal for a variety of groups, it remains a technologically challenging task.…”

Section: Discussionmentioning

confidence: 99%

Mammalian electrophysiology on a microfluidic platform

Ionescu-Zanetti

Shaw

Seo

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

170

131

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The recent development of automated patch clamp technology has increased the throughput of electrophysiology but at the expense of visual access to the cells being studied. To improve visualization and the control of cell position, we have developed a simple alternative patch clamp technique based on microfluidic junctions between a main chamber and lateral recording capillaries, all fabricated by micromolding of polydimethylsiloxane (PDMS). PDMS substrates eliminate the need for vibration isolation and allow direct cell visualization and manipulation using standard microscopy. Microfluidic integration allows recording capillaries to be arrayed 20 m apart, for a total chamber volume of <0.5 nl. The geometry of the recording capillaries permits high-quality, stable, whole-cell seals despite the hydrophobicity of the PDMS surface. Using this device, we are able to demonstrate reliable whole-cell recording of mammalian cells on an inexpensive microfluidic platform. Recordings of activation of the voltage-sensitive potassium channel Kv2.1 in mammalian cells compare well with traditional pipette recordings. The results make possible the integration of whole-cell electrophysiology with easily manufactured microfluidic lab-on-a-chip devices.microfluidics ͉ patch clamp ͉ drug screening ͉ single-cell assay P ipette-based recording of membrane currents is the mainstay in the characterization of cellular ion channels (1). Traditional patch clamp recording is accomplished in a vibration-free environment by using a micromanipulator to position the tip of a glass pipette against the membrane of a cell. Carefully applied negative pressure through the pipette tip causes the membrane to invaginate into the pipette and causes a gigaohm seal to form between the pipette and cell. For whole-cell recording, a second application of negative pressure or electrical current ruptures the captured membrane, providing continuity between the pipette electrode and the cell's cytoplasm. In voltage clamp mode, the membrane is clamped to a preset potential, and the current required to maintain this potential is recorded. Current recordings with different electrical protocols and in the presence of different reagents are used to characterize ion channel properties.Despite the success of the traditional patch clamp technique, it requires complex and expensive setups and remains highly laborious. Thus, the patch clamp technique is limiting in proteomics and drug development screens, which demand highthroughput automated measurements (2). To address this need, chip-based patch clamp devices using micromachined substrates from glass (3), quartz (4), coated silicon nitride (5), and treated elastomers (6) have been proposed. These devices are being developed into commercial platforms for pharmaceutical drug screening and drug safety testing (7-9). All chip-based devices developed to date use a planar geometry, where the recording pore is etched in a horizontal membrane dividing the top cell compartment from the bottom recording electrode compartment (1...

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Section: Discussionmentioning

confidence: 99%

Mammalian electrophysiology on a microfluidic platform

Ionescu-Zanetti

Shaw

Seo

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

170

131

View full text Add to dashboard Cite

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“…Unfortunately, silicon substrates are opaque, precluding microscopy observation. The first report of a gigaohm-seal recording on small mammalian cells is credited to Behrends' group, 191 who created geometrically defined submicron apertures in glass substrates by ion (highly accelerated gold) track etching and were able to record from CHO cells with typical seal resistance of 1-10 GΩ. These devices also allow for recordings from ion channels on artificial lipid bilayers 192 or single membrane patches 193 and from single ion channels.…”

Section: Extracellular Electrode Arrays For Multisite Recording and Smentioning

confidence: 99%

Biology on a Chip: Microfabrication for Studying the Behavior of Cultured Cells

Tourovskaia

Folch

2003

Crit Rev Biomed Eng

173

140

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The ability to culture cells in vitro has revolutionized hypothesis testing in basic cell and molecular biology research and has become a standard methodology in drug screening and toxicology assays. However, the traditional cell culture methodology-consisting essentially of the immersion of a large population of cells in a homogeneous fluid medium-has become increasingly limiting, both from a fundamental point of view (cells in vivo are surrounded by complex spatiotemporal microenvironments) and from a practical perspective (scaling up the number of fluid handling steps and cell manipulations for high-throughput studies in vitro is prohibitively expensive). Micro fabrication technologies have enabled researchers to design, with micrometer control, the biochemical composition and topology of the substrate, the medium composition, as well as the type of neighboring cells surrounding the microenvironment of the cell. In addition, microtechnology is conceptually well suited for the development of fast, low-cost in vitro systems that allow for high-throughput culturing and analysis of cells under large numbers of conditions. Here we review a variety of applications of microfabrication in cell culture studies, with an emphasis on the biology of various cell types.

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“…The challenge in fabricating planar patch-clamp sample supports for biological applications is the tight, giga ohm seal needed between the biological membrane and the support [7][8][9]. In most planar supports the sample is only deposited onto the surface, compared to patch clamping, where the membrane is partially sucked into the pipette and hence, conformably covers a large surface.…”

Section: Design and Fabricationmentioning

confidence: 99%