Hourglass SiO2 coating increases the performance of planar patch-clamp.
Obtaining high-throughput electrophysiological recordings is an ongoing challenge in ion channel biophysics and drug discovery. One particular area of development is the replacement of glass pipettes with planar devices in order to increase throughput. However, successful patch-clamp recordings depend on a surface coating which ideally should promote and stabilize giga-seal formation. Here, we present data supporting the use of a structured SiO(2) coating to improve the ability of cells to form a "seal" with a planar patch-clamp substrate. The method is based on a correlation study taking into account structure and size of the pores, surface roughness and chip capacitance. The influence of these parameters on the quality of the seal was assessed. Plasma-enhanced chemical vapour deposition (PECVD) of SiO(2) led to an hourglass structure of the pore and a tighter seal than that offered by a flat, thermal SiO(2) surface. The performance of PECVD chips was validated by recording recombinant potassium channels, BK(Ca), expressed in stable HEK-293 cell lines and in inducible CHO cell lines and low conductance IRK1, and endogenous cationic currents from CHO cells. This multiparametric investigation led to the production of improved chips for planar patch-clamp applications which allow electrophysiological recordings from a wide range of cell lines.
Sordel
Thomas
T
Laboratoire Biopuces, CEA, 17 Rue des Martyrs, 38054 Grenoble Cedex 9, France.
Garnier-Raveaud
Stéphanie
S
Sauter
Fabien
F
Pudda
Catherine
C
Marcel
Frédérique
F
De Waard
Michel
M
Arnoult
Christophe
C
Vivaudou
Michel
M
Chatelain
François
F
Picollet-D'hahan
Nathalie
N
D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D006801:Humans; D007473:Ion Channels / Q000235:genetics / Q000502:physiology*; D008564:Membrane Potentials / Q000502:physiology; D018625:Microscopy, Atomic Force; D008855:Microscopy, Electron, Scanning; D018408:Patch-Clamp Techniques / Q000295:instrumentation / Q000379:methods*; D010957:Plasmids / Q000235:genetics; D015203:Reproducibility of Results; D012822:Silicon Dioxide / Q000737:chemistry*; D014162:Transfection
D007473:Ion Channels; D012822:Silicon Dioxide