Gigaohm seals, or ‘gigaseals’, are imperative to patch clamp electrophysiology to enable good electrical access to the cell and high-quality recordings. These seals form through chemical bonds and electrostatic forces between the cell membrane and the glass pipette in manual patch clamp, or in the case of planar patch clamp, between the cell membrane and chip substrate1. Planar patch clamp often requires the use of ‘seal enhancers’ to increase the resistances of these seals, with CaF2 being the most extensively used. It is hypothesised that high concentrations of extracellular Ca2+ and intracellular F- give rise to CaF2 precipitate at the solution interface, fostering seal formation2.
CaF2 as a seal enhancer, however, has limitations. F- is known to stimulate G-protein modulation of ion channels, altering channel properties3-6. Furthermore, use of F- is not optimal when recording from Ca2+-activated ion channels due to resultant unknown concentrations of free intracellular Ca2+.
In an effort to overcome these limitations, Metrion and Sophion collaborated to determine whether other insoluble salts can act as seal enhancers, or whether this property is unique to CaF2.
We report the development and optimisation of a TREK-1 functional assay using the Qube 384, an automated patch clamp platform capable of supporting high-throughput screening. The assay was optimized to identify both activators and inhibitors on the same plate, providing key mechanistic data for high value, limited supply screening libraries such as venom fractions used in this study (Targeted Venom Discovery Array, TVDA, Venomtech, UK).
Current clamp recordings provide more physiologically relevant measurements of ion channel activity (in comparison to voltage clamp), allowing the contribution of different ion channel sub-types to resting membrane potential to be evaluated.