Sub-millisecond ligand probing of cell receptors with multiple solution exchange.

ABSTRACT: The accurate knowledge of receptor kinetics is crucial to our understanding of cell signal transduction in general and neural function in particular. The classical technique of probing membrane receptors on a millisecond scale involves placing a recording micropipette with a membrane patch in front of a double-barrel (?-glass) application pipette mounted on a piezo actuator. Driven by electric pulses, the actuator can rapidly shift the ?-glass pipette tip, thus exposing the target receptors to alternating ligand solutions. However, membrane patches survive for only a few minutes, thus normally restricting such experiments to a single-application protocol. In order to overcome this deficiency, we have introduced pressurized supply microcircuits in the ?-glass channels, thus enabling repeated replacement of application solutions within 10-15 s. This protocol, which has been validated in our recent studies and takes 20-60 min to implement, allows the characterization of ligand-receptor interactions with high sensitivity, thereby also enabling a powerful paired-sample statistical design.

SUBMITTER: Sylantyev S 

PROVIDER: S-EPMC3743020 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature