Ca(V)2.2 (N-type) calcium channels are key regulators of neurotransmission. 
Evidence from knockout animals and localization studies suggest that Ca(V)2.2 
channels play a critical role in nociceptive transmission. Additionally, 
ziconotide, a selective peptide inhibitor of Ca(V)2.2 channels, is clinically 
used to treat refractory pain. However, the use of ziconotide is limited by its 
low therapeutic index, which is believed, at least in part, to be a consequence 
of ziconotide inhibiting Ca(V)2.2 channels regardless of the channel state. 
Subsequent efforts have focused on the discovery of state-dependent inhibitors 
that preferentially bind to the inactivated state of Ca(V)2.2 channels in order 
to achieve an improved safety profile relative to ziconotide. Much less 
attention has been paid to understanding the binding kinetics of these 
state-dependent inhibitors. Here, we describe a novel electrophysiology-based 
assay on an automated patch platform designed to differentiate Ca(V)2.2 
inhibitors based on their combined state dependence and kinetics. More 
specifically, this assay assesses inactivated state block, closed state block, 
and monitors the kinetics of recovery from block when channels move between 
states. Additionally, a use-dependent assay is described that uses a train of 
depolarizing pulses to drive channels to a similar level of inactivation for 
comparison. This use-dependent protocol also provides information on the 
kinetics of block development. Data are provided to show how these assays can be 
utilized to screen for kinetic diversity within and across chemical classes.