Automated patch clamp (APC) instruments enable efficient evaluation of electrophysiologic effects of drugs on human cardiac currents in heterologous expression systems. Differences in experimental protocols, instruments, and dissimilar site procedures affect the variability of IC 50 values characterizing drug block potency. this impacts the utility of Apc platforms for assessing a drug's cardiac safety margin. We determined variability of Apc data from multiple sites that measured blocking potency of 12 blinded drugs (with different levels of proarrhythmic risk) against four human cardiac currents (heRG [i Kr ], hCav1.2 [L-Type I ca ], peak hNav1.5, [Peak I na ], late hNav1.5 [Late I na ]) with recommended protocols (to minimize variance) using five APC platforms across 17 sites. IC 50 variability (25/75 percentiles) differed for drugs and currents (e.g., 10.4-fold for dofetilide block of hERG current and 4-fold for mexiletine block of hNav1.5 current). Within-platform variance predominated for 4 of 12 hERG blocking drugs and 4 of 6 hNav1.5 blocking drugs. hERG and hNav1.5 block. Bland-Altman plots depicted varying agreement across Apc platforms. A follow-up survey suggested multiple sources of experimental variability that could be further minimized by stricter adherence to standard protocols. Adoption of best practices would ensure less variable Apc datasets and improved safety margins and proarrhythmic risk assessments. The characterization of drug effects on human currents using automated patch clamp (APC) platforms employing single-or multi-hole planar patch techniques 1-5 has revolutionized the assessment of electrophysiologic (and potential proarrhythmic) effects of new drug candidates. This increasingly popular experimental approach

We have developed an automated patch clamp module for high-throughput ion channel screening, recording from 384 cells simultaneously. The module is incorporated into a laboratory pipetting robot and uses a 384-channel pipettor head for application of cells and compounds. The module contains 384 amplifier channels for fully parallel recordings using a digital amplifier. Success rates for completed experiments (1- to 4-point concentration-response curves for cells satisfying defined quality control parameters) of greater than 85% have been routinely achieved with, for example, HEK, CHO, and RBL cell lines expressing hNa1.7, hERG, Kir2.1, GABA, or glutamate receptors. Pharmacology experiments are recorded and analyzed using specialized software, and the pharmacology of hNa1.7 and hERG is described. Fast external solution exchange rates of <50 ms are demonstrated using Kir2.1. Short exposure times are achieved by stacking the external solutions inside the pipette of the robot to minimize exposure of the ligand on the receptor. This ensures that ligand-gated ion channels, for example, GABA and glutamate described in this report, can be reproducibly recorded. Stem cell-derived cardiomyocytes have also been used with success rates of 52% for cells that have a seal resistance of >200 MΩ, and recordings of voltage-gated Na and Ca are shown.

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BackgroundAn in vitro electrophysiological assay system, which can assess compound effects and thus show cardiotoxicity including arrhythmia risks of test drugs, is an essential method in the field of drug development and toxicology.MethodsIn this study, high-throughput electrophysiological recordings of human embryonic kidney (HEK 293) cells and Chinese hamster ovary (CHO) cells stably expressing human ether-a-go-go related gene (hERG) were performed utilizing an automated 384-well-patch-clamp system, which records up to 384 cells simultaneously. hERG channel inhibition, which is closely related to a drug-induced QT prolongation and is increasing the risk of sudden cardiac death, was investigated in the high-throughput screening patch-clamp system.ResultsIn the automated patch-clamp measurements performed here, Kv currents were investigated with high efficiency. Various hERG channel blockers showed concentration-dependent inhibition, the 50 % inhibitory concentrations (IC50) of those blockers were in good agreement with previous reports.ConclusionsThe high-throughput patch-clamp system has a high potential in the field of pharmacology, toxicology, and cardiac physiology, and will contribute to the acceleration of pharmaceutical drug development and drug safety testing.