Enhanced Control for Liquid Displacement Facilitates Assays for Positive Modulation
Ali Yehia, Haiyang Wei, Michael Donio
Fluxion Biosciences, South San Francisco, CA, USA,
Eurofins Pharma Bioanalytics Services US Inc., St Charles, MO, USA
Ion channel modulation is an important target for drug discovery. Common compounds for ion channel activity are positive allosteric modulators (PAMs), which by binding to the protein allosteric binding site can effectively enhance channel activity while in the presence of the agonist. The activity of the modulator is considered to be in direct relationship with its concentration, provided a standardized agonist baseline effect can be had with a constant concentration of ligand. With the ability to record electrical signals across membranes of multiple cells in parallel sites, automated patch clamp systems can record the activity of a channel’s agonist or antagonist in real time. However, due to the dual-channel binding and the need for repeatability of ligand response, screening for new PAMs is a delicate process, sometimes requiring quick additions of multiple compounds at the same recording site to gauge a proper comparison. Here we present data from a number of different nicotinic ion channels stably expressed in commercially-available cell lines. Using a microfluidic-based automated patch clamp system capable of fast and flexible serial displacement of agonists, modulators and antagonists, we qualify procedures that can ultimately simplify the final process of screening for PAMs as well as provide extended flexibility in ligand-gated channel studies. We present data from a variety of nicotinic receptors (α1β1δε, α3β4, α4β2, α4α6β2, and α7) showing current stability, plate to plate reproducibility, agonists, antagonists, open channel block, PAM effects, and recovery from desensitization.
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