Multielectrode Array (MEA) Assay for Profiling Electrophysiological Drug Effects in Human Stem Cell‐Derived Cardiomyocytes

Mike Clements1

1 Forest Farm, Whitchurch, Cardiff
Publication Name:  Current Protocols in Toxicology
Unit Number:  Unit 22.4
DOI:  10.1002/cptx.2
Online Posting Date:  May, 2016
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More relevant and reliable preclinical cardiotoxicity tests are required to improve drug safety and reduce the cost of drug development. Human stem cell‐derived cardiomyocytes (hSC‐CMs) provide a potential model for the development of superior assays for preclinical drug safety screening. One such hSC‐CM assay that has shown significant potential for enabling more predictive drug cardiac risk assessment is the MEA assay. The Multi‐electrode Array (MEA) assay is an electrophysiology‐based technique that uses microelectrodes embedded in the culture surface of each well to measure fluctuations in extracellular field potential (FP) generated from spontaneously beating hSC‐CMs. Perturbations to the recorded FP waveform can be used as an unbiased method of predicting the identity of ion channel(s) impacted on drug exposure. Here, a higher throughput MEA assay using hSC‐CMs in 48‐well MEA plates is described for profiling compound‐induced effects on cardiomyocyte electrophysiology. Techniques for preparing hSC‐CM monolayers in MEA plates and methods to contextualize MEA assay experimental results are also covered. © 2016 by John Wiley & Sons, Inc.

Keywords: cardiomyocyte; cardiotoxicity; multielectrode array; preclinical; proarrhythmia; stem cell

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Table of Contents

  • Introduction
  • Significance Statement
  • Basic Protocol 1: Higher‐Throughput Extracellular Field Potential Recordings from Spontaneously Beating hSC‐CM Monolayers
  • Support Protocol 1: Preparation of Cytiva Plus hSC‐CM Monolayers in a 48‐Well MEA Plate
  • Support Protocol 2: Predicting Risk Factor Based on Perturbation of FPDcF (Secondary Analysis)
  • Support Protocol 3: Predicting MOA From Multi‐Parameter Signal Analysis of Drug‐Induced Effects (Secondary Analysis)
  • Reagents and Solutions
  • Commentary
  • Literature Cited
  • Figures
  • Tables
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Basic Protocol 1: Higher‐Throughput Extracellular Field Potential Recordings from Spontaneously Beating hSC‐CM Monolayers

  • Positive control compounds:
  • Mexiletine (Tocris, cat. no. 2596); store at 2° to 8°C
  • Nifedipine (Sigma, cat. no. N7634‐1 G); store at 2° to 8°C
  • E‐4031 (Tocris, cat. no. 1808); store at 2° to 8°C
  • Dimethyl sulfoxide (DMSO; Sigma, cat. no. D2650); store at room temperature
  • RPMI 1640/B27 medium (see recipe)
  • Sterile distilled H 2O, 500 ml (Fresenius Kabi, cat. no. 22‐96‐985)
  • Cytiva Plus hSC‐CM monolayers plated in a 48‐well MEA plate, on day 5 post‐thaw (see protocol 2);store in a standard cell culture incubator at 37°C, 5% CO 2
  • Axion Integrated Studio (AxIS) software V2.0 or higher (Axion BioSystems)
  • Maestro MEA system (Axion BioSystem)

Support Protocol 1: Preparation of Cytiva Plus hSC‐CM Monolayers in a 48‐Well MEA Plate

  • 48‐well MEA plate (Axion BioSystems, M768‐KAP‐48, opaque; or M768‐TMEA‐48, transparent base)
  • 70% isopropanol
  • Fetal bovine serum (FBS), 500 ml (GIBCO/Life Technologies, cat. no. 26140‐079)
  • Sterile distilled H 2O, 500 ml (Fresenius Kabi, cat. no. 22‐96‐985)
  • Fibronectin solution (12.5 μg/ml; see recipe)
  • Cytiva Plus cardiomyocytes, 3.5 million viable cells/cryovial (GE Healthcare, cat. no. 29‐0918‐81); store in vapor phase of a liquid nitrogen storage unit at −140°C
  • Dry ice
  • RPMI 1640/B27 medium (see recipe)
  • Reagent A100, 200 μl (ChemoMetec, cat. no. 910‐0003)
  • Reagent B, 200 μl (ChemoMetec, cat. no. 910‐0002)
  • Laboratory tissues (e.g., Kimwipes)
  • Biological tissue culture hood
  • 37°C, 5% CO 2 cell culture incubator
  • 37°C water bath
  • 200‐ and 1000‐μl pipets
  • Centrifuge tubes, 50 ml (Corning, cat. no. 430290)
  • Hemacytometer or automated cell counter (e.g., NucleoCounter NC‐100 ChemoMetec)
  • 1.5‐ml tubes
  • 60‐ml bottles

Support Protocol 2: Predicting Risk Factor Based on Perturbation of FPDcF (Secondary Analysis)

  • Compound(s)
  • Data analysis and graphing software (e.g., Microsoft Excel)

Support Protocol 3: Predicting MOA From Multi‐Parameter Signal Analysis of Drug‐Induced Effects (Secondary Analysis)

  • Test compounds
  • Hierarchical clustering data analysis software (e.g., TIBCO Spotfire DecisionSite V9.1.1)
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Literature Cited

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