All-Optical Miniaturized Co-culture Assay of Voltage-Gated Ca2+ Channels

Viviana Agus, Harald Janovjak

Research output: Chapter in Book/Report/Conference proceedingChapter (Book)Otherpeer-review

Abstract

Light-activated proteins enable the reversible and spatiotemporal control of cellular events in optogenetics. Optogenetics is also rapidly expanding into the field of drug discovery where it provides cost-effective and noninvasive approaches for cell manipulation in high-throughput screens. Here, we present a prototypical cell-based assay that applies Channelrhodopsin2 (ChR2) to recapitulate physiological membrane potential changes and test for voltage-gated ion channel (VGIC) blockade. ChR2 and the voltage-gated Ca2+ channel 1.2 (CaV1.2) are expressed in individual HEK293 cell lines that are then co-cultured for formation of gap junctions and an electrical syncytium. This co-culture allows identification of blockers using parallel fluorescence plate readers in the 384-well plate format in an all-optical mode of operation. The assay is transferable to other VGICs by modularly combining new and existing cell lines and potentially also to other drug targets.

Original languageEnglish
Title of host publicationPhotoswitching Proteins
Subtitle of host publicationMethods and Protocols
EditorsDominik Niopek
Place of PublicationNew York, NY
PublisherHumana Press
Chapter17
Pages247-260
Number of pages14
ISBN (Electronic)9781071607558
ISBN (Print)9781071607541
DOIs
Publication statusPublished - 2020

Publication series

NameMethods in Molecular Biology
Volume2173
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Keywords

  • 384-well plate
  • All-optical
  • Ca1.2
  • Channelrhodopsin
  • FLIPR
  • High-throughput screening
  • Optogenetics
  • Syncytium
  • Voltage-gated ion channel

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