A multiplexed epitope barcoding strategy that enables dynamic cellular phenotypic screens

Takamasa Kudo, Keara Lane, Markus W. Covert*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

4 Scopus citations


Pooled genetic libraries have improved screening throughput for mapping genotypes to phenotypes. However, selectable phenotypes are limited, restricting screening to outcomes with a low spatiotemporal resolution. Here, we integrated live-cell imaging with pooled library-based screening. To enable intracellular multiplexing, we developed a method called EPICode that uses a combination of short epitopes, which can also appear in various subcellular locations. EPICode thus enables the use of live-cell microscopy to characterize a phenotype of interest over time, including after sequential stimulatory/inhibitory manipulations, and directly connects behavior to the cellular genotype. To test EPICode's capacity against an important milestone—engineering and optimizing dynamic, live-cell reporters—we developed a live-cell PKA kinase translocation reporter with improved sensitivity and specificity. The use of epitopes as fluorescent barcodes introduces a scalable strategy for high-throughput screening broadly applicable to protein engineering and drug discovery settings where image-based phenotyping is desired.

Original languageEnglish (US)
Pages (from-to)376-387.e8
JournalCell Systems
Issue number5
StatePublished - May 18 2022


  • epitope barcodes
  • fluorescent reporters
  • in situ genotyping
  • pooled screen
  • spatial multiplexing

ASJC Scopus subject areas

  • Pathology and Forensic Medicine
  • Cell Biology
  • Histology


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