Reliable Protocols for Flow Cytometry Analysis of Intracellular Proteins in Pluripotent Stem Cell Derivatives: A Fit-For-Purpose Approach

Linda Berg Luecke, Matthew Waas, Rebekah L. Gundry

Research output: Contribution to journalArticle

Abstract

Human pluripotent stem cell (hPSC) derivatives are valuable for a variety of research applications and have the potential to revolutionize approaches to personalized medicine. However, differentiation efficiency varies among cell lines and protocols. Therefore, methods to reliably determine cell type identity in cultures of hPSC derivatives in a manner that is consistent among laboratories are needed. While flow cytometry is apt for routine assessment of population heterogeneity, standardized protocols are not available for most cell types. This article describes a workflow for establishing a fit-for-purpose protocol for flow cytometric analysis of hPSC derivatives. Based on the application of this workflow, a standard operating procedure (SOP) was developed for the analysis of cardiac troponin in hPSC-derived cardiomyocytes (hPSC-CM). Throughout the article, important concepts related to antibody validation and gating strategies are presented to enable users to properly validate any antibody of interest and develop a rigorous SOP for their experimental needs.

Original languageEnglish (US)
Article numbere94
JournalCurrent Protocols in Stem Cell Biology
Volume50
Issue number1
DOIs
StatePublished - Jan 1 2019

Fingerprint

Pluripotent Stem Cells
Flow Cytometry
Workflow
Proteins
Precision Medicine
Troponin
Antibodies
Population Characteristics
Cardiac Myocytes
Cell Line
Research

Keywords

  • flow cytometry
  • hPSC-CM
  • heterogeneity
  • intracellular proteins
  • quality control

ASJC Scopus subject areas

  • Developmental Biology
  • Cell Biology

Cite this

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