Are These Cardiomyocytes? Protocol Development Reveals Impact of Sample Preparation on the Accuracy of Identifying Cardiomyocytes by Flow Cytometry

Overview

Journal Stem Cell Reports

Publisher Cell Press

Specialty Cell Biology

Date 2019 Jan 29

PMID 30686762

Citations 12

Authors

Matthew Waas

Ranjuna Weerasekera

Erin M Kropp

Marisol Romero-Tejeda

Ellen N Poon

Kenneth R Boheler

Paul W Burridge

Rebekah L Gundry

Affiliations

Soon will be listed here.

Abstract

Several protocols now support efficient differentiation of human pluripotent stem cells to cardiomyocytes (hPSC-CMs) but these still indicate line-to-line variability. As the number of studies implementing this technology expands, accurate assessment of cell identity is paramount to well-defined studies that can be replicated among laboratories. While flow cytometry is apt for routine assessment, a standardized protocol for assessing cardiomyocyte identity has not yet been established. Therefore, the current study leveraged targeted mass spectrometry to confirm the presence of troponin proteins in day 25 hPSC-CMs and systematically evaluated multiple anti-troponin antibodies and sample preparation protocols for their suitability in assessing cardiomyocyte identity. Results demonstrate challenges to interpreting data generated by published methods and inform the development of a robust protocol for routine assessment of hPSC-CMs. The data, workflow for antibody evaluation, and standardized protocol described here should benefit investigators new to this field and those with expertise in hPSC-CM differentiation.

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