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.
Original language | English (US) |
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Pages (from-to) | 395-410 |
Number of pages | 16 |
Journal | Stem cell reports |
Volume | 12 |
Issue number | 2 |
DOIs | |
State | Published - Feb 12 2019 |
Keywords
- cardiomyocytes
- flow cytometry
- mass spectrometry
- quality control
- standard operating protocol
- troponin
ASJC Scopus subject areas
- Genetics
- Biochemistry
- Cell Biology
- Developmental Biology