Protocol to measure calcium spikes in cardiomyocytes obtained from human pluripotent stem cells using a ready-to-use
Veronica Astro1, Gustavo Ramirez-Calderon1, Antonio Adamo1
1Biological and Environmental Science and Engineering Division, King Abdullah University of Science and Technology, Thuwal 23955-6900, Saudi Arabia.
Insights
This study optimized a protocol for deriving cardiomyocytes from human pluripotent stem cells (hPSCs). The method yields contracting cells for studying heart development and disease modeling in vitro.
Area of Science:
- Stem Cell Biology
- Cardiovascular Research
- Developmental Biology
Background:
- Human pluripotent stem cells (hPSCs) are crucial for studying early cardiogenesis and in vitro disease modeling.
- Optimized protocols are needed to reliably derive functional cardiomyocytes from hPSCs.
Purpose of the Study:
- To present an optimized, ready-to-use protocol for generating contracting cardiomyocytes from hPSCs.
- To detail key parameters and validation methods for hPSC-derived cardiomyocyte differentiation.
Main Methods:
- hPSC culture and cardiomyocyte differentiation using a commercial kit.
- Optimization of critical parameters: starting cell confluency and temperature.
- Characterization via immunofluorescence and flow cytometry.
- Quantification of cardiomyocyte function using live imaging of calcium spikes.
Main Results:
- Successful derivation of contracting cardiomyocytes from hPSCs using the optimized protocol.
- Identification of critical parameters influencing differentiation efficiency.
- Demonstration of robust functional assessment through calcium imaging.
Conclusions:
- The presented protocol provides an efficient method for obtaining hPSC-derived cardiomyocytes.
- This optimized protocol facilitates robust in vitro investigation of cardiogenesis and cardiac diseases.
- The method enables reliable functional characterization of differentiated cardiomyocytes.
Abstract:
The derivation of cardiomyocytes from human pluripotent stem cells (hPSCs) is a powerful tool to investigate early cardiogenesis and model diseases in vitro. Here, we present an optimized protocol to obtain contracting hPSCs-derived cardiomyocytes using a ready-to-use kit. We describe steps for hPSC culture and differentiation to cardiomyocytes including the identification of key parameters such as starting cell confluency and temperature. We then detail immunofluorescence, flow cytometry, and the quantification of cardiomyocytes' calcium spikes using live imaging. For complete details on the use and execution of this protocol, please refer to Astro et al.1.
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