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Medientyp:
E-Artikel
Titel:
Transfection of hPSC-Cardiomyocytes Using Viafect™ Transfection Reagent
Beteiligte:
Bodbin, Sara E.;
Denning, Chris;
Mosqueira, Diogo
Erschienen:
MDPI AG, 2020
Erschienen in:Methods and Protocols
Sprache:
Englisch
DOI:
10.3390/mps3030057
ISSN:
2409-9279
Entstehung:
Anmerkungen:
Beschreibung:
<jats:p>Twenty years since their first derivation, human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) have shown promise in disease modelling research, while their potential for cardiac repair is being investigated. However, low transfection efficiency is a barrier to wider realisation of the potential this model system has to offer. We endeavoured to produce a protocol for improved transfection of hPSC-CMs using the ViafectTM reagent by Promega. Through optimisation of four essential parameters: (i) serum supplementation, (ii) time between replating and transfection, (iii) reagent to DNA ratio and (iv) cell density, we were able to successfully transfect hPSC-CMs to ~95% efficiencies. Transfected hPSC-CMs retained high purity and structural integrity despite a mild reduction in viability, and preserved compatibility with phenotyping assays of hypertrophy. This protocol greatly adds value to the field by overcoming limited transfection efficiencies of hPSC-CMs in a simple and quick approach that ensures sustained expression of transfected genes for at least 14 days, opening new opportunities in mechanistic discovery for cardiac-related diseases.</jats:p>