Abstract
Tumorigenesis and attendant safety risks are significant concerns of induced pluripotent stem cell (iPSC)-based therapies. Thus, it is crucial to evaluate iPSC proliferation, differentiation, and tumor formation after transplantation. Several approaches have been employed for tracking the donor cells, including fluorescent protein and luciferase, but both have limitations. Here, we introduce a protocol using iRFP genetic labeling technology to track tumor formation of iPSCs in skeletal muscle after CRISPR/Cas9 gene editing.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Mendell JR, Shilling C, Leslie ND et al (2012) Evidence-based path to newborn screening for Duchenne muscular dystrophy. Ann Neurol 71:304–313
Trounson A, Mcdonald C (2015) Stem cell therapies in clinical trials: progress and challenges. Cell Stem Cell 17:11–22
Cunningham JJ, Ulbright TM, Pera MF et al (2012) Lessons from human teratomas to guide development of safe stem cell therapies. Nat Biotechnol 30:849–857
Chudakov DM, Matz MV, Lukyanov S et al (2010) Fluorescent proteins and their applications in imaging living cells and tissues. Physiol Rev 90:1103–1163
Hong H, Yang Y, Cai W (2011) Imaging gene expression in live cells and tissues. Cold Spring Harb Protoc 2011:pdb top103
Lyons SK, Patrick PS, Brindle KM (2013) Imaging mouse cancer models in vivo using reporter transgenes. Cold Spring Harb Protoc 2013:685–699
Lecoq J, Schnitzer MJ (2011) An infrared fluorescent protein for deeper imaging. Nat Biotechnol 29:715–716
Auld DS, Inglese J (2004) Interferences with luciferase reporter enzymes. In: Sittampalam GS, Coussens NP, Brimacombe K, Grossman A, Arkin M, Auld D, Austin C, Baell J, Bejcek B, JMM C, TDY C, Dahlin JL, Devanaryan V, Foley TL, Glicksman M, Hall MD, Haas JV, Inglese J, Iversen PW, Kahl SD, Kales SC, Lal-Nag M, Li Z, McGee J, McManus O, Riss T, Trask OJ Jr, Weidner JR, Wildey MJ, Xia M, Xu X (eds) Assay Guidance Manual. Eli Lilly & Company and the National Center for Advancing Translational Sciences, Bethesda, MD
Wang Y, Zhou M, Wang X et al (2014) Assessing in vitro stem-cell function and tracking engraftment of stem cells in ischaemic hearts by using novel iRFP gene labelling. J Cell Mol Med 18:1889–1894
Filonov GS, Piatkevich KD, Ting LM et al (2011) Bright and stable near-infrared fluorescent protein for in vivo imaging. Nat Biotechnol 29:757–761
Hock AK, Lee P, Maddocks OD et al (2014) iRFP is a sensitive marker for cell number and tumor growth in high-throughput systems. Cell Cycle 13:220–226
Acknowledgements
Funding: N. Weintraub and Y. Tang were partially supported by the American Heart Association: GRNT31430008, NIH-AR070029, NIH-HL086555, and NIH-HL134354.
Conflict of Interest: All authors declare that he/she has no conflict of interest.
This article does not contain any studies with human participants performed by any of the authors.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2020 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Jin, Y., Shen, Y., Weintraub, N.L., Tang, Y. (2020). Using iRFP Genetic Labeling Technology to Track Tumorogenesis of Transplanted CRISPR/Cas9-Edited iPSC in Skeletal Muscle. In: Basel, M., Bossmann, S. (eds) Cell Tracking. Methods in Molecular Biology, vol 2126. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0364-2_7
Download citation
DOI: https://doi.org/10.1007/978-1-0716-0364-2_7
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-0363-5
Online ISBN: 978-1-0716-0364-2
eBook Packages: Springer Protocols