Abstract
The thymus organ is composed of a three-dimensional (3D) network of adjoining epithelium and stromal cells. Bone marrow-derived T cell precursors, upon entering the thymus, interact with and migrate through this cellular network as they differentiate and mature. An essential component of the stroma is the thymic epithelial cells (TEC), which play a vital role in T cell development and induction of self-tolerance for adaptive immunity. TEC can be isolated from the embryonic and adult thymus by a series of gentle enzymatic digestions and characterized into discrete subpopulations based on their expression of surface markers by flow cytometry. Enrichment of adult TEC can be achieved by depletion of hematopoietic cells, allowing sufficient numbers to be purified for subsequent functional and molecular analysis. Although monolayer cultures have been used to study TEC phenotype and T cell interaction, methods that mimic the 3D thymic microenvironment, such as fetal and reaggregate thymic organ cultures, are more accurate for the analysis of TEC function and support more complete T cell development. Herein, we describe methods for the efficient isolation and enrichment of TEC for downstream analyses as well as the reaggregation of embryonic progenitor epithelium to form a functional thymus graft under the kidney capsule.
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Acknowledgments
The authors would like to thank Jade Homann for her expertise in kidney grafting and Joanna Lim for the embryonic photos, Professor Richard Boyd for general support, and the National Health and Medical Research Council and the Australian Stem Cell Centre for funding.
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Seach, N., Hammett, M., Chidgey, A. (2012). Isolation, Characterization, and Reaggregate Culture of Thymic Epithelial Cells. In: Randell, S., Fulcher, M. (eds) Epithelial Cell Culture Protocols. Methods in Molecular Biology, vol 945. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-125-7_15
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DOI: https://doi.org/10.1007/978-1-62703-125-7_15
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