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Fetal Thymic Organ Culture (FTOC) Optimized for Gamma-Delta T Cell Studies

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Immune Receptors

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2421))

Abstract

Fetal thymic organ culture (FTOC) provides a method for analyzing T cell development in a physiological context outside the animal. This technique enables studies of genetically altered mice that are embryonic or neonatal lethal, in addition to bypassing the complication of migration of successive waves of T cells out of the thymus. The hanging drop method involves depletion of thymocytes from host lobes using deoxyguanosine, followed by reconstitution with hematopoietic progenitors. This method has become standard for analysis of fetal liver precursors, bone marrow precursors, and early thymocytes. However, difficulties are encountered in the analysis of γδ T cell precursors using this method. We have developed a modification of FTOC in which partial depletion of hematopoietic precursors by shortened deoxyguanosine treatment, coupled with the use of TCRδ-deficient host lobes, enables engraftment and development of fetal γδTCR+ thymocytes. This method allows comparisons of development and functional differentiation of γδ T cell precursors between cells of different genotypes or treatments, in the context of a permissive thymic microenvironment.

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Acknowledgments

This work was supported by grants CIHR 201610PJT, NIH 1P01AI102853-06, and RGPIN 05333-14 to MKA, and JS was supported by an American Association of Immunologist Careers in Immunology award and a CIHR Postdoctoral Fellowship. TSHI was supported by Ontario Graduate Fellowships.

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Authors and Affiliations

  1. Biological Sciences, Department of Immunology, Sunnybrook Research Institute, University of Toronto, Toronto, ON, Canada

    Johanna S. Selvaratnam, Tracy S. H. In & Michele K. Anderson

Authors
  1. Johanna S. Selvaratnam
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  2. Tracy S. H. In
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  3. Michele K. Anderson
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Correspondence to Michele K. Anderson .

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Editors and Affiliations

  1. Pathology and Laboratory Medicine, Emory University School of Medicine and Emory Vaccine Center, Atlanta, GA, USA

    Jonathan Rast

  2. Department of Biological Sciences, Auburn University, Auburn, AL, USA

    Katherine Buckley

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Selvaratnam, J.S., In, T.S.H., Anderson, M.K. (2022). Fetal Thymic Organ Culture (FTOC) Optimized for Gamma-Delta T Cell Studies. In: Rast, J., Buckley, K. (eds) Immune Receptors. Methods in Molecular Biology, vol 2421. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1944-5_17

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  • DOI: https://doi.org/10.1007/978-1-0716-1944-5_17

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1943-8

  • Online ISBN: 978-1-0716-1944-5

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