UNIT 3.18 In Vitro Systems for the Study of T Cell Development: Fetal Thymus Organ Culture and OP9-DL1 Cell Coculture
Published Online: 1 MAR 2006
Copyright © 2006 by John Wiley & Sons, Inc.
Lab Protocol Title
Current Protocols in Immunology
How to Cite
Ramsdell, F., Zúñiga-Pflücker, J. C. and Takahama, Y. 2006. In Vitro Systems for the Study of T Cell Development: Fetal Thymus Organ Culture and OP9-DL1 Cell Coculture. Current Protocols in Immunology. 71:3.18.1–3.18.18.
- Published Online: 1 MAR 2006
- Published Print: FEB 2006
Most T cell development occurs within the thymus and includes a series of selection processes that are, in large part, still poorly understood. Studies of T cell development have been greatly advanced by the description of multiple phenotypic subsets of T cells and their maturational relationships. This unit describes a system for observing and modulating T cell development in vitro via the culture of entire mouse fetal thymic lobes. Methods are included for the isolation of fetal thymi and culture to allow for normal T cell development on either transwell plates or Gelfoam sponges. A method for depleting hematopoietic cells from thymic lobes using 2′-deoxyguanosine and subsequent reconstitution with precursor cells is also described. This protocol is valuable for the study of tolerance and T cell selection. A support protocol describing methods of altering and monitoring T cell development are outlined. In addition, methods for culturing fetal thymic lobes under high oxygen submersion conditions and for the preparation of reaggregate thymus organ cultures are provided. Finally, a simple and practical method that allows for the thymus-independent generation of T cells from defined sources of stem/progenitor cells by OP9-DL1 coculture is described.