Testing the Efficiency and Kinetics of Negative Selection Using Thymic Slices

Tyng An Zhou, Chia Lin Hsu, Ivan Lilyanov Dzhagalov*

*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

4 Scopus citations


Central tolerance is an efficient barrier to autoimmunity and negative selection of self-reactive thymocytes is one of its major manifestations. Because of its importance, negative selection has been studied extensively through numerous in vitro and in vivo approaches that have tremendously increased our understanding of the process. Recently, in situ experimental systems using thymus slices have been developed that combine some of the advantages of in vitro assays such as ease of manipulation and high throughput with the existence of three dimensional mature thymus microenvironment. These approaches offer unprecedented opportunity to study negative selection. Here, we describe how thymic slices can be used to measure the kinetics and magnitude of negative selection. Taking the OT-1/Ova system as an example, we provide detailed guidance on cutting thymic slices, labeling and overlaying thymocytes on them and reading out the extent of negative selection by flow cytometry. The system can easily be adapted to evaluate the effects of various mutations or treatments on negative selection or to study the behavior of different cells in the thymus through time-lapse imaging.

Original languageEnglish
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Number of pages15
StatePublished - 2020

Publication series

NameMethods in Molecular Biology
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029


  • Central tolerance
  • Flow cytometry
  • Negative selection
  • OT-1
  • Ova
  • Thymic slices
  • Thymocytes
  • Thymus
  • Vibratome


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