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Analysis of DC Functions Using CD205-DTR Knock-In Mice

  • Tomohiro Fukaya
  • Hideaki Takagi
  • Tomofumi Uto
  • Keiichi Arimura
  • Katsuaki SatoEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1423)

Abstract

Dendritic cells (DCs) are essential antigen-presenting cells (APCs) that consist of heterogeneous subsets, mainly classified as conventional DCs (cDCs) and plasmacytoid DCs (pDCs). CD205, an endocytic type I C-type lectin-like molecule that belongs to the mannose receptor family, is mainly expressed on CD8α+ cDCs. However, it is unclear how CD205+ cDCs control immune responses in vivo. To evaluate the contribution of CD205+ cDCs to the immune system, we engineered knock-in (KI) mice that express the diphtheria toxin receptor (DTR) under the control of the Cd205 gene, which allows the selective conditional ablation of CD205+ cDCs in vivo. Conditional ablation of CD205+ cDCs impaired the antigen-specific priming of CD8+ T cells to generate cytotoxic T lymphocytes (CTLs) mediated through cross presentation of soluble antigen. Upon microbial infection, CD205+ cDCs contributed to the cross priming of CD8+ T cells for generating antibacterial CTLs to efficiently eliminate pathogens. Here, we provide a protocol for the generation of bone marrow WT/CD205-DT chimeric mice, depletion of CD205+ DCs and assessment of depletion efficiency, and protocols for in vivo cross presentation assay, CTL generation assay, and antibacterial immunity assay.

Key words

Dendritic cells Cytotoxic T lymphocytes Cross priming Microbial infection Knock-in mice 

Notes

Acknowledgments

The author would like to thank Yumiko Sato for excellent technical assistance. This work was supported by Grants-in-Aid for Scientific Research from the Ministry of Education, Science and Culture of Japan (B) 25293117 (K.S.) and Japan Science and Technology Agency, Precursory Research for Embryonic Science and Technology (PRESTO) (K.S).

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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Tomohiro Fukaya
    • 1
  • Hideaki Takagi
    • 1
  • Tomofumi Uto
    • 1
  • Keiichi Arimura
    • 1
  • Katsuaki Sato
    • 1
    Email author
  1. 1.Division of Immunology, Department of Infectious Diseases, Faculty of MedicineUniversity of MiyazakiMiyazakiJapan

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