New Tools for Imaging of Immune Systems: Visualization of Cell Cycle, Cell Death, and Cell Movement by Using the Mice Lines Expressing Fucci, SCAT3.1, and Kaede and KikGR

  • Michio Tomura
Part of the Methods in Molecular Biology book series (MIMB, volume 1763)


Visualization of biological events in real time in vivo has become a crucial to understand immune responses. We have been established novel visualization tools for life of immune cells: proliferation, cell death, and migration. Fucci-transgenic mice allow us to visualize cell cycle phases by reciprocal expression of mKusabira-Orange2 in G1 phase and mAzami-Green in S/G2/M phase. Caspase-3 indicator SCAT3.1 knock-in mice visualize cell death by changing color. Photoconvertible proteins, Kaede and KikGR expressing mice track cell movement between organs by labeling immune cells as red color. Here, I will introduce how to use and visualize these mice. These techniques will help to understand immune system in the living whole body.

Key words

Cell cycle Fucci Photoconvertible protein Cell migration Kaede KikGR Cell death Caspase-3 SCAT3.1 



This work was supported in part by JSPS Grants-in-Aid for Scientific Research in Innovative Areas “Analysis and Synthesis of Multidimensional Immune Organ Network” (#24111007); JSPS Grants-in-Aid for Scientific Research (B) (#16H05087); Special Coordination Funds for Promoting Science and Technology of the Japanese Government.


  1. 1.
    Huang AY, Qi H, Germain RN (2004) Illuminating the landscape of in vivo immunity: insights from dynamic in situ imaging of secondary lymphoid tissues. Immunity 21:331–339PubMedGoogle Scholar
  2. 2.
    Sumen C, Mempel TR, Mazo IB et al (2004) Intravital microscopy: visualizing immunity in context. Immunity 21:315–329PubMedGoogle Scholar
  3. 3.
    Tomura M, Sakaue-Sawano A, Mori Y et al (2013) Contrasting quiescent G0 phase with mitotic cell cycling in the mouse immune system. PLoS One 8:e73801CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Tomura M, Mori YS, Watanabe R et al (2009) Time-lapse observation of cellular function with fluorescent probe reveals novel CTL-target cell interactions. Int Immunol 21:1145–1150CrossRefPubMedGoogle Scholar
  5. 5.
    Tomura M, Yoshida N, Tanaka J et al (2008) Monitoring cellular movement in vivo with photoconvertible fluorescence protein “Kaede” transgenic mice. Proc Natl Acad Sci U S A 105:10871–10876CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Tomura M, Hata A, Matsuoka S et al (2014) Tracking and quantification of dendritic cell migration and antigen trafficking between the skin and lymph nodes. Sci Rep 4:6030CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Sakaue-Sawano A, Kurokawa H, Morimura T et al (2008) Visualizing spatiotemporal dynamics of multicellular cell-cycle progression. Cell 132:487–498CrossRefPubMedGoogle Scholar
  8. 8.
    David R, McIlwain TB, Mak TW (2013) Caspase functions in cell death and disease. Cold Spring Harb Perspect Biol 5:a008656Google Scholar
  9. 9.
    Takemoto K, Nagai T, Miyawaki A et al (2003) Spatio-temporal activation of caspase revealed by indicator that is insensitive to environmental effects. J Cell Biol 160:235–243CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Nagai T, Miyawaki A (2004) A high-throughput method for development of FRET-based indicators for proteolysis. Biochem Biophys Res Commun 319:72–77CrossRefPubMedGoogle Scholar
  11. 11.
    Ando R, Hama H, Yamamoto-Hino M et al (2002) An optical marker based on the UV-induced green-to-red photoconversion of a fluorescent protein. Proc Natl Acad Sci U S A 99:12651–12656CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Tsutsui H, Karasawa S, Shimizu H et al (2005) Semi-rational engineering of a coral fluorescent protein into an efficient highlighter. EMBO Rep 6:233–238CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Tomura M, Honda T, Tanizaki H et al (2010) Activated regulatory T cells are the major T cell type emigrating from the skin during a cutaneous immune response in mice. J Clin Invest 120:883–893CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Tomura M, Itoh K, Kanagawa O (2010) Naive CD4+ T lymphocytes circulate through lymphoid organs to interact with endogenous antigens and upregulate their function. J Immunol 184:4646–4653CrossRefPubMedGoogle Scholar
  15. 15.
    Chtanova T, Hampton HR, Waterhouse LA et al (2014) Real-time interactive two-photon photoconversion of recirculating lymphocytes for discontinuous cell tracking in live adult mice. J Biophotonics 7:425–433. 201CrossRefPubMedGoogle Scholar
  16. 16.
    Kotani M, Kikuta J, Klauschen F et al (2013) Systemic circulation and bone recruitment of osteoclast precursors tracked by using fluorescent imaging techniques. J Immunol 190:605–612CrossRefPubMedGoogle Scholar
  17. 17.
    Shand FH, Ueha S, Otsuji M et al (2014) Tracking of intertissue migration reveals the origins of tumor-infiltrating monocytes. Proc Natl Acad Sci U S A 111:7771–7776CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Ikebuchi R, Teraguchi S, Vandenbon A et al (2016) A rare subset of skin-tropic regulatory T cells expressing Il10/Gzmb inhibits the cutaneous immune response. Sci Rep 6:35002CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Tomura M, Kabashima K (2013) Analysis of cell movement between skin and other anatomical sites in vivo using photoconvertible fluorescent protein “Kaede”-transgenic mice. Mol Dermatol Methods Mol Biol 961:279–286. Spring (Edited by Cristina Has and Cassian Sitaru)CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2018

Authors and Affiliations

  • Michio Tomura
    • 1
  1. 1.Laboratory of Immunology, Faculty of PharmacyOsaka Ohtani UniversityOsakaJapan

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