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Nat Protoc. 2015 Jul;10(7):1116-30. doi: 10.1038/nprot.2015.070. Epub 2015 Jun 25.

Intravital imaging of hair follicle regeneration in the mouse.

Author information

1
Department of Genetics, Cell Biology, and Dermatology, Yale Stem Cell Center, Yale Cancer Center, Yale School of Medicine, New Haven, Connecticut, USA.
2
Department of Laboratory Medicine, Yale School of Medicine, New Haven, Connecticut, USA.
3
1] Department of Genetics, Cell Biology, and Dermatology, Yale Stem Cell Center, Yale Cancer Center, Yale School of Medicine, New Haven, Connecticut, USA. [2] Department of Cell Biology, Yale School of Medicine, New Haven, Connecticut, USA. [3] Department of Dermatology, Yale School of Medicine, New Haven, Connecticut, USA. [4] Yale Stem Cell Center, Yale School of Medicine, New Haven, Connecticut, USA. [5] Yale Cancer Center, Yale School of Medicine, New Haven, Connecticut, USA.

Abstract

Hair follicles are mammalian skin organs that periodically and stereotypically regenerate from a small pool of stem cells. Hence, hair follicles are a widely studied model for stem cell biology and regeneration. This protocol describes the use of two-photon laser-scanning microscopy (TPLSM) to study hair regeneration within a living, uninjured mouse. TPLSM provides advantages over conventional approaches, including enabling time-resolved imaging of single hair follicle stem cells. Thus, it is possible to capture behaviors including apoptosis, proliferation and migration, and to revisit the same cells for in vivo lineage tracing. In addition, a wide range of fluorescent reporter mouse lines facilitates TPLSM in the skin. This protocol also describes TPLSM laser ablation, which can spatiotemporally manipulate specific cellular populations of the hair follicle or microenvironment to test their regenerative contributions. The preparation time is variable depending on the goals of the experiment, but it generally takes 30-60 min. Imaging time is dependent on the goals of the experiment. Together, these components of TPLSM can be used to develop a comprehensive understanding of hair regeneration during homeostasis and injury.

PMID:
26110716
PMCID:
PMC4632978
DOI:
10.1038/nprot.2015.070
[Indexed for MEDLINE]
Free PMC Article
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