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Methods Mol Biol. 2019;1968:183-194. doi: 10.1007/978-1-4939-9199-0_15.

Two-Photon Intravital Imaging of Leukocytes in the Trachea During Pneumococcal Infection.

Author information

1
Institute for Research in Biomedicine, Università della Svizzera italiana, Bellinzona, Switzerland.
2
Graduate School of Cellular and Molecular Sciences, Faculty of Medicine, University of Bern, Bern, Switzerland.
3
Institute for Research in Biomedicine, Università della Svizzera italiana, Bellinzona, Switzerland. santiago.gonzalez@irb.usi.ch.

Abstract

Two-photon intravital imaging (2P-IVM) of the murine trachea is a powerful technique for real-time imaging of immune cell recruitment and trafficking during airborne pathogen infections. Neutrophils are an important component of the innate immune response that are able to rapidly infiltrate the airway mucosa in response to Streptococcus pneumoniae infection. Here we describe a protocol to visualize in vivo neutrophil extravasation and cell dynamics in the tracheal tissue of a S. pneumoniae-infected mouse using 2P-IVM. To perform this protocol, we infected and imaged the trachea of a lysozyme M green fluorescent protein (LysM-GFP) mouse, in which neutrophils express GFP. Additionally, we used a custom-designed platform, which allowed the intubation and fixation of the trachea after surgical exposition, and we injected intravenously a fluorescently labeled dextran solution to visualize the blood vessels.

KEYWORDS:

Extravasation; Neutrophil; Streptococcus pneumoniae; Trachea; Two-photon intravital microscopy (2P-IVM)

PMID:
30929215
DOI:
10.1007/978-1-4939-9199-0_15
[Indexed for MEDLINE]

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