Abstract
CD4+ T cell responses have been shown to be protective in many respiratory virus infections. In the respiratory tract, CD4+ T cells include cells in the airway and parenchyma and cells adhering to the pulmonary vasculature. Here we discuss in detail the methods that are useful for characterizing CD4+ T cells in different anatomic locations in mouse lungs.
Keywords: Memory CD4+ T cell, Lung, Bronchoalveolar lavage fluid, Intranasal/intravascular antibody labeling, Flow cytometry
Background
To distinguish memory T cells in the circulation and tissues, a method for intravascular staining of T cells has been developed (Anderson et al., 2012). This method has been widely used to define memory T cells in many organs and tissues, including lungs, spleens and intestines. However, memory T cells in the respiratory tract are located in three unique anatomic locations, i.e., airway, parenchyma and pulmonary vasculature. Intravascular staining cannot distinguish cells in the airway and parenchyma, since they are both isolated from the circulation and intravascularly-administered antibodies will not stain these two populations. We designed a simultaneous intranasal/intravascular antibody labeling assay that can label and distinguish cells in all three locations using minimal amount of antibodies.
Materials and Reagents
Equipment
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Procedure
Notes
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Acknowledgments
This work was supported by the Thousand Talents Plan Award of China 2015 (J.Z.), the Municipal Healthcare Joint-Innovation Major Project of Guangzhou (201604020011) (J.Z.) and the N.I.H. (USA) (PO1 AI060699, S.P.). The protocol described herein was based on the following manuscript: Zhao et al. Immunity. 2016 Jun 21; 44(6):1379-91.
References
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