Isolation and Analysis of Stromal Cell Populations from Mouse Lymph Nodes

Yannick O Alexandre; Scott N Mueller

doi:10.21769/BioProtoc.2445

Isolation and Analysis of Stromal Cell Populations from Mouse Lymph Nodes

Bio Protoc. 2017 Aug 20;7(16):e2445. doi: 10.21769/BioProtoc.2445.

Authors

Yannick O Alexandre¹, Scott N Mueller^{1

2}

Affiliations

¹ Department of Microbiology and Immunology, the University of Melbourne, at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia.
² The Australian Research Council Centre of Excellence in Advanced Molecular Imaging, the University of Melbourne, Melbourne, Australia.

Abstract

Our protocol describes a simple procedure for isolating stromal cells from lymph nodes (LN). LN are disrupted then enzymatically digested with collagenase and dispase to produce a single cell suspension that can be stained with fluorescently labelled antibodies and analysed by flow cytometry. This protocol will enable identification of fibroblastic reticular cells (FRC), lymphatic endothelial cells (LEC), blood endothelial cells (BEC) as PNAd⁺ BEC that form LN high endothelial venules (HEV). This method can be applied to examine LN stromal cell responses during inflammatory events induced by infections or immunologic adjuvants and to subset most leukocytes found in LN.

Keywords: Endothelial cells; Fibroblastic reticular cells; Immune response; Lymph nodes; Lymphoid stromal cells; Virus infection.