Intravascular staining for discrimination of vascular and tissue leukocytes

By K. Anderson, K. Mayer-Barber, H. Sung, Lalit K. Beura, Britnie R. James, Justin J. Taylor, Lindor Qunaj, T. Griffith, V. Vezys, Daniel L. Barber and 1 more. Nature Protocols, 2014

730 citations recorded by the source corpus.

Characterization of the cellular participants in tissue immune responses is crucial to understanding infection, cancer, autoimmunity, allergy, graft rejection and other immunological processes. Previous reports indicate that leukocytes in lung vasculature fail to be completely removed by perfusion. Several studies suggest that intravascular staining may discriminate between tissue-localized and blood-borne cells in the mouse lung. Here we outline a protocol for the validation and use of intravascular staining to define innate and adaptive immune cells in mice. We demonstrate application of this protocol to leukocyte analyses in many tissues and we describe its use in the contexts of lymphocytic choriomeningitis virus and Mycobacterium tuberculosis infections or solid tumors. Intravascular staining and organ isolation usually takes 5–30 min per mouse, with additional time required for any subsequent leukocyte isolation, staining and analysis. In summary, this simple protocol should help enable interpretable analyses of tissue immune responses.

Subfields: Immunology

View Paper: https://www.semanticscholar.org/paper/35b4d9682b8cf0618b09f279b8fabd73255ac2e4

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In Immunology

Citations in the corpus

Human T cell development, localization, and function throughout life (2018, background): “Phenotypically, mouse Trm cells express the early T cell activation marker CD69 (Anderson et al., 2014; Mueller and Mackay, 2016; Turner et al., 2014), which can promote retention through sequestration of the sphingosine-1-S1PR1 (Mackay et al.”
Human tissue-resident memory T cells are defined by core transcriptional and functional signatures in lymphoid and mucosal sites (2017, background): “In mice, tissue retention demonstrated by parabiosis (Jiang et al., 2012; Steinert et al., 2015) and in vivo antibody labeling (Anderson et al., 2014; Turner et al., 2014) identified phenotypic markers associated with tissue residence, including CD69 and CD103.”
High‐Dimensional Single‐Cell Mapping of Central Nervous System Immune Cells Reveals Distinct Myeloid Subsets in Health, Aging, and Disease (2018, neutral): “In order to label intra-vascular cells within the steady state CNS we injected PE conjugated anti-CD45 (7.5 mg in PBS) i.v. into adult C57BL/6 mice and euthanized them after 3 minutes, performed transcardiac perfusion and harvested the CNS, as previously described (Anderson et al., 2014).”
Developmental Relationships of Four Exhausted CD8+ T Cell Subsets Reveals Underlying Transcriptional and Epigenetic Landscape Control Mechanisms (2020, methodology): “Ab) staining to assess blood accessibility (Anderson et al., 2014).”
Quantifying memory CD8 T cells reveals regionalization of immunosurveillance (2015, methodology): “We next focused our analyses on the i.v. Ab+ BMP in liver and kidney, which includes cells within sinusoids and glomeruli (Anderson et al., 2014).”