Isolation and high‐dimensional phenotyping of gastrointestinal immune cells
| Autor: | Gustavo B. Menezes, Alan Moreira de Araujo, Maria Alice Freitas-Lopes, Thais Garcias Moreira, Bruna Araújo David, Nicole E. Paul, Stephen Rubino, Rafael M. Rezende |
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| Jazyk: | angličtina |
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Immunology Cell Culture Techniques Cell Separation Biology Immunophenotyping 03 medical and health sciences Mice 0302 clinical medicine Immune system Antigen Leukocytes Immunology and Allergy Animals Mass cytometry Microbiome Intestinal Mucosa Cells Cultured Original Articles Isolation (microbiology) Flow Cytometry Mice Inbred C57BL 030104 developmental biology Liver biology.protein Lymph Nodes Antibody Cytometry 030215 immunology |
| Popis: | The gastrointestinal immune system plays a pivotal role in the host relationship with food antigens, the homeostatic microbiome and enteric pathogens. Here, we describe how to collect and process liver and intestinal samples to efficiently isolate and analyse resident immune cells. Furthermore, we describe a step-by-step methodology showing how to high-dimensionally immunophenotype resident leucocytes using cytometry by time-of-flight, providing a well-characterized antibody platform that allows the identification of every leucocyte subset simultaneously. This protocol also includes instructions to purify and cultivate primary murine hepatocytes, a powerful tool to assess basic cell biology and toxicology assays. Gut and liver samples from the same mouse can be collected, processed and stained in less than 6 hr. This protocol enables the recovery of several populations of purified and viable immune cells from solid and fibrous organs, preventing unwanted loss of adherent cells during isolation. |
| Databáze: | OpenAIRE |
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