Mice were injected intravenously (i.v.) with 1 ug CD45-APC (clone 30-F11) or CD45-FITC (clone 30-F11) antibody 3 minutes prior to euthanasia. Cells from the lungs were processed as previously described (32 (link)). Lungs were harvested and made into single-cell suspensions as previously described (33 (link), 34 (link)). Cells from the lung were stained for extracellular surface molecules with antibodies specific to CD90.2 (clone 53–2.1), CD4 (clone GK1.5), CD8 (clone 53–6.7), CD49d (clone R1–2), CD11a (clone M17/4), CD103 (clone 2E7), CD69 (clone H1.2F3), CD62L (MEL-14), CD122 (5H4), and CXCR3 (clone CXCR3–173) for 30 minutes at 4°C and fixed with fix/lyse solution (eBioscience) for 10 minutes at room temperature. RSV-specific CD8 T cells were identified using a M282 tetramer, which is made in our laboratory. Stained cells were run on an LSRFortessa (BD Biosciences, San Jose, CA) and analyzed with FlowJo software (Tree Star, Ashland, OR).
Protocol detail
Lung Immune Cell Profiling via Flow Cytometry
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Antibodies
Antibody
Cd103
Cd122
Cd62l
Cd8 t cells
Cd90
Cells
Clone
Cxcr3
Euthanasia
Fitc
Lung
Mice
Tetramer
Tree
Corresponding Organization :
Other organizations : University of Iowa
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Variable analysis
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- CD45-APC (clone 30-F11) or CD45-FITC (clone 30-F11) antibody
- Cells from the lungs
- RSV-specific CD8 T cells
- Mice were injected intravenously (i.v.) with 1 ug of the antibody 3 minutes prior to euthanasia
- Cells from the lung were stained for extracellular surface molecules with antibodies specific to CD90.2 (clone 53–2.1), CD4 (clone GK1.5), CD8 (clone 53–6.7), CD49d (clone R1–2), CD11a (clone M17/4), CD103 (clone 2E7), CD69 (clone H1.2F3), CD62L (MEL-14), CD122 (5H4), and CXCR3 (clone CXCR3–173) for 30 minutes at 4°C and fixed with fix/lyse solution (eBioscience) for 10 minutes at room temperature
- RSV-specific CD8 T cells were identified using a M282 tetramer, which is made in our laboratory
- Not mentioned
- Not mentioned
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