OMIP-061
20‐Color Flow Cytometry Panel for High‐Dimensional Characterization of Murine Antigen‐Presenting Cells
Cytometry PART A, Volume 95, Issue 12, 1226-1230 (2019)
Anthony T. DiPiazza, Juliane P. Hill, Barney S. Graham, Tracy J. Ruckwardt
Description: This 20‐color flow cytometry panel was designed to resolve the cellular heterogeneity of antigen‐presenting cells and was optimized for lymph node tissue. Reagents were carefully selected and optimized for identification of B cells (B220), neutrophils (Ly6G), monocytes and macrophages (Ly6C, CD169, F4/80), and dendritic cells (XCR1, CD172a, CD11c, I‐A/I‐E, CD24, CD64, pDCA‐1, CD103, CD11b). Inclusion of additional functional markers involved in cell migration (CCR7), co‐stimulation (CD80), and adhesion (ICAM‐1) enabled further phenotypic characterization. Finally, this panel has been tested and is compatible with fluorescently labeled antigens such as Alexa Fluor 488 (Ax488) for the study of antigen‐bearing cells in vivo.
Species: Mouse
Tissue: Lymph node
Cell Subset: B cell, Dc, Mdc/cdc, Pdc, Myeloid cells, Macrophage, Monocyte, Neutrophil
Keyword: Cell migration
Machine: BD FACSymphony A5
Panel details
| Marker | Species | Clone | Fluorophore | Purpose | Co-expressions groups | Antigen Density |
|---|---|---|---|---|---|---|
| Dead cells | Mouse | Live/Dead Fix Blue | Viability | |||
| CD172a | Mouse | P84 | PerCP-eFluor 710 | cDC2, other myeloid cells | cDC | Med |
| CD11c | Mouse | N418 | Brilliant Violet 421 | macrophage and DC | Macrophage, cDC | Low |
| CD103 | Mouse | M290 | Brilliant Violet 510 | Lung migratory cDC1 | cDC | High |
| CD24 | Mouse | M1/69 | Brilliant Violet 605 | cDC lineage | Macrophage, B cell, cDC | High |
| CD3 | Mouse | 17A2 | Brilliant Violet 750 | T and NKT cells | High | |
| CD80 | Mouse | 16-10A1 | PE-CF594 | Co-stimulation | cDC | Low |
| F4/80 | Mouse | BM8 | PE-Cy5 | Macrophage subsets | DC, Macrophage | Low |
| CD169 | Mouse | 3D6.112 | PE-Cy7 | Macrophage subsets | Macrophage | Med |
| CD45 | Mouse | 30-F11 | Brilliant Ultraviolet 661 | Hematopoietic cell lineage | Neutrophil, Macrophage, cDC, pDC, B cell | High |
| Ly-6G | Mouse | 1A8 | Brilliant Ultraviolet 737 | Neutrophil | Neutrophil | High |
| CD317 | Mouse | 927 | APC | pDC | pDC | High |
| CD11b | Mouse | M1/70 | APC-R700 | Macrophages and dendritic cells | Macrophage, DC, Neutrophil | Med |
| ICAM-1 | Mouse | 3E2 | Brilliant Ultraviolet 395 | Adhesion | cDC, Macrophage | High |
| MHC II | Mouse | M5/114 | Brilliant Violet 711 | Activation | B cell, cDC | High |
| CCR7 | Mouse | 4B12 | PE | Trafficking | cDC | Low |
| XCR1 | Mouse | ZET | Brilliant Violet 650 | pDC (plasmacytoid DC) | pDC, cDC | High |
| Ly-6C | Mouse | HK1.4 | Brilliant Violet 570 | Monocytes/macrophages, neutrophils, moDC | Macrophage | Med |
| CD45R (B220) | Mouse | RA3-6B2 | Brilliant Ultraviolet 496 | B cells and pDCs | B cell, pDC | Med |
| CD64 | Mouse | X54-5/7.1 | Brilliant Violet 786 | Monocyte/macrophage, moDC lineage | Macrophage, cDC | Med |
