OMIPs Directory

OMIP-032

Two multi-color immunophenotyping panels for assessing the innate and adaptive immune cells in the mouse mammary gland

Cytometry PART A, Volume 89, Issue 6, 527–530 (2016)
Ashleigh Unsworth, Robin Anderson, Nicole Haynes, Kara Britt

Description: A multi-color antibody panel was designed and optimized to identify and characterize 10 leukocyte subpopulations from both the innate and adaptive arms of the immune system. Markers to detect B-cells (CD45.2+ CD19+), T-cells (CD45.2+ TCRβ+), natural killer cells (CD45.2+ TCRβ− CD49b+ NKp46+), and myeloid cells (CD45.2+ CD11b+) were employed. Myeloid cells were further differentiated as dendritic cells (CD45.2+ CD11b+ CD11c+ MHCII+), neutrophils (CD45.2+ CD11b+ Ly6G+), macrophages (CD45.2+ CD11b+ Ly6G− Ly6Clow), and monocytes (CD45.2+ CD11b+ Ly6G− Ly6Chigh). T-cell populations were sub-divided into T-helper cells (CD45.2+ TCRβ+ CD8− CD4+), and cytotoxic T-cells (CD45.2+ TCRβ+ CD4− CD8+). T-cell memory/effector status was determined using CD62L and CD44 to distinguish between effector (CD44+ CD62L−), memory (CD44+ CD62L+) and naïve (CD44− CD62L+) T cells. This panel was established for the analysis of collagenase digested mouse (Balb/C) mammary gland, spleen and tumor samples, as well as RBC-lysed whole blood.

Species: Mouse

Tissue: Mammary gland, Peripheral blood, Spleen, Tumor

Cell Subset: Neutrophil, B cell, T cell, T helper, Ctl, Dc, Nk, Myeloid cells, Macrophage, Monocyte

Keyword: Collagenase digestion, Immunoprofiling, Lysed whole blood (lwb)

Machine: BD LSR Fortessa X20

Panel details

Marker Species Clone Fluorophore Purpose Co-expressions groups Antigen Density
MHC II Mouse M5/114.15.2 Brilliant Violet 711 DC B cell, DC High
Dead cells Mouse Propidium Iodide Viability
Dead cells Mouse Fluoro-Gold Viability
CD11c Mouse HL3 APC DC monocyte, DC Low
CD11b Mouse M1/70 PE Myeloid cell monocyte, DC Med
Ly-6C Mouse AL-21 Brilliant Violet 421 Monocyte, macrophage monocyte, macrophage Low
Mannose Receptor Mouse C068C2 FITC alternatively activated macrophage macrophage Med
CD4 Mouse Gk1.5 APC-Cy7 Th T cell High
TCR beta Mouse H57-597 PE-Cy7 T cell T cell Med
CD49b Mouse DX5 FITC NK NK Med
CD44 Mouse IM7 Brilliant Violet 605 T cell differentiation T cell, B cell Med
CD62L Mouse Mel-14 Brilliant Violet 510 T cell differentiation T cell, B cell, NK, monocyte Med
CD19 Mouse 1D3 APC B cell B cell Med
NKp46 Mouse 29A1.4 Brilliant Violet 421 NK NK Low
Ly-6G Mouse 1A8 PE-Cy7 Neutrophil neutrophil Med
CD45.2 Mouse 104 PE leukocyte NK, monocyte, DC, T cell, B cell, neutrophil High
CD45.2 Mouse 104 APC-Cy7 Leukocyte T cell, B cell, NK, monocyte, DC, neutrophil High
CD8a Mouse 53-6.7 Brilliant Violet 711 cytotoxic T cell T cell High