Benefit from the possibility of gentle tissue dissociation with preserved cellular integrity. You can then isolate and analyze macrophages and myeloid-derived suppressor cells (MDSCs).
Mouse macrophages are extensively studied in tissue and tumor samples. For reliable isolation of mouse macrophages, gentle dissociation of lymphoid tissue is key. Mouse spleen can be dissociated into single-cell suspensions using our Spleen Dissociation Kit, mouse in combination with our gentleMACS™ Dissociator with Heaters. Mechanical dissociation with enzymatic degradation of the extracellular matrix maintains the structural integrity of tissues.
Watch how simple it is to dissociate mouse spleen tissue with the gentleMACS Dissociator.
The MDSC Phenotyping Cocktail Kit, anti-human, REAfinity has been developed to allow standardized and simplified flow cytometric analysis of subpopulations of MDSCs described so far. For greater flexibility, the product configuration also allows extension of the analysis panel with additional specificities of interest (e.g. CD10, CD66b, CD15, and LOX-1).
A) Gating strategy
B) Identification of low density PMN cells (PMN-MDSCs)
C) Identification of M-MDSCs:
D) Identification of e-MDSCs:
Human PBMCs were stained using the MDSC Phenotyping Cocktail Kit, anti-human, REAfinity and analyzed by flow cytometry using the MACSQuant®Analyzer 10. A) To exclude red blood cells and identify leukocytes, a first gate on CD45+cells was set (region R1). Next, to eliminate doublets, a gate on single cells in Forward scatter-A (A=area) versus Forward scatter-H (H=height) was set (region R2). These cells were further distinguished from debris via Forward scatter-A and Side scatter-A (region R3). Afterwards, a gate on viable cells (7-AAD–cells) was set (region R4). B) Cells from region R4 were further separated into 3 subsets: side scatterhigh(SSChigh) cells, which correspond to low density PMN-MDSCs (region R5), CD14+cells (region R6) and SSClowCD14–cells (region R7). Low density PMN-MDSCs contained in region R5 were further separated into 4 subsets based on the expression of CD16 and CD11b (gates R8 to R11). C) Cells stained with MDSC Control Cocktail from R6 were displayed with CD14 versus REA Control-FITC, and a region enclosing >99.5% of cells was set (region R12). This region was transferred to the cells labeled with MDSC Staining Cocktail, depicting M-MDSCs as CD14+HLA-DR–. D) Cells stained with MDSC Control Cocktail from region R7 were displayed with REA Control-FITC versus REA-Control-PE, and a region above the background staining, containing –CD33int.
The CD163 MicroBead Kit, human has been developed for the enrichment of cells based on the expression of the CD163 antigen. CD163 is most abundantly expressed by mature tissue macrophages and peripheral blood monocytes.
CD163+cells were isolated from human peripheral blood mononuclear cells (PBMCs) (A) or ovarian carcinoma samples (B) using the CD163 MicroBead Kit, an MS Column, and a MiniMACS™ Separator. Cells were fluorescently stained with CD45-VioBlue®as well as with CD14-FITC and CD163-VioBlue (A) or CD206-FITC and CD163-PE (B) and analyzed by flow cytometry using the MACSQuant®Analyzer. Cell debris and dead cells were excluded from the analysis based on scatter signals and propidium iodide fluorescence.
Isolation of F4/80+ macrophages. F4/80+ cells were isolated from spleen single-cell suspension using Anti-F4/80 MicroBeads UltraPure, two MS Columns, and an OctoMACS™ Separator. The cells were fluorescently stained with CD45-VioBlue®, CD11b-APC, and Anti-F4/80-FITC and analyzed by flow cytometry using the MACSQuant® Analyzer. Cell debris and dead cells were excluded from the analysis based on scatter signals and 7-AAD fluorescence.
We offer various reagents and kits for the isolation of mouse myeloid cells from tissues and tumors. Download the poster and find a comprehensive overview of all myeloid cell markers and products for convenient isolation and analysis.
Working with MDSCs? The Myeloid-Derived Suppressor Cell Isolation Kit, mouse has been developed for the isolation of CD11b+Gr-1+ myeloid cells from mouse lymphoid tissue.
Find out more about products for mouse myeloid cell research in tissues and tumors in this poster.
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