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Macsquant cytometer

Manufactured by FlowJo

The MACSQuant cytometer is a flow cytometry instrument designed for cellular analysis. It is capable of detecting and quantifying various cellular properties, such as size, complexity, and the expression of specific molecular markers. The core function of the MACSQuant cytometer is to provide accurate and reliable data on cell populations within a sample.

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3 protocols using macsquant cytometer

1

Quantifying CD38 Expression in Lung Cancer

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Flow cytometry was employed to verify the varying levels of CD38 expression by the lung cancer cell lines. Daratumumab served as the primary antibody, at a concentration of 20 μg of antibody per 1 mL of solution, while goat anti-human AlexaFluor488 secondary antibody was utilized. Proper controls of cells alone, primary antibody alone, secondary antibody alone, and a nonspecific IgG antibody were employed. Staining and flow cytometry analysis followed standard protocols.24 (link) Analysis was performed using the MACSQuant cytometer and FlowJo (V.10) software.
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2

Multiparametric Flow Cytometry for Skin and Lymph Node DC and Treg Analysis

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For DC analysis in the skin and sdLNs, cells were stained with different combinations of the following antibodies: MHC-II-Vioblue, CD11c-PerCPVio700, CD11b-VioGreen, CD11b-PerCPVio700, XCR1-Viobright-FITC, EPCAM-PE, CD86-PE-Vio770, CD86-APC-Vio770, PD-L2-PE-Vio770 (all from Miltenyi Biotec), CD11c-APC-Cy7 (BD Biosciences), XCR1-BV510 (Biolegend). Dead cells were excluded from analysis using Zombie dye (Biolegend) of appropriate color depending of the antibodies used.
For Tregs analysis, cells were stained with combinations of the following antibodies: anti-mouse CD4-FITC (Miltenyi), CD25-APC-Cy7, CD62L-PE-Cy7 (all from BD Biosciences, Le Pont de Claix, France), Latency-associated peptide (LAP)-PE and Foxp3-APC (from e-Bioscience, Paris, France), or control isotypes. Intracellular staining was performed after fixation and permeabilization, using Foxp3 Perm Kit (e-Bioscience, Paris, France). Dead cells were excluded from analysis using Zombie dye aqua (Biolegend).
Flow cytometry was performed on a MACSquant cytometer and analyzed using FlowJo software.
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3

Keratinocyte and Fibroblast Staining and TCRγδ Analysis

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Keratinocytes and fibroblasts were fixed, permeabilized (BD Cytofix/Cytoperm kit), and stained with antibodies against human vimentin (BD Pharmingen clone RV202), KRT15 (Abam clone LHK15) labeled with a chromophore preconjugated to Fab (Zenon mouse IgG labeling kit) or human CD200 (BD Biosciences; 552475). Data was collected on a dual-laser flow cytometer (BD FACSCalibur) followed by FlowJo 10 (TreeStar) software analysis.
To measure TCRγδ expression, healed wounds (~WD20) from wild type and TLRKO3 mice were minced and digested at 37 C in a buffer containing RPMI 1640, 1.67 collagenase3 Wunsch units/mL Liberase TL (Roche Life Sciences, Indianapolis, IN) and .01% DNAse (Sigma-Aldrich, St Louis, MO) for 75 minutes. Following digestion, samples were washed and filtered (40μm) to obtain a single cell suspension. Cells were stained with propidium iodide (Miltenyi Biotec, San Diego, CA) and TCRγδ (GL3) antibody (Miltenyi Biotec, San Diego, CA) followed by analysis with MACSQuant cytometer and FlowJo software.
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