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Clone wm59

Manufactured by BioLegend

The Clone WM59 is a monoclonal antibody specific for the human melanoma-associated antigen MCAM/CD146. It can be used for the detection and quantification of MCAM/CD146-expressing cells by flow cytometry.

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3 protocols using clone wm59

1

Isolation and Quantification of FAP and SC

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Muscle specimens underwent digestion separately at 37°C in 0.2% collagenase for 90 minutes followed by 0.4% dispase for 30 minutes. The resulting suspensions were filtered through a 40 μm cell strainer, then pelleted and resuspended in FACS buffer (5% Fetal Bovine Serum in PBS). FAP and SC quantity were reported as a percentage of nondebris live cells, as determined using LIVE/DEAD Fixable Aqua Dead Cell Stain Kit (ThermoFisher) and their respective surface markers. Cells were sorting and quantified using FACS BD Aria II. FAPs were defined as CD31‐/CD45‐/CD29‐/CD56‐/PDGFRα+/CD184‐, and SCs were defined as CD31‐/CD45‐/CD29+/CD56+/PDGFRα‐/CD184+ as previously described.29, 30 Gates were set based on each antibodies' fluorescence minus one for every run. The antibodies used for FACS are as follows at concentration of 1 μg/mL per 1 × 106 cells/mL: CD56 (Biolegend, Clone HCD56, #318304, FITC), CD29 (BD bioscience, Clone MAR4, #743783, Pacific Blue), CD184 (Biolegend, Clone 12G5, #306510, APC), PDGFRα (Biolegend, Clone 16A1, #323508, APC/CY7), CD31 (Biolegend, Clone WM59, #303122, BV605), and CD45 (Biolegend, Clone 2D1, #368524, BV605).
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2

Fetal Pericyte Surface Characterization

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Surface staining of fetal pericytes was performed with CD31-AF488 (1:100 dilution; clone WM59, BioLegend, #303110) and CD146-AF647 (1:100 dilution; clone P1H12, BioLegend, #361014) antibodies. Zombie Violet fixable viability kit (BioLegend, #423113) was used for fixable live/dead staining as per manufacturer instructions. FACS acquisitions was performed on BD FACSCanto II (BD Biosciences), using BD FACSDIVA v 6.0. All FACS data were analyzed using FlowJo v10.0 software.
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3

Fetal Pericyte Surface Characterization

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Surface staining of fetal pericytes was performed with CD31-AF488 (1:100 dilution; clone WM59, BioLegend, #303110) and CD146-AF647 (1:100 dilution; clone P1H12, BioLegend, #361014) antibodies. Zombie Violet fixable viability kit (BioLegend, #423113) was used for fixable live/dead staining as per manufacturer instructions. FACS acquisitions was performed on BD FACSCanto II (BD Biosciences), using BD FACSDIVA v 6.0. All FACS data were analyzed using FlowJo v10.0 software.
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