Cells were washed with phosphate buffered saline (PBS) and stripped with 0.025% trypsin and 0.24 mM EDTA. Cells (1 × 106 cells) were pelleted and then were resuspended in Hank’s Balanced Salt Solution (HBSS, Life Technologies, Carlsbad, CA, USA) or PBS containing 2% fetal calf serum (FCS) and incubated with 0.25 μg of anti-human nectin-4 monoclonal antibody (Clone 337516, R&D Systems, Minneapolis, MN, USA) on ice for at least 30 min. Next, the cells were washed in PBS containing 2% FCS and incubated with anti-mouse immunoglobulin G (IgG)-Alexa 488 diluted 1:2,000 (Life Technologies) on ice for at least 30 min. Finally, the cells were washed with PBS containing 2% FCS, and the fluorescence intensity was measured by a FACSCalibur (BD Biosciences, San Jose, CA, USA). Data were analyzed by FlowJo software.
Anti human nectin 4 monoclonal antibody
Manufactured by R&D Systems
Sourced in United States
Anti-human nectin-4 monoclonal antibody is a laboratory reagent used for the detection and quantification of nectin-4, a cell adhesion molecule, in biological samples. This antibody can be used in various immunoassay techniques, such as ELISA, Western blotting, and flow cytometry, to measure nectin-4 levels in cell lysates, tissue extracts, or other relevant samples.
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2 protocols using anti human nectin 4 monoclonal antibody
1
Quantification of Nectin-4 Expression
2
Quantification of Nectin-4 Expression
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Cells were washed with phosphate buffered saline (PBS) and trypsinized in 0.025% trypsin/0.24 mM ethylenediaminetetraacetic acid (EDTA). They were pelleted by centrifugation at 3,300 × g for 1 min, and then were resuspended in Hank's balanced salt solution (HBSS; Life Technologies) containing 2% FCS and incubated on ice for 30 min with anti-human Nectin-4 monoclonal antibody (Clone 337516, R&D Systems, Minneapolis, MN, USA), anti-human SLAM antibody [A12 (7D4); BioLegend, San Diego, CA, USA], and anti-CD46 antibody (M177; HyCult Biotech, Uden, The Netherlands). Next, the cells were washed in PBS containing 2% FCS, and incubated on ice for a further 30 min with Alexa 488-conjugated anti-mouse IgG (Life Technologies). Finally, the cells were washed with PBS containing 2% FCS, and the intensity of fluorescence was measured by a FACSCalibur (BD Biosciences, San Jose, CA, USA). To derive the relative level of Nectin-4 expression, the mean fluorescent intensity (MFI) of cells stained with anti-Nectin-4 antibody was calculated using Flowjo software ver 9.7.5 (TreeStar, San Carlos, CA).
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