For the quantitative determination of human cytokine concentrations, enzyme-linked immunosorbent assay was performed using The Quantikine Human HGF Immunoassay (R&D Systems, Inc. Minneapolis, MN, USA), Human CCL2/MCP-1 Immunoassay (R&D Systems, Inc.), Human CCL7/MCP-3 Immunoassay (R&D Systems, Inc.), Human Total MMP-3 Immunoassay (R&D Systems, Inc.), and Human CXCL12/SDF-1 Immunoassay (R&D Systems, Inc.) according to the manufacturer’s instructions. The ELISA data were obtained from four independent SHED, DPSCs, and BMMSCs, respectively, each of which was performed four times.
Human ccl2 mcp 1 immunoassay
Manufactured by R&D Systems
Sourced in United States, Germany
The Human CCL2/MCP-1 Immunoassay is a quantitative sandwich enzyme-linked immunosorbent assay (ELISA) designed for the measurement of CCL2/MCP-1 levels in human cell culture supernates, serum, and plasma. The assay employs an antibody specific for human CCL2/MCP-1 coated on a microplate. Standards and samples are pipetted into the wells, and any CCL2/MCP-1 present is bound by the immobilized antibody. After washing away any unbound substances, an enzyme-linked polyclonal antibody specific for CCL2/MCP-1 is added to the wells. Following a wash to remove any unbound antibody-enzyme reagent, a substrate solution is added to the wells, and color develops in proportion to the amount of CCL2/MCP-1 bound in the initial step.
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Lab products found in correlation
Human il 6 immunoassay, by R&D Systems (2 mentions)
Human cxcl12 sdf 1 immunoassay, by R&D Systems (1 mentions)
Human chemokine array kit, by R&D Systems (1 mentions)
Chemidoc, by Bio-Rad (1 mentions)
Mouse il 6 immunoassay, by R&D Systems (1 mentions)
Z1 image microscope, by Zeiss (1 mentions)
Axiovision windows software version 4, by Zeiss (1 mentions)
5 protocols using human ccl2 mcp 1 immunoassay
1
Quantitative Cytokine Analysis by ELISA
2
Chemokine Profiling in Breast Cancer Cells
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In total, 200 μg of protein was extracted from cell lysates from MDAMB231 cells to detect the expression of 31 chemokines using the human chemokine array kit (#ARY017, R&D Systems). Images of membranes were acquired with ChemiDoc (BioRad) and dots were quantified using QuickSpots (Western vision) software. At the same time point, supernatants of MDAMB231 and HCC1395 cells were collected to detect MCP-1 levels with the human CCL2/MCP-1 Immunoassay (#SCP00, R&D Systems).
See Supplementary Methods for details regarding RNA-seq analysis.
See Supplementary Methods for details regarding RNA-seq analysis.
3
Quantification of Peritoneal Macrophages
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CD68-, ER-HR3-, CD86-, CD206-and CD163-positive macrophages were identified and counted using a Zeiss Z1 image microscope and Axiovision Windows software version 4.4 (Carl Zeiss, Oberkochen, Germany), as described previously. 9, 22, 23 Macrophages were counted in at least 10 random 750 × 500-μm areas at × 200 magnification of the peritoneal wall and are expressed in terms of counts per square millimeter (/mm 2 ).
Enzyme-Linked Immunosorbent Assay MCP-1 and IL-6 protein levels in cell culture supernatant or in serum were measured using Human CCL2/MCP-1 Immunoassay (R&D, Minneapolis, MN, USA) and mouse IL-6 immunoassay (R&D) for IL-6, in accordance with the manufacturer's instructions. Samples were frozen at the time of collection and were stored at -80 °C. Samples were not subjected to freeze-thaw cycles.
Enzyme-Linked Immunosorbent Assay MCP-1 and IL-6 protein levels in cell culture supernatant or in serum were measured using Human CCL2/MCP-1 Immunoassay (R&D, Minneapolis, MN, USA) and mouse IL-6 immunoassay (R&D) for IL-6, in accordance with the manufacturer's instructions. Samples were frozen at the time of collection and were stored at -80 °C. Samples were not subjected to freeze-thaw cycles.
4
Plasma Cytokine and Chemokine Quantification
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Plasma concentrations of TNFα (Human TNFα/TNFSF1A Immunoassay R&D Systems, Inc., Wiesbaden, Germany), interleukin 6 (IL-6) (Human IL-6 Immunoassay R&D Systems, Inc., Wiesbaden, Germany), monocyte chemotactic protein-1 (MCP-1) (Human MCP-1/CCL2 Immunoassay R&D Systems, Inc., Wiesbaden, Germany), and CD40L (Human soluble CD40 Ligand Immunoassay R&D Systems, Inc., Wiesbaden, Germany) were determined by sandwich-type immunoassay according to the manufacturer instructions. All concentration analysis was performed on an ELISA-Reader-Lab Systems Multiskan RC (Lab Systems, Finland). Genesis Lite Software and ELISA Multiskan RC were used for data acquisition and evaluation.
5
Quantification of Inflammatory Biomarkers
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Plasma concentrations of TNFa (Human TNFa/TNFSF1A Immunoassay R&D Systems, Inc., Wiesbaden, Germany), IL-6 (Human IL-6 Immunoassay R&D Systems, Inc., Wiesbaden, Germany), MCP-1 (Human MCP-1/CCL2 Immunoassay R&D Systems, Inc., Wiesbaden, Germany), soluble CD40L (Human soluble CD40 Ligand Immunoassay R&D Systems, Inc., Wiesbaden, Germany) were determined by sandwich-type immunoassay according to the manufacturer's instructions. All concentration analysis was performed on an ELISA-Reader -Lab Systems Multiskan RC (Lab systems, Finland). Genesis Lite Software, ELISA Multiskan RC was used for data acquisition and evaluation.
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