Human MCP-1 ELISA Assay
The Human MCP-1 ELISA Assay is For Research Use Only
Size: 1×96 wells
Sensitivity: 15 pg/mL
Dynamic Range: 31.25 – 2000 pg/ml
Incubation Time: 3.5 hours
Sample Type: Serum, Plasma, Cell Culture
Sample Size: 100 µl
Alternative Names: Monocyte chemoattractant protein-1, monocyte chemotactic protein-1
Monocyte chemoattractant protein-1 (MCP-1) called also monocyte chemotactic protein-1. MCP-1 is an 8.6kDa protein and belongs to the family of chemotactic cytokines known as Chemokines. MCP-1 is expressed by monocytes, vascular endothelial cells, smooth muscle cells, glomerular mesangial cell, osteoblastic cells, and human pulmonary type-2-like epithelial cells in culture. MCP-1 exhibits chemotactic activity for monocytes/macrophages, basophils, T lymphocytes, particularly memory T cells and natural killer cells and neural stem cells. Elevated levels of MCP-1 are detected during inflammation and immune responses. MCP-1 activity is also important in wound healing as shown by delayed wound repair in MCP-1deficient mice. MCP-1 is implicated in the pathogenesis of disease states that involve monocyte/macrophage infiltrates, such as psoriasis, rheumatoid arthritis and recruitment of monocytes to the arterial wall in atherosclerosis. In addition to its chemotactic function, MCP-1 also induces the expression of IL-10 from macrophages, which favors a TH2 immune response. MCP-1 is a glycopeptide containing one potential N-linked glycosylation site. The extent of MCP-1 glycosylation influences its chemotactic potency and half-life in vivo.
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The Eagle Biosciences Human Monocyte Chemoattractant Protein-1 (MCP-1) ELISA Assay Kit employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for MCP-1 has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any MCP-1 present is bound by the immobilized antibody. Following incubation unbound samples are removed during a wash step, and then a detection antibody specific for MCP-1 is added to the wells and binds to the combination of capture antibody-MCP-1 in sample. Following a wash to remove any unbound combination, and enzyme conjugate is added to the wells. Following incubation and wash steps a substrate is added. A colored product is formed in proportion to the amount of MCP-1 present in the sample. The reaction is terminated by addition of acid and absorbance is measured at 450nm. A standard curve is prepared from seven MCP-1 standard dilutions and MCP-1 sample concentration determined.
- Prepare all reagents and working standards as directed in the previous sections.
- Determine the number of microwell strips required to test the desired number of samples plus appropriate number of wells needed for running blanks and standards. Remove extra microwell strips from holder and store in foil bag with the desiccant provided at 2-8°C sealed tightly.
- Add 100 μL of Standard, control, or sample, per well. Cover with the adhesive strip provided. Incubate for 1.5 hours at 37°C.
- Aspirate each well and wash, repeating the process three times for a total of four washes. Wash by filling each well with Wash Buffer (350 μL) using a squirt bottle, manifold dispenser or auto-washer. Complete removal of liquid at each step is essential to good performance. After the last wash, remove any remaining Wash Buffer by aspirating or decanting. Invert the plate and blot it against clean paper towels.
- Add 100 μL of the working solution of Biotin-Conjugate to each well. Cover with a new adhesive strip and incubate 1 hour at 37°C.
- Repeat the aspiration/wash.
- Add 100 μL of the working solution of Streptavidin-HRP to each well. Cover with a new adhesive strip and incubate for 30 minutes at 37°C Avoid placing the plate in direct light.
- Repeat the aspiration/wash.
- Add 100 μL of Substrate Solution to each well. Incubate for 10-20 minutes at 37°C. Avoid placing the plate in direct light.
- Add 100 μL of Stop Solution to each well. Gently tap the plate to ensure thorough mixing.
- Determine the optical density of each well immediately, using a microplate reader set to 450 nm.(optionally 630nm as the reference wave length;610-650nm is acceptable)
Please note: All documents above are for reference use only and should not be used in place of the documents included with this physical product. If digital copies are needed, please contact us.
Headley, S.A., Chapman, D.J., Germain, M.J, et al. The effects of 16-weeks of prebiotic supplementation nand aerobic exercise training of inflammatory markers, oxidative stress, uremic toxins, and the microbiota in pre-dialysis kidney patients: a randomized controlled trial-protocol paper. BMC Nephrology (2020) https://doi.org/10.1186/s12882-020-02177-x
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