Human sICAM-1 ELISA Assay

$405.00

The Human sICAM-1 ELISA Assay (enzyme-linked immunoassay kit) is intended for the quantitative determination of human Intercellular Adhesion Molecule 1 (sICAM-1) concentrations in cell culture supernates, serum, and plasma. The Human Intercellular Adhesion Molecule 1 (sICAM-1) ELISA Assay Kit is for research use only and not to be used in diagnostic procedures.

SKU: ICM31-K01 Categories: , ,

Human sICAM-1 ELISA Assay

The Human sICAM-1 ELISA Assay is For Research Use Only

Size: 1×96 wells
Sensitivity: 31 pg/mL
Dynamic Range: 62.5 – 2000 pg/ml
Incubation Time: 3.5 hours
Sample Type: Serum, Plasma, Cell Culture
Sample Size: 100 µl
Alternative Names: Soluble Intercellular Adhesion Molecule 1, CD54, Cluster of differentiation 54


Assay Background

Intercellular Adhesion Molecule 1 (ICAM-1), also known as CD54, is a nearly ubiquitous transmembrane glycoprotein that plays a key role in leukocyte migration and activation. Human ICAM-1 contains five Ig-like domains in its extracellular domain (ECD) and associates into non-covalently linked dimers. Soluble forms of monomeric and dimeric ICAM-1 (sICAM-1) can be generated via proteolytic cleavage by cathepsin G, elastase, MMP-9, MMP-14/MT1-MMP, and TACE/ADAM17. In the mouse, alternate splicing generates isoforms that lack particular Ig-like domains and are differentially sensitive to proteolysis. Within the ECD, human ICAM-1 shares 53% amino acid sequence identity with mouse and rat ICAM-1.

The principal binding partners of ICAM-1 are the leukocyte integrins LFA-1 (CD11a/CD18) and Mac-1 (CD11b/CD18) (9 – 11). The multivalency of dimeric ICAM-1 increases its strength of interaction with LFA-1. ICAM-1 also binds several non-integrin ligands including CD43/sialophorin, fibrinogen, hyaluronan, rhinoviruses, and Plasmodium falciparum-infected erythrocytes. At sites of inflammation, ICAM-1 is upregulated on endothelial and epithelial cells where it mediates the adhesion and paracellular migration of leukocytes expressing activated LFA-1 and Mac-1. ICAM-1 ligation prolongs antigen presentation by dendritic cells and promotes T cell proliferation and cytokine release. ICAM-1 activation also participates in angiogenesis, wound healing, and bone metabolism.

Soluble ICAM-1 has been reported in serum, cerebrospinal fluid, urine, and bronchoalveolar lavage fluid. Elevated levels of sICAM-1 in these fluids are associated with cardiovascular disease, type 2 diabetes, organ transplant dysfunction, oxidant stress, abdominal fat mass, hypertension, liver disease, and certain malignancies. sICAM-1 promotes angiogenesis and serves as an indicator of vascular endothelial cell activation or damage (41, 42). It also functions as an inhibitor of transmembrane ICAM-1 mediated activities such as monocyte adhesion to activated endothelial cells and sensitivity of tumor cells to NK cell-mediated lysis.


Related Products

Human CD102 sICAM-2 ELISA Assay
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Human CD54 ELISA Assay

Additional Information

Assay Principle


The Eagle Biosciences Human Intercellular Adhesion Molecule 1 (sICAM-1) ELISA Assay Kit employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for sICAM-1 has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any sICAM-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 sICAM-1 is added to the wells and binds to the combination of capture antibody- sICAM-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 sICAM-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 sICAM-1 standard dilutions and sICAM-1 sample concentration determined.

Assay Procedure


  1. Prepare all reagents and working standards as directed in the previous sections.
  2. 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.
  3. Add 100µL of Standard, control, or sample, per well. Cover with the adhesive strip provided. Incubate for 1.5 hours at 37°C.
  4. 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.
  5. 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.
  6. Repeat the aspiration/wash.
  7. 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.
  8. Repeat the aspiration/wash.
  9. Add 100 µL of Substrate Solution to each well. Incubate for 10-20 minutes at 37°C. Avoid placing the plate in direct light.
  10. Add 100 µL of Stop Solution to each well. Gently tap the plate to ensure thorough mixing.
  11. 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)

Manual

Product Manual


Publications

References


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