The mouse S100A9/Calprotectin ELISA will detect murine S100A9 homodimers as well as higher molecular oligomers, including the S100A8/A9 heterocomplex.
S100 proteins are low molecular weight calcium binding proteins. The family consists of 22 members to date that are expressed in several tissues and cell types and have diverse functions. Murine S100A9, also called calgranulin B or MRP14, has a molecular weight of approximately 13kD. S100 proteins consist of 2 (calcium binding) EF-hands located on the termini flanked by hydrophobic hinge regions. They are expressed in a cell type specific manner and with specific subcellular localization. Intracellular proteins can exist as dimers. S100A9 can form homodimers but is often associated with S100A8 (MRP8) to form heterodimers, also well known as calprotectin. Intracellular function of S100 proteins include modulation of enzyme activity, calcium homeostasis, cell differentiation and survival. Extracellularly, they participate in signal transduction and interact with cell surface receptors like RAGE, TLRs and others. S100A9 is constitutively expressed by myeloid cells nut not by lymphocytes. It is highly expressed in resting neutrophils, keratinocytes, macrophages and epithelial cells in active inflammatory disease. S100A9 is a potent chemoattractant for neutrophils and monocytes. The expression can be induced by several mediators like LPS, TNFα, IL-1, IL10 an TGFβ depending on the expressing cell type. The protein is predominantly associated with (chronic) inflammation. Elevated serum levels are found among others in patients with cystic fibrosis, inflammatory bowel diseases, rheumatoid arthritis, cardiovascular disease and malignancies.
The mouse S100A9 ELISA kit is to be used for the in vitro quantitative determination of mouse S100A9 in serum and plasma samples. The mouse S100A9 ELISA will detect murine S100A9 homodimers as well as higher molecular oligomers, including S100A8/A9 multimers.
The mouse S100A9 ELISA is a ready-to-use solid-phase enzyme-linked immunosorbent assay based on the sandwich principle with a working time of 3½ hours. The efficient format of a plate with twelve disposable 8-well strips allows free choice of batch size for the assay.
Samples and standards are incubated in microtiter wells coated with antibodies recognizing mouse S100A9. Biotinylated tracer antibody will bind to the captured mouse S100A9. Streptavidin-peroxidase conjugate will bind to the biotinylated tracer antibody. Streptavidin-peroxidase conjugate will react with the substrate, tetramethylbenzidine (TMB). The enzyme reaction is stopped by the addition of oxalic acid. The absorbance at 450 nm is measured with a spectrophotometer. A standard curve is obtained by plotting the absorbance (linear) versus the corresponding concentrations of the mouse S100A9 standards (log). The mouse S100A9 concentration of samples, which are run concurrently with the standards, can be determined from the standard curve.
Normal mouse plasma samples containing baseline levels of mouse S100A9, were spiked with mouse S100A9, in concentrations of 5 and 50 ng/ml. Samples with and without mouse S100A9, were incubated for 1 hour at room temperature. Samples were measured using the ELISA. Values for mouse S100A9, ranged between 80% and 91% for EDTA samples.