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|Baculovirus-Insect Cell lysate that Human S100A8 / CAGA / p8 transfected / overexpressed for Western blot (WB) positive control. The whole cell lysate is provided in 1X Sample Buffer (1X modified RIPA buffer+1X SDS loading buffer).|
|A DNA sequence encoding the human S100A8 (NP_002955.2) (Met 1-Glu 93) was expressed, fused with a polyhistidine tag at the C-terminus.|
|The recombinant human S100A8 consists of 103 amino acids and predicts a molecular mass of 12.2 kDa. It migrates as an approximately 14.6 kDa band in SDS-PAGE under reducing conditions.|
|Cell lysate was prepared by homogenization in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.|
|Modified RIPA Lysis Buffer: 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1% Sodium deoxycholate, 1mM PMSF.|
|12.5% SDS-PAGE Stained with Coomassie Blue after protein purification.|
|Samples are stable for up to twelve months from date of receipt.|
|1. Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube. 2. Re-dissolve the pellet using 200μL pure water and boil for 2-5 min. 3. Store the lyophilized cell lysate at 4℃. After re-dissolution, recommend to aliquot it into smaller quantities and store at -80℃.|
|1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).|
|Store at 4℃. After re-dissolution, aliquot and store at -80℃.|
|Western blot (WB): Use at an assay dependent dilution.|
Other Applications: Not tested.
Optimal dilutions/concentrations should be determined by the end user.
S100A8 is a member of the S100 protein family containing 2EF-hand calcium-binding motifs. S100 proteins are involved in the regulation of a number of cellular processes such as cell cycle progression and differentiation. Altered expression of S100A8 protein is associated with various diseases and cancers. S100A8 may have an immunoregulatory role by contributing to the regulation of fetal-maternal interactions. It may play a protective role and its absence may allow infiltration by maternal cells, a process eventually manifesting as resorption. The heterodimeric S100 protein complex S100A8/A9 which has been shown to be involved in inflammatory and neoplastic disorders. The complex can induce cell proliferation, or apoptosis, inflammation, collagen synthesis, and cell migration. S100A8/A9 has emerged as important pro-inflammatory mediator in acute and chronic inflammation. More recently, increased S100A8 and S100A9 levels were also detected in various human cancers, presenting abundant expression in neoplastic tumor cells as well as infiltrating immune cells. On the one hand, S100A8/A9 is a powerful apoptotic agent produced by immune cells, making it a very fascinating tool in the battle against cancer. It spears the risk to induce auto-immune response and may serve as a lead compound for cancer-selective therapeutics. In contrast, S100A8/A9 expression in cancer cells has also been associated with tumor development, cancer invasion or metastasis. Altogether, its expression and potential cytokine-like function in inflammation and in cancer suggests that S100A8/A9 may play a key role in inflammation-associated cancer.