The proliferation of TF-1 cells treated with increasing concentrations of hIL-6 was assessed. After 48 hour treatment with hIL-6, cells were incubated with a tetrazolium salt and the OD450 - OD650 was determined.
The purity of recombinant hIL-6 was determined by SDS-PAGE of 6 µg reduced (+) and non-reduced (-) recombinant hIL-6 and staining overnight with Coomassie Blue.
Western blot analysis of extracts from TF-1 cells, untreated or treated with hIL-6 for 10 minutes, using Phospho-Stat1 (Tyr701) Antibody #9171 (upper) and Stat1 Antibody #9172 (lower).
With carrier: Lyophilized from a 0.22 μm filtered solution of PBS, pH 7.2 containing 20 μg BSA per 1 μg hIL-6. Carrier free: Lyophilized from a 0.22 μm filtered solution of PBS, pH 7.2.
Stable in lyophilized state at -20°C for 1 year after receipt. Sterile stock solutions reconstituted with carrier protein are stable at 4°C for 2 months and at -20°C for 6 months. Avoid repeated freeze-thaw cycles.Maintain sterility. Storage at -20°C should be in a manual defrost freezer.
>98% as determined by SDS-PAGE of 6 μg reduced (+) and non-reduced (-) recombinant hIL-6. All lots are greater than 98% pure.
Based on amino acid sequencing, greater than 60% of recombinant hIL-6 has a Met on the amino-terminal Val30 (sequence of MVPPG) and has a calculated MW of 20,943. The remainder starts at pro 32 (sequence of PPGED) or Pro33 (sequence of PGEDS). DTT-reduced protein migrates as a 23 kDa polypeptide and non-reduced protein migrates as a 22 kDa polypeptide due to intramolecular cystines.
Recombinant human IL-6 (hIL-6) Val30-Met212 (Accession #NM_000600) was produced in E. coli at Cell Signaling Technology.
IL-6 is a potent inducer of the acute phase response and is produced by T cells, macrophages, fibroblasts, endothelial and other cells (1,2). IL-6 induces proliferation and differentiation and acts on B cells, T cells, thymocytes, and others. IL-6 in concert with TGFβ is important for developing Th17 responses. IL-6 binds to IL-6Rα and through this association induces gp130 homodimerization (1). gp130 homodimerization triggers the Jak/Stat cascade and the SHP2/Erk MAP kinase cascade (1,3,4). IL-6 also forms a complex with an IL-6Rα splice variant that is non-membrane associated (3).The IL-6/soluble IL-6Rα complex can then activate the gp130 signaling pathway on cells that express gp130 but not IL-6Rα (3). IL-6, through increasing expression of proangiogenic VEGF, may contribute to metastatic breast cancer (5).
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