The ability of hIL-6Rα to enhance IL-6 mediated inhibition of M1 cell proliferation was assessed. M1 Cells were treated with increasing concentrations of hIL-6Rα in the presence of 20 ng/ml Human Interleukin-6 (hIL-6) #8904. After 72 hours, cells were incubated with a tetrazolium salt and the OD450 - OD650 was determined.
The purity of recombinant hIL-6Rα was determined by SDS-PAGE of 6 µg reduced (+) and nonreduced (-) recombinant IL-6Rα and staining overnight with Coomassie Blue.
With carrier: Lyophilized from a 0.22 μm filtered solution of hIL-6Rα in 20 mM Tris, pH 7.2 containing 20 μg BSA per 1 μg
hIL-6Ra. Carrier free: Lyophilized from a 0.22 μm filtered solution of hIL-6Rα in 20 mM Tris, pH 7.2.
Stable in lyophilized state at 4°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.
>90% as determined by SDS-PAGE of 6 μg reduced (+) and nonreduced (-) recombinant hIL-6Rα. All lots are greater than 90% pure.
Molecular Characterization: Recombinant hIL-6Rα has a calculated MW of 37,871. DTT-reduced and non-reduced protein migrate as 66 kDa polypeptides. Lower mobility and heterogeneity in SDS-PAGE are due to glycosylation. The expected amino-terminal LAPRR of recombinant hIL-6Rα was verified by amino acid sequencing.
The activity of hIL-6Rα was determined by assessing its ability to enhance IL-6 mediated inhibition of M1 cells proliferation. The ED50 of each lot is between 2 and 15 ng/ml.
Less than 0.01 ng endotoxin/1 μg hIL-6Rα.
Recombinant Human Interleukin-6 Receptor α (hIL-6Rα) Leu20 - Asp358 (Accession #NP_000556) was expressed in human 293 cells at Cell Signaling Technology.
The IL-6 receptor is a heterodimeric complex that consists of a ligand-binding IL-6 receptor α (IL-6Rα) subunit and a signaling component, gp130 (1). Binding of IL-6 to IL-6Rα results in dimerization of receptor with gp130 and subsequent STAT3 activation (1). IL-6Rα is cleaved from the cell surface by ADAM17 (1,2). In humans, soluble IL-6Rα is also generated via alternatively spliced mRNA (1,3). Soluble IL-6Rα binds to IL-6 and can stimulate signaling via membrane bound gp130 in a process known as “trans-signaling” (1). It is through trans-signaling that IL-6 stimulates cells that do not express membrane bound IL-6Rα (1).
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