The purity of recombinant hHis6IL-25 was determined by SDS-PAGE of 6 µg reduced (+) and nonreduced (-) recombinant hHis6IL-25 and staining overnight with Coomassie Blue.
IL-5 production from human PBMC costimulated with IL-2 and increasing concentrations of hHis6IL-25 was assessed. PBMC were treated with TSLP (100 ng/ml, 24 hr) and then costimulated with IL-2 (10 ng/ml) and increasing concentrations of hHis6IL-25. After 72 hr, cell supernatants were harvested and assayed for IL-5 by ELISA and the OD450 was determined.
With carrier: Lyophilized from a 0.22 μm filtered solution of hHis6IL-25 in 20 mM Tris, pH 7.2 containing 150 mM NaCl and 20 μg BSA per 1 μg hHis6IL-25. Carrier free: Lyophilized from a 0.22 μm filtered solution of hHis6IL-25 in 20 mM Tris, pH 7.2 containing 150 mM NaCl.
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.
>97% as determined by SDS-PAGE of 6 μg reduced (+) and nonreduced (-) recombinant hHis6IL-25. 30% migrates as monomer under nonreducing conditions. All lots are greater than 97% pure.
Recombinant N-terminally His6-tagged hIL-25 has a calculated MW of 19,318. DTT-reduced protein migrates as a 26-32 kDa polypeptide. The nonreduced cystine-linked homodimer migrates as a 40-46 kDa protein. Lower mobility and heterogeneity in SDS-PAGE are due to glycosylation. The expected amino terminus of recombinant hHis6IL-25 was verified by amino acid sequencing.
The bioactivity of recombinant hHis6IL-25 was determined by its ability to induce IL-5 production from TSLP-primed PBMC in the presence of IL-2. The ED50 of each lot is between 5-15 pg/ml.
Less than 0.01 ng endotoxin/1 μg hHis6IL-25.
Recombinant human His6IL-25 (hHis6IL-25) Tyr33-Gly177 (Accession #NP_073626) was expressed in human 293 cells at Cell Signaling Technology.
IL-25, also known as IL-17E, is a member of the IL-17 superfamily of cytokines. IL-25 is expressed in epithelial cells, CD4+ T cells, mast cells, and eosinophils (1). Many cell types are responsive to IL-25, including T cells, macrophages, and epithelial cells (1). The receptor for IL-25 consists of a heterodimer of IL-17RA and IL-17RB (1,2). IL-25 promotes Th2 type immune responses by induction of IL-5, IL-4, and IL-13 and may contribute to allergic inflammation and asthma (1-3). IL-25 has also been shown in research to promote Th9 cell activation and induces apoptosis in breast cancer cells (4,5).
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