Human Interleukin-1 receptor antagonist Recombinant
Categories: Interleukin-1 familyRecombinant Human Cytokines$70.00 – $900.00
Description
Accession
P18510
Source
Optimized DNA sequence encoding Humaninterleukin-1 receptor antagonist mature chain was expressed in Escherichia Coli.
Molecular weight
Human interleukin-1 receptor antagonist is generated by the proteolytic removal of the signal peptide and propeptide.The molecule has a calculated molecular mass of approximately 17kDa. RecombinantIL-1RA is a monomer protein consisting of 153 amino acid residue subunits, and migrates as an approximately 17kDa protein under reducing conditions in SDS-PAGE.
Purity
>95%, as determined by SDS-PAGE and HPLC
Biological Activity
The ED(50) was determined by the ability to inhibit the IL-1α mediated cell proliferation in a murine helper T cell line,for this effect, in the presence of 10pg/mL of IL-1α, is typically - 2 ng/mL.
Protein Sequence
MEICRGLRSH LITLLLFLFH SETICRPSGR KSSKMQAFRI WDVNQKTFYL RNNQLVAGYL QGPNVNLEEK IDVVPIEPHA LFLGIHGGKM CLSCVKSGDE TRLQLEAVNI TDLSENRKQD KRFAFIRSDS GPTTSFESAA CPGWFLCTAM EADQPVSLTN MPDEGVMVTK FYFQEDE
Endotoxin
Endotoxin content was assayed using a LAL gel clot method. Endotoxin level was found to be less than 0.1 ng/µg(1EU/µg).
Presentation
Recombinant Interleukin-1 receptor antagonist was lyophilized from a 0.2 μm filtered PBS solution pH7.4 .
Reconstitution
A quick spin of the vial followed by reconstitution in distilled water to a concentration not less than 0.1 mg/mL. This solution can then be diluted into other buffers.
Storage
The lyophilized protein is stable for at least years from date of receipt at -20° C. Upon reconstitution, this cytokine can be stored in working aliquots at2° -8° C for one month, or at -20° C for six months, with a carrier protein without detectable loss of activity. Avoid repeated freeze/thaw cycles.
Usage
This cytokine product is for research purposes only.It may not be used for therapeutics or diagnostic purposes.
Methods
Treatment of Cell Cultures
- Human recombinant (r)IL-1ß (, , 10 ng/ml) was applied and maintained for different time periods (from 10 min to 48 h) before harvesting the cells for RNA isolation or western blot analysis.
- Medium was collected to perform enzyme-linked immunosorbent assay (ELISA).
- In some experiments lipo-polysaccharide (LPS; 100 ng/ml, , ), rIL-6 (10 ng/ml; athmann , , ), tumor necrosis factor α (TNFα; 1 ng/ml, , ) and High mobility group box 1 (HMGB1; 40 nM; HMGh , , ) alone or together with IL-1 ß were applied and maintained in the medium for 24 before harvesting the cells for RNA isolation.
- In some experiments cells stimulated with LPS were treated with LPS-RS (a TLR4 antagonist from the photosynthetic bacterium
Rhodobacter sphaeroides , , 10 µg/ml), applied 1 h before LPS. - Human IL-1 receptor antagonist (IL-1Ra; 1 µg/ml, , ) was used to neutralize IL-1β activity (applied 1 h before IL-1β).
- As previously…
Treatment of cell cultures
- Human recombinant (r)IL-1β (, , 10 ng/mL) was applied and maintained for 24 h before harvesting the cells for RNA isolation, western blot analysis or for immunocytochemistry.
- In some experiments different time periods of IL-1β exposure (ranging from 10 min to 48 h) were used and rIL-6 (10 ng/mL , , ), tumor necrosis factor α (TNFα; 1 ng/mL, , ) and high mobility group box 1 (HMGB1; 40nM; HMGh , , ) alone or together with IL-1β were applied and maintained in the medium for 24 h before harvesting the cells for RNA isolation.
- Human IL-1receptor antagonist (IL-1Ra; 1 μg/mL, , ) was used to neutralize IL-1β activity (applied 1 h before IL-1β).
- As previously shown [
Effect of pro-inflammatory agonists on the release of inflammatory cytokines by neutrophils.
- Neutrophils were incubated for 2 and 24 h in presence of PBS (vehicle), fMLP (10−7 M), LPS (1 µg/ml) or TNF-α (10 ng/ml) for the subsequent quantification of VEGF, IL-1RA (2 h) and IL-8 (24 h) release by ELISA.
