Technical Data
T9160-10M
Tumor Necrosis Factor alpha, Mutant, Recombinant, Human (TNFa)
10ug
50ug
Growth Factors, Cytokines Storage: -20°CShipping: Blue Ice
TNF is secreted by macrophages, monocytes, neutrophils, T-cells, NK-cells following their stimulation by bacterial LPS . Cells expressing CD4 secrete TNF-alpha while CD8 cells secrete little or no TNF-alpha. The synthesis of TNF-alpha is induced by many different stimuli including interferons, IL2 , GM-CSF.

Description:
The clinical use of the potent anti-tumor activity of TNF-alfa has been limited by the proinflammatory side effects including fever, dose-limiting hypotension, hepatotoxicity, intravascular thrombosis, and hemorrhage. Designing clinically applicable TNF-a mutants with low systemic toxicity has been an intense pharmacological interest. Human TNF- , which binds to the murine TNF-R55 but not to the mouse TNF-R75, exhibits retained anti-tumor activity and reduced systemic toxicity in mice compared with murine TNF-a, which binds to both murine TNF receptors. Based on these results, many TNF- mutants that selectively bind to TNF-R55 have been designed. These mutants displayed cytotoxic activities on tumor cell lines in vitro, and exhibited lower systemic toxicity in vivo. Recombinant Human TNF-alfa Variant/Mutant compared with the wild-type, has an amino acid sequence deletion from a.a. 1-7, and the following a.a. substitutes Arg8, Lys9, Arg10 and Phe157 which is proven to have more activity and with less inflammatory side effect in vivo. Recombinant Human TNF-a Variant produced in E.Coli is a single, non-glycosylated, polypeptide chain containing 151 amino acids and having a molecular mass of 16598 Dalton. The Human TNF-alpha Variant is purified by standard chromatographic techniques.

Amino Acid Sequence:
The sequence of the first five N-terminal amino acids was determined and was found to be MRKRKPVAHV VANPQAEGQL QWLNRRANAL LANGVELRDN QLVVPSEGLY LIYSQVLFKG QGCPSTHVLL THTISRIAVS YQTKVNLLSA IKSPCQRETP EGAEAKPWYE PIYLGGVFQL EKGDRLSAEI NRPDYLDFAE SGQVYFGIIAF.

Dimers and Aggregates:
Less than 1% as determined by silver-stained SDS-PAGE gel analysis.

Biological Activity:
Recombinant Human TNF-alpha is Variant fully biologically active when compared to the wild type. The ED50 as determined by the cytolysis of murine L929 cells in the presence of Actinomycin D is < 0.05ng/ml, corresponding to a Specific Activity of 1 x 108 IU/mg.

Endotoxin:
0.1ng/ug (IEU/ug) of Human TNF-alpha.

Protein Content:
Protein quantitation was carried out by two independent methods:
1. UV spectroscopy at 280 nm.
2. Analysis by RP-HPLC, using a calibrated solution of TNF-alfa as a Reference Standard.

Solubility:
It is recommended to reconstitute the lyophilized Recombinant Human TNF-alpha Variant in sterile 18M -cm H2O not less than 100µg/ml, which can then be further diluted to other aqueous solutions.

Stability:
Lyophilized Recombinant Human TNF-alpha Variant although stable at room temperature for 3 weeks, should be stored desiccated below -180C. Upon reconstitution Human TNF-alpha Variant should be stored at 40C between 2-7 days and for future use below -180C. For long term storage it is recommended to add a carrier protein (0.1% HSA or BSA). Please prevent freeze-thaw cycles.

Molecular Weight:
16.6kD
 Source: E. coli
Purity: 95% as determined by RP-HPLC, anion-exchange FPLC and/or reducing and non-reducing SDS-PAGE Silver Stained gel.
Concentration: As reported
Form: Supplied as a lyophilized powder in 0.5X PBS pH 7.0.

Important Note: This product as supplied is intended for research use only, not for use in human, therapeutic or diagnostic applications without the expressed written authorization of United States Biological.
1. Who might be predisposed to the development of serious side effects when treated with TNF-alpha antagonist?
Clin Exp Rheumatol 2006 Mar-Apr;24(2):211; author reply 211
2. The effect of periodontal therapy on TNF-alpha, IL-6 and metabolic control in type 2 diabetics.
J Dent Hyg 2006 Spring;80(2):7
3. Cell to cell contact through ICAM-1-LFA-1 and TNF-alpha synergistically contributes to GM-CSF and subsequent cytokine synthesis in DBA/2 mice induced by 1,3-beta-D-Glucan SCG.
J Interferon Cytokine Res 2006 Apr;26(4):235-47
4. TNF-alpha is necessary for induction of coronary artery inflammation and aneurysm formation in an animal model of Kawasaki disease.
J Immunol 2006 May 15;176(10):6294-301
5. Interleukin-17 acts i ndependently of TNF-alpha under arthritic conditions.
J Immunol 2006 May 15;176(10):6262-9
6. Peptide YY attenuates STAT1 and STAT3 activation induced by TNF-alpha in acinar cell line AR42J.
J Am Coll Surg 2006 May;202(5):788-96

Intended for research use only. Not for use in human, therapeutic, or diagnostic applications.