Recombinant Human TNF-alpha Hyperactive Mutant Protein

Catalog #: 8599-TA Datasheet

Mutant form with approximately 5-fold greater activity

Carrier Free

Catalog #	Availability	Size / Price	Qty
8599-TA-010/CF
8599-TA-025/CF
8599-TA-100/CF

With Carrier

Catalog #	Availability	Size / Price	Qty
8599-TA-010

Recombinant Human TNF-alpha Hyperactive Mutant Protein Bioactivity

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All TNF-alpha Proteins and Enzymes

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Summary Product Datasheets Carrier Free Data Images Reconstitution Calculator Background Related Research Areas

Recombinant Human TNF-alpha Hyperactive Mutant Protein Summary

Product Specifications

Purity

>95%, by SDS-PAGE with silver staining.

Endotoxin Level

<0.10 EU per 1 μg of the protein by the LAL method.

Activity

Measured in a cytotoxicity assay using L‑929 mouse fibroblast cells in the presence of the metabolic inhibitor actinomycin D. Matthews, N. and M.L. Neale (1987) in Lymphokines and Interferons, A Practical Approach. Clemens, M.J. et al. (eds): IRL Press. 221. The ED₅₀ for this effect is 2.5-12.5 pg/mL.

Source

E. coli-derived human TNF-alpha protein
Lys87-Leu233 (Leu233Phe), with an N-terminal Met-Arg-Lys-Arg

Accession #

P01375

N-terminal Sequence
Analysis

Met

Structure / Form

Non-covalent trimer

Predicted Molecular Mass

17 kDa

SDS-PAGE

16 kDa, reducing conditions

Product Datasheets

8599-TA (with carrier)

Product Datasheet

8599-TA/CF (carrier free)

Product Datasheet

Carrier Free

What does CF mean?

CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins. Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration. The carrier free version does not contain BSA.

What formulation is right for me?

In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard. In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.

8599-TA

Formulation	Lyophilized from a 0.2 μm filtered solution in PBS with BSA as a carrier protein.
Reconstitution	Reconstitute at 100 μg/mL in PBS.
Shipping	The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage:	Use a manual defrost freezer and avoid repeated freeze-thaw cycles. 12 months from date of receipt, -20 to -70 °C as supplied. 1 month, 2 to 8 °C under sterile conditions after reconstitution. 3 months, -20 to -70 °C under sterile conditions after reconstitution.

8599-TA/CF

Formulation	Lyophilized from a 0.2 μm filtered solution in PBS.
Reconstitution	Reconstitute at 100 μg/mL in PBS.
Shipping	The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage:	Use a manual defrost freezer and avoid repeated freeze-thaw cycles. 12 months from date of receipt, -20 to -70 °C as supplied. 1 month, 2 to 8 °C under sterile conditions after reconstitution. 3 months, -20 to -70 °C under sterile conditions after reconstitution.

Scientific Data

Bioactivity

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Recombinant Human TNF-alpha Hyperactive Mutant (Catalog # 8599-TA) induces cytotoxicity in the L-929 mouse fibroblast cell line in the presence of the metabolic inhibitor actinomycin D. The ED₅₀ for this effect is 2.5-12.5 pg/mL. This is an approximate 10-fold activity increase over the wild-type TNF-alpha (Catalog # 210-TA).

Background: TNF-alpha

Tumor necrosis factor alpha (TNF-alpha ), also known as cachectin and TNFSF1A, is the prototypic ligand of the TNF superfamily. It is a pleiotropic molecule that plays a central role in inflammation, immune system development, apoptosis, and lipid metabolism (1, 2). Human TNF-alpha consisits of a 35 amino acid (aa) cytoplasmic domain, a 21 aa transmembrane segment, and a 177 aa extracellular domain (ECD) (3). Within the ECD, human TNF-alpha shares 97% aa sequence identity with rhesus and 71%-92% with bovine, canine, cotton rat, equine, feline, mouse, porcine, and rat TNF-alpha. TNF-alpha is produced by a wide variety of immune, epithelial, endothelial, and tumor cells (1, 2). TNF-alpha is assembled intracellularly to form a noncovalently linked homotrimer which is expressed on the cell surface (4). Cell surface TNF-alpha can induce the lysis of neighboring tumor cells and virus infected cells, and it can generate its own downstream cell signaling following ligation by soluble TNFR I (2, 5). Shedding of membrane bound TNF-alpha by TACE/ADAM17 releases the bioactive cytokine, a 55 kDa soluble trimer of the TNF-alpha extracellular domain (6-8). TNF-alpha binds the ubiquitous 55-60 kDa TNF RI (9, 10) and the hematopoietic cell-restricted 80 kDa TNF RII (11, 12), both of which are also expressed as homotrimers (1, 2, 13). Both type I and type II receptors bind TNF-alpha with comparable affinity (14), although only TNF RI contains a cytoplasmic death domain which triggers the activation of apoptosis. Soluble forms of both types of receptors are released and can neutralize the biological activity of TNF-alpha (15). This recombinant protein contains an N-terminal insertion and a Leu233Phe substitution which have been shown to increase the bioactivity of TNF-alpha in vitro and in vivo (16).

References

Zelova, H. and J. Hosek (2013) Inflamm. Res. 62:641.
Juhasz, K. et al. (2013) Expert Rev. Clin. Immunol. 9:335.
Pennica, D. et al. (1984) Nature 312:724.
Tang, P. et al. (1996) Biochemistry 35:8216.
Perez, C. et al. (1990) Cell 63:251.
Black, R.A. et al. (1997) Nature 385:729.
Moss, M.L. et al. (1997) Nature 385:733.
Gearing, A.J.H. et al. (1994) Nature 370:555.
Schall, T.J. et al. (1990) Cell 61:361.
Loetscher, H. et al. (1990) Cell 61:351.
Dembic, Z. et al. (1990) Cytokine 2:231.
Smith, C.A. et al. (1990) Science 248:1019.
Loetscher, H. et al. (1991) J. Biol. Chem. 266:18324.
Pinckard, J.K. et al. (1997) J. Biol. Chem. 272:10784.
Engelmann, H. et al. (1990) J. Biol. Chem. 265:1531.
Yan, Z. et al. (2006) Cytotherapy 8:415.

Long Name

Tumor Necrosis Factor alpha

Entrez Gene IDs

7124 (Human); 21926 (Mouse); 24835 (Rat); 397086 (Porcine); 280943 (Bovine); 403922 (Canine); 102139631 (Cynomolgus Monkey); 100033834 (Equine); 493755 (Feline); 100009088 (Rabbit)

Alternate Names

APC1 protein; Cachectin; Cachetin; DIF; TNF; TNF, monocyte-derived; TNFA; TNF-A; TNFalpha; TNF-alpha; TNF-alphacachectin; TNFATNF, macrophage-derived; TNFG1F; TNFSF1A; TNFSF2; TNFSF2TNF superfamily, member 2; tumor necrosis factor (TNF superfamily, member 2); tumor necrosis factor alpha; Tumor necrosis factor ligand superfamily member 2; tumor necrosis factor; tumor necrosis factor-alpha