Recombinant Human TNF-alpha Avi-tag Protein, CF

Catalog #: AVI10728 Datasheet

Biotinylated

Catalog #	Availability	Size / Price	Qty
AVI10728-025

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Recombinant Human TNF-alpha Avi-tag Protein, CF Summary

Learn more about Avi-tag Biotinylated Proteins

Product Specifications

Purity

>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.

Endotoxin Level

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

Activity

The biotin to protein ratio is greater than 0.7 as determined by the HABA assay.

Measured by its binding ability in a functional ELISA. When Recombinant Human TNF RI/TNFRSF1A Fc Chimera (Catalog # 372-RI) is coated at 0.025 μg/mL (100 μL/well), Biotinylated Recombinant Human TNF‑ alpha Avi-tag binds with an ED₅₀ of 2-12 ng/mL. 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 20-200 pg/mL.

Source

Chinese Hamster Ovary cell line, CHO-derived human TNF-alpha protein

Avi-tag	Human TNF alpha (Val77-Leu233) Accession # P01375.1
N-terminus	C-terminus

Accession #

P01375.1

N-terminal Sequence
Analysis

Gly of Avi-tag

Structure / Form

Monomer, biotinylated via Avi-tag

Predicted Molecular Mass

19 kDa

SDS-PAGE

15-22 kDa, under reducing conditions

Product Datasheets

AVI10728

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.

AVI10728

Formulation	Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose.
Reconstitution	Reconstitute at 500 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.

Scientific Data

Binding Activity

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When Recombinant Human TNF RI/TNFRSF1A Fc Chimera (372-RI) is coated at 0.025 μg/mL (100 μL/well), Biotinylated Recombinant Human TNF‑alpha Avi-tag binds with an ED₅₀ of 2-12 ng/mL.

SDS-PAGE

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2 μg/lane of Recombinant Human TNF-alpha Avi-tag (Catalog # AVI10728) was resolved with SDS-PAGE under reducing (R) and non-reducing (NR) conditions and visualized by Coomassie® Blue staining, showing bands at 15-22 kDa.

Background: TNF-alpha

Tumor necrosis factor alpha (TNF-alpha ), also known as cachectin and TNFSF2, 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). Our Avi-tag Biotinylated human TNF-alpha features biotinylation at a single site contained within the Avi-tag, a unique 15 amino acid peptide. Protein orientation will be uniform when bound to streptavidin-coated surface due to the precise control of bionylation and the rest of the protein is unchanged so there is no interference in the protein bioactivity.

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.

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