Recombinant human TNF-alpha protein

Summary

• High purity human protein (Uniprot number: P01375)
• >98%, by SDS-PAGE quantitative densitometry
• Source: Expressed in E. coli
• 17.3 kDa (Non covalent trimer)
• Animal origin-free (AOF) and carrier protein-free
• Manufactured in Cambridge, UK
• Lyophilized from Tris/NaCl
• Resuspend in water at >100 µg/mL, prepare single-use aliquots, add carrier protein if desired, and store frozen at -20°C or -80°C

Featured applications

• Inflammation regulation
• Immune response modulation
• Cell survival regulation
• Apoptosis regulation
• Metabolic function regulation
• Autoimmune disease and inflammatory disorders

Bioactivity

TNF-α activity is determined using the TNF-α-responsive firefly luciferase reporter assay. Transfected HEK293T cells are treated in triplicate with a serial dilution of TNF-α for 3 hours. Firefly luciferase activity is measured and normalised to the control Renilla luciferase activity. Data from Qk083 lot #204609. EC50 = 1.1 ng/mL (63 pM).

Purity

Recombinant TNF-α migrates as a major band at approximately 17 kDa (monomer) in reduced (R) and non-reduced (NR) conditions. The dimeric and trimeric forms are also observed at approximately 33 and 55 kDa, respectively. No contaminating protein bands are present.The purified recombinant protein (3 µg) was resolved using 15% w/v SDS-PAGE in reduced (+β-mercaptoethanol, R) and non-reduced (NR) conditions and stained with Coomassie Brilliant Blue R250. Data from Qk083 lot #204609.

Protein background

Tumor necrosis factor-alpha (TNF-alpha) is a multifunctional cytokine that plays a central role in inflammation, immunity, and various physiological processes [1]. It is primarily produced by activated macrophages and other immune cells, although it can also be synthesized by a variety of other cell types, including endothelial cells, fibroblasts, and adipocytes [2,3].

TNF-alpha exerts its effects by binding to two distinct receptors: TNF receptor 1 (TNFR1) and TNF receptor 2 (TNFR2). Both receptors are widely expressed on many cell types throughout the body [4]. TNFR1 activation triggers pro-inflammatory responses, apoptosis, and immune cell recruitment, while TNFR2 signaling is involved in tissue regeneration, immune regulation, and cell survival [5,7].

One of the primary functions of TNF-alpha is its role in the initiation and regulation of inflammation. Upon activation, TNF-alpha stimulates the production of other pro-inflammatory cytokines, such as interleukin-1 (IL-1) and interleukin-6 (IL-6), as well as chemokines that attract immune cells to sites of infection or injury [6]. This pro-inflammatory cascade helps to eliminate pathogens and promote tissue repair. However, dysregulated TNF-alpha signaling can lead to chronic inflammation and tissue damage, contributing to the pathogenesis of various autoimmune diseases, such as rheumatoid arthritis, inflammatory bowel disease, and psoriasis.

TNF-alpha plays a crucial role in the regulation of immune responses. It modulates the activation, proliferation, and differentiation of immune cells, including T cells, B cells, and natural killer (NK) cells. TNF-alpha is also involved in the regulation of adaptive immunity by influencing antigen presentation and T cell activation [1-3,8,9].

TNF-alpha has diverse effects on various physiological processes, including metabolism, angiogenesis, and tissue remodeling. It is implicated in the pathogenesis of obesity, insulin resistance, and cardiovascular diseases. TNF-alpha contributes to the host defence against tumors by promoting inflammation and activating immune responses against cancer cells [10]. TNF-alpha has emerged as an important therapeutic target in the treatment of inflammatory and autoimmune diseases. Biologic agents that inhibit TNF-alpha, such as monoclonal antibodies and soluble receptors, have revolutionized the management of conditions like rheumatoid arthritis, Crohn’s disease, and psoriasis, providing relief for patients who do not respond to conventional therapies. However, the blockade of TNF-alpha can also lead to adverse effects, including increased susceptibility to infections and the development of paradoxical inflammatory reactions [11,12].

Background references

1. Abbas, A. K., Lichtman, A. H., & Pillai, S. (2017). Cellular and Molecular Immunology (9th ed.). Elsevier.
2. Croft, M., & Siegel, R. M. (2017). Beyond TNF: TNF superfamily cytokines as targets for the treatment of rheumatic diseases. Nature Reviews Rheumatology, 13(4), 217-233. doi.org/10.1038/nrrheum.2017.22
3. Tracey, D., Klareskog, L., & Sasso, E. H. (2019). Sounding the alarm: A primer on the immunobiology of the alert cytokine, TNF-α. Arthritis Research & Therapy, 21(1), 1-10. doi: 10.1016/j.pharmthera.2007.10.001
4. Aggarwal, B. B. (2003). Signalling pathways of the TNF superfamily: a double-edged sword. Nature Reviews Immunology, 3(9), 745-756. doi.org/10.1038/nri1184
5. Locksley, R. M., Killeen, N., & Lenardo, M. J. (2001). The TNF and TNF receptor superfamilies: integrating mammalian biology. Cell, 104(4), 487-501. doi: 10.1016/s0092-8674(01)00237-9
6. Balkwill, F., & Mantovani, A. (2001). Inflammation and cancer: back to Virchow? Lancet, 357(9255), 539–545. doi: 10.1016/S0140-6736(00)04046-0
7. Parameswaran, N., & Patial, S. (2010). Tumor necrosis factor-α signaling in macrophages. Critical Reviews in Eukaryotic Gene Expression, 20(2), 87–103. doi: 10.1615/critreveukargeneexpr.v20.i2.10
8. Tracey, D., Klareskog, L., Sasso, E. H., Salfeld, J. G., & Tak, P. P. (2008). Tumor necrosis factor antagonist mechanisms of action: a comprehensive review. Pharmacology & Therapeutics, 117(2), 244–279. doi: 10.1016/j.pharmthera.2007.10.001
9. Aggarwal, B. B. (2003). Signalling pathways of the TNF superfamily: a double-edged sword. Nature Reviews Immunology, 3(9), 745–756. doi.org/10.1038/nri1184
10. Choy, E. H. (2002). Clinical significance of tumor necrosis factor α antagonists. Expert Opinion on Drug Safety, 1(3), 229–241. PMID: 12149200
11. Popivanova, B. K., et al. (2008). Blocking TNF-α in mice reduces colorectal carcinogenesis associated with chronic colitis. Journal of Clinical Investigation, 118(2), 560–570. doi: 10.1172/JCI32453
12. Feldmann, M., & Maini, R. N. (2003). TNF defined as a therapeutic target for rheumatoid arthritis and other autoimmune diseases. Nature Medicine, 9(9), 1245–1250. doi.org/10.1038/nm939