Recombinant Human Tumor Necrosis Factor (TNF) is produced in E. coli and includes a C-terminal 6xHis tag. This partially expressed protein corresponds to amino acids 77-233 and achieves a high purity of over 95% as determined by SDS-PAGE analysis. It demonstrates biological activity with an ED50 of 10-40 pg/ml in a cytotoxicity assay using L929 mouse fibroblast cells. The endotoxin level is maintained below 1.0 EU/µg, verified by the LAL method.
TNF appears to be one of the most critical cytokines involved in systemic inflammation and serves as part of the body's immune response. The protein plays what seems to be a key role in regulating immune cells and inducing apoptotic cell death, inflammation, and inhibition of tumorigenesis. Given its central position in inflammatory processes, TNF has become an important molecule in the study of inflammatory diseases and cancer research. This makes it a valuable target for therapeutic interventions, though the complexity of its signaling pathways may present challenges.
Potential Applications
Note: The applications listed below are based on what we know about this protein's biological functions, published research, and experience from experts in the field. However, we haven't fully tested all of these applications ourselves yet. We'd recommend running some preliminary tests first to make sure they work for your specific research goals.
1. Cytotoxicity Assays for TNF Pathway Research
This recombinant human TNF protein works well as a positive control or standard in cytotoxicity assays for studying TNF-mediated cell death pathways. The established ED50 of 10-40 pg/ml using L-929 mouse fibroblast cells provides a validated reference point for dose-response studies. Researchers can apply this protein to investigate the mechanisms of TNF-induced apoptosis and necrosis across various cell lines. The high purity and low endotoxin levels should ensure reliable and reproducible results in cell-based assays, though individual cell types may respond differently.
2. Anti-TNF Antibody Development and Validation
The C-terminal 6xHis tag makes it relatively straightforward to immobilize this TNF protein onto nickel-coated surfaces for antibody screening and characterization studies. This recombinant protein can serve as an antigen for generating monoclonal or polyclonal antibodies against human TNF. Since the biological activity has been confirmed, antibodies developed against this protein will likely recognize the native, functional form of TNF. Researchers can then use this protein in ELISA, Western blot, and other immunoassays to validate antibody specificity and binding affinity.
3. TNF Receptor Binding Studies
This biologically active TNF protein works well in receptor binding assays to study the interaction between TNF and its receptors TNFR1 and TNFR2. The confirmed biological activity suggests proper protein folding and functional receptor binding capability. Researchers can apply this protein in competition binding assays, surface plasmon resonance studies, or flow cytometry-based binding experiments. The His-tag allows for easy purification and detection in pull-down assays to identify TNF-interacting proteins, though some binding partners may be context-dependent.
4. Inflammatory Response Modeling in Cell Culture
This recombinant TNF can stimulate inflammatory responses in primary cells or cell lines for mechanistic studies of inflammation. The validated cytotoxic activity demonstrates the protein's ability to trigger TNF signaling pathways, making it suitable for studying downstream inflammatory mediator production. Researchers can use this protein to investigate NF-κB activation, cytokine cascades, and other inflammatory signaling mechanisms in controlled in vitro systems. The low endotoxin content helps ensure that observed effects are specifically due to TNF activity rather than bacterial contamination, though some cellular responses may still vary between experimental conditions.
5. Drug Screening and Inhibitor Development
This biologically active TNF protein serves as a useful tool for screening potential TNF inhibitors or modulators in preclinical research. The established cytotoxicity assay parameters provide a standardized platform for evaluating the efficacy of small molecules or biologics that target TNF signaling. Researchers can apply this protein in high-throughput screening assays to identify compounds that block TNF-induced cell death or inflammatory responses. The His-tag makes protein handling and assay development easier for automated screening platforms, though hit validation will likely require additional confirmation in more complex systems.
