Recombinant Tetanus Neurotoxin Type Light Chain Protein, CF
Recombinant Tetanus Neurotoxin Type Light Chain Protein, CF Summary
Product Specifications
Pro2-Gly430, with an N-terminal Met and 6-His tag
Analysis
Product Datasheets
Carrier Free
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.
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.
6535-ZN
| Formulation | Supplied as a 0.2 μm filtered solution in Tris, NaCl and Tween® 20. |
| 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.
|
Assay Procedure
- Assay Buffer: 50 mM HEPES, 0.05% (v/v) Tween® 20, pH 7.5
- Recombinant C. tetani TeNT Light Chain (rTeNT-LC) (Catalog # 6535-ZN)
- Substrate: Recombinant GFP/SNAP25B/VAMP-2 (Catalog # 7375-SV)
- SDS-PAGE and silver stain reagents or equivalent or Western blot with appropriate antibodies
- Dilute Substrate to 100 µg/mL in Assay Buffer.
- Dilute rTeNT-LC to 4 µg/mL in Assay Buffer.
- Combine equal volumes of diluted Substrate with diluted rTeNT-LC. Prepare two controls by combining equal volumes of diluted Substrate with Assay Buffer.
- Incubate reaction vials at 37 °C for 1 hour. Incubate one control at 37 °C and the other at -20 °C for 1 hour.
- After incubation, combine reaction mixtures and controls with reducing SDS-PAGE sample buffer at a 1:1 (reaction mixture:sample buffer) ratio (v/v) to stop reactions.
- Analyze the cleavage products by SDS-PAGE (Load 40 µL of the mixture from step 5 per lane, 1 µg Substrate per lane) followed by silver staining and/or Western blot.
- rTeNT-LC: 40 ng
- Substrate: 1 µg
Reconstitution Calculator
Background: TeNT Light Chain
Tetanus toxin (TeNT) is a potent neurotoxin produced by Clostridium tetani. TeNT is similar in structure and function to the botulinum toxins produced by other Clostridium species (1, 2). TeNT is among the most toxic protein toxins known for humans. TeNT is synthesized as an inactive single chain precursor that undergoes proteolytic cleavage to create light and heavy chains that are linked by a disulfide bond. The 50 kDa light chain contains a metalloprotease domain whereas the 100 kDa heavy chain contains a receptor binding domain and a domain required for translocation across the cell membrane (3). Once inside a neuronal cell the zinc metalloprotease domain is able to cleave synaptobrevin to cause motor neuron disinhibition, resulting in the spastic paralysis characteristic of tetanus (4). The E. coli expressed recombinant TeNT light chain is an active protease. In the absence of heavy chains, however, it lacks toxicity because it cannot enter into host cells.
- Eisel, U. et al. (1986) EMBO J. 5:2495.
- Montecucco, C. and S. Giampietro (1993) Trends Biochem. Sci. 18:324.
- Schiavo, G. et al. (1992) Nature 359:832.
- Schiavo, G. et al. (1992) EMBO J. 11:3577.
Product Specific Notices
Coomassie is a registered trademark of Imperial Chemical Industries Ltd. Tween is a registered trademark of ICI Americas.Citation for Recombinant Tetanus Neurotoxin Type Light Chain Protein, CF
R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.
1 Citation: Showing 1 - 1
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Postsynaptic synucleins mediate endocannabinoid signaling
Authors: Albarran, E;Sun, Y;Liu, Y;Raju, K;Dong, A;Li, Y;Wang, S;S�dhof, TC;Ding, JB;
Nature neuroscience
Species: N/A, Mouse
Sample Types: Recombinant Protein, Whole Tissue
Applications: Bioassay
FAQs
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