Recombinant Human Trappin-2/Elafin Protein, CF Summary
Product Specifications
Ala23-Gln117, with a C-terminal 10-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.
1747-PI
| Formulation | Supplied as a 0.2 μm filtered solution in Tris and NaCl. |
| 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
- Activation Buffer: 50 mM MES, 50 mM NaCl, pH 5.5
- Assay Buffer: 50 mM Tris, 1.0 M NaCl, 0.05% (w/v) Brij-35, pH 7.5
- Recombinant Human Trappin-2/Elafin (rhTrappin-2) (Catalog # 1747-PI)
- Recombinant Mouse Neutrophil Elastase/ELA2 (rmELA2) (Catalog # 4517-SE)
- Recombinant Mouse Active Cathepsin C/DPPI (rmCathepsin C) (Catalog # 2336-CY)
- Substrate: MeOSuc-Ala-Ala-Pro-Val-AMC (Bachem, Catalog # I-1270), 10 mM stock in DMSO
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Dilute rmELA2 to 50 µg/mL in Activation Buffer containing 50 μg/mL rmCathepsin C.
- Incubate 50 µg/mL rmELA2 at 37 °C for 2 hours to activate.
- Prepare a dilution curve of rhTrappin-2 (MW: 16,000 Da) in Assay Buffer. Make serial dilutions of: 3200, 1600, 800, 600, 400, 200, 100, and 10 nM.
- Dilute activated rmELA2 to 10 µg/mL with Assay Buffer.
- Combine each dilution of rhTrappin-2 with rmELA2 in equal volumes. Include two rmELA2 controls containing equal volumes of rmELA2 and Assay Buffer without any rhTrappin-2.
- Incubate reactions at room temperature for 10 minutes.
- Dilute each reaction five-fold with Assay Buffer.
- Dilute Substrate to 200 µM in Assay Buffer.
- Load 50 µL of the diluted incubated curve into a black well plate, and start the reaction by adding 50 µL of 200 µM Substrate.
- Read at excitation and emission wavelengths of 380 nm and 460 nm (top read), respectively, in kinetic mode for 5 minutes.
- Derive the 50% inhibiting concentration (IC50) for rhTrappin-2 by plotting RFU/min (or specific activity) vs. concentration with 4-PL fitting.
- The specific activity of rmELA2 at each point may be determined using the following equation (if desired):
|
Specific Activity (pmol/min/µg) = |
Adjusted Vmax* (RFU/min) x Conversion Factor** (pmol/RFU) |
| amount of enzyme (µg) |
*Adjusted for Substrate Blank
**Derived using calibration standard 7-Amino, 4-Methyl Coumarin (AMC) (Sigma, Catalog # A-9891).
- rhTrappin-2: 160, 80, 40, 30, 20, 10, 5 and 0.5 nM
- rmELA2: 0.05 µg
- Substrate: 100 µM
Reconstitution Calculator
Background: Trappin-2/Elafin
Trappin-2 is the human member of the trappin gene family that contains SLPI (1). Trappin-2 consists of an N-terminal transglutaminase substrate domain (residues 23‑60) and a C-terminal four-disulfide core or whey acidic protein (WAP) domain (residues 72‑117). Elafin or ESI (elastase-specific inhibitor) and SKALP (skin‑derived anti‑leucoproteinase) are alternative names for Trappin-2 and reflect its protease targets. However, elafin and SKALP sometimes correspond only to the processed form that contains the C-terminal WAP domain of the molecule, which can be isolated naturally. The recombinant human Trappin-2 corresponds to the full‑length form (residues 23‑117), which migrates as two protein bands under SDS-PAGE due to an unidentified mechanism. In addition to its ability to inhibit human neutrophil elastase, it can also be used as a substrate for transglutaminases.
Citations for Recombinant Human Trappin-2/Elafin 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.
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Citations: Showing 1 - 6
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Concerted actions by MMPs, ADAMTS and serine proteases during remodeling of the cartilage callus into bone during osseointegration of hip implants
Authors: J Cassuto, A Folestad, J Göthlin, H Malchau, J Kärrholm
Bone Rep, 2020-09-11;13(0):100715.
Species: Human
Sample Types: Plasma
Applications: ELISA Detection -
Elafin drives poor outcome in high-grade serous ovarian cancers and basal-like breast tumors.
Authors: Labidi-Galy S, Clauss A, Ng V, Duraisamy S, Elias K, Piao H, Bilal E, Davidowitz R, Lu Y, Badalian-Very G, Gyorffy B, Kang U, Ficarro S, Ganesan S, Mills G, Marto J, Drapkin R
Oncogene, 2014-01-27;34(3):373-83.
Species: Human
Sample Types: Whole Cells
Applications: Bioassay -
Trappin-2/elafin modulate innate immune responses of human endometrial epithelial cells to PolyI:C.
Authors: Drannik AG, Nag K, Yao XD, Henrick BM, Sallenave JM, Rosenthal KL
PLoS ONE, 2012-04-24;7(4):e35866.
Species: Human
Sample Types: Buffer
Applications: Enzyme Assay -
Anti-HIV-1 activity of elafin is more potent than its precursor's, trappin-2, in genital epithelial cells.
Authors: Drannik AG, Nag K, Yao XD, Henrick BM, Jain S, Ball TB, Plummer FA, Wachihi C, Kimani J, Rosenthal KL
J. Virol., 2012-02-15;86(8):4599-610.
Species: Human
Sample Types: Vaginal Lavage Fluid
Applications: Bioassay -
Trappin-2 promotes early clearance of Pseudomonas aeruginosa through CD14-dependent macrophage activation and neutrophil recruitment.
Authors: Wilkinson TS, Dhaliwal K, Hamilton TW, Lipka AF, Farrell L, Davidson DJ, Duffin R, Morris AC, Haslett C, Govan JR, Gregory CD, Sallenave JM, Simpson AJ
Am. J. Pathol., 2009-03-05;174(4):1338-46.
Species: Mouse
Sample Types: In Vivo
Applications: In Vivo -
Mannan-binding lectin-associated serine protease 3 cleaves synthetic peptides and insulin-like growth factor-binding protein 5.
Authors: Cortesio CL, Jiang W
Arch. Biochem. Biophys., 2006-03-03;449(1):164-70.
Applications: Enzyme Assay
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