Serpin A4 is a member of the Serpin superfamily of the serine protease inhibitors (1). Known also as kallistatin or proteinase inhibitor 4, Serpin A4 has been shown to function as a specific inhibitor of tissue kallikrein and as an inhibitor of angiogenesis and tumor growth (2, 3).
Recombinant Human Serpin A4/Kallistatin Protein, CF
R&D Systems | Catalog # 1669-PI
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Key Product Details
- R&D Systems NS0-derived Recombinant Human Serpin A4/Kallistatin Protein (1669-PI)
- Quality control testing to verify active proteins with lot specific assays by in-house scientists
- All R&D Systems proteins are covered with a 100% guarantee
Source
NS0
Accession Number
Applications
Inhibition Activity
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Product Specifications
Source
Mouse myeloma cell line, NS0-derived human Serpin A4/Kallistatin protein
Gln21-Pro427, with a C-terminal 10-His tag
Gln21-Pro427, with a C-terminal 10-His tag
Purity
>90%, by SDS-PAGE under reducing conditions and visualized by silver stain.
Endotoxin Level
<1.0 EU per 1 μg of the protein by the LAL method.
N-terminal Sequence Analysis
No results obtained: Gln21 predicted
Predicted Molecular Mass
48 kDa
SDS-PAGE
57 kDa and 61 kDa, reducing conditions
Activity
Measured by its ability to inhibit KLK1 cleavage of a fluorogenic peptide substrate, Pro-Phe-Arg-7-amido-4-methylcoumarin (PFR-AMC).
The IC50 value is <40 nM, as measured under the described conditions.
The IC50 value is <40 nM, as measured under the described conditions.
Reviewed Applications
Read 3 reviews rated 4.7 using 1669-PI in the following applications:
Formulation, Preparation, and Storage
1669-PI
| Formulation | Lyophilized from a 0.2 μm filtered solution in MES and NaCl. |
| Reconstitution | Reconstitute at 100 μg/mL in sterile 50 mM Tris, 10 mM CaCl2 and 150 mM NaCl, pH 7.5. |
| 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.
|
Calculators
Background: Serpin A4/Kallistatin
References
- Silverman, G.A. et al. (2001) J. Biol. Chem. 276:33293.
- Chen, V.C. et al. (2000) J. Biol. Chem. 275:40371.
- Miao, R.Q. et al. (2002) Blood 100:3245.
Alternate Names
Kallistatin
Entrez Gene IDs
5267 (Human)
Gene Symbol
SERPINA4
UniProt
Additional Serpin A4/Kallistatin Products
Product Documents for Recombinant Human Serpin A4/Kallistatin Protein, CF
Certificate of Analysis
To download a Certificate of Analysis, please enter a lot or batch number in the search box below.
Note: Certificate of Analysis not available for kit components.
Product Specific Notices for Recombinant Human Serpin A4/Kallistatin Protein, CF
For research use only
Citations for Recombinant Human Serpin A4/Kallistatin Protein, CF
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4.7 out of 5
3 Customer Ratings
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Application: Enzymatic activity in vitroVerified Customer | Posted 07/02/2018
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Protocols
View specific protocols for Recombinant Human Serpin A4/Kallistatin Protein, CF (1669-PI):
Materials
- Activation Buffer: 50 mM Tris, 10 mM CaCl2, 150 mM NaCl, pH 7.5 (TCN)
- Inhibition Buffer: 25 mM Tris, 150 mM NaCl, pH 7.5
- Assay Buffer: 50 mM CHES, 250 mM NaCl, pH 10.0
- Recombinant Human Serpin A4/Kallistatin (rhSerpin A4) (Catalog # 1669-PI)
- Recombinant Human Kallikrein 1 (rhKLK1) (Catalog # 2337-SE)
- Bacterial Thermolysin (Thermolysin) (Catalog # 3097-ZN)
- 1,10-Phenanthroline (Sigma, Catalog # 320056), 0.6 M stock in DMSO
- Substrate: Pro-Phe-Arg-7-amido-4-methylcoumarin (Bachem, Catalog # I-1295), 10 mM stock in DMSO
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Activate rhKLK1 at 100 µg/mL with 1 µg/mL Thermolysin in Activation Buffer.
- Incubate at 37 °C for 1 hour.
- Add 1,10-Phenathroline at a final concentration of 10 mM to stop activation reaction.
- Prepare a curve of rhSerpin A4 (47710 Da) in Inhibition Buffer. Make the following serial dilutions: 8000, 4000, 2000, 1000, 625, 415, 200, and 75 nM.
- Dilute activated rhKLK1 to 5 µg/mL in Inhibition Buffer.
- Combine 20 µL of 5 µg/mL rhKLK1 with 20 µL of rhSerpin A4 curve dilutions. Include two controls of 20 µL Inhibition Buffer with 20 µL of 5 µg/mL rhKLK1.
- Incubate curve at 37 °C for 1 hour.
- Dilute curve 12.5 fold by adding 460 µL of Assay Buffer to each dilution.
- Dilute Substrate to 200 µM in Assay Buffer.
- In a plate load 50 µL of the rhSerpin A4 curve, and start the reaction by adding 50 µL of 200 µM Substrate to wells.
- 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) of rhSerpin A4 by plotting RFU/min (or specific activity) vs. concentration with 4-PL fitting.
- The specific activity for rhKLK1 at each point may be determined using the following formula (if needed):
|
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 (Sigma, Catalog # A-9891).
Per Well:
- rhKLK1: 0.010 µg
- rhSerpin A4 curve: 160, 80, 40, 20, 12.5, 8.3, 4 and 1.5 nM
- Substrate: 100 µM
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