Recombinant Human Serpin F2/alpha 2-Antiplasmin Protein, CF

Catalog # Availability Size / Price Qty
1470-PI-010
Product Details
Citations (4)
FAQs
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Recombinant Human Serpin F2/alpha 2-Antiplasmin Protein, CF Summary

Purity
>95%, 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.
Activity
Measured by its ability to inhibit trypsin cleavage of a fluorogenic peptide substrate, Mca-RPKPVE-Nval-WRK(Dnp)-NH2 (Catalog # ES002). The IC50 value is <2.0 nM, as measured under the described conditions.
Source
Mouse myeloma cell line, NS0-derived human Serpin F2/alpha 2-Antiplasmin protein
Met28-Lys491, with a C-terminal 10-His tag
Accession #
N-terminal Sequence
Analysis
Met28
Predicted Molecular Mass
53 kDa
SDS-PAGE
67 kDa, reducing conditions

Product Datasheets

Carrier Free

What does CF mean?

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.

What formulation is right for me?

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.

1470-PI

Formulation Lyophilized from a 0.2 μm filtered solution in Tris and NaCl.
Reconstitution Reconstitute at 100 μg/mL in sterile 25 mM Tris and 150 mM NaCl, pH 7.5.
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.
  • 6 months from date of receipt, -20 to -70 °C as supplied.
  • 3 months, -20 to -70 °C under sterile conditions after reconstitution.

Assay Procedure

Materials
  • Assay Buffer: 50 mM Tris, 10 mM CaCl2, 150 mM NaCl, 0.05% (w/v) Brij-35, pH 7.5 (TCNB)
  • Recombinant Human Serpin F2/ alpha 2‑Antiplasmin (rhSerpin F2 (Catalog # 1470-PI)
  • Trypsin (Sigma, Catalog # T-1426)
  • Substrate: MCA-Arg-Pro-Lys-Pro-Val-Glu-NVAL-Trp-Arg-Lys(DNP)-NH2 (Catalog # ES002), 2 mM stock in DMSO
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute Trypsin to 0.25 µg/mL in Assay Buffer and KEEP ON ICE.
  2. Prepare a curve of rhSerpin F2 (MW: 53,056 Da) in Assay Buffer. Make the following serial dilutions: 600, 200, 100, 50, 30, 20, 15, 10, 5 and 1 nM.
  3. Combine equal volumes of 0.25 µg/mL Trypsin and rhSerpin F2 serial dilutions. Include two enzyme controls of equal volumes of Assay Buffer and 0.25 µg/mL Trypsin.
  4. Incubate reaction mixtures at 37 °C for 15 minutes.
  5. Dilute incubated reaction mixtures by 1/5 in Assay Buffer.
  6. Dilute Substrate to 20 µM with Assay Buffer.
  7. In a plate load 50 µL of the diluted reaction mixtures, and start the reaction by adding 50 µL of 20 µM Substrate to wells.
  8. Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively, in kinetic mode for 5 minutes.
  9. Derive the 50% inhibition concentration (IC50) for rhSerpin F2 by plotting RFU/min (or specific activity) vs. concentration with 4-PL fitting.
  10. The specific activity for Trypsin 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 MCA-Pro-Leu-OH (Bachem, Catalog # M-1975).

Per Well:
  • Trypsin: 0.00125 µg
  • rhSerpin F2 curve: 30, 10, 5, 2.5, 1.5, 1, 0.75, 0.5, 0.25 and 0.05 nM
  • Substrate: 10 µM
Reconstitution Calculator

Reconstitution Calculator

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Background: Serpin F2/alpha 2-Antiplasmin

Serpin F2 is a member of the Serpin superfamily and the primary physiological inhibitor of the serine protease plasmin, which is responsible for the dissolution of fibrin clots (1, 2). In addition to plasmin, Serpin F2 is also an efficient inhibitor of trypsin and chymotrypsin (3). Liver and kidney are major sites of Serpin F2 production and other tissues such as muscle, intestine, central nervous system, and placenta also express its mRNA at a moderate level. The tissue expression pattern of Serpin F2 indicates that it is a key regulator of plasmin-mediated proteolysis in these tissues (4). Human Serpin F2 is synthesized as a 491 amino acid precursor with a 27 amino acid signal peptide. The secreted protein has a short propeptide (residues 28-39) and a mature chain (residues 40-491). The presence of the propeptide did not affect its ability to inhibit plasmin but reduced its cross-linking ability to fibrin (5).

References
  1. Tone, M. et al. (1987) J. Biochem. 102:1033.
  2. Silverman, G.A. et al. (2001) J. Biol. Chem. 276:33293.
  3. Potempa, J. et al. (1988) Science 241:699.
  4. Menoud, P.-A. et al. (1996) J. Clin. Invest. 97:2478.
  5. Sumi, Y. et al. (1989) J. Biochem. 106:703.
Entrez Gene IDs
5345 (Human); 18816 (Mouse)
Alternate Names
AAPclade F (alpha-2 antiplasmin; alpha 2-Antiplasmin; Alpha-2-AP; Alpha-2-PI; Alpha-2-plasmin inhibitor; pigment epithelium derived factor), member 2; Serpin F2; serpin peptidase inhibitor, clade F (alpha-2 antiplasmin, pigment epitheliumderived factor), member 2

Citations for Recombinant Human Serpin F2/alpha 2-Antiplasmin 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.

4 Citations: Showing 1 - 4
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  1. Lowering Low-Density Lipoprotein Particles in Plasma Using Dextran Sulphate Co-Precipitates Procoagulant Extracellular Vesicles
    Authors: JW Wang, YN Zhang, SK Sze, SM van de Weg, F Vernooij, AH Schoneveld, SH Tan, HH Versteeg, L Timmers, CSP Lam, DPV de Kleijn
    Int J Mol Sci, 2017;19(1):.
    Applications: Bioassay
  2. Extracellular Vesicle Proteins Associated with Systemic Vascular Events Correlate with Heart Failure: An Observational Study in a Dyspnoea Cohort.
    Authors: Zhang Y, Vernooij F, Ibrahim I, Ooi S, Gijsberts C, Schoneveld A, Sen K, den Ruijter H, Timmers L, Richards A, Jong C, Mazlan I, Wang J, Lam C, de Kleijn D
    PLoS ONE, 2016;11(1):e0148073.
    Species: Human
    Sample Types: Plasma
    Applications: ELISA Developmet
  3. Enzymatic properties of human kallikrein-related peptidase 12 (KLK12).
    Authors: Memari&lt;/LastName&gt;&lt;ForeNam N&lt;/Initial, Memari N, Jiang W, Diamandis EP, Luo LY
    Biol. Chem., 2007;388(4):427-35.
    Species: Human
    Sample Types: Recombinant Protein
    Applications: Enzyme Assay
  4. 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;449(1):164-70.
    Species: N/A
    Sample Types: Protein
    Applications: Enzyme Assay

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Fluorogenic Peptide Substrates

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