Recombinant Human Active Coagulation Factor XIV/ProteinC, CF

Catalog # Availability Size / Price Qty
4998-SE-010
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Recombinant Human Active Coagulation Factor XIV/ProteinC, 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 cleave the fluorogenic peptide substrate Boc-VPR-AMC (Catalog # ES011). The specific activity is >275 pmol/min/µg, as measured under the described conditions.
Source
Chinese Hamster Ovary cell line, CHO-derived human Coagulation Factor XIV/Protein C protein
Ala43-Pro461, with a C-terminal 10-His tag
The isolated protein was activated by thrombin/thrombomodulin complex and further purified.
Accession #
N-terminal Sequence
Analysis
Ala43 (light chain) & Leu212 (heavy chain)
Structure / Form
Disulfide-linked heterodimer
Predicted Molecular Mass
19 kDa (light chain), 29 kDa (heavy chain)
SDS-PAGE
22-25 kDa and 40-50 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.

4998-SE

Formulation Supplied as a 0.2 μm filtered solution in Tris, NaCl and CaCl2.
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 opening.

Assay Procedure

Materials
  • Assay Buffer: 50 mM Tris, 0.15 M NaCl, 10 mM CaCl2, 0.01% (w/v) Brij 35, pH 8.5
  • Recombinant Human Active Coagulation Factor XIV/Protein C (rhPROC) (Catalog # 4998-SE)
  • Substrate: Boc-Val-Pro-Arg-Amc (Catalog # ES011)
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute rhPROC to 2 ng/µL in Assay Buffer.
  2. Dilute Substrate to 200 µM in Assay Buffer.
  3. Load into plate 50 µL of 2 ng/µL rhPROC, and start the reaction by adding 50 µL of 200 μM Substrate. As a Substrate Blank combine 50 µL of 200 μM Substrate and 50 µL of Assay Buffer.
  4. Read at excitation and emission wavelengths of 380 nm and 460 nm (top read), respectively in kinetic mode for 5 minutes.
  5. Calculate specific activity:

     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).

Per Well:
  • rhPROC: 0.1 µg
  • Substrate: 100 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: Coagulation Factor XIV/Protein C

Protein C (PROC) is a vitamin K-dependent serine protease synthesized in liver as a single-chain precursor (1). The N-terminus consists of a signal peptide (aa 1-32) and a propeptide (aa 33-42). The mature chain (aa 43-461) is converted to two disulfide-linked chains (light: aa 43-199 and heavy: 200‑461) and both forms are inactive. The light chain consists of Gla (gamma-carboxy-glutamate) domain and two EGF-like domains. The heavy chain consists of an activation peptide (aa 200‑211) and serine protease domain (aa 212-450). Present in plasma at 3 to 5 mg/L, PROC plays a key role in anticoagulation. Physiologically, the inactive forms of PROC are converted to the active form by thrombin, which releases the activation peptide. The active PROC cleaves factors VIIIa and Va to inactivate them. This anticoagulation activity can be enhanced by a presence of a cofactor such as Protein S. In hereditary thrombophilia, PROC deficiency is caused by a genetic mutation which affects PROC activity. A severe recessive form may result in massive thrombosis fatal to patient.

References
  1. Shen, L. and Dahlbäck, B. (2004) in Handbook of Proteolytic Enzymes, Barrett, A.J. et al. eds. pp. 1673.
Long Name
Vitamin K-dependent Protein C
Entrez Gene IDs
5624 (Human); 19123 (Mouse); 25268 (Rat)
Alternate Names
Anticoagulant protein C; APC; Autoprothrombin IIA; Blood coagulation factor XIV; Coagulation Factor XIV; EC 3.4.21; EC 3.4.21.69; PC; PROC; PROC1; protein C (inactivator of coagulation factors Va and VIIIa); Protein C; vitamin K-dependent protein C

FAQs

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Fluorogenic Peptide Substrates

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