Recombinant Human u-Plasminogen Activator/Urokinase, CF

Catalog # Availability Size / Price Qty
1310-SE-010
Recombinant Human u-Plasminogen Activator/Urokinase, CF Data
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Product Details
Citations (8)
FAQs
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Recombinant Human u-Plasminogen Activator/Urokinase, CF Summary

Purity
>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
Endotoxin Level
<1.0 EU per 1 μg of the protein by the LAL method.
Activity
Measured by its ability to cleave a peptide substrate, N-carbobenzyloxy-Gly-Gly-Arg-7-amido-4-methylcoumarin (Z-GGR-AMC). The specific activity is >2,000 pmol/min/µg, as measured under the described conditions.
Source
Mouse myeloma cell line, NS0-derived human u-Plasminogen Activator (uPA)/Urokinase protein
Met1-Leu431 with a C-terminal 10-His tag
Accession #
N-terminal Sequence
Analysis
Ser21, Ile179 & Lys156
Predicted Molecular Mass
18 kDa (long A chain), 3 kDa (short A chain), 30 kDa (B chain)
SDS-PAGE
18 kDa and 32 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.

1310-SE

Formulation Supplied as a 0.2 μm filtered solution in HEPES, 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, 0.01% (v/v) Tween® 20, pH 8.5
  • Recombinant Human u‑Plasminogen Activator (uPA)/Urokinase (rhuPA)  (Catalog # 1310-SE)
  • Substrate: Z-Gly-Gly-Arg-AMC (Bachem, Catalog # I-1140), 10 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 rhuPA to 1 ng/µL in Assay Buffer.
  2. Dilute Substrate to 200 µM in Assay Buffer.
  3. Load 50 µL of the 1 ng/µL rhuPA into a black well plate, and start the reaction by adding 50 µL of 200 µM Substrate. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of 200 µM Substrate without any rhuPA.
  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 (Sigma, Catalog # A-9891)

Per Well:
  • rhuPA: 0.05 µg
  • Substrate: 100 µM

Data Image

Enzyme Activity View Larger

Recombinant Human u-Plasminogen Activator/Urokinase (Catalog # 1310-SE) is measured by its ability to cleave a peptide substrate, N-carbobenzyloxy-Gly-Gly-Arg-7-amido-4-methylcoumarin (Z-GGR-AMC).

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: u-Plasminogen Activator (uPA)/Urokinase

uPA is a serine protease with an extremely limited substrate specificity, cleaving the sequence Cys-Pro-Gly-Arg560-Val561-Val-Gly-Gly-Cys in plasminogen to form plasmin (1). uPA is a potent marker of invasion and metastasis in a variety of human cancers associated with breast, stomach, colon, bladder, ovary, brain and endometrium (2). For example, the combination (both low vs. either or both high) of uPA and its inhibitor, plasminogen activator inhibitor-1 (PAI-1), outperforms the single factors as well as other traditional prognostic factors with regard to risk group assessment for breast cancer, particularly in node-negative breast cancer (3). The human uPA is initially synthesized as 431 amino acid precursor with a N-terminal signal peptide (20 residues) (4‑6). The single chain molecule is processed into a disulfide-linked two-chain molecule. The B chain starting at Ile179 corresponds to the catalytic domain. Two forms of the A chain exist, one starting at Ser21 (the long form) and the other at Lys156 (the short form). The resulting two-chain forms have different molecular weights (MW). The B chain is common for both forms whereas the long and short A chains are unique to the high and low MW forms, respectively. The long A chain contains an EGF-like domain, which is responsible for binding of the uPA receptor (uPAR). Both high and low MW forms exist in the purified recombinant human uPA.

References
  1. Ellis, V. (2004) in Handbook of Proteolytic Enzymes. Barrett, A.J. et al. eds., Academic Press, San Diego, pp.1677.
  2. Duffy, M.J. (2002) Biochem. Soc. Trans. 30:207.
  3. Harbeck, N. et al. (2002) Clin. Breast Cancer 3:196.
  4. Riccio, A. et al. (1985) Nucleic Acids Res. 13:2785.
  5. Nagai, M. et al. (1985) Gene 36:183.
  6. Jacobs, P. et al. (1985) DNA 4:139.
Long Name
Urokinase-type Plasminogen Activator
Entrez Gene IDs
5328 (Human); 18792 (Mouse)
Alternate Names
ATF; EC 3.4.21; EC 3.4.21.73; plasminogen activator, urokinase; PLAU; uPA; u-PA; uPlasminogen Activator; u-Plasminogen Activator; urinary; Urokinase; urokinase-type plasminogen activator

Citations for Recombinant Human u-Plasminogen Activator/Urokinase, 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.

8 Citations: Showing 1 - 8
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  1. Design, Synthesis, and Testing of Potent, Selective Hepsin Inhibitors via Application of an Automated Closed-Loop Optimization Platform.
    Authors: Pant S, Mukonoweshuro A, Desai B, Ramjee M, Selway C, Tarver G, Wright A, Birchall K, Chapman T, Tervonen T, Klefstrom J
    J Med Chem, 2018;61(10):4335-4347.
    Applications: Enzyme Assay
  2. Plasminogen-Dependent Collagenolytic Properties of Staphylococcus aureus in Collagen Gel Cultures of Human Corneal Fibroblasts
    Authors: K Sugioka, A Kodama-Tak, T Sato, K Okada, J Murakami, AM Park, H Mishima, Y Shimomura, S Kusaka, T Nishida
    Invest. Ophthalmol. Vis. Sci., 2018;59(12):5098-5107.
    Species: Human
    Sample Types: Whole Cells
    Applications: Bioassay
  3. Masked Chimeric Antigen Receptor for Tumor-Specific Activation
    Authors: X Han, PD Bryson, Y Zhao, GE Cinay, S Li, Y Guo, N Siriwon, P Wang
    Mol. Ther, 2017;25(1):274-284.
    Species: Human
    Sample Types: Whole Cells
    Applications: Bioassay
  4. Senescent peritoneal mesothelium creates a niche for ovarian cancer metastases
    Cell Death Dis, 2016;7(12):e2565.
    Species: Human
    Sample Types: Whole Cells
    Applications: Bioassay
  5. An acidic microenvironment sets the humoral pattern recognition molecule PTX3 in a tissue repair mode.
    Authors: Doni A, Musso T, Morone D, Bastone A, Zambelli V, Sironi M, Castagnoli C, Cambieri I, Stravalaci M, Pasqualini F, Laface I, Valentino S, Tartari S, Ponzetta A, Maina V, Barbieri S, Tremoli E, Catapano A, Norata G, Bottazzi B, Garlanda C, Mantovani A
    J Exp Med, 2015;212(6):905-25.
    Species: Mouse
    Sample Types: Protein
    Applications: Bioassay
  6. Imaging active urokinase plasminogen activator in prostate cancer.
    Authors: Lebeau A, Sevillano N, Markham K, Winter M, Murphy S, Hostetter D, West J, Lowman H, Craik C, VanBrocklin H
    Cancer Res, 2015;75(7):1225-35.
    Species: Human
    Sample Types: Cell Culture Supernates
    Applications: Bioassay
  7. Gelsolin induces colorectal tumor cell invasion via modulation of the urokinase-type plasminogen activator cascade.
    Authors: Zhuo, Jingli, Tan, Ee Hong, Yan, Benedict, Tochhawng, Lalchhan, Jayapal, Manikand, Koh, Shiuan, Tay, Hwee Kee, Maciver, Sutherla, Hooi, Shing Ch, Salto-Tellez, Manuel, Kumar, Alan Pre, Goh, Yaw Chon, Lim, Yaw Chyn, Yap, Celestia
    PLoS ONE, 2012;7(8):e43594.
    Species: N/A
    Sample Types: N/A
    Applications: ELISA (Standard)
  8. Loss of osteoclasts contributes to development of osteosarcoma pulmonary metastases.
    Authors: Endo-Munoz L, Cumming A, Rickwood D, Wilson D, Cueva C, Ng C, Strutton G, Cassady AI, Evdokiou A, Sommerville S, Dickinson I, Guminski A, Saunders NA
    Cancer Res., 2010;70(18):7063-72.
    Species: Mouse
    Sample Types: Whole Cells
    Applications: Bioassay

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