Recombinant Human t-Plasminogen Activator/tPA Protein, CF

• 6 months from date of receipt, -20 to -70 °C as supplied.
• 3 months, -20 to -70 °C under sterile conditions after opening.

• Assay Buffer: 50 mM Tris, 0.01% Tween-20, pH 8.5
• Recombinant Human t‑Plasminogen Activator/tPA  (rhPLAT) (Catalog # 7449-SE)
• Fluorogenic Peptide 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 rhPLAT to 2 ng/µL in Assay Buffer.
2. Dilute Substrate to 200 µM in Assay Buffer.
3. Load 50 µL of the 2 ng/µL rhPLAT 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 rhPLAT.
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:

     *Adjusted for Substrate Blank
     **Derived using calibration standard 7-Amino, 4-Methyl Coumarin (AMC) (Sigma, Catalog # A9891).

• rhPLAT: 0.1 µg
• Substrate: 100 µM