Recombinant Human Kallikrein 13 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.

• Activation Buffer: 0.1 M Tris, pH 8.0
• Assay Buffer: 50 mM Tris, 150 mM NaCl, 0.05% Brij-35 (v/v), pH 8.0
• Recombinant Human Kallikrein 13 (rhKLK13) (Catalog # 2625-SE)
• Lysyl-Endopeptidase (Wako BioProducts, Catalog # 129-02541)
• Substrate: Boc-Val-Pro-Arg-AMC (Catalog # ES011), 100 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 rhKLK13 to 100 µg/mL in Activation Buffer.
2. Dilute Lysyl-Endopeptidase to 0.8 µg/mL in Activation Buffer.
3. Activate rhKLK13 by adding 1 µL of 0.8 µg/mL Lysyl-Endopeptidase to 40 µL of 100 µg/mL rhKLK13 for a final concentration of 0.02 µg/mL for Lysyl-Endopeptidase.
4. Incubate at 37 °C for 30 minutes.
5. Dilute activated rhKLK13 to 2 ng/µL in Assay Buffer.
6. Dilute Substrate to 200 µM in Assay Buffer.
7. Load 50 µL of 2 ng/µL rhKLK13 in a plate, and start the reaction by adding 50 µL of 200 µM Substrate. Include a Substrate Blank containing 50 µL of Assay Buffer and 50 µL of 200 uM Substrate.
8. Read at excitation and emission wavelengths of 380 nm and 460 nm (top read), respectively in kinetic mode for 5 minutes.
9. Calculate specific activity:

     *Adjusted for Substrate Blank
     **Derived using calibration standard 7-Amino, 4-Methyl Coumarin (AMC) (Sigma, Catalog # A-9891).

• rhKLK13: 0.1 µg
• Substrate: 100 µM