Recombinant Human Kallikrein 7 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: 50 mM Tris, 10 mM CaCl₂, 150 mM NaCl, 0.05% (w/v) Brij-35, pH 7.5 (TCNB)
• Assay Buffer: 50 mM Tris, 150 mM NaCl, pH 8.5
• Recombinant Human Kallikrein 7 (rhKLK7) (Catalog # 2624-SE)
• Bacterial Thermolysin (Thermolysin) (Catalog # 3097-ZN)
• EDTA (Sigma, Catalog # E-4884)
• Substrate: MCA-Arg-Pro Lys-Pro-Val-Glu-Nval-Trp-Arg-Lys(Dnp)-NH₂ (Catalog # ES002)
• F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
• Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent

1. Dilute rhKLK7 to 200 μg/mL in Activation Buffer.
2. Dilute Thermolysin to 20 μg/mL in Activation Buffer.
3. Combine equal volumes of 200 μg/mL rhKLK7 and 20 μg/mL Thermolysin.
4. Incubate at 37 °C for 2 hours.
5. Stop reaction with an equal volume 100 mM EDTA diluted in Assay Buffer (C_f = 50 mM).
6. Dilute activated rhKLK7 to 4 ng/μL in Assay Buffer.
7. Dilute Substrate to 20 μM in Assay Buffer.
8. Load 50 μL of 4 ng/μL rhKLK7 in a plate, and start the reaction by adding 50 μL of 20 μM Substrate to each well. Include a Substrate Blank containing 50 μL Assay Buffer and 50 μL of 20 μM Substrate.
9. Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively in kinetic mode for 5 minutes.
10. Calculate specific activity:

     *Adjusted for Substrate Blank

     **Derived using calibration standard MCA-Pro-Leu-OH (Bachem, Catalog # M-1975).

• rhKLK7: 0.2 μg
• Substrate: 10 μM