Recombinant Human MMP-14/MT1-MMP (NS0-expressed) Protein, CF

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

• Assay Buffer: 50 mM Tris, 0.15 M NaCl, 10 mM CaCl₂, 5 µM ZnCl₂, 0.05% (w/v) Brij-35, pH 7.5
• Recombinant Human MMP‑14/MT1‑MMP (rhMMP-14) (Catalog # 918-MPN)
• Recombinant Human Trypsin 3 (rhTrypsin 3) (Catalog # 3714-SE)
• AEBSF (Catalog # 5175), 100 mM stock in deionized water
• Substrate: MCA-Lys-Pro-Leu-Gly-Leu-DPA-Ala-Arg-NH₂ (Catalog # ES010), 2 mM stock in DMSO
• F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
• Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent

1. Activate rhMMP-14 at 100 µg/mL with 0.1 µg/mL rhTrypsin 3 in Assay Buffer.
2. Incubate at 37 °C for 1 hour.
3. Stop reaction with 1 mM AEBSF and incubate for 15 minutes at room temperature.
4. Dilute activated rhMMP-14 to 1 µg/mL in Assay Buffer.
5. Dilute Substrate to 80 µM in Assay Buffer.
6. In a plate, load 50 µL of 1 µg/mL rhMMP-14 to wells.  For a Substrate Blank, load 50 µL of Assay Buffer.
7. Start the reaction by adding 50 µL of 80 µM Substrate to wells.
8. Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively, in kinetic mode for 5 minutes.
9. Calculate specific activity:

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

• rhMMP-14: 0.050 µg
• Substrate: 40 µM