Recombinant Rat MMP-8 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, 10 mM CaCl₂, 150 mM NaCl, 0.05% (v/v) Brij-35, pH 7.5 (TCNB)
• Recombinant Rat MMP‑8  (rrMMP-8) (Catalog # 3245-MP)
• p-aminophenylmercuric acetate (APMA) (Sigma, Catalog # A-9563), 100 mM stock in DMSO
• ZnCl₂, 1 M 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. Dilute rrMMP-8 to 100 µg/mL in Assay Buffer.
2. Activate rrMMP-8 by adding APMA and ZnCl₂ to final concentrations of 1 mM and 25 µM respectively.
3. Incubate at room temperature for 4 hours.
4. Dilute activated rrMMP-8 to 2 ng/μL in Assay Buffer.
5. Dilute Substrate to 20 µM in Assay Buffer.
6. Load 50 µL of the 2 ng/µL rrMMP-8 into in plate, and start the reaction by adding 50 µL of 20 µM Substrate. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of 20 µM Substrate.
7. Read at excitation and emission wavelengths of 320 nm and 405 nm, respectively, in kinetic mode for 5 minutes.
8. Calculate specific activity:

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

• rrMMP-8: 0.10 µg
• Substrate: 10 µM