Recombinant Human MMP-1 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% (w/v) Brij-35, pH 7.5 (TCNB)
• Recombinant Human MMP-1 (rhMMP-1) (Catalog # 901-MP)
• p-aminophenylmercuric acetate (APMA), (Sigma, Catalog # A-9563), 100 mM stock in DMSO
• Substrate: MCA-Lys-Pro-Leu-Gly-Leu-DPA-Ala-Arg-NH₂ (R&D Systems, Catalog # ES010)
• F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
• Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent

1. Dilute rhMMP-1 to 50 µg/mL in Assay Buffer.
2. Activate 50 µg/mL rhMMP-1 by adding APMA to a final concentration of 1 mM.
3. Incubate at 37 °C for 2 hours.
4. Dilute activated rhMMP-1 to 1 ng/µL in Assay Buffer.
5. Dilute Substrate to 20 µM in Assay Buffer.
6. Load into a black well plate 50 µL of 1 ng/µL rhMMP-1 and start the reaction by adding 50 µL of 20 µM Substrate. Include a Substrate Blank containing 50 µL Assay Buffer, 50 µL Substrate, and no rhMMP-1.
7. Read at excitiation 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).

• rhMMP-1: 0.050 µg
• Substrate: 10 µM