Recombinant Human Cathepsin D 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: 0.1 M NaOAc, 0.2 M NaCl, pH 3.5
• Recombinant Human Cathepsin D (rhCathepsin D) (Catalog # 1014-AS)
• Substrate: MCA-Pro-Leu-Gly-Leu-DPA-Ala-Arg-NH₂ (Catalog # ES001), 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 rhCathepsin D to 20 µg/mL in Assay Buffer.
2. Aliquot 50 µL of 20 µg/mL rhCathepsin D.
3. Incubate at 37 °C for 30 minutes.
4. Dilute incubated rhCathepsin D to 1 ng/µL in Assay Buffer.
5. Dilute Substrate to 60 µM in Assay Buffer.
6. Load 50 µL of 1 ng/µL rhCathepsin D in a plate, and start the reaction by adding 50 µL of 60 µM Substrate. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of 60 µM Substrate.
7. Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), 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).

• rhCathepsin D: 0.050 µg
• Substrate: 30 µM