Recombinant Human Carbonic Anhydrase I 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: 12.5 mM Tris, 75 mM NaCl, pH 7.5
• Recombinant Human Carbonic Anhydrase I/CA1 (rhCA1) (Catalog # 2180-CA)
• Substrate: 4-Nitrophenyl acetate (4-NPA) (Sigma, Catalog # N8130), 100 mM stock in acetone
• 96-well Clear Plate (Costar, Catalog # 92592)
• Plate reader (Model: SpectraMax Plus by Molecular Devices) or equivalent

1. Dilute rhCA1 to 100 ng/µL in Assay Buffer.
2. Dilute Substrate to 2 mM in Assay Buffer.
3. Load 50 µL of 100 ng/µL rhCA1 into a plate, and start the reaction by adding 50 µL of 2 mM Substrate.
4. Include a Substrate Blank containing 50 µL of Assay Buffer and 50 µL of Substrate.
5. Read at a wavelength of 400 nm (bottom read) in kinetic mode for 5 minutes.
6. Calculate specific activity:

     *Adjusted for Substrate Blank
     **Derived using calibration standard 4-Nitrophenol (Sigma, Catalog # 241326).

• rhCA1: 5 µg
• Substrate: 1 mM