Recombinant Mouse Carboxyl Ester Lipase/CEL Protein, CF

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

• Assay Buffer: 25 mM Tris, pH 7.0
• Reading Buffer: 25 mM Tris, pH 8.0
• Recombinant Mouse Carboxyl Ester Lipase/CEL (rmCEL) (Catalog # 5658-CE)
• Substrate: p-Nitrophenyl butyrate (PNPB) (Sigma, Catalog # N9876), 100 mM stock in acetone
• 96-well Clear Plate (Catalog # DY990)
• Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent

1. Dilute rmCEL to 0.2 ng/µL in Assay Buffer.
2. Dilute Substrate to 2 mM in Assay Buffer.
3. Load into the wells of a clear microplate 50 μL of 0.2 ng/µL rmCEL, and start the reaction by adding 50 µL of Substrate. Include a Blank Control containing 50 µL of Assay Buffer and 50 µL of Substrate without any rmCEL.
4. Incubate 10 minutes at room temperature.
5. Quickly add 100 µL of Reading Buffer to all wells (this will not stop the reaction).
6. Read immediately at a wavelength of 410 nm (bottom read) in endpoint mode.
7. Calculate specific activity:

     *Adjusted for Substrate Blank

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

• rmCEL: 0.010 µg
• Substrate: 0.5 mM