Human embryonic kidney cell, HEK293-derived human PON1 protein Leu16-Leu355 with an N-terminal 6-His tag and a C-terminal Fc tag
>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
<1.0 EU per 1 μg of the protein by the LAL method.
N-terminal sequence Analysis
Predicted Molecular Mass
69-79 kDa, under reducing conditions
Measured by its ability to hydrolyze phenyl acetate. The specific activity is >5000 pmol/min/μg, as measured under the described conditions.
Recombinant Human PON1 His-tag Fc Chimera Protein, CF Scientific Data Examples
Recombinant Human PON1 His-tag Fc Chimera Protein Enzyme Activity
Recombinant Human PON-1 His-tag Fc Chimera (Catalog # 10175-PO) is measured by its ability to hydrolyze phenyl acetate. The activity (orange) is over 3-fold higher than the competitor's PON-1 (green).
Recombinant Human PON1 His-tag Fc Chimera Protein SDS-PAGE
2 μg/lane of Recombinant Human PON1 His-tag Fc Chimera was resolved with SDS-PAGE under reducing (R) and non-reducing (NR) conditions and visualized by Coomassie® blue staining, showing a band at ~74 kDa under reducing conditions.
Formulation, Preparation and Storage
What does CF mean?
CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our
Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant
protein to be stored at a more dilute concentration.
The carrier free version does not contain BSA.
What formulation is right for me?
In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or
as an ELISA standard.
In contrast, the carrier free protein is recommended for applications, in which the presence of BSA
Supplied as a 0.2 μm filtered solution in Tris, CaCl2, NaCl, and Glycerol.
The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage:
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
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, 5 mM CaCl2, pH 8.0
Recombinant Human PON1 His-tag Fc Chimera (rhPON1) (Catalog 10175-PO)
Substrate: Phenyl Acetate (7.88 M) (Sigma, Catalog # 108723)
Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
Dilute rhPON1 to 10 µg/mL in Assay Buffer.
Dilute Phenyl Acetate to 40 mM in deionized water.
Load 50 µL of 10 µg/mL rhPON1 in a plate, and start the reaction by adding 50 µL of Substrate. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of Substrate.
Read plate at a wavelength of 270 nm (absorbance) in kinetic mode for 5 minutes.
Calculate specific activity:
Specific Activity (pmol/min/µg) =
Adjusted Vmax* (OD/min) x well volume (L) x 1012 pmol/mol
ext. coeff** (M-1cm-1) x path corr.*** (cm) x amount of enzyme (µg)
*Adjusted for Substrate Blank **Using the extinction coefficient 1310 M-1cm-1 ***Using the path correction 0.32 cm Note: the output of many spectrophotometers is in mOD
rhPON1: 0.5 µg
Phenyl Acetate: 20 mM
Serum paraoxonase 1 (PON-1) is a member of the paraoxonase family which has three members: PON1, PON2, PON3. PON-1 is a lactonase (1,2) but has significant promiscuity (2,3) that allows hydrolysis of a variety of substrates including organophosphate triesters (including many pesticides), arylesters, cyclic carbamates, glucuronides and consequently also pharmaceuticals such as statins. PON-1 is a calcium-dependent, secreted, 43 kDa protein with a 6-bladed propeller structure and an active site gorge containing a His-His catalytic dyad (4). It is a homodimer and retains its hydrophobic signal sequence in the N-terminal region which enables its association with HDL, resulting in its localization predominantly in the plasma (5,6). Low serum PON1 and dysfunctional HDL is associated with many diseases with an inflammatory component including diabetes mellitus (7,8), rheumatoid arthritis, hepatic and renal diseases, psoriasis, and macular degeneration (9,10). PON1 shows a variety of atheroprotective properties (6,11,12) by metabolizing inflammatory lipid peroxides (13). PON-1 activity is low in coronary heart disease patients (14) and contributes to an increased risk of a major adverse cardiovascular event (15). PON-1 also protects against toxicity of organophosphorus compounds used as pesticides (16,17) thought to increase risk of neurodegenerative disorders such as Parkinson's (18) and Amyotrophic Lateral Sclerosis (19).
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Which Brands are Currently Available on bio-techne.com? R&D Systems, Tocris Bioscience and ProteinSimple branded products are available to purchase through bio‑techne.com. ProteinSimple branded instruments and Simple Plex Assays are available to quote. ProteinSimple branded instrument consumables and ACD branded products will be available on bio-techne.com in the near future. Novus Biologicals branded products are not currently available on bio‑techne.com and can be found at novusbio.com.
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