Recombinant Human Alkaline Phosphatase/ALPL Protein, CF

R&D Systems | Catalog # 2909-AP

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Key Product Details

  • R&D Systems NS0-derived Recombinant Human Alkaline Phosphatase/ALPL Protein (2909-AP)
  • Quality control testing to verify active proteins with lot specific assays by in-house scientists
  • All R&D Systems proteins are covered with a 100% guarantee

Source

NS0

Accession Number

P05186

Applications

Enzyme Activity
View all Alkaline Phosphatase/ALPL Proteins and Enzymes »

Product Specifications

Source

Mouse myeloma cell line, NS0-derived human Alkaline Phosphatase/ALPL protein
Leu18-Ser502, with a C-terminal 6-His tag

Purity

>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.

Endotoxin Level

<1.0 EU per 1 μg of the protein by the LAL method.

N-terminal Sequence Analysis

Leu18

Predicted Molecular Mass

54 kDa

SDS-PAGE

65-75 kDa, reducing conditions

Activity

Measured by its ability to cleave a fluorogenic substrate, 4-Methylumbelliferyl phosphate (4-MUP).
The specific activity is >30,000 pmol/min/µg, as measured under the described conditions.

Formulation, Preparation, and Storage

2909-AP
Formulation Supplied as a 0.2 μm filtered solution in HEPES, NaCl, MgCl2 and ZnCl2.
Shipping 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.

Background: Alkaline Phosphatase/ALPL

Four distinct genes encode alkaline phosphatases (APs) in humans (1). The ALPL gene encodes the liver/bone/kidney isozyme, also known as the tissue‑nonspecific AP (TNAP). In comparison, ALPI, ALPP and ALPPL2 encode intestinal, placental and placental-like or germ cell APs, respectively. The serum levels of human APs are useful tumor markers (2). There are many mutations in the ALPL gene, leading to different forms of hypophosphatasia, characterized by poorly mineralized cartilage and bones (3). The native ALPL is a glycosylated homodimer attached to the membrane through a GPI-anchor. The C-terminal pro peptide (residues 503 to 524) is not present in the mature form.

References

  1. Le Du, M.-H. and J.L. Millan (2002) J. Biol. Chem. 277:49808.
  2. Millan, J.L. and W.H. Fishman (1995) Crit. Rev. Clin. Lab. Sci. 32:1.
  3. Di Mauro, S. et al. (2002) J. Bone Miner. Res. 17:1383.

Long Name

Alkaline Phosphatase Liver

Alternate Names

Akp2, AP-TNAP, HOPS, TNAP, TNSALP

Entrez Gene IDs

249 (Human); 11647 (Mouse)

Gene Symbol

ALPL

UniProt

P05186

Additional Alkaline Phosphatase/ALPL Products

Product Documents for Recombinant Human Alkaline Phosphatase/ALPL Protein, CF

Certificate of Analysis

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Product Specific Notices for Recombinant Human Alkaline Phosphatase/ALPL Protein, CF

For research use only

Citations for Recombinant Human Alkaline Phosphatase/ALPL Protein, CF

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Protocols

View specific protocols for Recombinant Human Alkaline Phosphatase/ALPL Protein, CF (2909-AP):

Materials
  • Assay Buffer: 50 mM Tris, 1 mM MgCl2, pH 9.0
  • Recombinant Human Alkaline Phosphatase/ALPL/TNAP (rhTNAP) (Catalog # 2909-AP)
  • Substrate: 4-Methumbelliferly phosphate (4-MUP) (Calbiochem, Catalog # 474431), 10 mM in deionized water
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute rhTNAP to 0.01 ng/µL in Assay Buffer.
  2. Dilute Substrate 50 µM in Assay Buffer.
  3. In a plate load 50 µL of 0.01 ng/µL rhTNAP to wells. Include a Substrate Blank of 50 µL of Assay Buffer.
  4. Start the reaction by adding 50 µL of 50 µM Substrate to wells.
  5. Read at excitation and emission wavelengths of 365 nm and 445 nm (top read), respectively in kinetic mode for 5 minutes.
  6. Calculate specific activity:

     Specific Activity (pmol/min/µg) =

 Adjusted Vmax* (RFU/min) x Conversion Factor** (pmol/RFU)
amount of enzyme (µg)

     *Adjusted for Substrate Blank

     **Derived using calibration standard 4-Methumbelliferone (Sigma, Catalog # M1381).

Per Well:
  • rhTNAP: 0.0005 µg
  • Substrate: 25 µM

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