Recombinant Mouse Alkaline Phosphatase/ALPL Protein, CF

Catalog #: 2910-AP Datasheet / COA / SDS

Catalog #	Availability	Size / Price	Qty
2910-AP-010

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Summary Product Datasheets Carrier Free Data Images Reconstitution Calculator Background Related Research Areas

Recombinant Mouse Alkaline Phosphatase/ALPL Protein, CF Summary

Product Specifications

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.

Activity

Measured by its ability to cleave a fluorogenic substrate, 4-Methylumbelliferyl phosphate (4-MUP). The specific activity is >40,000 pmol/min/µg, as measured under the described conditions.

Source

Mouse myeloma cell line, NS0-derived mouse Alkaline Phosphatase/ALPL protein
Phe18-Gly503, with a C-terminal 6-His tag

Accession #

P09242

N-terminal Sequence
Analysis

Phe18

Predicted Molecular Mass

54 kDa

SDS-PAGE

77 kDa, reducing conditions

Product Datasheets

2910-AP

Product Datasheet

COA

SDS

Carrier Free

What does CF mean?

CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins. 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 could interfere.

2910-AP

Formulation	Supplied as a 0.2 μm filtered solution in Tris and NaCl.
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.

Assay Procedure

Materials

Assay Buffer: 50 mM Tris, 1 mM MgCl₂, pH 9.0
Recombinant Mouse Alkaline Phosphatase/ALPL (rmALPL) (Catalog # 2910-AP)
Substrate: 4-Methylumbelliferlyphosphate (4-MUP) (Sigma, Catalog # M8883)
F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent

Dilute rmALPL to 0.02 ng/µL in Assay Buffer.
Dilute Substrate to 50 µM in Assay Buffer.
Load 50 µL of 0.02 ng/µL rmALPL into a plate, and start the reaction by adding 50 µL of 50 µM Substrate. Include a Substrate Blank containing 50 µL of Assay Buffer and 50 µL of Substrate.
Read at excitation and emission wavelengths of 365 nm and 445 nm (top read), respectively, in kinetic mode for 5 minutes.
Calculate specific activity:

Specific Activity (pmol/min/µg) =	Adjusted V_max* (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:

rmALPL: 0.001 µg
Substrate: 25 µM

Reconstitution Calculator

Background: Alkaline Phosphatase/ALPL

Several distinct genes encode alkaline phosphatases (APs) in mice with different tissue-specific expression patterns. The Alpl gene, also known as Akp2, encodes the liver/bone/kidney isozyme, also known as the tissue-nonspecific AP (TNAP) (1). The Alpl gene is a key regulator of bone mineralization in mice (2). A variety of mutations in the human ALPL gene leads 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 504 to 524) is not present in the mature form.

References

Terao, M. and B. Mintz (1987) Proc. Natl. Acad. Sci. USA 84:7051.
Hessle, L. et al. (2002) Proc. Natl. Acad. Sci. USA 99:9445.
Di Mauro, S. et al. (2002) J. Bone Miner. Res. 17:1383.

Long Name

Alkaline Phosphatase Liver

Entrez Gene IDs

249 (Human); 11647 (Mouse)

Alternate Names

Akp2; Alkaline phosphatase liver/bone/kidney isozyme; alkaline phosphatase, liver/bone/kidney; alkaline phosphatase, tissue-nonspecific isozyme; alkaline phosphomonoesterase; ALPL; APTNAP; AP-TNAP; EC 3.1.3.1; FLJ40094; FLJ93059; glycerophosphatase; HOPS; liver/bone/kidney-type alkaline phosphatase; MGC161443; tissue-nonspecific ALP; TNAP; TNSALP; TNSALPMGC167935