Recombinant Human Fetuin A/AHSG Protein, CF

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Product Details

Summary
Reactivity HuSpecies Glossary
Applications Inhibition Activity
Format
Carrier-Free

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Recombinant Human Fetuin A/AHSG Protein, CF Summary

Details of Functionality
Measured by its ability to inhibit calcium phosphate precipitation. Schinke, T. et al. (1996) J. Biol. Chem. 271:20789. The IC50 value is <50 μg/mL, as measured under the described conditions.
Source
Mouse myeloma cell line, NS0-derived human Fetuin A/AHSG protein
Ala19-Val367, with a C-terminal 10-His tag
Accession #
N-terminal Sequence
Ala19
Protein/Peptide Type
Recombinant Enzymes
Gene
AHSG
Purity
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain
Endotoxin Note
<1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

Theoretical MW
39 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
SDS-PAGE
58 kDa, reducing conditions
Publications
Read Publications using
1184-PI in the following applications:

Packaging, Storage & Formulations

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 reconstitution.
Buffer
Lyophilized from a 0.2 μm filtered solution in Tris and NaCl.
Purity
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain
Reconstitution Instructions
Reconstitute at 500 μg/mL in sterile 25 mM Tris and 150 mM NaCl, pH 7.5.
Assay Procedure
  • Assay Buffer: 50 mM Hepes, pH 7.5
  • Recombinant Human Fetuin A/AHSG (rhFetuin A) (Catalog # 1184-PI)
  • Calcium chloride (Sigma, Catalog # C3881), 100 mM stock in deionized water
  • Sodium phosphate, pH 7.5 (Sigma, Catalog # S9638), 100 mM stock in deionized water
  • Acetic acid (Sigma, Catalog # 242853), 100 mM stock in deionized water
  • Malachite Green Phosphate Detection Kit (Catalog # DY996)
  • 96 well Clear Plate (Costar, Catalog # 92592)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute sodium phosphate to 3.2 mM in Assay buffer.
  2. Dilute calcium chloride to 9.6 mM in Assay Buffer.
  3. Dilute rhFetuin A to 500, 250, 125, 62.5, 31.3, 15.6, 7.8, 3.9, 1.95, and 0.97 µg/mL in Assay Buffer containing 3.2 mM sodium phosphate.
  4. Combine 50 µL of dilute calcium chloride with 50 µL of each dilution of rhFetuin A. Include an enzyme blank containing 50 µL calcium chloride and 50 µL dilute sodium phosphate buffer.
  5. Incubate reactions at 37 °C for 100 minutes.
  6. Centrifuge reactions in a tabletop microcentrifuge at maximum speed for 5 minutes.
  7. Carefully remove the supernatant from each tube, and discard.
  8. Resuspend each pellet in 200 µL of 100 mM acetic acid. Vortex briefly.
  9. Incubate suspension at room temperature for 15 minutes.
  10. Add 800 µL of deionized water to each tube.
  11. Load 50 µL of each reaction into wells of a 96 well plate.
  12. Add 25 µL of the Malachite Green Reagent A to all wells.  Mix briefly.
  13. Add 75 µL of deionized water to all wells. Mix briefly.
  14. Add 25 µL of the Malachite Green Reagent B to all wells.  Mix and incubate for 20 minutes at room temperature.
  15. Read plate at 620 nm (absorbance) in endpoint mode.
  16. Derive the 50% inhibiting concentration (IC50) of rhFetuin A by plotting absorbance vs. reaction concentration of rhFetuin A with 4‑PL fitting.
Per Incubation:
  • rhFetuin A curve: 250, 125, 62.5, 31.3, 15.6, 7.8, 3.9, 1.95, 0.97 and 0.49 µg/mL
  • Calcium chloride: 4.8 mM
  • Sodium phosphate: 1.6 mM

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Recombinant Human Fetuin A/AHSG Protein, CF

  • A2HS
  • AHS
  • AHSG
  • alpha-2-HS-glycoprotein
  • alpha-2-Z-globulin
  • Ba-alpha-2-glycoprotein
  • FETUAba-alpha-2-glycoprotein
  • Fetuin A
  • fetuin-A
  • HSGA

Background

Human Fetuin A, also known as alpha 2-Heremans-Schmid glycoprotein, is encoded by the AHSG gene. It is a major plasma protein and a member of the cystatin superfamily of protease inhibitors (1, 2). It is expressed by hepatocytes, the principal cell source, and by monocyte/macrophages (3). The major form of plasma Fetuin A corresponds to a disulfide bond-linked two chains derived from the single chain (4). Human Fetuin A has a number of functions. It is a negative acute-phase protein with normal circulating levels in adults (300‑600 μg/mL), which fall significantly (30‑50%) during injury and infection (5). It enhances entry of cationic inhibitors into macrophages (6). It inhibits both insulin receptor autophosphorylation and undesirable calcification (6, 7). The purified recombinant human Fetuin A corresponds to the single chain, which can be converted to the two-chain form by recombinant human Furin (Catalog # 1503-SE) in vitro. However, the conversion does not enhance its inhibitory activity against recombinant human Cathpesin V, a cysteine protease.
  1. Kellemann, J. et al. (1989) J. Biol. Chem. 264:14121.
  2. Dziegielewska, K.M. et al. (1990) J. Biol. Chem. 265:4354.
  3. Dziegielewska, K.M. et al. (1996) Histochem. Cell Biol. 106:319.
  4. Gejyo, F. and K. Schmid (1981) Biochim. Biophys. Acta. 671:78.
  5. Wang, H. et al. (1998) Proc. Natl. Acad. Sci. USA 95:14429.
  6. Mathews, S.T. et al. (2000) Mol. Cell Endocrinol. 164:87.
  7. Schäfer, C. et al. (2003) J. Clin. Invest. 112:357.

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1184-PI
Species: Hu
Applications: Inhibition Activity

Publications for Fetuin A/AHSG (1184-PI)(2)

We have publications tested in 2 confirmed species: Human, N/A.

We have publications tested in 2 applications: ELISA (Standard), Single Molecule Capture Standa.


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ELISA (Standard)
(1)
Single Molecule Capture Standa
(1)
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(1)
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(1)
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Bioinformatics

Gene Symbol AHSG
Uniprot