Recombinant Human Adenosine Deaminase/ADA Protein, CF

R&D Systems | Catalog # 7048-AD

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Key Product Details

  • R&D Systems Sf 21 (baculovirus)-derived Recombinant Human Adenosine Deaminase/ADA Protein (7048-AD)
  • 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

Sf 21 (baculovirus)

Accession Number

P00813

Applications

Enzyme Activity
View all Adenosine Deaminase/ADA Proteins and Enzymes »

Product Specifications

Source

Spodoptera frugiperda, Sf 21 (baculovirus)-derived human Adenosine Deaminase/ADA protein
Met1-Leu363, 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

Inconclusive result, Met predicted. Protein identity confirmed by MS analysis of tryptic fragments

Predicted Molecular Mass

42 kDa

SDS-PAGE

40-42 kDa, reducing conditions

Activity

Measured by the ability to catalyze the hydrolytic deamination of adenosine to inosine.
The specific activity is >44,000 pmol/min/μg, as measured under the described conditions.

Formulation, Preparation, and Storage

7048-AD
Formulation Supplied as a 0.2 μm filtered solution in Tris, NaCl and Glycerol.
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: Adenosine Deaminase/ADA

Adenosine Deaminase (ADA, adenosine aminohydrolase) is one of the key enzymes of purine nucleotide catabolism. It catalyses the hydrolytic deamination of adenosine and deoxy‑adenosine to inosine and deoxyinosine (1, 2). ADA is expressed in virtually all tissues and is expressed at high levels in T-lymphocytes. Adenosine Deaminase deficiency can cause a form of SCID (severe combined immunodeficiency) and lymphopenia in both B- and T-cell lineages (3, 4). ADA can be used as a sensitive diagnostic marker for tuberculous pleuritis (5). Although it is primarily a cytosolic enzyme, ADA is known to be a positive regulator of T-cell co‑activation due to its binding to CD26 at the cell surface. The interaction of ADA with CD26 regulates lymphocyte-epithelial cell adhesion (6).

References

  1. Wolfenden, R.V. et al. (1969) Biochemistry 6:2412.
  2. Lowenstein, J.M. (1972) Physiol. Rev. 52:382.
  3. Giblett, E.R. et al. (1972) Lancet 2:1067.
  4. Coleman, M.S. et al. (1978) J. Biol. Chem. 253:1619.
  5. Baba, K. et al. (2008) PLoS ONE 3:e2788.
  6. Gines, S. et al. (2002) Biochem. J. 361:203.

Long Name

Adenosine Aminohydrolase

Alternate Names

ADA, ADA1

Entrez Gene IDs

100 (Human); 11486 (Mouse); 24165 (Rat)

Gene Symbol

ADA

UniProt

P00813

Additional Adenosine Deaminase/ADA Products

Product Documents for Recombinant Human Adenosine Deaminase/ADA Protein, CF

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

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Product Specific Notices for Recombinant Human Adenosine Deaminase/ADA Protein, CF

For research use only

Citations for Recombinant Human Adenosine Deaminase/ADA Protein, CF

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Protocols

View specific protocols for Recombinant Human Adenosine Deaminase/ADA Protein, CF (7048-AD):

Materials
  • Assay Buffer: 50 mM HEPES, pH 7.5
  • Recombinant Human Adenosine Deaminase/ADA (rhADA) (Catalog # 7048-AD)
  • Substrate: Adenosine (Sigma, Catalog # A9251), 10 mM stock in deionized water
  • UV plate (Costar, Catalog # 3635)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Thaw the Substrate stock and then warm it at 37 °C for 20 minutes. Mix well after incubation.
  2. Dilute rhADA to 0.1 ng/μL in Assay Buffer.
  3. Dilute Substrate to 200 μM in Assay Buffer.
  4. Load into a plate 50 μL of 0.1 ng/μL rhADA and start the reaction by adding 50 μL of 200 μM Substrate. For Substrate Blanks, load 50 μL of Assay Buffer and 50 μL of 200 μM Substrate.
  5. Read plate at a wavelength of 265 nm (bottom read) in kinetic mode for 5 minutes.
  6. Calculate specific activity:

     Specific Activity (pmol/min/µg) =

 Adjusted Vmax* (OD/min) x -1 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 8500 M-1cm-1 
     ***Using the path correction 0.32 cm
     Note: the output of many spectrophotometers is in mOD Per Well:
  • rhADA: 0.005 μg
  • Substrate: 100 μM

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