Recombinant Human IDO Protein, CF

Indoleamine 2,3-dioxygenase
Catalog # Availability Size / Price Qty
6030-AO-010
R&D Systems Recombinant Proteins and Enzymes
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Product Details
Citations (3)
FAQs
Reviews (1)

Recombinant Human IDO Protein, CF Summary

Product Specifications

Purity
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain.
Endotoxin Level
<0.10 EU per 1 μg of the protein by the LAL method.
Activity
Measured by its ability to oxidize L-tryptophan to N-formyl-kynurenine. The specific activity is >500 pmol/min/μg, as measured under the described conditions.
Source
E. coli-derived human Indoleamine 2,3-dioxygenase/IDO protein
MHHHHHHHuman IDO
(Ala2-Gly403)
Accession # P14902
N-terminusC-terminus
Accession #
N-terminal Sequence
Analysis
Met
Structure / Form
Monomer
Predicted Molecular Mass
46 kDa
SDS-PAGE
42 kDa, reducing conditions

Product Datasheets

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6030-AO

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.

6030-AO

Formulation Supplied as a 0.2 μm filtered solution in Sodium Acetate, NaCl and Glycerol.
Shipping The product is shipped with dry ice or equivalent. 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, -70 °C as supplied.
  • 3 months, -70 °C under sterile conditions after opening.

Assay Procedure

Materials
  • Assay Buffer: 50 mM MES, pH 6.5
  • 0.405 M Tris, pH 8.0
  • Recombinant Human Indoleamine 2,3‑dioxygenase/IDO (rhIDO) (Catalog # 6030-AO)
  • Ascorbic Acid (Sigma, Catalog # 255564), 500 mM stock in deionized water
  • L-Tryptophan (Sigma, Catalog # T0254), 10 mM stock in deionized water
  • Catalase (Sigma, Catalog # C30), 100,000 units/mL stock in Assay Buffer
  • Methylene Blue (Sigma, Catalog # 28514), 10 mM stock in deionized water
  • 96-well Clear Plate (Costar, Catalog # 92592)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Prepare the Substrate Mixture.
    1. Dilute Ascorbic Acid to 80 mM in 0.405 M Tris, pH 8.0.
    2. Prepare a mixture of 800 μM L-Tryptophan, 9000 units/mL catalase, and 40 μM Methyene Blue in Assay Buffer.
    3. Mix equal volumes of 1a and 1b for final concentrations of 40 mM Ascorbic Acid, 400 μM L-Tryptophan, 4500 units/mL Catalase, and 20 μM Methylene Blue.
  2. Dilute rhIDO to 16 ng/μL in Assay Buffer.
  3. Load into a plate 50 μL of 16 ng/μL rhIDO, and start the reaction by adding 50 μL of Substrate Mixture.
  4. Include a Substrate Blank containing 50 μL of Assay Buffer and 50 μL of Substrate Mixture.
  5. Read at 321 nm in kinetic mode for 5 minutes.

     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 3750 M-1cm-1 
     ***Using the path correction 0.32 cm
     Note: the output of many spectrophotometers is in mOD. Per Well:
  • rhIDO: 0.800 µg
  • Ascorbic Acid: 20 mM
  • L-Tryptophan: 200 μM
  • Catalase: 225 units
  • Methylene Blue: 10 µM
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Background: Indoleamine 2,3-dioxygenase/IDO

Indoleamine 2,3-dioxygenase (IDO) is a heme-containing intracellular dioxygenase catalyzing the degradation of the essential amino acid L-tryptophan to N‑formyl‑kynurenine (1). This degradation is the first and rate-limiting step of the L-kynurenine pathway (2). IDO is widely expressed in dendritic cells, macrophages, microglia, eosinophils, fibroblasts, endothelial cells, and most tumor cells. In immune cells, its expression is mainly induced by cytokines such as IFN‑ gamma, IFN‑ alpha, IFN‑ beta, and IL‑10. IDO has an antimicrobial function due to its decreasing the availability of the essential amino acid tryptophan in inflammatory environments (3). Recent studies have demonstrated that IDO induces immunosuppression during infection, pregnancy, transplantation, autoimmunity, and neoplasia (3‑5).

References
  1. Lewis-Ballester, A. et al. (2009) Proc. Natl. Acad. Sci. USA. 106:17371.
  2. Costantino, G. (2009) Expert Opin. Ther. Targets 13:247.
  3. Xu, H. et al. (2008) Immunol. Lett. 121:1.
  4. Lob, S. et al. (2009) Nat. Rev. Cancer 9:445.
  5. Curti, A. et al. (2009) Blood 113:2394.
Entrez Gene IDs
3620 (Human); 15930 (Mouse)
Alternate Names
3dioxygenase; EC 1.13.11.52; IDO; IDO1; IDOIDO-1; INDO; INDOindole 2,3-dioxygenase; Indoleamine 2; indoleamine 2,3-dioxygenase 1; Indoleamine 2,3-dioxygenase; indoleamine-pyrrole 2,3 dioxygenase; Indoleamine-pyrrole 2,3-dioxygenase

Citations for Recombinant Human IDO Protein, CF

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

3 Citations: Showing 1 - 3
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  1. Iron activates microglia and directly stimulates indoleamine-2,3-dioxygenase activity in the N171-82Q mouse model of Huntington's disease
    Authors: DW Donley, M Realing, JP Gigley, JH Fox
    PLoS ONE, 2021;16(5):e0250606.
    Species: Human
    Sample Types: Recombinant Protein
    Applications: Bioassay
  2. Influence of Indoleamine-2,3-Dioxygenase and Its Metabolite Kynurenine on gammadelta T Cell Cytotoxicity against Ductal Pancreatic Adenocarcinoma Cells
    Authors: H Jonescheit, HH Oberg, D Gonnermann, M Hermes, V Sulaj, C Peters, D Kabelitz, D Wesch
    Cells, 2020;9(5):.
    Species: N/A
    Sample Types:
  3. Dendritic Cells Treated with Exogenous Indoleamine 2,3-Dioxygenase Maintain an Immature Phenotype and Suppress Antigen-specific T cell Proliferation
    Authors: E Bracho-San, A Hassanzade, MA Brusko, MA Wallet, BG Keselowsky
    J Immunol Regen Med, 2019;5(0):.
    Species: Human
    Sample Types: Amino Acid
    Applications: Bioassay

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Reviews for Recombinant Human IDO Protein, CF

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Recombinant Human IDO Protein, CF
By Jason Frazier on 05/14/2016
Application: Enzymatic activity in vitro
Reason for Rating: Worked exactly as anticipated for in vitro enzyme activity assay