Recombinant Human alpha-L-Iduronidase/IDUA Protein, CF Summary
Ala26-Pro653 (Ala26Thr), with a C-terminal 10-His tag
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.
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.
|Formulation||Supplied as a 0.2 μm filtered solution in Sodium Acetate, 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.
- Assay Buffer: 50 mM NaOAc, 150 mM NaCl, 0.02% Brij-35 (w/v) pH 3.5
- Developing Buffer: 0.1 M Tris, pH 9.0
- Recombinant Human alpha ‑L‑Iduronidase/IDUA (rhIDUA) (Catalog # 4119-GH)
- Substrate: 4-methylumberlliferyl-alpha -L-iduronide (Glycosynth, Catalog # 44076), 20 mM stock in DMSO
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Dilute rhIDUA to 0.2 µg/mL in Assay Buffer. Minimize the number of dilution steps to obtain the best activity results.
- Dilute Substrate to 200 µM in Assay Buffer.
- Combine equal volumes of 0.2 µg/mL rhIDUA and 200 µM Substrate. Include a Substrate Blank containing Assay Buffer and Substrate.
- Incubate for 10 minutes at room temperature.
- Dilute mixtures to 0.005 µg/mL in Developing Buffer.
- Load 100 µL of the diluted mixtures into a plate.
- Read at excitation and emission wavelengths of 365 nm and 445 nm (top read), respectively in endpoint mode.
- Calculate specific activity:
Specific Activity (pmol/min/µg) =
|Adjusted Fluorescence* (RFU) x Conversion Factor** (pmol/RFU)|
|Incubation time (min) x amount of enzyme (µg)|
*Adjusted for Substrate Blank
**Derived using calibration standard 4-methylumbelliferone (Sigma, Catalog # M1381).Per Well:
- rhIDUA: 0.0005 µg
- Substrate: 5 µM
alpha -L-Iduronidase encoded by the IDUA gene is an important enzyme required for the lysosomal degradation of glycosaminoglycans (GAGs). It hydrolyzes the non‑reducing terminal alpha -L-iduronic acid residues in GAGs including dermatan sulfate and heparan sulfate. Mutations in IDUA that result in enzymatic deficiency lead to the autosomal recessive disease mucopolysaccharidosis type I (MPS I) (1). MPS I causes progressive cellular, tissue and organ damage, and several clinical studies using enzyme replacement therapy have shown promising benefits (2).
Citations for Recombinant Human alpha-L-Iduronidase/IDUA 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.
Citations: Showing 1 - 2
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Dose-Dependent Prevention of Metabolic and Neurologic Disease in Murine MPS II by ZFN-Mediated Inï¿½Vivo Genome Editing
Authors: K Laoharawee, RC DeKelver, KM Podetz-Ped, M Rohde, S Sproul, HO Nguyen, T Nguyen, SJ St Martin, L Ou, S Tom, R Radeke, KE Meyer, MC Holmes, CB Whitley, T Wechsler, RS McIvor
Mol. Ther., 2018;26(4):1127-1136.
Sample Types: Tissue Homogenates
Shutdown of ER-associated degradation pathway rescues functions of mutant iduronate 2-sulfatase linked to mucopolysaccharidosis type II
Authors: Y Osaki, A Saito, S Kanemoto, M Kaneko, K Matsuhisa, R Asada, T Masaki, K Orii, T Fukao, S Tomatsu, K Imaizumi
Cell Death Dis, 2018;9(8):808.
Sample Types: Whole Cells
Applications: Enzyme Assay
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