Quantitative Assay, suitable for detection of Prx I, II, III, IV, V via Spectrophotometric UV/Vis at 340nm, sufficient for 100 reactions, ultra accurate, ready-to-use, quick protocol, hyper sensitive, high reproducibility
Thioredoxin is a small (12 kDa) redox active thiol protein that is present in prokaryotic and eukaryotic cells. Thioredoxin maintains a thiol redox pool of cells and is known to be oxidized in conditions of oxidative stress. Mammalian thioredoxin has 3 structural cysteines at positions 62, 69 and 72; and two cysteines at positions 32 and 35 that take part in redox reactions. Once oxidized this protein is re-reduced by thioredoxin reductase and utilizes NADPH as a reducing equivalent. Mammalian thioredoxin is oxidized in response to oxidative stress and can remain in various redox states. For example, a strong oxidant such as H2O2 can fully oxidize thioredoxin which can remain in various redox states as shown in Fig 2. The Thioredoxin Redox State Assay will determine the relative abundance of oxidized and reduced thioredoxin in cellular lysates, or tissue homogenates. The thiol group is derivatized with an Iodoacetic Acid (IAA) and the oxidized and reduced thioredoxin are separated based on their relative mobility in a native polyacrylamide gel. The oxidized and reduced bands are visualized by western analysis as demonstrated in Fig 1. The bands can be quantified by densitometric determination of relative intensity of oxidized and reduced bands. Fluorescent labeling can also be done for direct measurement of these bands using secondary antibody conjugated with appropriate fluorescent label.
Intracellular redox state is an important regulator of gene expression, signal transduction and antioxidant defense system. The redox state of thioredoxin is a key indicator of redox environment of the cell. In a reduced state thioredoxin can induce gene expression, activate transcription factors and can detoxify peroxides via peroxiredoxins. However, increased oxidative stress could oxidize thioredoxin resulting in loss of function that is dependent on reduced thioredoxin. Therefore, it is important to determine the relative levels of reduced and oxidized thioredoxin in cells or tissues in response to a stress. Thioredoxin redox state can also be used as a marker of oxidative stress in cancer research, cardiovascular research, neurodegenerative disease research, and in toxicological research. This assay along with our Thioredoxin Activity Assay Kit (Cat #TR-7011K) provides a complete understanding of the response of thioredoxin systems to a specific oxidant or other thioredoxin modulating stimuli.
The Thioredoxin Redox State Assay Kit by Redoxica has undergone extensive testing with a multitude of lysate types, has been published in many scientific journals and after a decade since its market introduction has been established as the gold standard in redox state detection. We designed the assay by utilizing the core principles of biochemistry to ensure maximum accuracy, sensitivity, and reproducibility accompanied by a quick and easy to use protocol.
|Unit of Measure||Assays|
|Shelf Life||24 MONTHS|
|Made In||United States of America|
|Components||• Thioredoxin Redox State Assay Buffer (2 x 10 mL bottles, Store at 2-8°C)
• Iodoacetic Acid (IAA) (2 x 120 mg ampoules, Store at -20°C, use within 2 hours of reconstitution, not recommended for storage)
• Spin Columns (40 columns and elution tubes, Store at: 2-8°C)
• Thioredoxin (Trx) Antibody (80 μl vial, Sufficient for 10 mini gels, Store at 2-8°C)
|Required Materials Not Included||• Electrophoresis apparatus
• Electroblotting apparatus
• Western blotting reagents
|Synonyms||Thioredoxin Redox State Assay Kit, TRX, TXN, TRDX, TRX1, Surface-Associated Sulphydryl Protein, SASP, ATL-Derived Factor, ADF, TXN Delta, Thioredoxin Delta, RP11-427L11.1, MGC61975, DKFZp686B1993|
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STORAGE & SAFETY
Recommended to store IAA at -20°C (KEEP FROZEN, DESICCATED, LIGHT SENSITIVE) and all other kit components at 2-8°C (KEEP REFRIGERATED). For component storage upon reconsistution please refer to the product insert for more information. Avoid repeated freez
Precautions & Disclaimer
For Laboratory Research and Development Use Only. Not for drug, household or other uses.
SHIPPING & HANDLING
Each kit utilizes a variety of packaging materials including borosilicate glass, PET, polybags, magnetic board, crosslinked foam, PVC.
12"" X 12"" X 10""