NFkB Secreted Alkaline Phosphatase Reporter Assay Kit (Colorimetric)

Novus Biologicals, part of Bio-Techne | Catalog # NBP2-25286

Novus Biologicals, part of Bio-Techne
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Product Specifications

Kit Type

Assay Kit (Colorimetric)

Reactivity

Human

Description

  • SEAP is secreted into the media so no cell lysis is required for the detection of its enzymatic activity.
  • A secreted indicator function permits multiple kinetics experiments using only one culture through sequential sampling of the medium, and also allows the cells to be used for other purposes, such as RNA extraction, Western blot analysis, and other assays
  • SEAP is stable in serum, allowing it to be used as a reporter assay in vivo.
  • SEAP catalyzes the hydrolysis of pNitrophenyl phosphate (PNPP) producing a yellow product that can be read in a spectrophotometer or ELISA reader at 405 nm.

    • Scientific Data Images for NFkB Secreted Alkaline Phosphatase Reporter Assay Kit (Colorimetric)

    Enzyme Activity: NFkB Secreted Alkaline Phosphatase Reporter Assay Kit (Colorimetric) [NBP2-25286]

    Enzyme Activity: NFkB Secreted Alkaline Phosphatase Reporter Assay Kit (Colorimetric) [NBP2-25286]

    Enzyme Activity: NFkB Secreted Alkaline Phosphatase Reporter Assay Kit (Colorimetric) [NBP2-25286] - Standard curve for the SEAP protein provided in the NF-KB SEAPorter(TM) Assay Kit. A serial dilution of SEAP protein was added to a 96-well microtiter plate and the standard curve was generated according to the Kit assay protocol.

    Kit Contents for NFkB Secreted Alkaline Phosphatase Reporter Assay Kit (Colorimetric)

    • 10X PNPP Buffer (4 mL)
    • 10X Sample Dilution Buffer (10 mL)
    • 96-well Microplate (4 EA)
    • Microplate seal (4 EA)
    • pCMV/SEAP plasmid at 0.5 ug/uL (2 x 10 ug)
    • pNF-kappa B/SEAP plasmid at 0.5 ug/uL (2 x 10 ug)
    • PNPP Substrate (8 x 5 mg)
    • SEAP Standard (200 ug/mL)

    Preparation and Storage

    Shipping

    The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

    Stability & Storage

    Storage of components varies. See protocol for specific instructions.

    Background: NFkB Secreted Alkaline Phosphatase Reporter

    Placental alkaline phosphatase (PLAP) is one of the most stable isoenzymes, only existing in the placenta of higher primates. These characteristics make placental alkaline phosphatase suitable to use as a reporter gene for the analysis of promoter activity and gene expression in cell culture and animal serum. The natural form of PLAP is membrane anchored. The recombinant form of placental alkaline phosphatase (secreted alkaline phosphatase, SEAP) is used for reporter gene function. SEAP is designed by inserting a translational terminator after amino acid 489 (Berger, et al., Gene 66 (1): 10 (1988). This mutation converts the membrane-bound PLAP protein into the secreted protein SEAP. As a major transcription factor NF-KB plays a key role in regulating genes responsible for the innate and adaptive immune responses. In unstimulated cells, the NF-KB dimers are held in cytoplasm by IKBs that masks the nuclear localization signals of NF-KB. Upon cell stimulation, which leads to IKB degradation, NF-KB quickly translocates to the nucleus and activates various genes that have DNA-binding sites for NF-KB. Our NF-KB SEAPORTER Assay Kit is designed to measure NF-KB activation using SEAP protein secreted to the culture media as a read-out.

    Alternate Names

    SEAP assay

    Additional NFkB Secreted Alkaline Phosphatase Reporter Products

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    Product Specific Notices for NFkB Secreted Alkaline Phosphatase Reporter Assay Kit (Colorimetric)

    This product is for research use only and is not approved for use in humans or in clinical diagnosis. Kits are guaranteed for 6 months from date of receipt.

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