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UGT-Glo Assay

by Promega
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  • The UGT-Glo™ Assay provides a luminescent method for measuring UDP glucuronosyltransferase (UGT) activity. The UGT-Glo™ Assay is designed to measure UGT activity from a variety of sources, such as microsomes containing recombinantly expressed enzymes or microsomal preparations derived from mammalian tissues, and to test the effects of various chemicals on UGT activity.

    The assay involves incubating UGT with a proluciferin substrate; a portion of the substrate gets conjugated with UDP, while the remainder is unmodified. Upon the addition of d-Cysteine, the unconjugated proluciferin is converted into luciferin and, in a coupled reaction with luciferase/luciferin, is converted into light. Conjugated proluciferin remains intact and does not contribute to the luminescence. Thus, the signal generated is inversely correlated with UGT activity present in the sample.

    The UGT-Glo™ Assay contains two proluciferin substrates: the UGT Multienzyme Substrate, which is compatible with a wide range of UGTs, and the UGT1A4 Substrate, which reacts specifically with UGT1A4. The kit also contains Luciferin Detection Reagent and Reconstitution Buffer, UGT Buffer, d-Cysteine and UDPGA.


    Conversion of UGT Multienzyme Substrate

    Conversion of UGT Multienzyme Substrate by UGT enzymes.
    Conversion of UGT Multienzyme Substrate by UGT enzymes.

     

     

    UGT enzymes attach a glucuronic acid moiety to the proluciferin substrate. During the detection step, the unconjugated proluciferin is converted to luciferin by cyclization with D-cysteine and then converted into light by luciferase. Glucuronidated proluciferin does not get converted into luciferin, and thus does not produce light. Light output is inversely proportional to UGT enzymatic activity.

    Features and Benefits

    • The luminescent format eliminates the need for time-consuming analyses such as HPLC and LC/MS.
    • The simple "add and read" protocol makes the assay amenable to higher throughput screening in multiwell plates.
    • Use less enzyme and scale down reaction volumes; save on reagent costs.

    Applications include screening drugs and new chemical entities for their capacity to modulate UGT activity in native or recombinant fractions and measuring recombinant UGT activities in membrane fractions from heterologous expression systems, such as insect cells and E. coli.

  • Protocols

    Complete Protocol

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    UGT-Glo™ Assay Technical Bulletin

    PDF (1 MB)

  • Certificate of Analysis

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    Storage Conditions

    -30C TO -10C

    Use Restrictions

    For Research Use Only. Not for Use in Diagnostic Procedures.

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