Key features and details
- Detection method: Fluorescent
- Platform: Microplate reader
- Sample type: Cell culture extracts, Tissue Extracts
Product nameDeubiquitinase Assay Kit
Sample typeCell culture extracts, Tissue Extracts
Deubiquitinase Assay Kit (ab241002) provides a straight-forward and general measure of deubiquitinase activity by utilizing a fluorescent deubiquitinase substrate to detect activity as low as 0.25 µU with purified enzyme.
Fl-Substrate -------> Cleaved Substrate + Fluorescence
This product is manufactured by BioVision, an Abcam company and was previously called K485 Deubiquitinase Activity Assay Kit (Fluorometric). K485-100 is the same size as the 100 test size of ab241002.
Storage instructionsStore at -20°C. Please refer to protocols.
Components 100 tests 1 M DTT 1 x 100µl AMC Standard (1 mM) 1 x 100µl DUB Assay Buffer 1 x 25ml DUB Positive Control 1 vial DUB Substrate (in DMSO) 1 x 25µl White 96-well Half-Area Plate 1 unit
RelevanceCell activity and viability is tightly regulated by controlling the production and degradation of the thousands of different proteins in the cell. The proteasome is responsible for the majority of cellular protein degradation; however, drugs targeting the proteasome can have side effects caused by the lack of specificity associated with inhibiting the proteasome itself. Altering the ubiquitination state of target proteins is thus appealing as an alternative approach. Modification of the ubiquitin-mediated proteasome pathway has been shown to be a valid mechanism for treating a variety of diseases, all of which involve dysregulation of cellular proteostasis. As such, it is imperative that these ubiquitination signals also be reversible. The enzymes responsible for cleavage, and hence removal of ubiquitin from ubiquitinated proteins, are known as de-ubiquitinating enzymes, or DUBs. They are proteases that hydrolyze the isopeptide bond between an ubiquitin moiety and a lysine residue on its target protein. By removing the ubiquitin molecule, the protein escapes the fate of proteasomal degradation and remains a viable factor in the cell.
AMC Standard Curve
Time course using positive control DUB as described.
Rat tissue samples (10 mg each) were resuspended in DUB Assay Buffer with DTT (100 l), homogenized, and clarified by centrifugation. The DUB activities for Rat Kidney and Liver lysates, in mU/mg, were determined to be 0.76 and 3.31, respectively. Assays were performed following the kit protocol.
ab241002 has been referenced in 1 publication.
- Zhu QZ et al. DJ-1 activates the noncanonical NF-κB pathway via interaction with Cezanne to inhibit the apoptosis and promote the proliferation of Ishikawa cells. Mol Biol Rep 48:6075-6083 (2021). PubMed: 34374892