HT Ubiquitin Cellular Assays
Measuring cellular ubiquitylation in a plate-based format accelerates the process of ubiquitin pathway drug discovery. We have developed several platforms that enable rapid, quantitative monitoring of cellular ubiquitylation.
LifeSensors’ UbiQuant™ S ELISA and UbiQuant™ assays
These quantitative plate based assays are industry-leading platforms for measuring ubiquitylation of a target protein in cells. These assays utilized TUBEs (Tandem Ubiquitin Binding Entities) to detect ubiquitylated substrates, and are individually optimized for a specific substrate protein.
LifeSensors’ UbiQuant™ ELISA is intended for the accurate determination of the concentration of total ubiquitin (poly- + mono-) in cell and tissue lysates. This assay is useful for monitoring the effects of broad-spectrum compounds such as proteasome inhibitors on total ubiquitin levels.
Conceptual schematic of the UbiQuant ELISA assay. Ubiquitlyated proteins are captured by an immobilized ubiquitin affinity matrix. Subsequently, unbound proteins are washed away, and total free ubiquitin and ubiquitin conjugates are quantified using an anti-ubiquitin antibody and traditional ELISA detection reagents.
Sample data from the UbiQuant ELISA assay. A serial dilution of ubiquitin was analyzed by the UbiQuant ELISA assay. UbiQuant S
The UbiQuant S ELISA provides a plate based method of quantifying the ubiquitilation of your protein of interest. This is accomplished by by capturing all ubiquitylated proteins, and then probing your protein of interest with a highly specific antibody. There is a more effective version of this assay, UbiQuant AlphaLISA. However, this product is not sold off the shelf, but is performed as a service. Contact us to learn more about AlphaLISA.
Conceptual Schematic of the UbiQuant S ELISA assay.Polyubiquitlyated proteins are captured by an immobilized ubiquitin affinity matrix. Subsequently, unbound proteins are washed away, and ubiquitylated protein of interest bound to the matrix is identified using a POI-specific antibody and traditional ELISA detection reagents.
Sample data from UbiQuant S ELISA and AlphaLISA assaysJurkat cells were treated with bortezomib (BTZ) or BTZ in combination with selective USP7 inhibitor P0217564 (564). Following treatment and cell lysis, Ubiquitylation of USP7 was measured by the UbiQuant S ELISA and AlphaLISA assays.