UBIQAPTURE-Q®kit

Uses: Isolation and detection of the full range of ubiquitinylated protein conjugates (mono-/multi-/poly-ubiquitin modified, lysine linkage independent) from specific cell/tissue lysates of interest, especially low abundance (regulatory) proteins involved in the ubiquitin-proteasome pathway. Capture and analysis of specific ubiquitinylated protein conjugates of interest from particular cell/tissue lysates. Purification/pull down of ubiquitinylated proteins from cell free in vitro assays; for example to isolate Ub-p53 species from ubiquitinylation assays mediated by HeLa S100 lysate (Prod. No. BML-SW8750) . Release of free proteins in their active/native form by cleavage of ubiquitin/ubiquitin chains from the UBI-QAPTURE-Q ® matrix using deubiquitinylating enzymes such as USP2 (Prod. No. BML-UW9850) . Release of ubiquitinylated proteins in their active/native form by elution from the UBI-QAPTURE-Q ® matrix using, for example, high salt buffer. Note : Protocol provided covers applications 1-3. Assay set-up can be readily modified for alternative applications by inclusion, omission or substitution of specific components. Kit is not suitable for use with purified proteins.

UBI-QAPTURE-Q ® matrix (Prod. No. BML-UW0125) 0.5 ml settled resin provided; 50% suspension. Control ubiquitinylated-protein lysate (Ub-lysate) (Prod. No. BML-UW0130) 500 μg (100μl), 5mg/ml in 50mM HEPES, pH 7.6, 1mM DTT. Mono- and polyubiquitinylated conjugates monoclonal antibody (UBCJ2) (HRP conjugate) (Ubiquitin-conjugates specific HRP-linked antibody solution) (Prod. No. ENZ-ABS840HRP) 25μl.