Tandem Ubiquitin Binding Entities (TUBEs) capitalize on the linkage of multiple ubiquitin interacting motifs (UIMs) to generate reagents with high affinity for polyubiquitin. Biotin K63-TUBE binds K63-linked polyubiquitin with a 1000 to 10,000-fold preference for K63 over other linkage types and provides a sensitive and cost-effective tool for determining the abundance of K63-linked polyubiquitin in the cell, or identifying the linkage-type associated with a particular protein of interest. Based on an engineered peptide developed by Sims et al., Biotin K63-TUBE consists of multiple ubiquitin interacting motifs separated by rigid linkers, presumably increasing its affinity to extended K63 polyubiquitin chains. Biotin K63-TUBE is useful for detection and characterization of K63-linked polyubiquitinated proteins by Far Western blotting, affinity purification from lysates and biological fluids, and in situ detection. Far Westerns with Biotin K63-TUBE require only minor modifications to existing immunoblot protocols, the most notable of which is to avoid subjecting the blot to denaturing conditions, e.g. boiling or autoclaving, which are common treatments used in Western blotting protocols for ubiquitin antibodies. Biotin K63-TUBE substitutes for the anti-ubiquitin IgG as the primary detection reagent. Secondary detection is then accomplished with an avidin or streptavidin enzyme conjugate.
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|Molecular Weight||22 kDa|
|Storage||-80°C, avoid freeze/thaw cycles|