Our new Human Phospho-Ubiquitin (S65) Antibody detects mono- and poly-Ubiquitin chains that are phosphorylated on serine 65 (see data below). It has no cross-reactivity with non-phosphorylated Ubiquitin, Ubiquitin phosphorylated on serine 57 or tyrosine 59, or phosphorylated Parkin. Start exploring the role of phosphorylated Ubiquitin!
Detection of Phospho-Ubiquitin (S65) by Western Blot. Tetraubiquitin chains of each indicated linkage type were incubated for 0-4 hours in reactions containing recombinant PINK1 Kinase (Cat# AP-180) and ATP. At indicated times a portion of each reaction was removed and terminated with SDS-PAGE sample buffer. SDS-PAGE gels (10-20%) were used to resolve approximately 150 ng of Ubiquitin tetramer from each reaction. Western blots were developed using either Rabbit Anti-Human Phospho-Ubiquitin (S65) Affinity-Purified Polyclonal Antibody (Catalog # A-110; upper panels) or Mouse Anti-Human Ubiquitin Monoclonal Antibody (Catalog # MAB701; lower panels).