Mutations in EFEMP1 in Patients with Juvenile Open-Angle Glaucoma

Ubiquitin (Ub) is a posttranslational modifier, and total Ub (UbT) is always in dynamic equilibrium among free Ub (UbF), activated Ub (UbA), and conjugated Ub (UbC) in the forms of mono-Ub, thioester-bond-linked Ub, and peptide-bond-linked Ub, respectively. In this study, we developed a simple method to simultaneously determine the levels of UbT, UbF + UbA, and UbC in a single immunoblot and demonstrated its reliability and reproducibility by determining [UbT], [UbF + UbA], and [UbC] in various mouse tissues and cultured cells.