Simultaneous quantification of total and conjugated ubiquitin levels in a single immunoblot.
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.
School of Life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Republic of Korea.
Choongseob
C
Oh
Jong Hyuk
JH
Yoon
Soonyong
S
Park
Yung Joon
YJ
Yoo
D000818:Animals; D015153:Blotting, Western / Q000379:methods*; D002460:Cell Line; D006801:Humans; D051379:Mice; D010455:Peptides / Q000032:analysis; D015203:Reproducibility of Results; D013440:Sulfides / Q000032:analysis; D025801:Ubiquitin / Q000032:analysis*
D010455:Peptides; D013440:Sulfides; D025801:Ubiquitin
Ubiquitin; Ubiquitination; Western blot