DIABLA: a new screening method for the discovery of protein targets.
Dynamic isoelectric/anisotropy binding ligand assay (DIABLA) is a new method to identify proteins in a complex sample that bind to a molecule of interest. This is accomplished by first using capillary isoelectric focusing (cIEF) to separate the proteins in a capillary based on their isoelectric point. This separation is performed while the compound being tested is present in the separation buffer. When the proteins are focused, the entire capillary is scanned to identify regions of nonzero anisotropy, which are locations where the test compound is interacting with a focused protein band. DIABLA was demonstrated by observing the binding of fluorescein-tagged progesterone to an MCF-7 breast cancer cell lysate. The proteins were tagged with rhodamine to permit their observation and then focused in the presence of the tagged progesterone. Anisotropy measurements show that progesterone binds to six different proteins bands in the sample.
Montgomery
Robbie
R
Department of Chemistry, Southern Illinois University, Carbondale, Illinois 62901, USA.
Shay
Hanna
H
McCarroll
Matthew
M
Tolley
Luke
L
D000818:Animals; D016880:Anisotropy; D045744:Cell Line, Tumor; D006801:Humans; D007525:Isoelectric Focusing / Q000295:instrumentation / Q000379:methods*; D011374:Progesterone / Q000737:chemistry / Q000378:metabolism; D011506:Proteins / Q000032:analysis*
D011506:Proteins; D011374:Progesterone