title: Isoelectric focusing technology quantifies protein signaling in 25 cells.
abstract: A previously undescribed isoelectric focusing technology allows cell signaling to be quantitatively assessed in <25 cells. High-resolution capillary isoelectric focusing allows isoforms and individual phosphorylation forms to be resolved, often to baseline, in a 400-nl capillary. Key to the method is photochemical capture of the resolved protein forms. Once immobilized, the proteins can be probed with specific antibodies flowed through the capillary. Antibodies bound to their targets are detected by chemiluminescence. Because chemiluminescent substrates are flowed through the capillary during detection, localized substrate depletion is overcome, giving excellent linearity of response across several orders of magnitude. By analyzing pan-specific antibody signals from individual resolved forms of a protein, each of these can be quantified, without the problems associated with using multiple antibodies with different binding avidities to detect individual protein forms.
authors.affiliation: Cell Biosciences Inc., 1050 Page Mill Road, Palo Alto, CA 94304, USA.
authors.forename: Roger A
authors.initials: RA
authors.lastname: O'Neill
authors.forename: Arunashree
authors.initials: A
authors.lastname: Bhamidipati
authors.forename: Xiahui
authors.initials: X
authors.lastname: Bi
authors.forename: Debabrita
authors.initials: D
authors.lastname: Deb-Basu
authors.forename: Linda
authors.initials: L
authors.lastname: Cahill
authors.forename: Jason
authors.initials: J
authors.lastname: Ferrante
authors.forename: Erik
authors.initials: E
authors.lastname: Gentalen
authors.forename: Marc
authors.initials: M
authors.lastname: Glazer
authors.forename: John
authors.initials: J
authors.lastname: Gossett
authors.forename: Kevin
authors.initials: K
authors.lastname: Hacker
authors.forename: Celeste
authors.initials: C
authors.lastname: Kirby
authors.forename: James
authors.initials: J
authors.lastname: Knittle
authors.forename: Robert
authors.initials: R
authors.lastname: Loder
authors.forename: Catherine
authors.initials: C
authors.lastname: Mastroieni
authors.forename: Michael
authors.initials: M
authors.lastname: Maclaren
authors.forename: Thomas
authors.initials: T
authors.lastname: Mills
authors.forename: Uyen
authors.initials: U
authors.lastname: Nguyen
authors.forename: Nineveh
authors.initials: N
authors.lastname: Parker
authors.forename: Audie
authors.initials: A
authors.lastname: Rice
authors.forename: David
authors.initials: D
authors.lastname: Roach
authors.forename: Daniel
authors.initials: D
authors.lastname: Suich
authors.forename: David
authors.initials: D
authors.lastname: Voehringer
authors.forename: Karl
authors.initials: K
authors.lastname: Voss
authors.forename: Jade
authors.initials: J
authors.lastname: Yang
authors.forename: Tom
authors.initials: T
authors.lastname: Yang
authors.forename: Peter B
authors.initials: PB
authors.lastname: Vander Horn
mesh_terms: D045744:Cell Line, Tumor; D006801:Humans; D007525:Isoelectric Focusing / Q000379:methods*; D007526:Isoelectric Point; D010766:Phosphorylation; D020033:Protein Isoforms / Q000378:metabolism; D011506:Proteins / Q000378:metabolism*; D012680:Sensitivity and Specificity; D015398:Signal Transduction
chemical_list: D020033:Protein Isoforms; D011506:Proteins