Isoelectric focusing technology quantifies protein signaling in 25 cells.
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.
O'Neill
Roger A
RA
Cell Biosciences Inc., 1050 Page Mill Road, Palo Alto, CA 94304, USA.
Bhamidipati
Arunashree
A
Bi
Xiahui
X
Deb-Basu
Debabrita
D
Cahill
Linda
L
Ferrante
Jason
J
Gentalen
Erik
E
Glazer
Marc
M
Gossett
John
J
Hacker
Kevin
K
Kirby
Celeste
C
Knittle
James
J
Loder
Robert
R
Mastroieni
Catherine
C
Maclaren
Michael
M
Mills
Thomas
T
Nguyen
Uyen
U
Parker
Nineveh
N
Rice
Audie
A
Roach
David
D
Suich
Daniel
D
Voehringer
David
D
Voss
Karl
K
Yang
Jade
J
Yang
Tom
T
Vander Horn
Peter B
PB
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
D020033:Protein Isoforms; D011506:Proteins