Ultrasensitive pathway phosphoproteomics

Award Information
Agency: Department of Health and Human Services
Branch: N/A
Contract: 1R43GM081956-01A1
Agency Tracking Number: GM081956
Amount: $194,424.00
Phase: Phase I
Program: SBIR
Awards Year: 2008
Solicitation Year: 2008
Solicitation Topic Code: N/A
Solicitation Number: PHS2007-2
Small Business Information
DUNS: 616743261
HUBZone Owned: Y
Woman Owned: Y
Socially and Economically Disadvantaged: Y
Principal Investigator
 () -
Business Contact
Phone: (650) 504-0376
Email: ciwang_99@yahoo.com
Research Institution
DESCRIPTION (provided by applicant): Site-specific protein phosphorylation is essential for signal transduction in mammalian cells, and perturbation in phosphoprotein levels is a hallmark of cancerous transformation. Unfortunately there are no sensitive me thods for detecting multiple phosphorylation events in parallel, so researchers are forced to run multiple Western blot or ELISA experiments, expending precious samples, in order to dissect these signaling pathways. We describe here a novel, high throughpu t, ultrasensitive method for detecting multiple phosphoproteins simultaneously. The method consists of a sandwich immuno- assay comprised of multiple pan-specific capture antibodies and a cocktail of detection antibodies against both phosphorylated and non -phosphorylated epitopes of the target proteins. Each detection antibody is labeled with a unique DNA tag. Real-time PCR is then used to quantify all of the DNA tags in parallel, thus revealing the quantities of the cognate (phospho-)proteins in the biolog ical samples. We show that this immuno-PCR method is more sensitive than ELISA and can detect endogenous proteins at sub-picomolar levels in real biological samples. In this grant application, we propose to develop and validate this concept by developing a highly sensitive, multiplexed assay to measure the phosphorylation states of signaling proteins in the BCR-ABL pathway. This platform will allow researchers to measure numerous phosphoproteins from minimal (~50 ul) sample volumes. Because all steps use mi crotiter plates and simple manipulations, this technology will allow researchers, with minimal effort and standard equipment, to decipher changes in phosphorylation patterns resulting from numerous biological stimuli.

* Information listed above is at the time of submission. *

Agency Micro-sites

SBA logo
Department of Agriculture logo
Department of Commerce logo
Department of Defense logo
Department of Education logo
Department of Energy logo
Department of Health and Human Services logo
Department of Homeland Security logo
Department of Transportation logo
Environmental Protection Agency logo
National Aeronautics and Space Administration logo
National Science Foundation logo
US Flag An Official Website of the United States Government