? Top-down proteome profiling and protein characterization to measure reproducibly 1500 protein features in human whole salivary fluids will be performed to assess the suitability of saliva for cancer detection and diagnosis. Plasma or blood has been the accepted biofluid of choice for measuring levels of proteins and other biomolecules for clinical testing, but there may be advantages for other biomedia which can be more readily obtained for clinical sampling. Saliva has a rich proteome that is derived from the salivary glands, the linings of the oral cavity, and blood. Saliva is readily available and easier to collect compared to other body fluids. Compared to plasma, with highly abundant albumin, transferrin, and IgGs, the salivary proteome presents fewer challenges for accessing the entire protein complement. For these reasons, the use of saliva for cancer diagnostic purposes presents an attractive option. Protein pre-fractionation strategies will be developed to partition the salivary proteome for further characterization by top-down proteomics. Three methods will be investigated: separation of proteins by their hydrophobicity via HPLC; solution isoelectric focusing to partition the complex mixture into 5-7 separate sub-samples on the basis of their pl; affinity selection of N-linked glycoproteins by a hydrazide resin method, and the formerly glycosylated proteins will be released for subsequent MS analysis. The goal will be to resolve and observe more lower abundance proteins than without pre-fractionation. A new method based on a combined laser desorption and electrospray ionization, termed electrospray-assisted laser desorption/ionization (ELDI), will be further developed to provide top-down MS/MS-generated sequence tags to identify the relevant protein markers of cancer. The overall platform of protein pre-fractionation and MALDI/ELDI-MS top-down proteomics will be applied to saliva samples from healthy control and oral cancer subjects. The goal will be to identify 1500 protein features reproducibly in the saliva samples from the healthy subjects and compare these characteristics to the saliva proteins measured in oral cancer samples. Relevance to public health: Salivary fluid may be an attractive sample for the identification of markers of diseases, such as cancer. Top-down proteomics will be applied to compare proteins in saliva from healthy individuals and cancer subjects. ? ? ?
|Peng, Ivory X; Ogorzalek Loo, Rachel R; Shiea, Jentaie et al. (2008) Reactive-electrospray-assisted laser desorption/ionization for characterization of peptides and proteins. Anal Chem 80:6995-7003|
|Peng, Ivory X; Shiea, Jentaie; Ogorzalek Loo, Rachel R et al. (2007) Electrospray-assisted laser desorption/ionization and tandem mass spectrometry of peptides and proteins. Rapid Commun Mass Spectrom 21:2541-6|