Published Study Validates New Protein Enrichment Approach For Low-Abundance Biomarker Detection

Hercules, CA — April 20, 2011 — University of Minnesota researchers found that Bio-Rad Laboratories' ProteoMiner protein enrichment kit enhanced identification of changes to low-abundance proteins and detection of post-translationally modified (PTM) proteins in human saliva. These findings offer promise for improving differential proteomic analyses and biomarker studies aimed at identifying disease-specific proteins and their PTM variants in various types of biological samples and fluids. The study was published in the Dec. 13, 2010, issue of the Journal of Proteome Research.

ven when highly sensitive mass spectrometers are used to analyze complex biological samples and bodily fluids, high-abundance proteins obscure the detection of lower-abundance proteins and their post-translational modifications," said Sri Bandhakavi, who led the study at the University of Minnesota in 2010. (Bandhakavi is now a senior scientist at Bio-Rad.) "These lower-abundance proteins and PTMs are often of most interest to researchers, given their association with specific disease or physiological states."

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A single blood drop could detect heart disease, cancer

A University of Victoria researcher hopes to change the nature of testing forheart disease, cancer and drug toxicity using a highly sensitive and fast machine that would only require a single drop of blood from a patient.

Called a mass spectrometer, this machine determines the weight of protein molecules in the blood, and would allow researchers to determine if key marker proteins related to heart disease or cancer are present. The mass spectrometer being used in this research is among the most sensitive spectrometers that are commercially available, and is currently the only one of its kind in Canada.

Dr. Christoph Borchers at the University of Victoria-Genome BC Proteomics Centre will use the Agilent ion funnel 6490 mass spectrometer to develop methodologies for early diagnostic tests. These tests will detect and measure biomarkers, which are proteins in a patient’s blood that can signal early and subtle health changes. Dr. Borchers hopes to apply the technology to develop inexpensive, fast, and reproducible biomarker tests for early diagnosis of cardiovascular disease (CVD), the leading cause of death in the Western hemisphere.

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Strategy for purification and mass spectrometry identification of SELDI peaks corresponding to low-abundance plasma and serum proteins.

Abstract

Analysis by SELDI-TOF-MS of low abundance proteins makes it possible to select peaks as candidate biomarkers. Our aim was to define a purification strategy to optimise identification by MS of peaks detected by SELDI-TOF-MS from plasma or serum, regardless of any treatment by a combinatorial peptide ligand library (CPLL). We describe 2 principal steps in purification. First, choosing the appropriate sample containing the selected peak requires setting up a databank that records all the m/z peaks detected from samples in different conditions. Second, the specific purification process must be chosen: separation was achieved with either chromatographic columns or liquid-phase isoelectric focusing, both combined when appropriate with reverse-phase chromatography. After purification, peaks were separated by gel electrophoresis and the candidate proteins analyzed by nano-liquid-chromatography-MS/MS. We chose 4m/z peaks (9400, 13571, 13800 and 15557) selected for their differential expression between two conditions, as examples to explain the different strategies of purification, and we successfully identified 3 of them. Despite some limitations, our strategy to purify and identify peaks selected from SELDI-TOF-MS analysis was effective.

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Clinical proteomics for liver disease: a promising approach for discovery of novel biomarkers

Hepatocellular carcinoma (HCC) is the fifth most common cancer and advanced hepatic fibrosis is a major risk factor for HCC. Hepatic fibrosis including liver cirrhosis and HCC are mainly induced by persistent hepatitis B or C virus infection, with approximately 500 million people infected with hepatitis B or C virus worldwide. Furthermore, the number of patients with non-alcoholic fatty liver disease (NAFLD) has recently increased and NAFLD can progress to cirrhosis and HCC. These chronic liver diseases are major causes of morbidity and mortality, and the identification of non-invasive biomarkers is important for early diagnosis. Recent advancements in quantitative and large-scale proteomic methods could be used to optimize the clinical application of biomarkers. Early diagnosis of HCC and assessment of the stage of hepatic fibrosis or NAFLD can also contribute to more effective therapeutic interventions and an improve prognosis. Furthermore, advancements of proteomic techniques contribute not only to the discovery of clinically useful biomarkers, but also in clarifying the molecular mechanisms of disease pathogenesis by using body fluids, such as serum, and tissue samples and cultured cells. In this review, we report recent advances in quantitative proteomics and several findings focused on liver diseases, including HCC, NAFLD, hepatic fibrosis and hepatitis B or C virus infections.

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