伯乐公司-2D样品制备.ppt

To alleviate the dynamic range problem in serum plasma, many products utilizing immunodepletion have been developed (remove the top 7, 14, 20 proteins) Immunodepletion suffers from many limitations Reliant upon predefined set of antibodies- limits options available and the samples that can be treated with the products Dilutes the sample, need to concentrate samples afterwards Inadvertently lose associated proteins when high abundant proteins are removed Expensive, due to the use of antibodies Novel and simple protein enrichment and depletion tool for biological samples Reduces high abundant proteins Enriches medium low abundant proteins Improves upon limitations of immunodepletion products Decreases the amount of high-abundance proteins without losing associated proteins Enriches and concentrates low-abundance proteins that cannot be detected through traditional methods Not dependent on a predefined set of antibodies Can be used to decrease the dynamic range of the protein concentration in a variety of biological fluid Different kinds of protein in one cell Microsome Mitochondria Nucleus Cytoplasm Isolation of Subcellular Fractions Isolation of mouse liver nuclear proteins Cytoplasmic proteins Nuclei proteins pH 3-10 NL, 17 cm ReadyStrip IPG strips, 8 – 16% gel Protocols/Kits for Membrane Proteins Total protein Membrane I Membrane II Signal Temp-dep. differential solubilization (Triton X-114) Carbonate buffer extraction Triton X-100 differential solubility Caveoli-associated proteins ProteoMinerTM TechnologyDig Deeper in the Proteome Proteomics Biomarker Discovery Proteomics is seen as a potentially powerful tool for identifying biomarkers - Changes in protein content can provide valuable information - The proteome is highly dynamic and far more complex than the genome Protein changes as a response to changes in the organism’s biological state can be measured in tissue samples or body fluids. Plasma and serum are used most frequently - They