定量蛋白质组学【荐】.pptVIP

Expression Proteomics Expression proteomics:quality and quantity of the proteins expressed of the cell. Technology: 1.Isolation: SDSgel, HPLC (high performance liquid chromatography), CE (capillary electrophoresis) 2.Identification: mass spectrometry 3.Quantitation: ICAT, DIGE Function Proteomics Function proteomics: the function of the proteins, mainly proteins interaction. Technology: 1.yeast two hybrid 2.phage display 3.TAP(tandem affinity purification) 4. Bioinformatics Quantitative proteome Definition The global analysis of protein expression, a complementary method to study steady-state gene expression and perturbation-induced changes. Relative quantitative methods A. 2DE-based B. LC-based 2-DE 原理 首先根据等电点的不同将蛋白质分离； 再根据分子量大小的不同，将蛋白质分离。 Protein level: 2DE Differential In-gel Electrophoresis (DIGE) DIGE289-63up-36down LC-based approach Peptide level: shotgun approach Protein level: PF-2D approach Shotgun approach: Peptide level 1. Label: chemical probes 2. Separation: 2D-LC/MS/MS (SCX-RP) 3. Identification: mass spectrometry and database searching algorithm Quantitation Principle (Peptide level) Quantitation method: 1. Peptide abundance peak area peak height signal/noise ratio 2. Protein spectrum number Isotope-coded affinity tags (ICAT)in vitro ITRAQ * Quantitative proteomics Introduction Aims of Quantitative Proteomics Drug target discovery Drug effects on organisms Biomarkers identification/validation Disease mechanisms Function identification of Genes Development mechanisms 2DE flow chart ESI-MS/MS MAIDI-TOF Data Processing Shotgun flow chart SCX RP Data Processing lit Gygi,S.P.,et al. Nat Biotech, 1999, 17, 994-9