第二章 样品制备.pdfVIP

Proteome Study Workflow Sample preparation Sample preparation Sample separation Sample separation Protein identification Protein identification Protein quantification Protein quantification Protein function/protein interactions assay Chapter 2 Sample preparation Sample preparation is the first and key Sample preparation is the first and key step for successful proteome study step for successful proteome study It is difficult because It is difficult because  Too many different kinds of proteins  Too many different kinds of proteins  Different expression abundance  Different expression abundance  Different character  Different character Sample Preparation Method(s) Selection According to the goal (Try the best to get the need proteins) As many kinds of proteins as possible  Only some interested proteins  Should consider the combination with later separation method  2DE: the amount of SDS  HPLC: the amount of detergents/ampholytes Sample preparation No universal method Need to be optimized for each sample General recommendations Keep it simple to increase reproducibility 2.1 Principle for Sample Preparation A. Thorough, consistent method resulting in complete solubilisation, disaggregation, denaturation and reduction of all proteins under electrophoresis conditions Basic principles 1. Keep it simple to increase reproducibility 2. Minimize possibility of protein losses and modifications  keep sample treatment by a minimum step  keep sample as cold as possible 