title: Direct digestion of proteins in living cells into peptides for proteomic analysis.
abstract: To analyze the proteome of an extremely low number of cells or even a single cell, we established a new method of digesting whole cells into mass-spectrometry-identifiable peptides in a single step within 2 h. Our sampling method greatly simplified the processes of cell lysis, protein extraction, protein purification, and overnight digestion, without compromising efficiency. We used our method to digest hundred-scale cells. As far as we know, there is no report of proteome analysis starting directly with as few as 100 cells. We identified an average of 109 proteins from 100 cells, and with three replicates, the number of proteins rose to 204. Good reproducibility was achieved, showing stability and reliability of the method. Gene Ontology analysis revealed that proteins in different cellular compartments were well represented.
authors.affiliation: Department of Chemistry, Institute of Biomedical Sciences, Fudan University, Shanghai, 200433, China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Chen
authors.forename: Guoquan
authors.initials: G
authors.lastname: Yan
authors.forename: Mingxia
authors.initials: M
authors.lastname: Gao
authors.forename: Xiangmin
authors.initials: X
authors.lastname: Zhang
mesh_terms: D002853:Chromatography, Liquid / Q000379:methods; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods; D010455:Peptides / Q000032:analysis*; D011506:Proteins / Q000032:analysis / Q000737:chemistry*; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results; D014357:Trypsin / Q000737:chemistry; D014407:Tumor Cells, Cultured
chemical_list: D010455:Peptides; D011506:Proteins; D014357:Trypsin