title: Modified filter-aided sample preparation (FASP) method increases peptide and protein identifications for shotgun proteomics.
abstract: BACKGROUND
  Mass spectrometry (MS)-based protein identification depends mainly on protein extraction and digestion. Although sodium dodecyl sulfate (SDS) can preclude enzymatic digestion and interfere with MS analysis, it is still the most widely used surfactant in these steps. To overcome these disadvantages, a SDS-compatible proteomic technique for SDS removal prior to MS-based analyses was developed, namely filter-aided sample preparation (FASP).
  
  
  METHODS
  Herein, based on the effectiveness of sodium deoxycholate and a detergent removal spin column, we developed a modified FASP (mFASP) method and compared its overall performance, total number of peptides and proteins identified for shotgun proteomic experiments with that of the FASP method.
  
  
  RESULTS
  Identification of 4570 ± 392 and 9139 ± 317 peptides and description of 862 ± 46 and 1377 ± 33 protein groups with two or more peptides from the ovarian cancer cell line A2780 was accomplished by FASP and mFASP methods, respectively. The mFASP method (21.2 ± 0.2%) had higher average peptide to protein coverage than FASP method (13.2 ± 0.5%). More hydrophobic peptides were identified by mFASP than by FASP, as indicated by the GRAVY score distribution.
  
  
  CONCLUSIONS
  The reported method enables reliable and efficient identification of proteins and peptides in whole-cell extracts containing SDS. The new approach allows for higher throughput (the simultaneous identification of more proteins), a more comprehensive investigation of proteins, and potentially the discovery of new biomarkers. Copyright © 2016 John Wiley & Sons, Ltd.
authors: Zhejiang Cancer Hospital, Hangzhou, Zhejiang, P.R. China.
authors: Mao-Wei
authors: MW
authors: Ni
authors: Zhejiang Province Key Laboratory of Anti-Cancer Drug Research, Zhejiang University, Hangzhou, 310058, Zhejiang, China.|Institute of Drug Metabolism and Analysis, College of Pharmaceutical Sciences, Hangzhou, 310058, Zhejiang, China.
authors: Lu
authors: L
authors: Wang
authors: Zhejiang Cancer Hospital, Hangzhou, Zhejiang, P.R. China.
authors: Wei
authors: W
authors: Chen
authors: Zhejiang Cancer Hospital, Hangzhou, Zhejiang, P.R. China.
authors: Han-Zhou
authors: HZ
authors: Mou
authors: Zhejiang Academy of Traditional Chinese Medicine, Development and Research Center of Official Silkworm Resources, Hangzhou, Zhejiang, P.R. China.
authors: Jie
authors: J
authors: Zhou
authors: Zhejiang Cancer Hospital, Hangzhou, Zhejiang, P.R. China.
authors: Zhi-Guo
authors: ZG
authors: Zheng
mesh_terms: D045744:Cell Line, Tumor; D003840:Deoxycholic Acid; D003902:Detergents; D005374:Filtration; D006801:Humans; D010446:Peptide Fragments / Q000032:analysis* / Q000737:chemistry; D011506:Proteins / Q000032:analysis* / Q000737:chemistry; D020543:Proteome / Q000032:analysis / Q000737:chemistry; D040901:Proteomics / Q000379:methods*; D012967:Sodium Dodecyl Sulfate; D014357:Trypsin
chemical_list: D003902:Detergents; D010446:Peptide Fragments; D011506:Proteins; D020543:Proteome; D003840:Deoxycholic Acid; D012967:Sodium Dodecyl Sulfate; D014357:Trypsin