Quick 96FASP for high throughput quantitative proteome analysis

摘要: Abstract Filter aided sample preparation (FASP) is becoming a central method for proteomic cleanup and peptide generation prior to LC-MS analysis. We previously adapted this 96-well filter plate, applied prepare protein digests from cell lysate body fluid samples in high throughput quantitative manner. While the 96FASP approach scalable can handle multiple simultaneously, two key advantages compared single FASP, it also time-consuming. The centrifugation-based liquid transfer on plate takes 3–5 times longer than filter. To address limitation, we now present quick (named q96FASP) that, relying use of membranes with large MWCO size (~ 30 kDa), significantly reduces centrifugal times. show that q96FASP allows derived whole lysates fluids quality similar FASP method. Processing wells parallel, observed excellent experimental repeatability by label-free quantitation approach. conclude promises be promising cost- time-effective shotgun proteomics will particularly useful scale biomarker discovery studies. Significance High processing particular interests proteomics. developed appealing, however time-consuming context (~ 1.5 h per spin). This study presents truly based cut-off which shortens spin time ~ 20 min. our knowledge, first multi-well entirely comparable conventional FASP. thoroughly examined types plates performed side-by-side comparisons Two samples, UTI (urinary tract infection)-associated Klebsiella pneumoniae human urine, were tested demonstrated its capability q96FSAP makes plate-based more appealing projects, could broadly