title: Rapid sample processing for LC-MS-based quantitative proteomics using high intensity focused ultrasound.
abstract: A new sample processing workflow that uses high intensity focused ultrasound to rapidly reduce and alkylate cysteines, digest proteins and then label peptides with (18)O was developed for quantitative proteomics applications. Each step was individually refined to minimize reaction times, peptide loses and undesired byproducts or modifications. When this novel workflow was used, mouse plasma proteins were successfully denatured, alkylated, in-solution digested, and (18)O-labeled in <10 min for subsequent analysis by liquid chromatography-electrospray ionization high resolution mass spectrometry. Performance was evaluated in terms of the number of mouse plasma peptides and proteins identified in a shotgun approach and the quantitative dynamic range. The results were compared with previously published results obtained using conventional sample preparation methods and were found to be similar. Advantages of the new method include greatly simplified and accelerated sample processing, as well as being readily amenable to automation.
authors.affiliation: Biological Science Division and Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, Washington 99352, USA.
authors.forename: Daniel
authors.initials: D
authors.lastname: López-Ferrer
authors.forename: Tyler H
authors.initials: TH
authors.lastname: Heibeck
authors.forename: Konstantinos
authors.initials: K
authors.lastname: Petritis
authors.forename: Kim K
authors.initials: KK
authors.lastname: Hixson
authors.forename: Weijun
authors.initials: W
authors.lastname: Qian
authors.forename: Matthew E
authors.initials: ME
authors.lastname: Monroe
authors.forename: Anoop
authors.initials: A
authors.lastname: Mayampurath
authors.forename: Ronald J
authors.initials: RJ
authors.lastname: Moore
authors.forename: Mikhail E
authors.initials: ME
authors.lastname: Belov
authors.forename: David G
authors.initials: DG
authors.lastname: Camp
authors.forename: Richard D
authors.initials: RD
authors.lastname: Smith
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002853:Chromatography, Liquid / Q000379:methods*; D051379:Mice; D008969:Molecular Sequence Data; D040901:Proteomics; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*