Rapid Sample Processing For LC-MS Based Quantitative Proteomics Using High Intensity Focused Ultrasounds

Journal Article · Mon Sep 01 00:00:00 EDT 2008 · Journal of Proteome Research, 7(9):3860-3867

DOI:https://doi.org/10.1021/pr800161x· OSTI ID:947042

Lopez-Ferrer, Daniel; Heibeck, Tyler H.; Petritis, Konstantinos; Hixson, Kim K.; Qian, Weijun; Monroe, Matthew E.; Mayampurath, Anoop M.; Moore, Ronald J.; Belov, Mikhail E.; Camp, David G.; Smith, Richard D.

A new sample processing workflow that uses high intensity focused ultrasound to rapidly reduce and alkylate cysteines, digest proteins and then label peptides with 18O was developed for quantitative proteomics applications. Each step was individually refined to minimize reaction times, peptide loses and undesired by-products or modifications. By using this novel workflow, mouse plasma proteins were successfully denatured, alkylated, in-solution digested, and 18O labelled 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.

• Research Organization: Pacific Northwest National Laboratory (PNNL), Richland, WA (US)
• Sponsoring Organization: USDOE
• DOE Contract Number: AC05-76RL01830
• OSTI ID: 947042
• Report Number(s): PNNL-SA-59400; KP1601010
• Journal Information: Journal of Proteome Research, 7(9):3860-3867, Journal Name: Journal of Proteome Research, 7(9):3860-3867 Journal Issue: 9 Vol. 7
• Country of Publication: United States
• Language: English