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Ultra-Fast Sample Preparation for High-Throughput Proteomics

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Sample Preparation in Biological Mass Spectrometry

Abstract

Sample preparation oftentimes can be the Achilles Heel of any analytical process, and in the field of proteomics, preparing samples for mass spectrometric analysis is no exception. Current goals, concerning proteomic sample preparation on a large scale, include efforts toward improving reproducibility, reducing the time of processing and ultimately the automation of the entire workflow. This chapter reviews an array of recent approaches applied to bottom-up proteomics sample preparation to reduce the processing time down from hours to minutes. The current state-of-the-art approaches in the field use different energy inputs such as microwave, ultrasound or pressure to perform the four basic steps in sample preparation: protein extraction, denaturation, reduction/alkylation, and digestion. No single energy input for enhancement of proteome sample preparation has become the universal gold standard. Instead, a combination of different energy inputs tends to produce the best results. This chapter further describes the future trends in the field such as the hyphenation of sample preparation with downstream detection and analysis systems. Finally, a detailed protocol describing the combined use of both pressure cycling technology and ultrasonic energy inputs to hasten proteomic sample preparation is presented.

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Abbreviations

ESI:

Electrospray ionization

FDR:

False discovery rate

HIFU:

High intensity focused ultrasound

HPP:

High pressure processing

IAM:

Iodoacetamide

IT:

Ion trap

MAPED:

Microwave-assisted protein enzymatic digestion

MS:

Mass spectrometry

MS/MS:

Tandem mass spectrometry

MW:

Molecular weight

NA:

Not assigned

PCT:

Pressure cycling technology

RP:

Reversed phase

TCEP:

Tris(2-carboxyethyl) phosphine

TOF:

Time-of-flight

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Acknowledgments

Portions of this work were supported by the NIH National Center for Research Resources (NCRR, RR018522), NIH National Cancer Institute (R21 CA12619-01), and the Pacific Northwest National Laboratory’s (PNNL) Laboratory Directed Research and Development Program. This research was enabled in part by capabilities developed under support from the U.S. Department of Energy (DOE) Office of Biological and Environmental Research and the NCRR, and was conducted in the Environmental Molecular Sciences Laboratory, a DOE national scientific user facility located at the Pacific Northwest National Laboratory (PNNL) in Richland, WA. PNNL is a multiprogram national laboratory operated by Battelle for the DOE under Contract No. DE-AC05-76RLO 1830.

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Authors and Affiliations

  1. Pacific Northwest National Laboratory, Biological Science Division, Richland, WA, USA

    Daniel Lopez-Ferrer

  2. Caprion Proteomics U.S., LLC, 1455 Adams Drive, Menlo Park, CA, 94025, USA

    Daniel Lopez-Ferrer

  3. Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, WA, 99352, USA

    Kim K. Hixson

  4. Biological Science Division, Pacific Northwest National Laboratory, Richland, WA, 99352, USA

    Mikhail E. Belov & Richard D. Smith

Authors
  1. Daniel Lopez-Ferrer
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  2. Kim K. Hixson
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  3. Mikhail E. Belov
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  4. Richard D. Smith
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Corresponding author

Correspondence to Daniel Lopez-Ferrer .

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Editors and Affiliations

  1. , Department of Genetics and Complex Disea, Harvard School of Public Health, Harvard, Huntington Ave., SPH-1 665, Boston, 02115, Massachusetts, USA

    Alexander R. Ivanov

  2. , Research and Development, Pressure BioSciences, Inc., Norfolk Ave. 14, South Easton, 02375, Massachusetts, USA

    Alexander V. Lazarev

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Lopez-Ferrer, D., Hixson, K.K., Belov, M.E., Smith, R.D. (2011). Ultra-Fast Sample Preparation for High-Throughput Proteomics. In: Ivanov, A., Lazarev, A. (eds) Sample Preparation in Biological Mass Spectrometry. Springer, Dordrecht. https://doi.org/10.1007/978-94-007-0828-0_8

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