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Proteomic Challenges: Sample Preparation Techniques for Microgram-Quantity Protein Analysis from Biological Samples

Department of Chemistry and Biochemistry, Integrated Biomedical Sciences Program, and the Harper Cancer Research Institute, 251 Nieuwland Science Hall, University of Notre Dame, Notre Dame, IN 46556, USA
Author to whom correspondence should be addressed.
Academic Editor: Johannes Haybaeck
Int. J. Mol. Sci. 2015, 16(2), 3537-3563;
Received: 21 November 2014 / Accepted: 29 January 2015 / Published: 5 February 2015
(This article belongs to the Collection Molecular Mechanisms of Human Liver Diseases)
PDF [1882 KB, uploaded 5 February 2015]


Proteins regulate many cellular functions and analyzing the presence and abundance of proteins in biological samples are central focuses in proteomics. The discovery and validation of biomarkers, pathways, and drug targets for various diseases can be accomplished using mass spectrometry-based proteomics. However, with mass-limited samples like tumor biopsies, it can be challenging to obtain sufficient amounts of proteins to generate high-quality mass spectrometric data. Techniques developed for macroscale quantities recover sufficient amounts of protein from milligram quantities of starting material, but sample losses become crippling with these techniques when only microgram amounts of material are available. To combat this challenge, proteomicists have developed micro-scale techniques that are compatible with decreased sample size (100 μg or lower) and still enable excellent proteome coverage. Extraction, contaminant removal, protein quantitation, and sample handling techniques for the microgram protein range are reviewed here, with an emphasis on liquid chromatography and bottom-up mass spectrometry-compatible techniques. Also, a range of biological specimens, including mammalian tissues and model cell culture systems, are discussed. View Full-Text
Keywords: proteins; mass-spectrometry; mass-limited samples; microgram samples proteins; mass-spectrometry; mass-limited samples; microgram samples

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Feist, P.; Hummon, A.B. Proteomic Challenges: Sample Preparation Techniques for Microgram-Quantity Protein Analysis from Biological Samples. Int. J. Mol. Sci. 2015, 16, 3537-3563.

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