Development of a high-throughput method for preparing human urine for two-dimensional electrophoresis

Development of a high-throughput method for preparing human urine for two-dimensional electrophoresis
11 May 2005
Received: 20 August 2004
Graeme Smith 1 *, Derek Barratt 2, Rachel Rowlinson 1, Janice Nickson 1, Robert Tonge 1
PROTEOMICS
Wiley InterScience
1Pathways, Global Sciences & Information, AstraZeneca, Macclesfield, Cheshire, UK
2Protein Science & Supply, Global Sciences & Information, AstraZeneca, Macclesfield, Cheshire, UK

email: Graeme Smith (graeme.smith@astrazeneca.com)
*Correspondence to Graeme Smith, Pathways, Global Sciences & Information, Mereside, AstraZeneca Alderley Park, Macclesfield, Cheshire, SK10 4TG, UK Fax: +44-1625-516443
Keywords
Biomarker ? Human urine ? Two-dimensional gel electrophoresis
Abstract
There is an increasing interest in analysing the human urinary proteome in the search for biomarkers. However, despite the ease of its collection, urine is a difficult fluid to analyse by two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) because of its dilute protein content and high salt levels. Here, we describe a method for high-throughput processing of urine for 2-D PAGE. Urine is filtered and applied to solid phase extraction columns. After washing, the urinary proteins are eluted and freeze dried. The lyophilised powder can then be resuspended in an appropriate buffer for downstream proteomic analysis.
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