Comprehensive human urine standards for comparability and standardization in clinical proteome analysis

Abstract

Purpose: Urine proteomics is emerging as a powerful tool for biomarker discovery. The purpose of this study is the development of a well-characterized ''real life'' sample that can be used as reference standard in urine clinical proteomics studies. Experimental design: We report on the generation of male and female urine samples that are extensively characterized by different platforms and methods (CE-MS, LC-MS, LC-MS/MS, 1-D gel analysis in combination with nano-LC MS/MS (using LTQ-FT ultra), and 2-DE-MS) for their proteome and peptidome. In several cases analysis involved a definition of the actual biochemical entities, i.e. proteins/peptides associated with molecular mass and detected PTMs and the relative abundance of these compounds. Results: The combination of different technologies allowed coverage of a wide mass range revealing the advantages and complementarities of the different technologies. Application of these samples in ''inter-laboratory'' and ''inter-platform'' data comparison is also demonstrated. Conclusions and clinical relevance: These well-characterized urine samples are freely available upon request to enable data comparison especially in the context of biomarker discovery and validation studies. It is also expected that they will provide the basis for the comprehensive characterization of the urinary proteome.

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“…Finally, some of the rodent markers were detected in human samples using the identical CE-MS platform. This employed concept is in line with current guidelines for clinical proteome analysis (33,34).…”

Profilin 1 is a Potential Biomarker for Bladder Cancer Aggressiveness

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Does urinary peptide content differ between COPD patients with and without inherited alpha1-antitrypsin deficiency?

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“…18 For analysis, urine was diluted in a solution containing 2 M urea, 10 mM NH 4 OH and 0.02% (w/v) sodium dodecyl sulfate. To remove higher molecular mass proteins, such as albumin and immunoglobulin G, each sample was ultrafiltered using a Centrisart ultracentrifugation filter device (20 kDa cutoff Vivaspin; Sartorius, Goettingen, Germany) at 3,000× g until 1.1 mL of filtrate was obtained.…”