Simple Tip-Based Sample Processing Method for Urinary Proteomic Analysis

David J. Clark, Yingwei Hu, Michael Schnaubelt, Yi Fu, Sean Ponce, Shao Yung Chen, Yangying Zhou, Punit Shah, Hui Zhang

Research output: Contribution to journalArticle

Abstract

Mass spectrometry-based urinary proteomics is one of the most attractive strategies to discover proteins for diagnosis, prognosis, monitoring, or prediction of therapeutic responses of urological diseases involving the kidney, prostate, and bladder; however, interfering compounds found in urine necessitate sample preparation strategies that are currently not suitable for urinary proteomics in the clinical setting. Herein, we describe the C4-tip method, comprising a simple, automated strategy utilizing a reverse-phase resin tip-based format and "on-tip" digestion to examine the urine proteome. We first determined the optimal conditions for protein isolation and protease digestion on the C4-tip using the standard protein bovine fetuin. Next, we applied the C4-tip method to urinary proteomics, identifying a total of 813 protein groups using LC-MS/MS, with identified proteins from the C4-tip method displaying a similar distribution of gene ontology (GO) cellular component assignments compared to identified proteins from an ultrafiltration preparation method. Finally, we assessed the reproducibility of the C4-tip method, revealing a high Spearman correlation R-value for shared proteins identified across all tips. Together, we have shown the C4-tip method to be a simple, robust method for high-throughput analysis of the urinary proteome by mass spectrometry in the clinical setting.

Original languageEnglish (US)
Pages (from-to)5517-5522
Number of pages6
JournalAnalytical chemistry
Volume91
Issue number9
DOIs
StatePublished - May 7 2019

    Fingerprint

ASJC Scopus subject areas

  • Analytical Chemistry

Cite this

Clark, D. J., Hu, Y., Schnaubelt, M., Fu, Y., Ponce, S., Chen, S. Y., Zhou, Y., Shah, P., & Zhang, H. (2019). Simple Tip-Based Sample Processing Method for Urinary Proteomic Analysis. Analytical chemistry, 91(9), 5517-5522. https://doi.org/10.1021/acs.analchem.8b05234