Application of a label-free, gel-free quantitative proteomics method for ecotoxicological studies of small fish species.

Journal Article (Journal Article)

Although two-dimensional electrophoresis (2D-GE) remains the basis for many ecotoxicoproteomic analyses, newer non-gel-based methods are beginning to be applied to overcome throughput and coverage limitations of 2D-GE. The overall objective of our research was to apply a comprehensive, liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomic approach to identify and quantify differentially expressed hepatic proteins from female fathead minnows exposed to fadrozole, a potent inhibitor of estrogen synthesis. Female fathead minnows were exposed to 0 (control), 0.04, and 1.0 μg of fadrozole/L of water for 4 days, and proteomic analysis was performed. Proteins were extracted and digested, and proteolytic peptides were separated via high-resolution one- or two-dimensional (1-D or 2-D) ultrapressure liquid chromatography (UPLC) and analyzed by tandem mass spectrometry. Mass spectra were searched against the National Center for Biotechnology Information (NCBI) ray-finned fish ( Actinopterygii ) database, resulting in identification of 782 unique proteins by single-dimension UPLC. When multidimensional LC analysis (2-D) was performed, an average increase of 1.9× in the number of identified proteins was observed. Differentially expressed proteins in fadrozole exposures were consistent with changes in liver function, including a decline in concentrations of vitellogenin as well as other proteins associated with endocrine function and cholesterol synthesis. Overall, these results demonstrate that a gel-free, label-free proteomic analysis method can successfully be utilized to determine differentially expressed proteins in small fish species after toxicant exposure.

Full Text

Duke Authors

Cited Authors

  • Ralston-Hooper, KJ; Turner, ME; Soderblom, EJ; Villeneuve, D; Ankley, GT; Moseley, MA; Hoke, RA; Ferguson, PL

Published Date

  • January 15, 2013

Published In

Volume / Issue

  • 47 / 2

Start / End Page

  • 1091 - 1100

PubMed ID

  • 23181269

Electronic International Standard Serial Number (EISSN)

  • 1520-5851

Digital Object Identifier (DOI)

  • 10.1021/es303170u


  • eng

Conference Location

  • United States