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Modern high-throughput methods for the proteome analysis are gradually replacing more traditional 2D gel-based techniques. Almost immediately after the introduction of high-throughput proteomics techniques in 2001, reproducibility of the results became an issue. Extensive discussion in the literature led to the conclusion that certain "undersampling" exhibited during measurements could be due to the stochastic nature of the data-dependent sampling, routinely used with current mass spectrometry equipment. At the same time, the effect of the acquisition of different charge states on the reproducibility and confidence of protein identifications, to the best of our knowledge, has never been properly evaluated. There exists the frequently voiced yet hardly documented opinion that +1 charge states should be rejected during data-dependent acquisition. The work presented here shows that inclusion of the +1 charge state in the data-dependent acquisition protocols can indeed lead to improved proteome coverage, reproducibility, and the confidence of protein identifications by high-throughput proteomics. It was also shown that contrary to the established opinion, gas-phase dissociation of singly charged peptide species results in rich fragmentation patterns containing both b- and y-ions allowing for successful and confident peptide identification.

Thousands of man-made chemicals are annually released into the environment by agriculture, transport, industries, and other human activities. In general, chemical analysis of environmental samples used to assess the pollution status of a specific ecosystem is complicated by the complexity of the mixture, and in some cases by the very low toxicity thresholds of chemicals present. In that sense, a proteomics approach, capable of detecting subtle changes in the level and structure of individual proteins within the whole proteome in response to the altered surroundings, has obvious applications in the field of ecotoxicology. In addition to identifying new protein biomarkers, it can also help to provide an insight into underlying mechanisms of toxicity. Despite being a comparatively new field with a number of caveats, proteomics applications have spread from microorganisms and plants to invertebrates and vertebrates, gradually becoming an established technology used in environmental research. This review article highlights recent advances in the field of environmental proteomics, mainly focusing on experimental approaches with a potential to understand toxic modes of action and to identify novel ecotoxicological biomarkers., (Copyright 2008 Wiley Periodicals, Inc.)