Mass spectrometric compatibility of Deep Purple and SYPRO Ruby total protein stains for high-throughput proteomics using large-format two-dimensional gel electrophoresis.

In order to identify putative biomarkers from two-dimensional (2D) gel electrophoresis it is necessary to use a visualization technique that is sensitive, has a large dynamic range and does not interfere with the identification of the protein. As mass spectrometry increases in sensitivity more pressure is placed on visualization techniques that facilitate proteomic workflows but do not interfere with downstream processing. Two stains reported to meet these requirements are SYPRO Ruby (Invitrogen) and Deep Purple (GE Healthcare). This study examined the compatibility of these stains with protein identification by selecting spots from replicate 2D gels of human plasma and subjecting these to protein identification using liquid chromatography/tandem mass spectrometry (LC/MS/MS). Using a test of two populations of proportions it was found that proteins were statistically more likely to be identified from gels stained with Deep Purple. Additionally, the identifications from Deep Purple stained gels are of higher quality because they are based on multiple peptides.

The discovery and early validation of novel plasma biomarkers in mild-to-moderate Alzheimer's disease patients responding to treatment with rosiglitazone.

Recent advances in clinical, pathological and neuroscience studies have identified disease-modifying therapeutic approaches for Alzheimer's disease that are now in clinical trials. This has highlighted the need for reliable and convenient biomarkers for both early disease diagnosis and a rapid signal of drug efficacy. We describe the identification and assessment of a number of candidate biomarkers in patients with Alzheimer's disease and the correlation of those biomarkers with rosiglitazone therapeutic efficacy, as represented by a change in the Alzheimer's Disease Assessment Scale-Cognitive (ADAS-Cog). Plasma from 41 patients with Alzheimer's disease were analysed by open platform proteomics at baseline and after receiving 8 mg rosiglitazone for 24 weeks. From a comparison of protein expression following treatment with rosiglitazone, 97 proteins were observed to be differentially expressed with a p-value<0.01. From this analysis and comparison to recently published data from our laboratory, a prioritized list of 10 proteins were analysed by immunoassay and/or functional assay in a wider set of samples from the same clinical study, representing a rosiglitazone dose response, in order to verify the changes observed. A number of these proteins appeared to show a correlation with change in ADAS-Cog at the higher treatment doses compared with the placebo. Alpha-2-macroglobulin, complement C1 inhibitor, complement factor H and apolipoprotein E expression showed a correlation with ADAS-Cog score at the higher doses (4 mg and 8 mg). These results are discussed in light of the pathology and other recently published data.