Determination of protein markers in human serum: Analysis of protein expression in toxic oil syndrome studies

Abstract

Toxic oil syndrome (TOS) is a disease that appeared in Spain in 1981. It affected more than 20 000 people and produced over 300 deaths in the first 2 years. In this paper, a prospective study on the differences in gene expression in sera between a control versus a TOS-affected population, both originally exposed to the toxic oil, is presented. Differential protein expression was analyzed by two-dimensional electrophoresis (2-DE). Several problems related with serum analysis by 2-DE were addressed in order to improve protein detection in the gel images. Three new commercial systems for albumin depletion were tested to optimize the detection of minor proteins that can be obscured by the presence of a few families of high abundance proteins (albumin, immunoglobulins). Other factors, such as the use of nonionic reductants or the presence of thiourea in the gels, were also tested. From these optimized images, a group of 329 major gel spots was located, matched and compared in serum samples. Thirty-five of these protein spots were found to be under- or overexpressed in TOS patients (> three-fold increase or decrease). Proteins in the differential spots were identified by matrix-assisted laser desorption/ionization-time of flight peptide map fingerprinting and database search. Several haptoglobin isoforms were found to be differentially expressed, showing expression phenotypes that could be related with TOS affection. Haptoglobin phenotypes have been previously reported to have important biological and clinical consequences and have been described as risk factors for several diseases.

Expression of differential antennal proteins in males and females of an important crop pest, Sesamia nonagrioides

“…A sample containing 100 mg of antennal proteins was diluted with rehydration buffer to a volume of 350 mL (Quero et al, 2004b), loaded on strips, and then actively rehydrated at 50 V for 10 h following the same program as previously described (Quero et al, 2004b). The sample was maintained at 8000 V until a total run of 50 kVh was achieved.…”

Cell viability and proteomic analysis in cultured neurons exposed to methylmercury

“…The normalized spot density in each gel was measured and compared with the spot density in other gels in order to detect differentially expressed proteins. 30 Mass spectrometry protein identification The spots of interest were excised and subjected to in-gel digestion with trypsin. Tryptic peptides were extracted from the gel with acetonitrile/water/trifluoroacetic acid and the extracts were evaporated to dryness and redissolved in acetonitrile/water 1/1, 1% acetic acid.…”

Review Article Toxic Oil Syndrome: Review of Immune Aspects of the Disease

“…Proteomic analysis, including two-dimensional electrophoresis and mass spectrometry/peptide mass fingerprinting, of pooled human sera indicated differential expression of Hp isoforms in TOS patients, compared to controls. The most frequent control phenotype was Hp2-2, while cases demonstrated increased frequencies of the Hp2-1s and Hp1-1s phenotyes (Quero et al, 2004).…”