We have identified over 100 protein and peptide components of normal human seminal fluid.

More than 10 million people are thought to be infected with Trypanosoma cruzi, primarily in the Americas. The clinical manifestations of Chagas' disease (CD) are variable, but most subjects remain asymptomatic for decades. Only 15 to 30% eventually develop terminal complications. All current diagnostic tests have limitations. New approaches are needed for blood bank screening as well as for improved diagnosis and prognosis. Sera from subjects with asymptomatic CD (n ‫؍‬ 131) were compared to those from uninfected controls (n ‫؍‬ 164) and subjects with other parasitic diseases (n ‫؍‬ 140), using protein array mass spectrometry. To identify biomarkers associated with CD, sera were fractionated by anion-exchange chromatography and bound to two commercial ProteinChip array chemistries: WCX2 and IMAC3. Multiple candidate biomarkers were found in CD sera (3 to 75.4 kDa). Algorithms employing 3 to 5 of these biomarkers achieved up to 100% sensitivity and 98% specificity for CD. The biomarkers most useful for diagnosis were identified and validated. These included MIP1 alpha, C3a anaphylatoxin, and unusually truncated forms of fibronectin, apolipoprotein A1 (ApoA1), and C3. An antipeptide antiserum against the 28.9-kDa C terminus of the fibronectin fragment achieved good specificity (90%) for CD in a Western blot format. We identified full-length ApoA1 (28.1 kDa), the major structural and functional protein component of high-density lipoprotein (HDL), as an important negative biomarker for CD, and relatively little full-length ApoA1 was detected in CD sera. This work provides proof of principle that both platform-dependent (i.e., mass spectrometry-based) and platform-independent (i.e., Western blot) tests can be generated using high-throughput mass profiling.

A range of low molecular weight compounds (<800 Da) have been examined by matrix-assisted laser desorption time-of-flight mass spectrometry to demonstrate the general analytical utility of this technique. The compound classes investigated included: carbohydrates, quaternary ammonium salts, sterols, nucleosides, purine and pyrimidine bases, amino acids, catecholamines, opioids, antibiotics, prostaglandins and a range of macrocyclic metal complexes of porphyrins and phthalocyanines. Many of the compounds tested are of biochemical, geochemical, industrial and/or therapeutic interest. Most organic analytes gave intense protonated molecules, but some were characterized by sodium adduct ions (i.e., [M + Na]'). The metal-containing compounds formed radical-cation molecular ions. In all instances the ions detected could be assigned, and excellent agreements between calculated and experimental mass values were obtained.Matrix-assisted laser desorption/ionization (MALDI) combined with time-of-flight (TOF) mass spectrometry has found general acclaim as an approach to syntheticand bio-polymer analysis. MALDI allows the direct analysis of mixtures, and hence, analyte impurities can usually be identified and characterized. Sub-picomole amounts of sample are adequate for multiple analyses, results are available in minutes, and most of the sample can be recovered directly from the target. The increasing availability of inexpensive commercial instruments has promoted widespread use of MALDI-TOFMS, particularly for biopolymers, but studies of compounds of low molecular weight (i.e., ca. 800 Da or less) have been re~tricted.'-~ Here we present experimental results for the MALDI analysis of a range of low molecular weight compounds including carbohydrates, quaternary ammonium salts, sterols, nucleosides, purine and pyrimidine bases, amino acids, catecholamines, opioids, antibiotics, prostaglandins and a range of macrocyclic metal complexes of porphyrins and phthalocyanines. Many of the compounds tested are of biochemical, geochemical, industrial and/or therapeutic significance. EXPERIMENTAL InstrumentationAnalyses were performed on a Finnigan Lasermat MALDI linear time-of-flight mass spectrometer (Finnigan MAT, Heme1 Hempstead, UK). Much of the work described in this paper was performed on a standard Lasermat, but later data were obtained on the instrument following upgrade to Lasermat 2000 specifications. The system uses a nitrogen laser (337 nm; 3 ns pulse duration), and the laser fluence is adjusted by a variable-beam attenuator. Samples were applied to stainless-steel slides with a circular sample application area of 2 mm diameter. The Lasermat software allows the user to irradiate one of four possible target regions * Author for correspondence within this area. Under standard operating conditions, and for the mass range 100-1000Da, the resolving power ( m / A m , full width at half maximum height) of the Lasermat 2000 is in the range ca. 100-200 (or 50-100 for the Lasermat). Materials and methodsWater was purified by a Millipor...

To unlock the full potential of disease modifying treatments, it is essential to develop early biomarkers for Alzheimer's disease (AD). For practical reasons, blood-based markers that could provide a signal at the stage of mild cognitive impairment (MCI) or even earlier would be ideal. Using the proteomic approach of isobaric tagging for relative and absolute quantitation (iTRAQ), we compared the plasma protein profiles of MCI, AD, and cognitively normal control subjects from two independent cohorts: the Sydney Memory and Ageing Study (261 MCI subjects, 24 AD subjects, 411 controls) and the Hunter Community Study (180 MCI subjects, 153 controls). The objective was to identify any proteins that are differentially abundant in MCI and AD plasma in both cohorts, since they might be of interest as potential biomarkers, or could help direct future mechanistic studies. Proteins representative of biological processes relevant to AD pathology, such as the complement system, the coagulation cascade, lipid metabolism, and metal and vitamin D and E transport, were found to differ in abundance in MCI. In particular, levels of complement regulators C1 inhibitor and factor H, fibronectin, ceruloplasmin, and vitamin D-binding protein were significantly decreased in MCI participants from both cohorts. Several apolipoproteins, including apolipoprotein AIV, B-100, and H were also significantly decreased in MCI. Most of these proteins have previously been reported as potential biomarkers for AD; however, we show for the first time that a significant decrease in plasma levels of two potential biomarkers (fibronectin and C1 inhibitor) is evident at the MCI stage.