1,720,981 research outputs found
Detection of thiazide-based diuretics in equine urine by liquid chromatography/mass spectrometry
No full textThiazide-based diuretics are included in the list of banned drugs in the horse-racing industry. One effect of their misuse is increased urine flow, contributing to dilution of other doping agents. Their determination is essential in ensuring compliance to horse-racing regulation. This study evaluates the feasibility of using liquid chromatography/mass spectrometry (LC/MS) with electrospray and atmospheric pressure chemical ionization interfaces to analyze thiazidic diuretics in equine urine samples. Existing LC and gas chromatography/MS methods are limited in their applicability to thiazide analysis. Sample preparation, analyte extraction, chromatographic separation, ion-source collision induced dissociation, solvent composition, ionization mode, and ion polarity are discussed. The practicality of LC/MS for this analysis is demonstrated with actual equine administration samples collected at specified time intervals. Detection limits were 270 ng/mL for chlorothiazide, 131 ng/mL for hydrochlorothiazide, and 384 ng/mL for trichlormethiazide
Hypothalamus proteomics from mouse models with obesity and anorexia reveals therapeutic targets of appetite regulation
No full textObjective: This study examined the proteomic profile of the hypothalamus in mice exposed to a high-fat diet (HFD) or with the anorexia of acute illness. This comparison could provide insight on the effects of these two opposite states of energy balance on appetite regulation.Methods: Four to six-week-old male C56BL/6J mice were fed a normal (control 1 group; n=7) or a HFD (HFD group; n=10) for 8 weeks. The control 2 (n=7) and lipopolysaccharide (LPS) groups (n=10) were fed a normal diet for 8 weeks before receiving an injection of saline and LPS, respectively. Hypothalamic regions were analysed using a quantitative proteomics method based on a combination of techniques including iTRAQ stable isotope labeling, orthogonal two-dimensional liquid chromatography hyphenated with nanospray ionization and high-resolution mass spectrometry. Key proteins were validated with quantitative PCR.Results: Quantitative proteomics of the hypothalamous regions profiled a total of 9249 protein groups (q<0.05). Of these, 7718 protein groups were profiled with a minimum of two unique peptides for each. Hierachical clustering of the differentiated proteome revealed distinct proteomic signatures for the hypothalamus under the HFD and LPS nutritional conditions. Literature research with in silico bioinformatics interpretation of the differentiated proteome identified key biological relevant proteins and implicated pathways. Furthermore, the study identified potential pharmacologic targets. In the LPS groups, the anorexigen pro-opiomelanocortin was downregulated. In mice with obesity, nuclear factor-?B, glycine receptor subunit alpha-4 (GlyR) and neuropeptide Y levels were elevated, whereas serotonin receptor 1B levels decreased.Conclusions: High-precision quantitative proteomics revealed that under acute systemic inflammation in the hypothalamus as a response to LPS, homeostatic mechanisms mediating loss of appetite take effect. Conversely, under chronic inflammation in the hypothalamus as a response to HFD, mechanisms mediating a sustained ‘perpetual cycle’ of appetite enhancement were observed. The GlyR protein may constitute a novel treatment target for the reduction of central orexigenic signals in obesity
Proteomic feature maps: a new visualization approach in proteomics analysis
No full textThe different steps of a proteomics analysis workflow generate a plethora of features for each extracted proteomic object (a protein spot in 2D gel electrophoresis (2-DE), or a peptide peak in liquid chromatography–mass spectrometry (LC–MS) analysis). Yet, the joint visualization of multiple object features on 2D gel-like maps is rather limited in currently available proteomics software packages. We introduce a new, simple, and intuitive visualization method that utilizes spheres to represent proteomic objects on proteomic feature maps, and exploits the spheres size and color to provide simultaneous visualization of user-selected feature pairs. Our contribution, a unified and flexible visualization mechanism that can be easily applied at any stage of a 2-DE or a LC–MS based differential proteomics study, is demonstrated and discussed using five representative scenarios. The joint visualization of proteomic object features and their spatial distribution is a powerful tool for inspecting and comparing the proteomics analysis results, attracting the users attention to useful information, such as differential expression trends and patterns, and even assisting in the evaluation and refinement of a proteomics experiment
Proteomic raw data - Discovery datasets
No full textThis dataset supports the thesis entitled 'Protemic discovery and validation of diagnostic plasma biomarkers for pulmonary tuberculosis' by Baquero.
List of peptides and proteins identified in three proteomic discovery experiments generated as a multi consensus report and individual reports. Relative protein expression is included.</span
Intact protein profiling in breast cancer biomarker discovery: protein identification issue and the solutions based on 3D protein separation, bottom-up and top-down mass spectrometry
No full textProteomics profiling of intact proteins based on MALDI-TOF MS and derived platforms has been used in cancer biomarker discovery studies. This approach suffers from a number of limitations such as low resolution, low sensitivity, and that no knowledge is available on the identity of the respective proteins in the discovery mode. Nevertheless, it remains the most high-throughput, untargeted mode of clinical proteomics studies to date. Here we compare key protein separation and MS techniques available for protein biomarker identification in this type of studies and define reasons of uncertainty in protein peak identity. As a result of critical data analysis, we consider 3D protein separation and identification workflows as optimal procedures. Subsequently, we present a new protocol based on 3D LC-MS/MS with top-down at high resolution that enabled the identification of HNRNP A2/B1 intact peptide as correlating with the estrogen receptor expression in breast cancer tissues. Additional development of this general concept toward next generation, top-down based protein profiling at high resolution is discussed
Towards paper-based point-of-care diagnostics fabricated by Laser Induced Forward Transfer
No full textWe report the Laser Induced Forward Transfer (LIFT) of antibodies from a liquid donor film onto paper receivers for application as point-of-care (POC) diagnostic sensors. Paper was chosen as the ideal receiver due to its inherent biocompatibility, wicking properties, wide availability and price, all of which make it an efficient and suitable platform for POC diagnostic sensors. A modified LIFT procedure, referred to as Dynamic Release Layer (DRL-LIFT), has been employed to ensure the viability of the biomolecules post-transfer. The laser used was a KrF excimer operating at 248 nm with a repetition rate of 1 Hz, pulse duration of ~10 ns, and delivering a maximum energy of ~400 mJ per pulse. Both enzyme-tagged and untagged IgG antibodies were LIFT-printed. The functionality and immunological reactivity of the LIFT-printed antibodies was confirmed by developing and demonstrating an Enzyme Linked Immunosorbent Assay and establishing the standard calibration curve for the LIFT-printed pixels. Additionally, it was shown that the localisation of the LIFT-printed pixels and immobilisation of the antibodies, a pre-requisite for paper-based diagnostic devices, was maintained throughout the wet-bench process which further justifies the spatial patterning ability of LIFT. This work demonstrates that LIFT is a technique capable of transferring antibodies onto a paper substrate accurately, reproducibly and with minimal loss of biochemical viability
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Combined proteomics and transcriptomics identifies carboxypeptidase B1 and nuclear factor KB (NF-KB) associated proteins as putative biomarkers of metastasis in low grade breast cancer
No full textCurrent prognostic factors are insufficient for precise risk-discrimination in breast cancer patients with low grade breast tumors, which, in disagreement with theoretical prognosis, occasionally form early lymph node metastasis. To identify markers for this group of patients, we employed iTRAQ-2DLC-MS/MS proteomics to 24 lymph node positive and 24 lymph node negative grade 1 luminal A primary breast tumors. Another group of 48 high-grade tumors (luminal B, triple negative, Her-2 subtypes) was also analyzed to investigate marker specificity for grade 1 luminal A tumors. From the total of 4405 proteins identified (FDR < 5%), the top 65 differentially expressed together with 30 previously identified and control markers were analyzed also at transcript level. Increased levels of carboxypeptidase B1 (CPB1), PDZ and LIM domain protein 2 (PDLIM2), and ring finger protein 25 (RNF25) were associated specifically with lymph node positive grade 1 tumors, whereas stathmin 1 (STMN1) and thymosin beta 10 (TMSB10) associated with aggressive tumor phenotype also in high grade tumors at both protein and transcript level. For CPB1, these differences were also observed by immunohistochemical analysis on tissue microarrays. Up-regulation of putative biomarkers in lymph node positive (versus negative) luminal A tumors was validated by gene expression analysis of an independent published data set (n = 343) for CPB1 (p = 0.00155), PDLIM2 (p = 0.02027) and RELA (p = 0.00015). Moreover, statistically significant connections with patient survival were identified in another public data set (n = 1678). Our findings indicate unique pro-metastatic mechanisms in grade 1 tumors that can include up-regulation of CPB1, activation of NF-?B pathway and changes in cell survival and cytoskeleton. These putative biomarkers have potential to identify the specific minor subpopulation of breast cancer patients with low grade tumors who are at higher than expected risk of recurrence and who would benefit from more intensive follow-up and may require more personalized therapy
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
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