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Elena Tianfei Yuan, Yoko Ino, Maho Kawaguchi, Yayoi Kimura, Hisashi Hirano, Emiko Kinoshita-Kikuta, Eiji Kinoshita, Tohru Koike
Phosphorylated peptides are attractive targets in the study of the phosphoproteome. Here, we introduce a simple and convenient micropipette-tip method for the separation of phosphorylated and nonphosphorylated peptides by using a phosphate-binding zinc(II) complex of 1,3-bis(pyridin-2-ylmethylamino)propan-2-olate (Phos-tag). A 200-μL micropipette tip containing 10 μL of swollen agarose beads functionalized with Phos-tag moieties was prepared. All steps in the phosphate-affinity separation (binding, washing, and elution) were conducted by using aqueous buffers at neutral pH values...
June 21, 2017: Electrophoresis
EunJi Sung, Oh Kwang Kwon, Jae-Mok Lee, Sangkyu Lee
Prostate cancer is the leading type of cancer diagnosed, and the most frequent cause of worldwide male cancer-related deaths annually. The limitations of current prostate cancer screening tests demand the identification of novel biomarkers for the early diagnosis of prostate cancer bone metastasis. In the present study, we performed a proteomic analysis of secreted proteins from the prostate cancer bone metastasis cell line, PC-3, and the normal prostate cell line, RWPE-1. We thus quantified 917 proteins, of which 68 were found to be secreted at higher levels by PC-3 than by RWPE-1 cells via LC-MS/MS...
June 19, 2017: Electrophoresis
Biswapriya Biswavas Misra, Johannes F Fahrmann, Dmitry Grapov
Data processing and analysis are major bottlenecks in high-throughput metabolomic experiments. Recent advancements in data acquisition platforms are driving trends toward increasing data size (e.g. petabyte scale) and complexity (multiple omic platforms). Improvements in data analysis software and in silico methods are similarly required to effectively utilize these advancements and link the acquired data with biological interpretations. Herein we provide an overview of recently developed and freely-available metabolomic tools, algorithms, databases and data analysis frameworks...
June 16, 2017: Electrophoresis
Sinwook Park, Daniel Capelin, Gadi Piriatinskiy, Tamar Lotan, Gilad Yossifon
Jellyfish stinging capsules known as nematocysts are explosive, natural-injection systems with high potential as a natural drug-delivery system. These organelles consist of a capsule containing a highly folded thin needle-like tubule and a matrix highly concentrated with charged constituents that enable the tubule to fire and penetrate a target. For the purpose of using these nematocysts as drug delivery system it is first required to purify sub-populations from heterogeneous population of capsules and to investigate each subpopulation's distinct function and characteristics...
June 14, 2017: Electrophoresis
Wei-Qiang Chen, Philipp Obermayr, Urh Černigoj, Jana Vidič, Tanja Panić-Janković, Goran Mitulović
Classical proteomics approaches involve enzymatic hydrolysis of proteins (either separated by polyacrylamide gels or in solution) followed by peptide identification using LC-MS/MS analysis. This method requires normally more than sixteen hours to complete. In the case of clinical analysis, it is of the utmost importance to provide fast and reproducible analysis with minimal manual sample handling. Herein we report the method development for online protein digestion on immobilized monolithic enzymatic reactors (IMER) to accelerate protein digestion, reduce manual sample handling, and provide reproducibility to the digestion process in clinical laboratory...
June 14, 2017: Electrophoresis
André P S Kpaibe, Randa Ben-Ameur, Gaëlle Coussot, Yoann Ladner, Jérôme Montels, Michèle Ake, Catherine Perrin
Snake venoms constitute a very promising resource for the development of new medicines. They are mainly composed of very complex peptides and proteins mixtures, which composition may vary significantly from batch to batch. This latter consideration is a challenge for routine quality control (QC) in the pharmaceutical industry. In this article, we report the use capillary zone electrophoresis for the development of an analytical fingerprint methodology to assess the quality of snake venoms. The analytical fingerprint concept is being widely used for the QC of herbal drugs but rarely for venoms QC so far...
June 13, 2017: Electrophoresis
Sepeedah Soltanian-Zadeh, Kruthika Kikkeri, Ayesha N Shajahan-Haq, Jeannine Strobl, Robert Clarke, Masoud Agah
Inherent electrical properties of cells can be beneficial to characterize different cell lines and their response to experimental drugs. This paper presents a novel method to characterize the response of breast cancer cells to drug stimuli through use of off-chip passivated-electrode insulator-based dielectrophoresis (OπDEP) and the application of AC electric fields. This work is the first to demonstrate the ability of OπDEP to differentiate between two closely related breast cancer cell lines, LCC1 and LCC9 while assessing their drug sensitivity to an experimental anti-cancer agent, Obatoclax...
June 13, 2017: Electrophoresis
Fabian Schulte, Markus Hardt, Karsten Niehaus
Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) is a key technique for the separation of complex protein samples to survey the protein inventory of prokaryotic micro-organisms. Although the preparation of proteins is a critical step in 2D-gel experiments, its effect on the outcome of proteome data is often underestimated. In this work, we show that the choice of protein isolation method can have a profound impact on the quality of 2D-gels of protein extracts from prokaryotes. Based on Xanthomonas campestris, a commercially relevant producer of the thickening agent xanthan, we validated a phenol extraction protocol for the purification of bacterial proteins that provides excellent 2D-gel separation...
June 12, 2017: Electrophoresis
Manuel Ferrer, Beata Anna Raczkowska, Mónica Martínez-Martínez, Coral Barbas, David Rojo
The analysis of the microbial metabolome is crucial to fully understand the symbiotic relation between humans and microbes. That is why an explosion of metabolomics took place in the area. So far, at least several hundreds of microbial metabolites have been shown to be statistically altered when humans undergo a plethora of commonly faced perturbations. NMR and MS, usually coupled with GC, LC and CE have revealed their identities. CE is a robust analytical platform for the analysis of polar and ionic metabolites that are essential in order to assess the cells' activity...
June 12, 2017: Electrophoresis
Juan Peris-Vicente, Khaled Tayeb-Cherif, Samuel Carda-Broch, Josep Esteve-Romero
The suitability of an analytical method to determine oxolinic acid, danofloxacin, ciprofloxacin and enrofloxacin in edible tissues, based on micellar liquid chromatography coupled with fluorescence detection, to be applied in chicken, turkey, duck, lamb, goat, rabbit and horse muscle, is described. The method was fully matrix-matched in-lab revalidated, for each antimicrobial drug and meat, following the guidelines of the EU Commission Decision 2002/657/EC. The permitted limits were the maximum residue limits stated by the EU Commission Regulation 37/2010...
June 9, 2017: Electrophoresis
Frantisek Svec
This review article is an updated and extended version of the previous review that has been published 14 years ago 1. Although history does not change, its interpretation may. Therefore, I took the liberty to revisit the original work and added some of my new thoughts that could be interesting for the generation of scientists who are entering the intriguing arena of monoliths and may want to know the roots of the field. The current monolithic columns that are well applicable for efficient separations, were first described less than 30 years ago, and their advent represented the initial step to their present widespread appearance...
June 5, 2017: Electrophoresis
María Molina-Calle, Verónica Sánchez de Medina, M Calderón-Santiago, Feliciano Priego-Capote, María D Luque de Castro
Black garlic is increasing its popularity in cuisine around the world; however, scant information exists on the composition of this processed product. In this study, polar compounds in fresh garlic and in samples taken at different times during the heat treatment process to obtain black garlic have been characterized by liquid chromatography coupled to tandem mass spectrometry in high resolution mode. Ninety-five compounds (mainly amino acids and metabolites, organosulfur compounds, and saccharides and derivatives) were tentatively identified in all the analyzed samples and classified as a function of the family they belong to...
May 31, 2017: Electrophoresis
Urška Vidic, Irena Trbojević-Akmačić, Urh Černigoj, Malena Albers, Jernej Gašperšič, Maja Pučić-Baković, Jana Vidič, Aleš Štrancar, Gordan Lauc
Fibrinogen (FIB) is a secretory glycoprotein synthesized by hepatocytes that has a key role in blood clotting. Its glycosylation has not been studied in detail and little is known about the biological variability of FIB N-glycosylation, mainly due to the lack of fast, simple, and robust approaches to purify FIB from blood plasma samples. In recent years, customised chromatographic monoliths have been used for a variety of biological applications due to their unique characteristics. Here we describe development and optimisation of monolithic supports bearing monoclonal anti-human fibrinogen antibodies in a single column as well as in multi-well plate formats with high FIB specificity and binding capacity for fast immunoaffinity purification of FIB from human blood samples...
May 30, 2017: Electrophoresis
Sigurd Delanghe, Alena Moerman, Anneleen Pletinck, Eva Schepers, Griet Glorieux, Wim Van Biesen, Joris R Delanghe, Marijn M Speeckaert
Protein carbamylation, a nonenzymatic posttranslational modification promoted during uremia, is linked to a poor prognosis. In the present study, carbamylation of serum albumin was assayed using the symmetry factor on a capillary electrophoresis instrument (Helena V8). The symmetry factor has been defined as the distance from the center line of the peak to the back slope, divided by the distance from the center line of the peak to the front slope, with all measurements made at 10% of the maximum peak height...
May 30, 2017: Electrophoresis
Gleb Zilberstein, Roman Zilberstein, Svetlana Zilberstein, Uriel Maor, Emmanuil Baskin, Shoumo Zhang, Pier Giorgio Righetti
A miniaturized chemical sensor is here described for analysis of environmental pollutants (VOC: volatile organic chemicals). It is used for remote detection of formaldehyde (FA) fumes in the atmosphere, and is based on the redox reaction between FA and silver nitrate. The sensor is worn as a bracelet and the data acquired are transferred via a blue-tooth channel to a smartphone. A dedicated software transforms the signal from a grey to a colour scale. The signal response has been assessed over low (20 to 120 ppb) as well as higher (1-15 ppm range) levels...
May 29, 2017: Electrophoresis
Alba Hykollari, Daniel Malzl, Shi Yan, Iain B H Wilson, Katharina Paschinger
The unusual nature of the N-glycans of the cellular slime mould Dictyostelium discoideum has been revealed by a number of studies, primarily based on examination of radiolabeled glycopeptides but more recently also by MS. The complexity of the N-glycomes of even glycosylation mutants is compounded by the occurrence of anionic modifications, which also present an analytical challenge. In this study, we have employed hydrophilic interaction anion exchange (HIAX) HPLC in combination with MALDI-TOF MS/MS to explore the anionic N-glycome of the M31 (modA) strain, which lacks endoplasmic reticulum α-glucosidase II, an enzyme conserved in most eukaryotes including Homo sapiens...
May 29, 2017: Electrophoresis
Yifan Huang, Shiyue Zhou, Jianhui Zhu, David M Lubman, Yehia Mechref
Early stage detection and cancer treatment demand the identification of reliable biomarkers. Over the past decades, efforts have been devoted to assess the variation of glycosylation level as well as the glycan structures of proteins in blood or serum, associated with the development and/or progression of several cancers, including liver. Herein, an LC-MS/MS-based analysis was conducted to define the glycosylation patterns of haptoglobin glycoprotein derived from sera collected from cirrhotic and hepatocellular carcinoma (HCC) patients...
May 25, 2017: Electrophoresis
Marcella Matos Cordeiro Borges, Hanna Leijoto de Oliveira, Keyller Bastos Borges
The assessment of selective molecularly imprinted solid-phase extraction (MISPE) for clean-up and extraction of cholesterol (CHO) from bovine and human serum samples was evaluated. Two molecular imprinted polymers (MIP) based on organic and inorganic/organic (hybrid) materials were obtained and compared regarding recovery and selectivity toward CHO. Optimum chromatographic separation between the CHO and its isobaric compound lathosterol (LATHO) has been achieved within 15 min using an Agilent Poroshell 120 EC-C18 column (120 mm × 3...
May 24, 2017: Electrophoresis
Vinicius Veri Hernandes, Coral Barbas, Danuta Dudzik
Metabolomics has been found to be applicable to a wide range of clinical studies, bringing a new era for improving clinical diagnostics, early disease detection, therapy prediction and treatment efficiency monitoring. A major challenge in metabolomics, particularly untargeted studies, is the extremely diverse and complex nature of biological specimens. Despite great advances in the field there still exist fundamental needs for considering pre-analytical variability that can introduce bias to the subsequent analytical process and decrease the reliability of the results and moreover confound final research outcomes...
May 19, 2017: Electrophoresis
Radhouene Doggui, Chaïma Abdelhafidh Sahli, Wassef Lotfi Aissa, Maroua Hammami, Maha Ben Sedrine, Rahma Mahjoub, Khemais Zouaoui, Rim Daboubi, Hajer Siala, Taieb Messaoud, Amina Bibi
HbA1c is used for monitoring diabetic balance. In this paper we report an assessment of the analytical performances of Capillarys 2 Flex Piercing (C2FP) for HbA1c measurement using CE (Capillary Electrophoresis). CLSI (Clinical and Laboratory Standard Institute) protocols are used for the evaluation of apparatus performances: precision, linearity, method comparison, truness and common interferences. HbA1c CVs average in intra-assay was 1.6% between run imprecision CV ranged from0.1 to 1.8%. The linearity was demonstrated between 4...
May 19, 2017: Electrophoresis
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