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Find video protocols related to scientific articles indexed in Pubmed.
Glycoproteomic analysis of prostate cancer tissues by SWATH mass spectrometry discovers N-acylethanolamine acid amidase and protein tyrosine kinase 7 as signatures for tumor aggressiveness.
Mol. Cell Proteomics
PUBLISHED: 04-16-2014
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The identification of biomarkers indicating the level of aggressiveness of prostate cancer (PCa) will address the urgent clinical need to minimize the general overtreatment of patients with non-aggressive PCa, who account for the majority of PCa cases. Here, we isolated formerly N-linked glycopeptides from normal prostate (n = 10) and from non-aggressive (n = 24), aggressive (n = 16), and metastatic (n = 25) PCa tumor tissues and analyzed the samples using SWATH mass spectrometry, an emerging data-independent acquisition method that generates a single file containing fragment ion spectra of all ionized species of a sample. The resulting datasets were searched using a targeted data analysis strategy in which an a priori spectral reference library representing known N-glycosites of the human proteome was used to identify groups of signals in the SWATH mass spectrometry data. On average we identified 1430 N-glycosites from each sample. Out of those, 220 glycoproteins showed significant quantitative changes associated with diverse biological processes involved in PCa aggressiveness and metastasis and indicated functional relationships. Two glycoproteins, N-acylethanolamine acid amidase and protein tyrosine kinase 7, that were significantly associated with aggressive PCa in the initial sample cohort were further validated in an independent set of patient tissues using tissue microarray analysis. The results suggest that N-acylethanolamine acid amidase and protein tyrosine kinase 7 may be used as potential tissue biomarkers to avoid overtreatment of non-aggressive PCa.
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Carbon-coated LiCrTiO4 electrode material promoting phase transition to reduce asymmetric polarization for lithium-ion batteries.
Phys Chem Chem Phys
PUBLISHED: 01-16-2014
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In this work, carbon-free and carbon-coated spinel LiCrTiO4 oxides were synthesized by a conventional solid state reaction. The lithium-ion diffusion coefficient and electronic conductivity of prepared electrode materials were systematically investigated using the galvanostatic intermittent titration technique (GITT), cyclic voltammetry (CV), and electrochemical impedance spectroscopy (EIS). The rate performances of the prepared materials were evaluated by galvanostatic charge-discharge. Carefully comparing the charge-discharge polarization potential of both materials, we unexpectedly discovered that the pristine LiCrTiO4 electrode demonstrated asymmetric polarization during the charging-discharging process, which is possibly attributed to the nonuniform electron conductivity between the endmember of a two-phase reaction, whereas carbon coating could level this phenomenon. Additionally, using an asymmetric core-shell model from the microscopic point of view can easily explain this common phenomenon. Meanwhile, this new research perspective can be extended to other active materials in lithium ion batteries.
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Mass spectrometric protein maps for biomarker discovery and clinical research.
Expert Rev. Mol. Diagn.
PUBLISHED: 10-21-2013
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Among the wide range of proteomic technologies, targeted mass spectrometry (MS) has shown great potential for biomarker studies. To extend the degree of multiplexing achieved by selected reaction monitoring (SRM), we recently developed SWATH MS. SWATH MS is a variant of the emerging class of data-independent acquisition (DIA) methods and essentially converts the molecules in a physical sample into perpetually re-usable digital maps. The thus generated SWATH maps are then mined using a targeted data extraction strategy, allowing us to profile disease-related proteomes at a high degree of reproducibility. The successful application of both SRM and SWATH MS requires the a priori generation of reference spectral maps that provide coordinates for quantification. Herein, we demonstrate that the application of the mass spectrometric reference maps and the acquisition of personalized SWATH maps hold a particular promise for accelerating the current process of biomarker discovery.
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Axonal and glial responses to a mid-thoracic spinal cord hemisection in the Macaca fascicularis monkey.
J. Neurotrauma
PUBLISHED: 05-13-2013
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A comprehensive understanding of the pathology of spinal cord injury (SCI) in non-human primates may facilitate greatly the development of new strategies to promote recovery in humans with SCI. Relatively few studies, however, have been conducted to systemically examine pathological changes in the monkey, a non-human primate, after SCI. We report axonal, glial, and fibrotic responses in the spinal cord of monkey Macaca fascicularis after a thoracic (T) 8-9 lateral hemisection. We examined these changes at three regions--i.e., the lesion epicenter, the peri-lesion area, and the lateral white matter of the intact, contralateral hemicord at 7 (subacute) and 30 (early chronic) days post-injury. The lateral hemisection resulted in a marked axon and myelin loss, along with tissue loss, at the lesion epicenter at both time points. Unexpectedly, axonal loss and myelin degeneration, along with reactive gliosis and microglia/macrophages activation, were also observed in the contralateral spared hemicord, indicating a spread of the initial damage to the contralateral side. In addition, activated microglia/macrophages were found both within the injury epicenter and the peri-lesion area, indicating that they participate in injury-induced immune responses that may exacerbate the secondary damage. In contrast to rodents, substantial reactive astrocytic responses at the lesion border were not observed in the monkey. Conversely, a deposit of robust fibrotic scar was observed at the injury epicenter, which filled the space originally created by the hemisection. Thus, understanding the pathology of monkey SCI may provide clinically relevant information in designing repair strategies targeting specific problems associated with human SCIs.
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Quantitative measurements of N-linked glycoproteins in human plasma by SWATH-MS.
Proteomics
PUBLISHED: 03-11-2013
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SWATH-MS is a data-independent acquisition method that generates, in a single measurement, a complete recording of the fragment ion spectra of all the analytes in a biological sample for which the precursor ions are within a predetermined m/z versus retention time window. To assess the performance and suitability of SWATH-MS-based protein quantification for clinical use, we compared SWATH-MS and SRM-MS-based quantification of N-linked glycoproteins in human plasma, a commonly used sample for biomarker discovery. Using dilution series of isotopically labeled heavy peptides representing biomarker candidates, the LOQ of SWATH-MS was determined to reach 0.0456 fmol at peptide level by targeted data analysis, which corresponds to a concentration of 5-10 ng protein/mL in plasma, while SRM reached a peptide LOQ of 0.0152 fmol. Moreover, the quantification of endogenous glycoproteins using SWATH-MS showed a high degree of reproducibility, with the mean CV of 14.90%, correlating well with SRM results (R(2) = 0.9784). Overall, SWATH-MS measurements showed a slightly lower sensitivity and a comparable reproducibility to state-of-the-art SRM measurements for targeted quantification of the N-glycosites in human blood. However, a significantly larger number of peptides can be quantified per analysis. We suggest that SWATH-MS analysis combined with N-glycoproteome enrichment in plasma samples is a promising integrative proteomic approach for biomarker discovery and verification.
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Multiple positive solutions for nonlinear fractional boundary value problems.
ScientificWorldJournal
PUBLISHED: 01-01-2013
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This paper is devoted to the existence of multiple positive solutions for fractional boundary value problem DC0+?u(t) = f(t, u(t), u(t)), 0 < t < 1, u(1) = u(1) = u(0) = 0, where 2 < ? ? 3 is a real number, DC0+? is the Caputo fractional derivative, and f : [0,1]×[0, +?) × R ? [0, +?) is continuous. Firstly, by constructing a special cone, applying Guo-Krasnoselskiis fixed point theorem and Leggett-Williams fixed point theorem, some new existence criteria for fractional boundary value problem are established; secondly, by applying a new extension of Krasnoselskiis fixed point theorem, a sufficient condition is obtained for the existence of multiple positive solutions to the considered boundary value problem from its auxiliary problem. Finally, as applications, some illustrative examples are presented to support the main results.
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Effect of dexamethasone on gliosis, ischemia, and dopamine extraction during microdialysis sampling in brain tissue.
Anal. Chem.
PUBLISHED: 09-15-2011
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Microdialysis sampling of the brain is an analytical technique with numerous applications in neuroscience and the neurointensive care of brain-injured human patients. Even so, implanting microdialysis probes into brain tissue causes a penetration injury that triggers gliosis (the activation and proliferation of glial cells) and ischemia (the interruption of blood flow). Thus, the probe samples injured tissue. Mitigating the effects of the penetration injury might refine the technique. The synthetic glucocorticoid dexamethasone is a potent anti-inflammatory and immunosuppressant substance. We performed microdialysis in the rat brain for 5 days, with and without dexamethasone in the perfusion fluid (10 ?M for the first 24 h and 2 ?M thereafter). On the first and fourth day of the perfusion, we performed dopamine no-net-flux measurements. On the fifth day, we sectioned and stained the brain tissue and examined it by fluorescence microscopy. Although dexamethasone profoundly inhibited gliosis and ischemia around the probe tracks it had only modest effects on dopamine no-net-flux results. These findings show that dexamethasone is highly effective at suppressing gliosis and ischemia but is limited in its neuroprotective activity.
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Ultrastructural insights into morphology and reproductive mode of Blastocystis hominis.
Parasitol. Res.
PUBLISHED: 06-08-2011
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To understand well the morphology and reproductive mode of Blastocystis hominis, with the help of transmission electron microscopy and scanning electron microscopy the ultrastructural details of B. hominis from fresh diarrheal specimens and cultured strains were observed. In both fecal samples and culture conditions, there were vacuolar and granular forms. In diarrhea, it exists in multivacuolar, avacuolar, and amoeboid forms. In the in vitro culture, vacuolar form could transform to granular form. The most commonly noticed structure on the cell surface was surface coat with diversity in appearance (the funiform, lamellar, filiform, and floccose in different thickness) and distributions. Three modes of reproduction were confirmed, they were binary fission, plasmotomy, and budding. Under the impact of hosts response, the ultrastructures of surface coat, nucleus, and mitochondrion-like organelle sometimes changed.
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A high-quality secretome of A549 cells aided the discovery of C4b-binding protein as a novel serum biomarker for non-small cell lung cancer.
J Proteomics
PUBLISHED: 01-11-2011
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Cancer secretomes are a promising source for biomarker discovery. The analysis of cancer secretomes still faces some difficulties mainly related to the intracellular contamination, which hinders the qualification and follow-up validations. This study aimed to establish a high-quality secretome of A549 cells by using the cellular proteome as a reference and to test the merits of this refined secretome for biomarker discovery for non-small cell lung cancer (NSCLC). Using one-dimensional gel electrophoresis followed by liquid-chromatography tandem mass spectrometry, we comprehensively investigated the secretome and the concurrent cellular proteome of A549 cells. A high-quality secretome consisting of 382 proteins was refined from 889 initial secretory proteins. More than 85.3% of proteins were annotated as secreted and 76.8% as extracellular or membrane-bound. The discriminative power of the lung-cancer associated secretome was confirmed by gene expression and serum proteomic data. The elevated level of C4b-binding Protein (C4BP) in NSCLC blood was verified by enzyme-linked immunosorbent assays (ELISA, p = 6.07e-6). Moreover, the serum C4BP level in 89 patients showed a strong association with the clinical staging of NSCLC. Our reference-experiment-driven strategy is simple and widely applicable, and may facilitate the identification of novel promising biomarkers of lung cancer.
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Capillary ultrahigh performance liquid chromatography with elevated temperature for sub-one minute separations of basal serotonin in submicroliter brain microdialysate samples.
Anal. Chem.
PUBLISHED: 11-09-2010
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Improving the time resolution in microdialysis coupled to high performance liquid chromatography (HPLC) requires that the volume of the separation system be decreased. A low-volume separation permits smaller microdialysate volumes to be injected without suffering a sensitivity loss from dilution. Thus, improved time resolution can be achieved with offline analysis simply by decreasing the separations system volume. For online (near real-time) analysis, there is a further requirement. The separation speed must be at least as fast as the sampling time. Here, the combined use of high column pressures and temperatures, sub-2-?m stationary phase particles, capillary columns, and sensitive, low dead-volume detection resulted in a retention time for the neurotransmitter serotonin of less than 1 min in a 500 nL dialysate sample volume. Two sensitive detectors, photoluminescence following electron transfer (PFET) and electrochemical, were used for the detection of subnanomolar concentrations of serotonin in brain microdialysate samples. The general principles developed are applicable to a wide range of separations with the additional advantages of increases in sample throughput and decreases in mobile phase usage.
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Proteomic mining in the dysplastic liver of WHV/c-myc mice--insights and indicators for early hepatocarcinogenesis.
FEBS J.
PUBLISHED: 08-27-2010
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Because of the asymptomatic process of carcinogenesis, the early detection of cancers such as hepatocellular carcinoma (HCC) is very challenging. Tumor-prone transgenic mouse models of oncogenesis can provide a stable and powerful tool for the analysis of cancer initiation, and are therefore promising for the discovery of early putative biomarkers of HCC. Using a label-free proteomic quantification strategy, we comprehensively investigated the protein expression profile in the livers of three 2-month-old WHV/c-myc mice at the dysplastic stage, with age-matched wt-C57 mice as controls. We identified 2781 proteins, 540 of which were differentially expressed. These proteins successfully characterized certain precancerous biological processes and alterations in transcriptional regulators before tumor onset. Two candidates, FK506-binding protein 4 (FKBP52) and ferritin heavy chain, were taken as examples for a search from the mouse model to clinical human tissues. Their levels in serum samples were determined by western blotting, and showed a noteworthy ability to distinguish between HCC and control cases. Immunohistochemical analysis with tissue microarrays confirmed the differential expression of FKBP52 between HCC and the paired controls (P < 0.001). The regulation of FKBP52 was also discovered to be relevant to HCC staging, with a dramatic decline at stage III (P < 0.05). The potentials of the candidate pool in this study were discussed in terms of delineating c-myc-induced hepatocarcinogenesis and facilitating biomarker discovery for early HCC diagnosis.
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Glial response and myelin clearance in areas of wallerian degeneration after spinal cord hemisection in the monkey Macaca fascicularis.
J. Neurotrauma
PUBLISHED: 05-22-2009
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Spinal cord injury (SCI) in mammals not only damages the focal area, but also leads to wallerian degeneration (WD) of axons and myelin distal to the injury. In the present study, we investigated cellular responses within areas of WD of a sensory pathway, the fasciculus gracilis, after a T8-9 lateral spinal hemisection in the adult monkey Macaca fascicularis. Spinal cord segments rostral and caudal to the injury at two clinically-relevant time points, 1 week and 4 weeks post-SCI, representing subacute and chronic stages, respectively, were examined. We observed marked axon degeneration in the areas of WD at the subacute stage, and minimal axonal neurofilament staining at the chronic stage. At the ultrastructural level, however, many degenerating axonal profiles remained at the chronic stage. Myelin breakdown was a much-delayed process. A large number of residual myelin sheaths was observed at the chronic stage. In contrast to rodents, a substantial astrogliotic response was not found in the WD regions up to 4 weeks post-injury. Microglia activation was evident in the WD areas at the subacute stage, and was enhanced at the chronic stage. However, the lack of round reactive microglia/macrophages in these regions suggests that microglial activation was either delayed or incomplete. Thus it appears that many pathological characteristics of WD in monkeys are much delayed compared to those in rodents, but are similar to those in humans. Our results suggest that non-human primate SCI models are useful for evaluating repair strategies before they are translated to clinical trials of human SCI.
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Culling structured hosts to eradicate vector-borne diseases.
Math Biosci Eng
PUBLISHED: 04-15-2009
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A compartmental model is developed, in the form of a nonautonomous system of delay differential equations subject to impulses at specific times, for mosquito-born disease control involving larvicides and insecticide sprays. Sufficient conditions in terms of the frequencies and rates of larvicides and insecticide sprays are derived, and numerical simulations are provided to illustrate the sharpness of these disease eradication conditions.
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Integrative proteomics and tissue microarray profiling indicate the association between overexpressed serum proteins and non-small cell lung cancer.
PLoS ONE
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Lung cancer is the leading cause of cancer deaths worldwide. Clinically, the treatment of non-small cell lung cancer (NSCLC) can be improved by the early detection and risk screening among population. To meet this need, here we describe the application of extensive peptide level fractionation coupled with label free quantitative proteomics for the discovery of potential serum biomarkers for lung cancer, and the usage of Tissue microarray analysis (TMA) and Multiple reaction monitoring (MRM) assays for the following up validations in the verification phase. Using these state-of-art, currently available clinical proteomic approaches, in the discovery phase we confidently identified 647 serum proteins, and 101 proteins showed a statistically significant association with NSCLC in our 18 discovery samples. This serum proteomic dataset allowed us to discern the differential patterns and abnormal biological processes in the lung cancer blood. Of these proteins, Alpha-1B-glycoprotein (A1BG) and Leucine-rich alpha-2-glycoprotein (LRG1), two plasma glycoproteins with previously unknown function were selected as examples for which TMA and MRM verification were performed in a large sample set consisting about 100 patients. We revealed that A1BG and LRG1 were overexpressed in both the blood level and tumor sections, which can be referred to separate lung cancer patients from healthy cases.
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Recommendations on bioanalytical method stability implications of co-administered and co-formulated drugs by Global CRO Council for Bioanalysis (GCC).
Bioanalysis
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An open letter written by the Global CRO Council for Bioanalysis (GCC) describing the GCC survey results on stability data from co-administered and co-formulated drugs was sent to multiple regulatory authorities on 14 December 2011. This letter and further discussions at different GCC meetings led to subsequent recommendations on this topic of widespread interest within the bioanalytical community over the past 2 years.
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Optimization for speed and sensitivity in capillary high performance liquid chromatography. The importance of column diameter in online monitoring of serotonin by microdialysis.
J Chromatogr A
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The speed of a separation defines the best time resolution possible in online measurements using chromatography. The desired time resolution multiplied by the flow rate of the stream of analyte being sampled defines the maximum volume of sample per injection. The best concentration sensitivity in chromatography is obtained by injecting the largest volume of sample that is consistent with achieving a satisfactory separation, and thus measurement accuracy. Taking these facts together, it is easy to understand that separation speed and concentration sensitivity are linked in this type of measurement. To address the problem of how to achieve the best sensitivity and shortest measurement time simultaneously, we have combined recent approaches to the optimization of the separation itself with an analysis of method sensitivity. This analysis leads to the column diameter becoming an important parameter in the optimization process. We use these ideas in one particular problem presented by online microdialysis sampling/liquid chromatography/electrochemical detection for measuring concentrations of serotonin in the dialysate. In this case the problem becomes the optimization of conditions to yield maximum signal for a given sample volume under the highest speed conditions with a certain required number of theoretical plates. It turns out that the observed concentration sensitivity at an electrochemical detector can be regulated by temperature, particle size, injection volume/column diameter, and void time. The theory was successfully used for optimization of neurotransmitter serotonin measurement by capillary HPLC when sampling from a microdialysis flow stream. The final conditions are: 150 ?m i.d., 3.1cm long columns with 1.7 ?m particle diameter working at a flow rate of 12 ?L/min, an injection volume of 500 nL, and a temperature of 343 K. The retention time for serotonin is 22.7s, the analysis time is about 36 s (which allows for determination of 3-methoxytyramine), and the sampling time is about 0.8 min with a perfusion flow rate of 0.6 ?L/min.
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JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.