Two-dimensional analysis of human lymphocyte proteins. III. Preliminary report on a marker for the early detection and diagnosis of infectious mononucleosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Willard, K.E.

1982-04-01

Two-dimensional gel electrophoretic patterns of human peripheral blood leukocytes from 12 patients with infectious mononucleosis were prepared by use of the ISO-DALT system. Before the two-dimensional separation, the leukocytes were purified by Ficoll-Paque gradient centrifugation and labeled overnight with (/sup 35/S) methionine. Quantitative increases in two proteins were detected in the patterns of infected leukocytes from the patients as compared with controls. Fluorescence-activated cell sorting of leukocytes from normal human peripheral blood before subsequent two-dimensional gel analysis revealed that the dramatic increase in one of these proteins (Inmono:2) could be due to shifts in the population ratios of lymphocytes, monocytes,more » and granulocytes. In contrast, the appearance in the infected leukocytes of a second protein, Inmono:1, could not be accounted for by cell-population shifts. Increased amounts of these two proteins have been found in every patient studied who had clinically detectable infectious mononucleosis. In addition, a patient who displayed symptoms of infectious mononucleosis but who did not have a positive result in the MONOSPOT test (Ortho) until three weeks after our analysis also demonstrated increased relative amounts of these proteins in his leukocyte pattern.« less

Microfluidic device having an immobilized pH gradient and PAGE gels for protein separation and analysis

DOEpatents

Sommer, Gregory J.; Hatch, Anson V.; Singh, Anup K.; Wang, Ying-Chih

2012-12-11

Disclosed is a novel microfluidic device enabling on-chip implementation of a two-dimensional separation methodology. Previously disclosed microscale immobilized pH gradients (IPG) are combined with perpendicular polyacrylamide gel electrophoresis (PAGE) microchannels to achieve orthogonal separations of biological samples. Device modifications enable inclusion of sodium dodecyl sulfate (SDS) in the second dimension. The device can be fabricated to use either continuous IPG gels, or the microscale isoelectric fractionation membranes we have also previously disclosed, for the first dimension. The invention represents the first all-gel two-dimensional separation microdevice, with significantly higher resolution power over existing devices.

Microfluidic device having an immobilized pH gradient and page gels for protein separation and analysis

DOEpatents

Sommer, Gregory J; Hatch, Anson V; Singh, Anup K; Wang, Ying-Chih

2014-05-20

Disclosed is a novel microfluidic device enabling on-chip implementation of a two-dimensional separation methodology. Previously disclosed microscale immobilized pH gradients (IPG) are combined with perpendicular polyacrylamide gel electrophoresis (PAGE) microchannels to achieve orthogonal separations of biological samples. Device modifications enable inclusion of sodium dodecyl sulfate (SDS) in the second dimension. The device can be fabricated to use either continuous IPG gels, or the microscale isoelectric fractionation membranes we have also previously disclosed, for the first dimension. The invention represents the first all-gel two-dimensional separation microdevice, with significantly higher resolution power over existing devices.

SU-F-T-159: Monte Carlo Simulation Studies of Three-Dimensional Dose Distribution for Polymer Gel Dosimeter and Radiochromic Gel Dosimeter in a Proton Beam

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, M; Kim, G; Jung, H

Purpose: The purpose of this simulation study is to evaluate the proton detectability of gel dosimeters, and estimate the three-dimensional dose distribution of protons in the radiochromic gel and polymer gel dosimeter compared with the dose distribution in water. Methods: The commercial composition ratios of normoxic polymer gel and LCV micelle radiochromic gel were included in this simulation study. The densities of polymer and radiochromic gel were 1.024 and 1.005 g/cm3, respectively. The 50, 80 and 140 MeV proton beam energies were selected. The dose distributions of protons in the polymer and radiochromic gel were simulated using Monte Carlo radiationmore » transport code (MCNPX 2.7.0, Los Alamos Laboratory). The water equivalent depth profiles and the dose distributions of two gel dosimeters were compared for the water. Results: In case of irradiating 50, 80 and 140 MeV proton beam to water phantom, the reference Bragg-peak depths are represented at 2.22, 5.18 and 13.98 cm, respectively. The difference in the water equivalent depth is represented to about 0.17 and 0.37 cm in the radiochromic gel and polymer gel dosimeter, respectively. The proton absorbed doses in the radiochromic gel dosimeter are calculated to 2.41, 3.92 and 6.90 Gy with increment of incident proton energies. In the polymer gel dosimeter, the absorbed doses are calculated to 2.37, 3.85 and 6.78 Gy with increment of incident proton energies. The relative absorbed dose in radiochromic gel (about 0.47 %) is similar to that of water than the relative absorbed dose of polymer gel (about 2.26 %). In evaluating the proton dose distribution, we found that the dose distribution of both gel dosimeters matched that of water in most cases. Conclusion: As the dosimetry device, the radiochromic gel dosimeter has the potential particle detectability and is feasible to use for quality assurance of proton beam therapy beam.« less

Screening trematodes for novel intervention targets: a proteomic and immunological comparison of Schistosoma haematobium, Schistosoma bovis and Echinostoma caproni

PubMed Central

HIGÓN, MELISSA; COWAN, GRAEME; NAUSCH, NORMAN; CAVANAGH, DAVID; OLEAGA, ANA; TOLEDO, RAFAEL; STOTHARD, J. RUSSELL; ANTÚNEZ, ORETO; MARCILLA, ANTONIO; BURCHMORE, RICHARD; MUTAPI, FRANCISCA

2011-01-01

SUMMARY With the current paucity of vaccine targets for parasitic diseases, particularly those in childhood, the aim of this study was to compare protein expression and immune cross-reactivity between the trematodes Schistosoma haematobium, S. bovis and Echinostoma caproni in the hope of identifying novel intervention targets. Native adult parasite proteins were separated by 2-dimensional gel electrophoresis and identified through electrospray ionisation tandem mass spectrometry to produce a reference gel. Proteins from differential gel electrophoresis analyses of the three parasite proteomes were compared and screened against sera from hamsters infected with S. haematobium and E. caproni following 2-dimensional Western blotting. Differential protein expression between the three species was observed with circa 5% of proteins from S. haematobium showing expression up-regulation compared to the other two species. There was 91% similarity between the proteomes of the two Schistosoma species and 81% and 78·6% similarity between S. haematobium and S. bovis versus E. caproni, respectively. Although there were some common cross-species antigens, species-species targets were revealed which, despite evolutionary homology, could be due to phenotypic plasticity arising from different host-parasite relationships. Nevertheless, this approach helps to identify novel intervention targets which could be used as broad-spectrum candidates for future use in human and veterinary vaccines. PMID:21729355

Evaluation of radiochromic gel dosimetry and polymer gel dosimetry in a clinical dose verification

NASA Astrophysics Data System (ADS)

Vandecasteele, Jan; De Deene, Yves

2013-09-01

A quantitative comparison of two full three-dimensional (3D) gel dosimetry techniques was assessed in a clinical setting: radiochromic gel dosimetry with an in-house developed optical laser CT scanner and polymer gel dosimetry with magnetic resonance imaging (MRI). To benchmark both gel dosimeters, they were exposed to a 6 MV photon beam and the depth dose was compared against a diamond detector measurement that served as golden standard. Both gel dosimeters were found accurate within 4% accuracy. In the 3D dose matrix of the radiochromic gel, hotspot dose deviations up to 8% were observed which are attributed to the fabrication procedure. The polymer gel readout was shown to be sensitive to B0 field and B1 field non-uniformities as well as temperature variations during scanning. The performance of the two gel dosimeters was also evaluated for a brain tumour IMRT treatment. Both gel measured dose distributions were compared against treatment planning system predicted dose maps which were validated independently with ion chamber measurements and portal dosimetry. In the radiochromic gel measurement, two sources of deviations could be identified. Firstly, the dose in a cluster of voxels near the edge of the phantom deviated from the planned dose. Secondly, the presence of dose hotspots in the order of 10% related to inhomogeneities in the gel limit the clinical acceptance of this dosimetry technique. Based on the results of the micelle gel dosimeter prototype presented here, chemical optimization will be subject of future work. Polymer gel dosimetry is capable of measuring the absolute dose in the whole 3D volume within 5% accuracy. A temperature stabilization technique is incorporated to increase the accuracy during short measurements, however keeping the temperature stable during long measurement times in both calibration phantoms and the volumetric phantom is more challenging. The sensitivity of MRI readout to minimal temperature fluctuations is demonstrated which proves the need for adequate compensation strategies.

Quantitative two-dimensional gel electrophoresis analysis of human fibroblasts transformed by ras oncogenes.

PubMed

Miller, M J; Maher, V M; McCormick, J J

1992-11-01

Quantitative two-dimensional gel electrophoresis was used to compare the cellular protein patterns of a normal foreskin-derived human fibroblasts cell line (LG1) and three immortal derivatives of LG1. One derivative, designated MSU-1.1 VO, was selected for its ability to grow in the absence of serum and is non-tumorigenic in athymic mice. The other two strains were selected for focus-formation following transfection with either Ha-ras or N-ras oncogenes and form high grade malignant tumors. Correspondence and cluster analysis provided a nonbiased estimate of the relative similarity of the different two-dimensional patterns. These techniques separated the gel patterns into three distinct classes: LG1, MSU-1.1 VO, and the ras transformed cell strains. The MSU-1.1 VO cells were more closely related to the parental LG1 than to the ras-transformed cells. The differences between the three classes were primarily quantitative in nature: 16% of the spots demonstrated statistically significant changes (P < 0.01, T test, mean ratio of intensity > 2) in the rate of incorporation of radioactive amino acids. The patterns from the two ras-transformed cell strains were similar, and variations in the expression of proteins that occurred between the separate experiments obscured consistent differences between the Ha-ras and N-ras transformed cells. However, while only 9 out of 758 spots were classified as different (1%), correspondence analysis could consistently separate the two ras transformants. One of these spots was five times more intense in the Ha-ras transformed cells than the N-ras.(ABSTRACT TRUNCATED AT 250 WORDS)

Quantitative and qualitative measure of intralaboratory two-dimensional protein gel reproducibility and the effects of sample preparation, sample load, and image analysis.

PubMed

Choe, Leila H; Lee, Kelvin H

2003-10-01

We investigate one approach to assess the quantitative variability in two-dimensional gel electrophoresis (2-DE) separations based on gel-to-gel variability, sample preparation variability, sample load differences, and the effect of automation on image analysis. We observe that 95% of spots present in three out of four replicate gels exhibit less than a 0.52 coefficient of variation (CV) in fluorescent stain intensity (% volume) for a single sample run on multiple gels. When four parallel sample preparations are performed, this value increases to 0.57. We do not observe any significant change in quantitative value for an increase or decrease in sample load of 30% when using appropriate image analysis variables. Increasing use of automation, while necessary in modern 2-DE experiments, does change the observed level of quantitative and qualitative variability among replicate gels. The number of spots that change qualitatively for a single sample run in parallel varies from a CV = 0.03 for fully manual analysis to CV = 0.20 for a fully automated analysis. We present a systematic method by which a single laboratory can measure gel-to-gel variability using only three gel runs.

Detection of phosphorylated forms of moloney murine leukemia virus major capsid protein p30 by immunoprecipitation and two-dimensional gel electrophoresis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ikuta, K.; Luftig, R.B.

1988-01-01

The authors detected phosphorylation of the major Moloney murine leukemia virus (M-MuLV) capsid polypeptide, p30, by using /sup 32/P/sub i/-labeled virions. This was observed both on two-dimensional polyacrylamide gels directly or on one-dimensional gels of viral lysates that had been immunoprecipitated with monospecific goat anti-p30 serum. The phosphorylation event had been difficult to detect because pp12 the major virion phosphoprotein incorporates almost all of the /sup 32/P label added to infected cells. When immunoprecipitates from M-MuLV lysates labeled with /sup 32/P/sub i/ were compared with those labeled with (/sup 35/S)methionine, it was calculated that the degree of phosphorylation at themore » p30 domain of Pr65/sup gag/ was only 0.22 to 0.54% relative to phosphorylation at the p12 domain. Two-dimensional gel electrophoresis of the /sup 32/P-labeled p30 immunoprecipitates showed that there were three phosphorylated p30 forms with isoelectric points (pIs) of 5.7, 5.8, and 6.0. These forms were generally more acidic than the (/sup 35/S) methionine-labeled p30 forms, which had pIs of 6.0, 6.1, 6.3 (the major constituent with > 80% of the label), and 6.6. The predominant phosphoamino acid of the major phosphorylated p30 form (pI 5.8) was phosphoserine. Further, tryptic peptide analysis of this p30 form showed that only one peptide was predominantly phosphorylated. Based on a comparison of specific labeling of p30 tryptic peptides with (/sup 14/C)sesrine, (/sup 35/S)methionine, and /sup 32/P/sub i/, we tentatively assigned the phosphorylation site to a 2.4-kilodalton NH/sub 2/-terminal peptide containing triple tandem serines spanning the region from amino acids 4 to 24.« less

Two-dimensional gel analysis of protein expression in ovarian tumors shows a low degree of intratumoral heterogeneity.

PubMed

Alaiya, A A; Franzén, B; Moberger, B; Silfverswärd, C; Linder, S; Auer, G

1999-01-01

The process of tumor progression leads to the emergence of multiple clones, and to the development of tumor heterogeneity. One approach to the study of the extent of such heterogeneity is to examine the expression of marker proteins in different tumor areas. Two-dimensional gel electrophoresis (2-DE) is a powerful tool for such studies, since the expression of a large number of polypeptide markers can be evaluated. In the present study, tumor cells were prepared from human ovarian tumors and analyzed by 2-DE and PDQUEST. As judged from the analysis of two different areas in each of nine ovarian tumors, the intratumoral variation in protein expression was low. In contrast, large differences were observed when the protein profiles of different tumors were compared. The differences in gene expression between pairs of malignant carcinomas were slightly larger than the differences observed between pairs of benign tumors. We conclude that 2-DE analysis of intratumoral heterogeneity in ovarian cancer tissue indicates a low degree of heterogeneity.

Electrophoretic extraction of proteins from two-dimensional electrophoresis gel spots

DOEpatents

Zhang, Jian-Shi; Giometti, C.S.; Tollaksen, S.L.

1987-09-04

After two-dimensional electrophoresis of proteins or the like, resulting in a polyacrylamide gel slab having a pattern of protein gel spots thereon, an individual protein gel spot is cored out from the slab, to form a gel spot core which is placed in an extraction tube, with a dialysis membrane across the lower end of the tube. Replicate gel spots can be cored out from replicate gel slabs and placed in the extraction tube. Molten agarose gel is poured into the extraction tube where the agarose gel hardens to form an immobilizing gel, covering the gel spot cores. The upper end portion of the extraction tube is filled with a volume of buffer solution, and the upper end is closed by another dialysis membrane. Upper and lower bodies of a buffer solution are brought into contact with the upper and lower membranes and are provided with electrodes connected to the positive and negative terminals of a dc power supply, thereby producing an electrical current which flows through the upper membrane, the volume of buffer solution, the agarose, the gel spot cores and the lower membrane. The current causes the proteins to be extracted electrophoretically from the gel spot cores, so that the extracted proteins accumulate and are contained in the space between the agarose gel and the upper membrane. 8 figs.

Investigation of Cu-, Zn- and Fe-containing human brain proteins using isotopic-enriched tracers by LA-ICP-MS and MALDI-FT-ICR-MS

NASA Astrophysics Data System (ADS)

Becker, J. Susanne; Zoriy, Miroslav; Pickhardt, Carola; Przybylski, Michael; Becker, J. Sabine

2005-04-01

Identification of metal-containing proteins and determination of Cu, Fe, Zn concentration in very small protein volumes is of increasing importance in protein research. Proteins containing metal ions were analyzed directly and simultaneously in separated protein spots in two-dimensional gels (2D gels) by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) as an element mass spectrometric technique. In order to study the formation of proteins containing Cu, Zn and Fe in a human brain sample, isotopic-enriched tracers (54Fe, 65Cu and 67Zn) were doped to two-dimensional gels of separated Alzheimer-diseased brain proteins after two-dimensional (2D) gel electrophoresis. The protein spots were screened systematically by LA-ICP-MS with respect to these metal ion intensities. 54Fe/56Fe, 65Cu/63Cu and 67Zn/64Zn isotope ratios in metal-containing proteins were measured directly by LA-ICP-MS. The isotope ratio measurements obtained by LA-ICP-MS indicate certain protein spots with a natural isotope composition of Cu, Zn and/or Fe. These proteins already contained the metal investigated in the original proteins and are stable enough to survive the reducing conditions during gel electrophoresis. On the other hand, proteins with a changed isotope ratio of metals in comparison to the isotope ratio in nature demonstrate the accumulation of tracers within the protein complexes during the tracer experiments in 2D gels. The identification of singular protein spots from Alzheimer-diseased brain separated by 2D gel electrophoresis was attempted by biopolymer mass spectrometry using MALDI-FTICR-MS after excision from the 2D gel and tryptic digestion.

Allelic analysis of low molecular weight glutenin subunits using 2-DGE in Korean wheat cultivars

USDA-ARS?s Scientific Manuscript database

Two-dimensional gel electrophoresis (2-DGE) was used to determine the allelic compositions of LMW-GS in 32 Korean wheat cultivars. Protein patterns generated by 2-DGE from each cultivar were compared to patterns from standard wheat cultivars for each allele. At the Glu-A3 locus, thirteen c, twelve ...

Simple and rapid system for two-dimensional gel electrophoresis technique: A laboratory exercise for high school students.

PubMed

Maurye, Praveen; Basu, Arpita; Biswas, Jayanta Kumar; Bandyopadhyay, Tapas Kumar; Naskar, Malay

2018-02-28

Polyacrylamide gel electrophoresis (PAGE) is the most classical technique favored worldwide for resolution of macromolecules in many biochemistry laboratories due to its incessant advanced developments and wide modifications. These ever-growing advancements in the basic laboratory equipments lead to emergence of many expensive, complex, and tricky laboratory equipments. Practical courses of biochemistry at high school or undergraduate levels are often affected by these complications. Two dimensional gel electrophoresis technique (2D-PAGE) used for resolving thousands of proteins in a gel is a combination of isoelectric focusing (first dimension gel electrophoresis technique) and sodium-dodecylsulphate PAGE (second dimension gel electrophoresis technique or SDS-PAGE). Two different laboratory equipments are needed to carry out effective 2D-PAGE technique, which also invites extra burden to the school laboratory. Here, we describe a low cost, time saving and simple gel cassette for protein 2D-PAGE technique that uses easily fabricated components and routine off-the-shelf materials. The performance of the apparatus was verified in a practical exercise by a group of high school students with positive outcomes. © 2018 by The International Union of Biochemistry and Molecular Biology, 2018. © 2018 The International Union of Biochemistry and Molecular Biology.

Drug transporter expression profiling in a three-dimensional kidney proximal tubule in vitro nephrotoxicity model.

PubMed

Diekjürgen, Dorina; Grainger, David W

2018-05-09

Given currently poor toxicity translational predictions for drug candidates, improved mechanistic understanding underlying nephrotoxicity and drug renal clearance is needed to improve drug development and safety screening. Therefore, better relevant and well-characterized in vitro screening models are required to reliably predict human nephrotoxicity. Because kidney proximal tubules are central to active drug uptake and secretion processes and therefore to nephrotoxicity, this study acquired regio-specific expression data from recently reported primary proximal tubule three-dimensional (3D) hyaluronic acid gel culture and non-gel embedded cultured murine proximal tubule suspensions used in nephrotoxicity assays. Quantitative assessment of the mRNA expression of 21 known kidney tubule markers and important proximal tubule transporters with known roles in drug transport was obtained. Asserting superior gene expression levels over current commonly used two-dimensional (2D) kidney cell culture lines was the study objective. Hence, we compare previously published gel-based 3D proximal tubule fragment culture and their non-gel suspensions for up to 1 week. We demonstrate that 3D tubule culture exhibits superior gene expression levels and profiles compared to published commonly used 2D kidney cell lines (Caki-1 and HK-2) in plastic plate monocultures. Additionally, nearly all tested genes retain mRNA expression after 7 days in both proximal tubule cultures, a limitation of 2D cell culture lines. Importantly, gel presence is shown not to interfere with the gene expression assay. Western blots confirm protein expression of OAT1 and 3 and OCT2. Functional transport assays confirm their respective transporter functions in vitro. Overall, results validate retention of essential toxicity-relevant transporters in this published 3D proximal tubule model over conventional 2D kidney cell cultures, producing opportunities for more reliable, sensitive, and comprehensive drug toxicity studies relevant to drug development and nephrotoxicity goals.

Ignition and combustion characteristics of metallized propellants, phase 2

NASA Technical Reports Server (NTRS)

Mueller, D. C.; Turns, S. R.

1994-01-01

Experimental and analytical investigations focusing on aluminum/hydrocarbon gel droplet secondary atomization and its effects on gel-fueled rocket engine performance are being conducted. A single laser sheet sizing/velocimetry diagnostic technique, which should eliminate sizing bias in the data collection process, has been designed and constructed to overcome limitations of the two-color forward-scatter technique used in previous work. Calibration of this system is in progress and the data acquisition/validation code is being written. Narrow-band measurements of radiant emission, discussed in previous reports, will be used to determine if aluminum ignition has occurred in a gel droplet. A one-dimensional model of a gel-fueled rocket combustion chamber, described in earlier reports, has been exercised in conjunction with a two-dimensional, two-phase nozzle code to predict the performance of an aluminum/hydrocarbon fueled engine. Estimated secondary atomization effects on propellant burnout distance, condensed particle radiation losses to the chamber walls, and nozzle two phase flow losses are also investigated. Calculations indicate that only modest secondary atomization is required to significantly reduce propellant burnout distances, aluminum oxide residual size, and radiation heat losses. Radiation losses equal to approximately 2-13 percent of the energy released during combustion were estimated, depending on secondary atomization intensity. A two-dimensional, two-phase nozzle code was employed to estimate radiation and nozzle two phase flow effects on overall engine performance. Radiation losses yielded a one percent decrease in engine Isp. Results also indicate that secondary atomization may have less effect on two-phase losses than it does on propellant burnout distance and no effect if oxide particle coagulation and shear induced droplet breakup govern oxide particle size. Engine Isp was found to decrease from 337.4 to 293.7 seconds as gel aluminum mass loading was varied from 0-70 wt percent. Engine Isp efficiencies, accounting for radiation and two phase flow effects, on the order of 0.946 were calculated for a 60 wt percent gel, assuming a fragmentation ratio of five.

Cell and organ printing 2: fusion of cell aggregates in three-dimensional gels.

PubMed

Boland, Thomas; Mironov, Vladimir; Gutowska, Anna; Roth, Elisabeth A; Markwald, Roger R

2003-06-01

We recently developed a cell printer (Wilson and Boland, 2003) that enables us to place cells in positions that mimic their respective positions in organs. However, this technology was limited to the printing of two-dimensional (2D) tissue constructs. Here we describe the use of thermosensitive gels to generate sequential layers for cell printing. The ability to drop cells on previously printed successive layers provides a real opportunity for the realization of three-dimensional (3D) organ printing. Organ printing will allow us to print complex 3D organs with computer-controlled, exact placing of different cell types, by a process that can be completed in several minutes. To demonstrate the feasibility of this novel technology, we showed that cell aggregates can be placed in the sequential layers of 3D gels close enough for fusion to occur. We estimated the optimum minimal thickness of the gel that can be reproducibly generated by dropping the liquid at room temperature onto a heated substrate. Then we generated cell aggregates with the corresponding (to the minimal thickness of the gel) size to ensure a direct contact between printed cell aggregates during sequential printing cycles. Finally, we demonstrated that these closely-placed cell aggregates could fuse in two types of thermosensitive 3D gels. Taken together, these data strongly support the feasibility of the proposed novel organ-printing technology. Copyright 2003 Wiley-Liss, Inc.

The Make 2D-DB II package: conversion of federated two-dimensional gel electrophoresis databases into a relational format and interconnection of distributed databases.

PubMed

Mostaguir, Khaled; Hoogland, Christine; Binz, Pierre-Alain; Appel, Ron D

2003-08-01

The Make 2D-DB tool has been previously developed to help build federated two-dimensional gel electrophoresis (2-DE) databases on one's own web site. The purpose of our work is to extend the strength of the first package and to build a more efficient environment. Such an environment should be able to fulfill the different needs and requirements arising from both the growing use of 2-DE techniques and the increasing amount of distributed experimental data.

An Optimized Trichloroacetic Acid/Acetone Precipitation Method for Two-Dimensional Gel Electrophoresis Analysis of Qinchuan Cattle Longissimus Dorsi Muscle Containing High Proportion of Marbling.

PubMed

Hao, Ruijie; Adoligbe, Camus; Jiang, Bijie; Zhao, Xianlin; Gui, Linsheng; Qu, Kaixing; Wu, Sen; Zan, Linsen

2015-01-01

Longissimus dorsi muscle (LD) proteomics provides a novel opportunity to reveal the molecular mechanism behind intramuscular fat deposition. Unfortunately, the vast amounts of lipids and nucleic acids in this tissue hampered LD proteomics analysis. Trichloroacetic acid (TCA)/acetone precipitation is a widely used method to remove contaminants from protein samples. However, the high speed centrifugation employed in this method produces hard precipitates, which restrict contaminant elimination and protein re-dissolution. To address the problem, the centrifugation precipitates were first grinded with a glass tissue grinder and then washed with 90% acetone (TCA/acetone-G-W) in the present study. According to our result, the treatment for solid precipitate facilitated non-protein contaminant removal and protein re-dissolution, ultimately improving two-dimensional gel electrophoresis (2-DE) analysis. Additionally, we also evaluated the effect of sample drying on 2-DE profile as well as protein yield. It was found that 30 min air-drying did not result in significant protein loss, but reduced horizontal streaking and smearing on 2-DE gel compared to 10 min. In summary, we developed an optimized TCA/acetone precipitation method for protein extraction of LD, in which the modifications improved the effectiveness of TCA/acetone method.

An Optimized Trichloroacetic Acid/Acetone Precipitation Method for Two-Dimensional Gel Electrophoresis Analysis of Qinchuan Cattle Longissimus Dorsi Muscle Containing High Proportion of Marbling

PubMed Central

Hao, Ruijie; Adoligbe, Camus; Jiang, Bijie; Zhao, Xianlin; Gui, Linsheng; Qu, Kaixing; Wu, Sen; Zan, Linsen

2015-01-01

Longissimus dorsi muscle (LD) proteomics provides a novel opportunity to reveal the molecular mechanism behind intramuscular fat deposition. Unfortunately, the vast amounts of lipids and nucleic acids in this tissue hampered LD proteomics analysis. Trichloroacetic acid (TCA)/acetone precipitation is a widely used method to remove contaminants from protein samples. However, the high speed centrifugation employed in this method produces hard precipitates, which restrict contaminant elimination and protein re-dissolution. To address the problem, the centrifugation precipitates were first grinded with a glass tissue grinder and then washed with 90% acetone (TCA/acetone-G-W) in the present study. According to our result, the treatment for solid precipitate facilitated non-protein contaminant removal and protein re-dissolution, ultimately improving two-dimensional gel electrophoresis (2-DE) analysis. Additionally, we also evaluated the effect of sample drying on 2-DE profile as well as protein yield. It was found that 30 min air-drying did not result in significant protein loss, but reduced horizontal streaking and smearing on 2-DE gel compared to 10 min. In summary, we developed an optimized TCA/acetone precipitation method for protein extraction of LD, in which the modifications improved the effectiveness of TCA/acetone method. PMID:25893432

A Robust Two-Dimensional Separation for Top-Down Tandem Mass Spectrometry of the Low-Mass Proteome

PubMed Central

Lee, Ji Eun; Kellie, John F.; Tran, John C.; Tipton, Jeremiah D.; Catherman, Adam D.; Thomas, Haylee M.; Ahlf, Dorothy R.; Durbin, Kenneth R.; Vellaichamy, Adaikkalam; Ntai, Ioanna; Marshall, Alan G.; Kelleher, Neil L.

2010-01-01

For fractionation of intact proteins by molecular weight (MW), a sharply improved two-dimensional (2D) separation is presented to drive reproducible and robust fractionation before top-down mass spectrometry of complex mixtures. The “GELFrEE” (i.e., gel-eluted liquid fraction entrapment electrophoresis) approach is implemented by use of Tris-glycine and Tris-tricine gel systems applied to human cytosolic and nuclear extracts from HeLa S3 cells, to achieve a MW-based fractionation of proteins from 5 to >100 kDa in 1 h. For top-down tandem mass spectroscopy (MS/MS) of the low-mass proteome (5–25 kDa), between 5 and 8 gel-elution (GE) fractions are sampled by nanocapillary-LC-MS/MS with 12 or 14.5 tesla Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometers. Single injections give about 40 detectable proteins, about half of which yield automated ProSight identifications. Reproducibility metrics of the system are presented, along with comparative analysis of protein targets in mitotic versus asynchronous cells. We forward this basic 2D approach to facilitate wider implementation of top-down mass spectrometry and a variety of other protein separation and/or characterization approaches. PMID:19747844

GESA--a two-dimensional processing system using knowledge base techniques.

PubMed

Rowlands, D G; Flook, A; Payne, P I; van Hoff, A; Niblett, T; McKee, S

1988-12-01

The successful analysis of two-dimensional (2-D) polyacrylamide electrophoresis gels demands considerable experience and understanding of the protein system under investigation as well as knowledge of the separation technique itself. The present work concerns the development of a computer system for analysing 2-D electrophoretic separations which incorporates concepts derived from artificial intelligence research such that non-experts can use the technique as a diagnostic or identification tool. Automatic analysis of 2-D gel separations has proved to be extremely difficult using statistical methods. Non-reproducibility of gel separations is also difficult to overcome using automatic systems. However, the human eye is extremely good at recognising patterns in images, and human intervention in semi-automatic computer systems can reduce the computational complexities of fully automatic systems. Moreover, the expertise and understanding of an "expert" is invaluable in reducing system complexity if it can be encapsulated satisfactorily in an expert system. The combination of user-intervention in the computer system together with the encapsulation of expert knowledge characterises the present system. The domain within which the system has been developed is that of wheat grain storage proteins (gliadins) which exhibit polymorphism to such an extent that cultivars can be uniquely identified by their gliadin patterns. The system can be adapted to other domains where a range of polymorpic protein sub-units exist. In its generalised form, the system can also be used for comparing more complex 2-D gel electrophoretic separations.

Laterally structured ripple and square phases with one and two dimensional thickness modulations in a model bilayer system.

PubMed

Debnath, Ananya; Thakkar, Foram M; Maiti, Prabal K; Kumaran, V; Ayappa, K G

2014-10-14

Molecular dynamics simulations of bilayers in a surfactant/co-surfactant/water system with explicit solvent molecules show formation of topologically distinct gel phases depending upon the bilayer composition. At low temperatures, the bilayers transform from the tilted gel phase, Lβ', to the one dimensional (1D) rippled, Pβ' phase as the surfactant concentration is increased. More interestingly, we observe a two dimensional (2D) square phase at higher surfactant concentration which, upon heating, transforms to the gel Lβ' phase. The thickness modulations in the 1D rippled and square phases are asymmetric in two surfactant leaflets and the bilayer thickness varies by a factor of ∼2 between maximum and minimum. The 1D ripple consists of a thinner interdigitated region of smaller extent alternating with a thicker non-interdigitated region. The 2D ripple phase is made up of two superimposed square lattices of maximum and minimum thicknesses with molecules of high tilt forming a square lattice translated from the lattice formed with the thickness minima. Using Voronoi diagrams we analyze the intricate interplay between the area-per-head-group, height modulations and chain tilt for the different ripple symmetries. Our simulations indicate that composition plays an important role in controlling the formation of low temperature gel phase symmetries and rippling accommodates the increased area-per-head-group of the surfactant molecules.

Fluctuations and symmetries in two-dimensional active gels.

PubMed

Sarkar, N; Basu, A

2011-04-01

Motivated by the unique physical properties of biological active matter, e.g., cytoskeletal dynamics in eukaryotic cells, we set up effective two-dimensional (2d) coarse-grained hydrodynamic equations for the dynamics of thin active gels with polar or nematic symmetries. We use the well-known three-dimensional (3d) descriptions (K. Kruse et al., Eur. Phys. J. E 16, 5 (2005); A. Basu et al., Eur. Phys. J. E 27, 149 (2008)) for thin active-gel samples confined between parallel plates with appropriate boundary conditions to derive the effective 2d constitutive relations between appropriate thermodynamic fluxes and generalised forces for small deviations from equilibrium. We consider three distinct cases, characterised by spatial symmetries and boundary conditions, and show how such considerations dictate the structure of the constitutive relations. We use these to study the linear instabilities, calculate the correlation functions and the diffusion constant of a small tagged particle, and elucidate their dependences on the activity or nonequilibrium drive.

Effects of cedar oil and silica gel treatment on dimensional stability and mechanical properties of Southern Yellow Pine boards

Treesearch

Xiping Wang; Christopher Adam Senalik; Robert Ross; Neal Bennett; Debbie Conner

2016-01-01

A laboratory study was conducted to investigate the effects of cedar oil and silica gel treatment on dimensional stability and mechanical performance of southern yellow pine (SYP) boards. Two hundred pieces of SYP and 100 pieces of red oak boards with a nominal dimension of 1 by 6 by 48 in. (25 by 152 by 1,219 mm) were selected for this study. The red oak boards were...

Verification on the Dose Profile Variation of a 3-D—NIPAM Polymer Gel Dosimeter

NASA Astrophysics Data System (ADS)

Hsieh, Bor-Tsung; Wu, Jay; Chang, Yuan-Jen

2013-04-01

A gel dosimeter is a three-dimensional (3-D) device that is used in radiotherapy. It is more efficient than traditional one-dimensional and two-dimensional dosimeters because it can be used in complicated radiation therapy applications. However, the achievement of temporal and spatial stabilities for gel dosimeters remains challenging in clinical applications because the fabrication process affects the polymerization reaction during irradiation. This study investigated the dose profile variation of an N-isopropyl acrylamide (NIPAM) polymer gel dosimeter by using the 3-D optical computed tomography scanner OCTOPUSTM 10X (MGS Research Inc.). Two acrylic containers (diameter=10, height=10, and diameter=15, height=15cm ) filled with polymer gel (gelatin: 5%, NIPAM: 5%, Bis: 3%, THPC: 5 mM) were irradiated by using intensity-modulated radiotherapy (SIEMENS Oncor Impression, 6 MV Photo beam). The treatment field was a 3 cm 3 cm square field, and the prescribed dose was 5 Gy. The results of the reconstruction line profile showed that the uncertainty of non-irradiated gel is less than 1.3% when a container with 10 cm diameters cooled in a refrigerator with a water bath. The maximum uncertainties of the irradiated gel at 24 h, 48 h, and 72 h post-irradiation were 2.9%, 2.9%, and 3.1%, respectively. However, the maximum uncertainty of the non-irradiated gel dosimeter increased to 3% when a container with 15 cm diameter was cooled in the same refrigerator. After irradiation, the maximum uncertainties of the irradiated gel at 24 h, 48 h, and 72 h post-irradiation were 13.1%, 13.7%, and 12.95%, respectively. The uncertainty differences for gels at different container sizes were attributed to the different cooling rates that were applied to the gels. The time required for large gel containers to cool in the refrigerator was more than 10 h, whereas the cooling process only took 4.2 h for gels in a small container. The time difference produced different temperature histories for gels and may result in changes in gel sensitivity. Given the thermally induced pre-radiation polymerization, the time difference resulted in a deviation in dose profiles. This study reports that thermal control during gel preparation should be carefully performed for clinical applications to achieve a more accurate dose distribution in 3-D image reconstruction.

Two-dimensional proteome reference maps for the soybean cyst nematode Heterodera glycines

USDA-ARS?s Scientific Manuscript database

Two-dimensional electrophoresis (2-DE) reference maps of Heterodera glycines were constructed. After in-gel digestion with trypsin, 803 spots representing 426 proteins were subsequently identified by LC-MS/MS. Proteins with annotated function were further categorized by Gene Ontology. Results showed...

Comparative proteome analysis of laboratory grown Brucella abortus 2308 and Brucella melitensis 16M.

PubMed

Eschenbrenner, Michel; Horn, Troy A; Wagner, Mary Ann; Mujer, Cesar V; Miller-Scandle, Tabbi L; DelVecchio, Vito G

2006-07-01

Brucella species are pathogenic agents that cause brucellosis, a debilitating zoonotic disease that affects a large variety of domesticated animals and humans. Brucella melitensis and Brucella abortus are considered major health threats because of their highly infectious nature and worldwide occurrence. The availability of the annotated genomes for these two species has allowed a comparative proteomics study of laboratory grown B. melitensis 16M and B. abortus 2308 by two-dimensional (2-D) gel electrophoresis and peptide mass fingerprinting. Computer-assisted analysis of the different 2-D gel images of strains 16M and 2308 revealed significant quantitative and qualitative differences in their protein expression patterns. Proteins involved in membrane transport, particularly the high affinity amino acids binding proteins, and those involved in Sec-dependent secretion systems related to type IV and type V secretion systems, were differentially expressed. Differential expression of these proteins may be responsible for conferring specific host preference in the two strains 2308 and 16M.

Optimization of Protein Extraction and Two-Dimensional Electrophoresis Protocols for Oil Palm Leaf.

PubMed

Daim, Leona Daniela Jeffery; Ooi, Tony Eng Keong; Yusof, Hirzun Mohd; Majid, Nazia Abdul; Karsani, Saiful Anuar Bin

2015-08-01

Oil palm (Elaeis guineensis) is an important economic crop cultivated for its nutritional palm oil. A significant amount of effort has been undertaken to understand oil palm growth and physiology at the molecular level, particularly in genomics and transcriptomics. Recently, proteomics studies have begun to garner interest. However, this effort is impeded by technical challenges. Plant sample preparation for proteomics analysis is plagued with technical challenges due to the presence of polysaccharides, secondary metabolites and other interfering compounds. Although protein extraction methods for plant tissues exist, none work universally on all sample types. Therefore, this study aims to compare and optimize different protein extraction protocols for use with two-dimensional gel electrophoresis of young and mature leaves from the oil palm. Four protein extraction methods were evaluated: phenol-guanidine isothiocyanate, trichloroacetic acid-acetone precipitation, sucrose and trichloroacetic acid-acetone-phenol. Of these four protocols, the trichloroacetic acid-acetone-phenol method was found to give the highest resolution and most reproducible gel. The results from this study can be used in sample preparations of oil palm tissue for proteomics work.

Unraveling the equine lymphocyte proteome: differential septin 7 expression associates with immune cells in equine recurrent uveitis.

PubMed

Degroote, Roxane L; Hauck, Stefanie M; Amann, Barbara; Hirmer, Sieglinde; Ueffing, Marius; Deeg, Cornelia A

2014-01-01

Equine recurrent uveitis is a spontaneous, lymphocyte-driven autoimmune disease. It affects horses worldwide and presents with painful remitting-relapsing inflammatory attacks of inner eye structures eventually leading to blindness. Since lymphocytes are the key players in equine recurrent uveitis, we were interested in potential changes of their protein repertoire which may be involved in disease pathogenesis. To create a reference for differential proteome analysis, we first unraveled the equine lymphocyte proteome by two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and subsequently identified 352 protein spots. Next, we compared lymphocytes from ERU cases and healthy horses with a two-dimensional fluorescence difference in gel electrophoresis approach. With this technique, we identified seven differentially expressed proteins between conditions. One of the significantly lower expressed candidates, septin 7, plays a role in regulation of cell shape, motility and migration. Further analyses revealed T cells as the main cell type with decreased septin 7 abundance in equine recurrent uveitis. These findings point to a possible pathogenetic role of septin 7 in this sight-threatening disease.

SU-E-T-753: Three-Dimensional Dose Distributions of Incident Proton Particle in the Polymer Gel Dosimeter and the Radiochromic Gel Dosimeter: A Simulation Study with MCNP Code

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, M; Kim, G; Ji, Y

Purpose: The purpose of this study is to estimate the three-dimensional dose distributions in the polymer and the radiochromic gel dosimeter, and to identify the detectability of both gel dosimeters by comparing with the water phantom in case of irradiating the proton particles. Methods: The normoxic polymer gel and the LCV micelle radiochromic gel were used in this study. The densities of polymer and the radiochromic gel dosimeter were 1.024 and 1.005 g/cm{sup 3}, respectively. The dose distributions of protons in the polymer and radiochromic gel were simulated using Monte Carlo radiation transport code (MCNPX, Los Alamos National Laboratory). Themore » shape of phantom irradiated by proton particles was a hexahedron with the dimension of 12.4 × 12.4 × 15.0 cm{sup 3}. The energies of proton beam were 50, 80, and 140 MeV energies were directed to top of the surface of phantom. The cross-sectional view of proton dose distribution in both gel dosimeters was estimated with the water phantom and evaluated by the gamma evaluation method. In addition, the absorbed dose(Gy) was also calculated for evaluating the proton detectability. Results: The evaluation results show that dose distributions in both gel dosimeters at intermediated section and Bragg-peak region are similar with that of the water phantom. At entrance section, however, inconsistencies of dose distribution are represented, compared with water. The relative absorbed doses in radiochromic and polymer gel dosimeter were represented to be 0.47 % and 2.26 % difference, respectively. These results show that the radiochromic gel dosimeter was better matched than the water phantom in the absorbed dose evaluation. Conclusion: The polymer and the radiochromic gel dosimeter show similar characteristics in dose distributions for the proton beams at intermediate section and Bragg-peak region. Moreover the calculated absorbed dose in both gel dosimeters represents similar tendency by comparing with that in water phantom.« less

Alterations in brain cerebral cortex proteome of rabies-infected cat.

PubMed

Kasempimolporn, Songsri; Lumlertdacha, Boonlert; Chulasugandha, Pannipa; Boonchang, Supatsorn; Sitprija, Visith

2014-07-01

Comparative proteome analysis using brain cerebral cortex tissues from cats and dogs infected with/without rabies virus were conducted using both two-dimensional gel-electrophoresis (2-DE) and 2-D fluorescence difference gel- electrophoresis (2D-DIGE) methods. The 2-DE gel images of all samples revealed >1,000 protein spots in each gel. Quantitative intensity analysis revealed the same overall protein pattern in certain regions of the gel, but the rabies-infected brains exhibited more protein spots than the non-infected controls. From approximately 880 protein spots detected by 2D-DIGE, 65 protein spots were increased and 46 were decreased. Eight of these protein spots were randomly selected and annotated by reference to previous known proteome data of rabid dog brains. They were similarly altered in both of the rabies-infected cats and dogs. A more detailed comparison of changes in proteomic profiles of brains between rabid cats and dogs should shed some light on the pathophysiological mechanism of rabies in domestic animals, as most rabies cases have been traceable to or believed to have originated from rabid dogs.

Protein Changes in Macrophages Induced by Plasma from Rats Exposed to 35-GHz Millimeter Waves

DTIC Science & Technology

2010-12-01

HumanEffectiveness Directorate, Air Force Research Laboratory, Brooks City-Base,Texas A macrophage assay and proteomic screening were used to...mW/cm2 until core temperature reached 41.0 8C. Two-dimensional polyacrylamide gel electrophoresis, image analysis, and Western blotting were used to...stimulation. Proteins of interest were identified using peptide mass fingerprinting. Compared to plasma from sham- exposed rats, plasma from

Three-dimensional multichannel aerogel of carbon quantum dots for high-performance supercapacitors.

PubMed

Lv, Lingxiao; Fan, Yueqiong; Chen, Qing; Zhao, Yang; Hu, Yue; Zhang, Zhipan; Chen, Nan; Qu, Liangti

2014-06-13

A three-dimensional (3D) carbon quantum dot (CQD) aerogel has been prepared by in situ assembling CQDs in the sol-gel polymerization of resorcinol (R) and formaldehyde (F) and subsequently pyrolyzing the formed CQD gel. Compared to the supercapacitor based on the CQD-free aerogel, the supercapacitor fabricated with the CQD aerogel showed 20-fold higher specific capacitance (294.7 F g(-1) at the current density of 0.5 A g(-1)) and an excellent stability over 1000 consecutive charge-discharge cycles.

Three-dimensional multichannel aerogel of carbon quantum dots for high-performance supercapacitors

NASA Astrophysics Data System (ADS)

Lv, Lingxiao; Fan, Yueqiong; Chen, Qing; Zhao, Yang; Hu, Yue; Zhang, Zhipan; Chen, Nan; Qu, Liangti

2014-06-01

A three-dimensional (3D) carbon quantum dot (CQD) aerogel has been prepared by in situ assembling CQDs in the sol-gel polymerization of resorcinol (R) and formaldehyde (F) and subsequently pyrolyzing the formed CQD gel. Compared to the supercapacitor based on the CQD-free aerogel, the supercapacitor fabricated with the CQD aerogel showed 20-fold higher specific capacitance (294.7 F g-1 at the current density of 0.5 A g-1) and an excellent stability over 1000 consecutive charge-discharge cycles.

Comparative proteomic analysis of the ribosomes in 5-fluorouracil resistance of a human colon cancer cell line using the radical-free and highly reducing method of two-dimensional polyacrylamide gel electrophoresis.

PubMed

Kimura, Kosei; Wada, Akira; Ueta, Masami; Ogata, Akihiko; Tanaka, Satoru; Sakai, Akiko; Yoshida, Hideji; Fushitani, Hideo; Miyamoto, Akiko; Fukushima, Masakazu; Uchiumi, Toshio; Tanigawa, Nobuhiko

2010-11-01

Many auxiliary functions of ribosomal proteins (r-proteins) have received considerable attention in recent years. However, human r-proteins have hardly been examined by proteomic analysis. In this study, we isolated ribosomal particles and subsequently compared the proteome of r-proteins between the DLD-1 human colon cancer cell line and its 5-fluorouracil (5-FU)-resistant sub-line, DLD-1/5-FU, using the radical-free and highly reducing method of two-dimensional polyacrylamide gel electrophoresis, which has a superior ability to separate basic proteins, and we discuss the role of r-proteins in 5-FU resistance. Densitometric analysis was performed to quantify modulated proteins, and protein spots showing significant changes were identified by employing matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry. Three basic proteins (L15, L37 and prohibitin) which were significantly modulated between DLD-1 and DLD-1/5-FU were identified. Two proteins, L15 and L37, showed down-regulated expression in DLD-1/5-FU in comparison to DLD-1. Prohibitin, which is not an r-protein and is known to be localized in the mitochondria, showed up-regulated expression in DLD-1/5-FU. These 3 proteins may be related to 5-FU resistance.

Three-Dimensional Bioprinting of Oppositely Charged Hydrogels with Super Strong Interface Bonding.

PubMed

Li, Huijun; Tan, Yu Jun; Liu, Sijun; Li, Lin

2018-04-04

A novel strategy to improve the adhesion between printed layers of three-dimensional (3D) printed constructs is developed by exploiting the interaction between two oppositely charged hydrogels. Three anionic hydrogels [alginate, xanthan, and κ-carrageenan (Kca)] and three cationic hydrogels [chitosan, gelatin, and gelatin methacrylate (GelMA)] are chosen to find the optimal combination of two oppositely charged hydrogels for the best 3D printability with strong interface bonding. Rheological properties and printability of the hydrogels, as well as structural integrity of printed constructs in cell culture medium, are studied as functions of polymer concentration and the combination of hydrogels. Kca2 (2 wt % Kca hydrogel) and GelMA10 (10 wt % GelMA hydrogel) are found to be the best combination of oppositely charged hydrogels for 3D printing. The interfacial bonding between a Kca layer and a GelMA layer is proven to be significantly higher than that of the bilayered Kca or bilayered GelMA because of the formation of polyelectrolyte complexes between the oppositely charged hydrogels. A good cell viability of >96% is obtained for the 3D-bioprinted Kca-GelMA construct. This novel strategy has a great potential for 3D bioprinting of layered constructs with a strong interface bonding.

The Proteome of Seed Development in the Model Legume Lotus japonicus1[C][W

PubMed Central

Dam, Svend; Laursen, Brian S.; Ørnfelt, Jane H.; Jochimsen, Bjarne; Stærfeldt, Hans Henrik; Friis, Carsten; Nielsen, Kasper; Goffard, Nicolas; Besenbacher, Søren; Krusell, Lene; Sato, Shusei; Tabata, Satoshi; Thøgersen, Ida B.; Enghild, Jan J.; Stougaard, Jens

2009-01-01

We have characterized the development of seeds in the model legume Lotus japonicus. Like soybean (Glycine max) and pea (Pisum sativum), Lotus develops straight seed pods and each pod contains approximately 20 seeds that reach maturity within 40 days. Histological sections show the characteristic three developmental phases of legume seeds and the presence of embryo, endosperm, and seed coat in desiccated seeds. Furthermore, protein, oil, starch, phytic acid, and ash contents were determined, and this indicates that the composition of mature Lotus seed is more similar to soybean than to pea. In a first attempt to determine the seed proteome, both a two-dimensional polyacrylamide gel electrophoresis approach and a gel-based liquid chromatography-mass spectrometry approach were used. Globulins were analyzed by two-dimensional polyacrylamide gel electrophoresis, and five legumins, LLP1 to LLP5, and two convicilins, LCP1 and LCP2, were identified by matrix-assisted laser desorption ionization quadrupole/time-of-flight mass spectrometry. For two distinct developmental phases, seed filling and desiccation, a gel-based liquid chromatography-mass spectrometry approach was used, and 665 and 181 unique proteins corresponding to gene accession numbers were identified for the two phases, respectively. All of the proteome data, including the experimental data and mass spectrometry spectra peaks, were collected in a database that is available to the scientific community via a Web interface (http://www.cbs.dtu.dk/cgi-bin/lotus/db.cgi). This database establishes the basis for relating physiology, biochemistry, and regulation of seed development in Lotus. Together with a new Web interface (http://bioinfoserver.rsbs.anu.edu.au/utils/PathExpress4legumes/) collecting all protein identifications for Lotus, Medicago, and soybean seed proteomes, this database is a valuable resource for comparative seed proteomics and pathway analysis within and beyond the legume family. PMID:19129418

Extraction and Assembly of Tissue-Derived Gels for Cell Culture and Tissue Engineering

PubMed Central

Uriel, Shiri; Labay, Edwardine; Francis-Sedlak, Megan; Moya, Monica L.; Weichselbaum, Ralph R.; Ervin, Natalia; Cankova, Zdravka

2009-01-01

Interactions with the extracellular matrix (ECM) play an important role in regulating cell function. Cells cultured in, or on, three-dimensional ECM recapitulate similar features to those found in vivo that are not present in traditional two-dimensional culture. In addition, both natural and synthetic materials containing ECM components have shown promise in a number of tissue engineering applications. Current materials available for cell culture and tissue engineering do not adequately reflect the diversity of ECM composition between tissues. In this paper, a method is presented for extracting solutions of proteins and glycoproteins from soft tissues and inducing assembly of these proteins into gels. The extracts contain ECM proteins specific to the tissue source with low levels of intracellular molecules. Gels formed from the tissue-derived extracts have nanostructure similar to ECM in vivo and can be used to culture cells as both a thin substrate coating and a thick gel. This technique could be used to assemble hydrogels with varying composition depending upon the tissue source, hydrogels for three-dimensional culture, as scaffolds for tissue engineering therapies, and to study cell–matrix interactions. PMID:19115821

Informatics and Statistics for Analyzing 2-D Gel Electrophoresis Images

PubMed Central

Dowsey, Andrew W.; Morris, Jeffrey S.; Gutstein, Howard B.; Yang, Guang-Zhong

2013-01-01

Despite recent progress in “shotgun” peptide separation by integrated liquid chromatography and mass spectrometry (LC/MS), proteome coverage and reproducibility are still limited with this approach and obtaining enough replicate runs for biomarker discovery is a challenge. For these reasons, recent research demonstrates that there is a continuing need for protein separation by two-dimensional gel electrophoresis (2-DE). However, with traditional 2-DE informatics, the digitized images are reduced to symbolic data through spot detection and quantification before proteins are compared for differential expression by spot matching. Recently, a more robust and automated paradigm has emerged where gels are directly aligned in the image domain before spots are detected across the whole image set as a whole. In this chapter, we describe the methodology for both approaches and discuss the pitfalls present when reasoning statistically about the differential protein expression discovered. PMID:20013375

[Accuracy Check of Monte Carlo Simulation in Particle Therapy Using Gel Dosimeters].

PubMed

Furuta, Takuya

2017-01-01

Gel dosimeters are a three-dimensional imaging tool for dose distribution induced by radiations. They can be used for accuracy check of Monte Carlo simulation in particle therapy. An application was reviewed in this article. An inhomogeneous biological sample placing a gel dosimeter behind it was irradiated by carbon beam. The recorded dose distribution in the gel dosimeter reflected the inhomogeneity of the biological sample. Monte Carlo simulation was conducted by reconstructing the biological sample from its CT image. The accuracy of the particle transport by Monte Carlo simulation was checked by comparing the dose distribution in the gel dosimeter between simulation and experiment.

Sera of IgA nephropathy patients contain a heterogeneous population of relatively cationic alpha-heavy chains.

PubMed

Shuib, A S; Chua, C T; Hashim, O H

1998-01-01

Sera of IgA nephropathy (IgAN) patients and normal subjects were analysed by two-dimensional (2-D) gel electrophoresis. Densitometric analysis of the 2-D gels of IgAN patients and normal subjects revealed that their protein maps were comparable. There was no shift of pI values in the major alpha-heavy chain spots. However, the volume of the alpha-heavy chain bands were differently distributed. Distribution was significantly lower at the anionic region in IgAN patients (mean anionic:cationic ratio of 1.184 +/- 0.311) as compared to normal healthy controls (mean anionic:cationic ratio of 2.139 +/- 0.538). Our data are in support of the previously reported findings that IgA1 of IgAN patients were lacking in sialic acid residues.

Increase in local protein concentration by field-inversion gel electrophoresis.

PubMed

Tsai, Henghang; Low, Teck Yew; Freeby, Steve; Paulus, Aran; Ramnarayanan, Kalpana; Cheng, Chung-Pui Paul; Leung, Hon-Chiu Eastwood

2007-09-26

Proteins that migrate through cross-linked polyacrylamide gels (PAGs) under the influence of a constant electric field experience negative factors, such as diffusion and non-specific trapping in the gel matrix. These negative factors reduce protein concentrations within a defined gel volume with increasing migration distance and, therefore, decrease protein separation efficiency. Enhancement of protein separation efficiency was investigated by implementing pulsed field-inversion gel electrophoresis (FIGE). Separation of model protein species and large protein complexes was compared between FIGE and constant field electrophoresis (CFE) in different percentages of PAGs. Band intensities of proteins in FIGE with appropriate ratios of forward and backward pulse times were superior to CFE despite longer running times. These results revealed an increase in band intensity per defined gel volume. A biphasic protein relative mobility shift was observed in percentages of PAGs up to 14%. However, the effect of FIGE on protein separation was stochastic at higher PAG percentage. Rat liver lysates subjected to FIGE in the second-dimension separation of two-dimensional polyarcylamide gel electrophoresis (2D PAGE) showed a 20% increase in the number of discernible spots compared with CFE. Nine common spots from both FIGE and CFE were selected for peptide sequencing by mass spectrometry (MS), which revealed higher final ion scores of all nine protein spots from FIGE. Native protein complexes ranging from 800 kDa to larger than 2000 kDa became apparent using FIGE compared with CFE. The present investigation suggests that FIGE under appropriate conditions improves protein separation efficiency during PAGE as a result of increased local protein concentration. FIGE can be implemented with minimal additional instrumentation in any laboratory setting. Despite the tradeoff of longer running times, FIGE can be a powerful protein separation tool.

Direct influence of culture dimensionality on human mesenchymal stem cell differentiation at various matrix stiffnesses using a fibrous self-assembling peptide hydrogel.

PubMed

Hogrebe, Nathaniel J; Gooch, Keith J

2016-09-01

Much is unknown about the effects of culture dimensionality on cell behavior due to the lack of biomimetic substrates that are suitable for directly comparing cells grown on two-dimensional (2D) and encapsulated within three-dimensional (3D) matrices of the same stiffness and biochemistry. To overcome this limitation, we used a self-assembling peptide hydrogel system that has tunable stiffness and cell-binding site density as well as a fibrous microarchitecture resembling the structure of collagen. We investigated the effect of culture dimensionality on human mesenchymal stem cell differentiation at different values of matrix stiffness (G' = 0.25, 1.25, 5, and 10 kPa) and a constant RGD (Arg-Gly-Asp) binding site concentration. In the presence of the same soluble induction factors, culture on top of stiff gels facilitated the most efficient osteogenesis, while encapsulation within the same stiff gels resulted in a switch to predominantly terminal chondrogenesis. Adipogenesis dominated at soft conditions, and 3D culture induced better adipogenic differentiation than 2D culture at a given stiffness. Interestingly, initial matrix-induced cell morphology was predictive of these end phenotypes. Furthermore, optimal culture conditions corresponded to each cell type's natural niche within the body, highlighting the importance of incorporating native matrix dimensionality and stiffness into tissue engineering strategies. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2356-2368, 2016. © 2016 Wiley Periodicals, Inc.

Dosimetry study of diagnostic X-ray using doped iodide normoxic polymer gels

NASA Astrophysics Data System (ADS)

Huang, Y. R.; Chang, Y. J.; Hsieh, L. L.; Liu, M. H.; Liu, J. S.; Chu, C. H.; Hsieh, B. T.

2014-11-01

In radiotherapy, polymer gel dosimeters are used for three-dimensional (3D) dose distribution. However, the doses are within the Gy range. In this study, we attempted to develop a low-dose 3D dosimeter within the mGy range for diagnostic radiology. The effect of the iodinated compound was used as a dose enhancement sensitizer to enhance the dose sensitivity of normoxic polymer gel dosimeters. This study aims to use N-isopropylacrylamide(NIPAM)-based and methacrylic acid (MAGAT)-based gels to evaluate the potential dose enhancement sensitizer, as well as to compare two gels that may be suitable for measuring diagnostic radiation doses. The suitable formulation of NIPAM gel [5% (w/w) gelatin, 5% (w/w) NIPAM, 3% (w/w) N,N‧-methylenebisacrylamide (BIS), 5 mM tetrakis (hydroxymethyl) phosphonium chloride (THPC), and 87% (w/w) deionized distilled water] and MAGAT gel (4% MAA, 9% gelatin, 87% deionized water, and 10 mM THPC) were used and loaded with clinical iodinated contrast medium agent (Iobitridol, Xenetix® 350). Irradiation was conducted using X-ray computed tomography. The irradiation doses ranged from 0 mGy to 80 mGy. Optical computed tomography was the employed gel measurement system. The results indicate that the iodinated contrast agent yields a quantifiable dose enhancement ratio. The dose enhancement ratios of NIPAM and MAGAT gels are 3.35±0.6 and 1.36±0.3, respectively. The developed NIPAM gel in this study could be suitable for measuring diagnostic radiation doses.

Determining Degradation and Synthesis Rates of Arabidopsis Proteins Using the Kinetics of Progressive 15N Labeling of Two-dimensional Gel-separated Protein Spots*

PubMed Central

Li, Lei; Nelson, Clark J.; Solheim, Cory; Whelan, James; Millar, A. Harvey

2012-01-01

The growth and development of plant tissues is associated with an ordered succession of cellular processes that are reflected in the appearance and disappearance of proteins. The control of the kinetics of protein turnover is central to how plants can rapidly and specifically alter protein abundance and thus molecular function in response to environmental or developmental cues. However, the processes of turnover are largely hidden during periods of apparent steady-state protein abundance, and even when proteins accumulate it is unclear whether enhanced synthesis or decreased degradation is responsible. We have used a 15N labeling strategy with inorganic nitrogen sources coupled to a two-dimensional fluorescence difference gel electrophoresis and mass spectrometry analysis of two-dimensional IEF/SDS-PAGE gel spots to define the rate of protein synthesis (KS) and degradation (KD) of Arabidopsis cell culture proteins. Through analysis of MALDI-TOF/TOF mass spectra from 120 protein spots, we were able to quantify KS and KD for 84 proteins across six functional groups and observe over 65-fold variation in protein degradation rates. KS and KD correlate with functional roles of the proteins in the cell and the time in the cell culture cycle. This approach is based on progressive 15N labeling that is innocuous for the plant cells and, because it can be used to target analysis of proteins through the use of specific gel spots, it has broad applicability. PMID:22215636

Two-dimensional porous anodic alumina for optoelectronics and photocatalytic application

NASA Astrophysics Data System (ADS)

Khoroshko, L. S.

2015-11-01

Fabrication of porous anodic alumina film structures using anodizing, sol-gel synthesis and photolithography is reported. The structures receive interest as planar waveguides due to strong photoluminescence of the embedded trivalent lanthanides. Mesoporous structures comprising sol-gel derived titania in porous anodic alumina play a role of effective catalyst for water purification.

Simple and Rapid System for Two-Dimensional Gel Electrophoresis Technique: A Laboratory Exercise for High School Students

ERIC Educational Resources Information Center

Maurye, Praveen; Basu, Arpita; Biswas, Jayanta Kumar; Bandyopadhyay, Tapas Kumar; Naskar, Malay

2018-01-01

Polyacrylamide gel electrophoresis (PAGE) is the most classical technique favored worldwide for resolution of macromolecules in many biochemistry laboratories due to its incessant advanced developments and wide modifications. These ever-growing advancements in the basic laboratory equipments lead to emergence of many expensive, complex, and tricky…

Modification of vital wheat gluten with phosphoric acid to produce high free solution capacity

USDA-ARS?s Scientific Manuscript database

Wheat gluten reacts with phosphoric acid in the presence of urea to produce natural superabsorbent gels. Fourier Transform Infra-red (FT-IR) spectroscopy and two-dimensional gel electrophoresis (2DE) reveal chemical changes from the reaction. Temperatures above 120°C and dry conditions create the op...

Application of a label-free, gel-free quantitative proteomics method for ecotoxicological studies of small fish species

EPA Science Inventory

Although two-dimensional electrophoresis (2D-GE) remains the basis for many ecotoxicoproteomic analyses, new, non gel-based methods are beginning to be applied to overcome throughput and coverage limitations of 2D-GE. The overall objective of our research was to apply a comprehe...

Gelling Properties of Fish/Pork Mince Mixtures.

PubMed

Liu, Ru; Zhao, Siming; Regenstein, Joe M; Liu, Qing; Yang, Hong; Xiong, Shanbai

2016-02-01

The gel properties of silver carp/pork mince mixtures were investigated as well as the protein structural changes and interactions during gelling using rheology, SEM, and FT-Raman spectroscopy. The breaking force values for gels containing 0% to 40% pork was significantly lower (P < 0.05) compared with gels containing 50% to 100% pork. Gels containing 70% to 100% pork had significantly higher (P < 0.05) breaking force values compared with gels containing 50% to 60% pork. Deformation values were more mixed. Dynamic rheological data suggested that mixing fish and pork at 3:7 could strengthen the gel network. The addition of 40% pork or above, significantly decreased (P < 0.05) the water retention of the gels compared with the 100% fish gels. The dimensional ordering of gels was also reduced by addition of pork. The reduced ordering was one of the reasons for the low water retention for fish/pork mixed gels. Raman spectral analysis confirmed that mixing fish and pork in 7:3 and 3:7 ratios could promote hydrophobic interactions such as bringing tyrosine residues into the intermolecular interface. The interactions in the 3:7 fish/pork mixed gels were favorable for forming a stronger gel. However, the interactions in the 7:3 fish/pork mixed gels were adverse. The water retention of gels was related to both molecular interactions and secondary structures of protein as well as the microstructure of the gels. © 2016 Institute of Food Technologists®

Analysis of bacterial populations in the environment using two-dimensional gel electrophoresis of genomic DNA and complementary DNA.

PubMed

Liu, Guo-Hua; Nakamura, Tatsuo; Amemiya, Takashi; Rajendran, Narasimmalu; Itoh, Kiminori

2011-01-01

Two-dimensional gel electrophoresis (2-DGE) mapping of genomic DNA and complementary DNA (cDNA) amplicons was attempted to analyze total and active bacterial populations within soil and activated sludge samples. Distinct differences in the number and species of bacterial populations and those that were metabolically active at the time of sampling were visually observed especially for the soil community. Statistical analyses and sequencing based on the 2-DGE data further revealed the relationships between total and active bacterial populations within each community. This high-resolution technique would be useful for obtaining a better understanding of bacterial population structures in the environment.

Changes induced by the Pepper mild mottle tobamovirus on the chloroplast proteome of Nicotiana benthamiana.

PubMed

Pineda, M; Sajnani, C; Barón, M

2010-01-01

We have analyzed the chloroplast proteome of Nicotiana benthamiana using two-dimensional gel electrophoresis and mass spectrometry followed by a database search. In order to improve the resolution of the two-dimensional electrophoresis gels, we have made separate maps for the low and the high pH range. At least 200 spots were detected. We identified 72 polypeptides, some being isoforms of different multiprotein families. In addition, changes in this chloroplast proteome induced by the infection with the Spanish strain of the Pepper mild mottle virus were investigated. Viral infection induced the down-regulation of several chloroplastidic proteins involved in both the photosynthetic electron-transport chain and the Benson-Calvin cycle.

Performance evaluation of an improved optical computed tomography polymer gel dosimeter system for 3D dose verification of static and dynamic phantom deliveries

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lopatiuk-Tirpak, O.; Langen, K. M.; Meeks, S. L.

2008-09-15

The performance of a next-generation optical computed tomography scanner (OCTOPUS-5X) is characterized in the context of three-dimensional gel dosimetry. Large-volume (2.2 L), muscle-equivalent, radiation-sensitive polymer gel dosimeters (BANG-3) were used. Improvements in scanner design leading to shorter acquisition times are discussed. The spatial resolution, detectable absorbance range, and reproducibility are assessed. An efficient method for calibrating gel dosimeters using the depth-dose relationship is applied, with photon- and electron-based deliveries yielding equivalent results. A procedure involving a preirradiation scan was used to reduce the edge artifacts in reconstructed images, thereby increasing the useful cross-sectional area of the dosimeter by nearly amore » factor of 2. Dose distributions derived from optical density measurements using the calibration coefficient show good agreement with the treatment planning system simulations and radiographic film measurements. The feasibility of use for motion (four-dimensional) dosimetry is demonstrated on an example comparing dose distributions from static and dynamic delivery of a single-field photon plan. The capability to visualize three-dimensional dose distributions is also illustrated.« less

A Chip-Capillary Hybrid Device for Automated Transfer of Sample Pre-Separated by Capillary Isoelectric Focusing to Parallel Capillary Gel Electrophoresis for Two-Dimensional Protein Separation

PubMed Central

Lu, Joann J.; Wang, Shili; Li, Guanbin; Wang, Wei; Pu, Qiaosheng; Liu, Shaorong

2012-01-01

In this report, we introduce a chip-capillary hybrid device to integrate capillary isoelectric focusing (CIEF) with parallel capillary sodium dodecyl sulfate – polyacrylamide gel electrophoresis (SDS-PAGE) or capillary gel electrophoresis (CGE) toward automating two-dimensional (2D) protein separations. The hybrid device consists of three chips that are butted together. The middle chip can be moved between two positions to re-route the fluidic paths, which enables the performance of CIEF and injection of proteins partially resolved by CIEF to CGE capillaries for parallel CGE separations in a continuous and automated fashion. Capillaries are attached to the other two chips to facilitate CIEF and CGE separations and to extend the effective lengths of CGE columns. Specifically, we illustrate the working principle of the hybrid device, develop protocols for producing and preparing the hybrid device, and demonstrate the feasibility of using this hybrid device for automated injection of CIEF-separated sample to parallel CGE for 2D protein separations. Potentials and problems associated with the hybrid device are also discussed. PMID:22830584

Design and Construction of an Optical Computed Tomography Scanner for Polymer Gel Dosimetry Application

PubMed Central

Zakariaee, Seyed Salman; Mesbahi, Asghar; Keshtkar, Ahmad; Azimirad, Vahid

2014-01-01

Polymer gel dosimeter is the only accurate three dimensional (3D) dosimeter that can measure the absorbed dose distribution in a perfect 3D setting. Gel dosimetry by using optical computed tomography (OCT) has been promoted by several researches. In the current study, we designed and constructed a prototype OCT system for gel dosimetry. First, the electrical system for optical scanning of the gel container using a Helium-Neon laser and a photocell was designed and constructed. Then, the mechanical part for two rotational and translational motions was designed and step motors were assembled to it. The data coming from photocell was grabbed by the home-built interface and sent to a personal computer. Data processing was carried out using MATLAB software. To calibrate the system and tune up the functionality of it, different objects was designed and scanned. Furthermore, the spatial and contrast resolution of the system was determined. The system was able to scan the gel dosimeter container with a diameter up to 11 cm inside the water phantom. The standard deviation of the pixels within water flask image was considered as the criteria for image uniformity. The uniformity of the system was about ±0.05%. The spatial resolution of the system was approximately 1 mm and contrast resolution was about 0.2%. Our primary results showed that this system is able to obtain two-dimensional, cross-sectional images from polymer gel samples. PMID:24761377

Comparative study between prokaryotes and eukaryotes by chemical iodination of ribosomal proteins.

PubMed

Bernabeu, C; Vázquez, D; Conde, F P

1979-04-25

Escherichia coli and Saccharomyces cerevisiae ribosomal proteins were chemically iodinated with 125I by chloramine T under conditions in which the proteins were denatured. The labelled proteins were subsequently separated by two-dimensional gel electrophoresis with an excess of untreated ribosomal proteins from the same species. The iodination did not change the electrophoretic mobility of the proteins as shown by the pattern of spots in the stained gel slabs and their autoradiography. The 125I radioactivity incorporated in the proteins was estimated by cutting out the gel spots from the two-dimensional electrophoresis gel slabs. The highest content of 125I was found in the ribosomal proteins L2, L11, L13, L20/S12, S4 and S9 from E. coli, and L2/L3, L4/L6/S7, L5, L19/L20, L22/S17, L29/S27, L35/L37 and S14/S15 from S. cerevisiae. Comparisons between the electrophoretic patterns of E. coli and S. cerevisiae ribosomal proteins were carried out by coelectrophoresis of labelled and unlabelled proteins from both species. E. coli ribosomal proteins L5, L11, L20, S2, S3 and S15/S16 were found to overlap with L15, L11/L16, L36/L37, S3, S10 and S33 from S. cerevisiae, respectively. Similar coelectrophoresis of E. coli 125I-labelled proteins with unlabelled rat liver and wheat germ ribosomal proteins showed the former to overlap with proteins L1, L11, L14, L16, L19, L20 and the latter with L2, L5, L6, L15, L17 from E. coli.

Chemistry and Processing of Nanostructured Materials

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fox, G A; Baumann, T F; Hope-Weeks, L J

2002-01-18

Nanostructured materials can be formed through the sol-gel polymerization of inorganic or organic monomer systems. For example, a two step polymerization of tetramethoxysilane (TMOS) was developed such that silica aerogels with densities as low as 3 kg/m{sup 3} ({approx} two times the density of air) could be achieved. Organic aerogels based upon resorcinol-formaldehyde and melamine-formaldehyde can also be prepared using the sol-gel process. Materials of this type have received significant attention at LLNL due to their ultrafine cell sizes, continuous porosity, high surface area and low mass density. For both types of aerogels, sol-gel polymerization depends upon the transformation ofmore » these monomers into nanometer-sized clusters followed by cross-linking into a 3-dimensional gel network. While sol-gel chemistry provides the opportunity to synthesize new material compositions, it suffers from the inability to separate the process of cluster formation from gelation. This limitation results in structural deficiencies in the gel that impact the physical properties of the aerogel, xerogel or nanocomposite. In order to control the properties of the resultant gel, one should be able to regulate the formation of the clusters and their subsequent cross-linking. Towards this goal, we are utilizing dendrimer chemistry to separate the cluster formation from the gelation so that new nanostructured materials can be produced. Dendrimers are three-dimensional, highly branched macromolecules that are prepared in such a way that their size, shape and surface functionality are readily controlled. The dendrimers will be used as pre-formed clusters of known size that can be cross-linked to form an ordered gel network.« less

Polyimide Aerogels with Three-Dimensional Cross-Linked Structure

NASA Technical Reports Server (NTRS)

Meador, Mary Ann B. (Inventor)

2016-01-01

A method for creating a three dimensional cross-linked polyimide structure includes dissolving a diamine, a dianhydride, and a triamine in a solvent, imidizing a polyamic acid gel by heating the gel, extracting the gel in a second solvent, supercritically drying the gel, and removing the solvent to create a polyimide aerogel.

Proteomics Analysis of the Nucleolus in Adenovirus-infected Cells

PubMed Central

Lam, Yun W.; Evans, Vanessa C.; Heesom, Kate J.; Lamond, Angus I.; Matthews, David A.

2010-01-01

Adenoviruses replicate primarily in the host cell nucleus, and it is well established that adenovirus infection affects the structure and function of host cell nucleoli in addition to coding for a number of nucleolar targeted viral proteins. Here we used unbiased proteomics methods, including high throughput mass spectrometry coupled with stable isotope labeling by amino acids in cell culture (SILAC) and traditional two-dimensional gel electrophoresis, to identify quantitative changes in the protein composition of the nucleolus during adenovirus infection. Two-dimensional gel analysis revealed changes in six proteins. By contrast, SILAC-based approaches identified 351 proteins with 24 proteins showing at least a 2-fold change after infection. Of those, four were previously reported to have aberrant localization and/or functional relevance during adenovirus infection. In total, 15 proteins identified as changing in amount by proteomics methods were examined in infected cells using confocal microscopy. Eleven of these proteins showed altered patterns of localization in adenovirus-infected cells. Comparing our data with the effects of actinomycin D on the nucleolar proteome revealed that adenovirus infection apparently specifically targets a relatively small subset of nucleolar antigens at the time point examined. PMID:19812395

Proteomics analysis of the nucleolus in adenovirus-infected cells.

PubMed

Lam, Yun W; Evans, Vanessa C; Heesom, Kate J; Lamond, Angus I; Matthews, David A

2010-01-01

Adenoviruses replicate primarily in the host cell nucleus, and it is well established that adenovirus infection affects the structure and function of host cell nucleoli in addition to coding for a number of nucleolar targeted viral proteins. Here we used unbiased proteomics methods, including high throughput mass spectrometry coupled with stable isotope labeling by amino acids in cell culture (SILAC) and traditional two-dimensional gel electrophoresis, to identify quantitative changes in the protein composition of the nucleolus during adenovirus infection. Two-dimensional gel analysis revealed changes in six proteins. By contrast, SILAC-based approaches identified 351 proteins with 24 proteins showing at least a 2-fold change after infection. Of those, four were previously reported to have aberrant localization and/or functional relevance during adenovirus infection. In total, 15 proteins identified as changing in amount by proteomics methods were examined in infected cells using confocal microscopy. Eleven of these proteins showed altered patterns of localization in adenovirus-infected cells. Comparing our data with the effects of actinomycin D on the nucleolar proteome revealed that adenovirus infection apparently specifically targets a relatively small subset of nucleolar antigens at the time point examined.

Two-Dimensional Gel Electrophoresis: Discovering Biomolecules for Environmental Bioremediation

NASA Astrophysics Data System (ADS)

Singh, Om V.; Chandel, Anuj K.

Environmental contamination has been viewed as an ecological malaise for which bioremediation can be prescribed as a “perfect medicine.” The solution to the problems with bioremediation lies in analyzing to what extent the microbes’ physiological machinery contributes to the degradation process and which biomolecules and their mechanisms are responsible for regulatory factors within the degradation system, such as protein, metabolite, and enzymatic chemical transformation. In the post-genomic era, recent advances in proteomics have allowed us to elucidate many complex biological mechanisms. Two-dimensional gel electrophoresis (2DE) in conjunction with mass spectrometry (MS) can be utilized to identify the biomolecules and their molecular mechanisms in bioremediation. A set of highly abundant global proteins over a pI range 4-7 was separated and compared by size fractionation (25-100 kDa) on 2DE. We identified a set of catabolic proteins, enzymes, and heat shock molecular chaperones associated with the regulatory network that was found to be overexpressed under phenol-stressed conditions. This chapter also offers optimized ideal directions for 2DE, followed by easy-to-follow directions for a protein identification strategy using MALDI-TOF and targeting novel proteins/enzymes for a universal set of experiments.

Stabilization of the Inverse Laplace Transform of Multiexponential Decay through Introduction of a Second Dimension

PubMed Central

Celik, Hasan; Bouhrara, Mustapha; Reiter, David A.; Fishbein, Kenneth W.; Spencer, Richard G.

2013-01-01

We propose a new approach to stabilizing the inverse Laplace transform of a multiexponential decay signal, a classically ill-posed problem, in the context of nuclear magnetic resonance relaxometry. The method is based on extension to a second, indirectly detected, dimension, that is, use of the established framework of two-dimensional relaxometry, followed by projection onto the desired axis. Numerical results for signals comprised of discrete T1 and T2 relaxation components and experiments performed on agarose gel phantoms are presented. We find markedly improved accuracy, and stability with respect to noise, as well as insensitivity to regularization in quantifying underlying relaxation components through use of the two-dimensional as compared to the one-dimensional inverse Laplace transform. This improvement is demonstrated separately for two different inversion algorithms, nonnegative least squares and non-linear least squares, to indicate the generalizability of this approach. These results may have wide applicability in approaches to the Fredholm integral equation of the first kind. PMID:24035004

Deduction of two-dimensional blood flow vector by dual angle diverging waves from a cardiac sector probe

NASA Astrophysics Data System (ADS)

Maeda, Moe; Nagaoka, Ryo; Ikeda, Hayato; Yaegashi, So; Saijo, Yoshifumi

2018-07-01

Color Doppler method is widely used for noninvasive diagnosis of heart diseases. However, the method can measure one-dimensional (1D) blood flow velocity only along an ultrasonic beam. In this study, diverging waves with two different angles were irradiated from a cardiac sector probe to estimate a two-dimensional (2D) blood flow vector from each velocity measured with the angles. The feasibility of the proposed method was evaluated in experiments using flow poly(vinyl alcohol) (PVA) gel phantoms. The 2D velocity vectors obtained with the proposed method were compared with the flow vectors obtained with the particle image velocimetry (PIV) method. Root mean square errors of the axial and lateral components were 11.3 and 29.5 mm/s, respectively. The proposed method was also applied to echo data from the left ventricle of the heart. The inflow from the mitral valve in diastole and the ejection flow concentrating in the aorta in systole were visualized.

A theoretical evaluation of aluminum gel propellant two-phase flow losses on vehicle performance

NASA Technical Reports Server (NTRS)

Mueller, Donn C.; Turns, Stephen R.

1993-01-01

A one-dimensional model of a hydrocarbon/Al/O2(gaseous) fueled rocket combustion chamber was developed to study secondary atomization effects on propellant combustion. This chamber model was coupled with a two dimensional, two-phase flow nozzle code to estimate the two-phase flow losses associated with solid combustion products. Results indicate that moderate secondary atomization significantly reduces propellant burnout distance and Al2O3 particle size; however, secondary atomization provides only moderate decreases in two-phase flow induced I(sub sp) losses. Despite these two-phase flow losses, a simple mission study indicates that aluminum gel propellants may permit a greater maximum payload than the hydrocarbon/O2 bi-propellant combination for a vehicle of fixed propellant volume. Secondary atomization was also found to reduce radiation losses from the solid combustion products to the chamber walls, primarily through reductions in propellant burnout distance.

A Theoretical Evaluation of Secondary Atomization Effects on Engine Performance for Aluminum Gel Propellants

NASA Technical Reports Server (NTRS)

Mueller, D. C.; Turns, S. R.

1994-01-01

A one-dimensional model of a gel-fueled rocket combustion chamber has been developed. This model includes the processes of liquid hydrocarbon burnout, secondary atomization. aluminum ignition, and aluminum combustion. Also included is a model of radiative heat transfer from the solid combustion products to the chamber walls. Calculations indicate that only modest secondary atomization is required to significantly reduce propellant burnout distances, aluminum oxide residual size and radiation heat wall losses. Radiation losses equal to approximately 2-13 percent of the energy released during combustion were estimated. A two-dimensional, two-phase nozzle code was employed to estimate radiation and nozzle two-phase flow effects on overall engine performance. Radiation losses yielded a 1 percent decrease in engine I(sub sp). Results also indicate that secondary atomization may have less effect on two-phase losses than it does on propellant burnout distance and no effect if oxide particle coagulation and shear induced droplet breakup govern oxide particle size. Engine I(sub sp) was found to decrease from 337.4 to 293.7 seconds as gel aluminum mass loading was varied from 0-70 wt percent. Engine I(sub sp) efficiencies, accounting for radiation and two-phase flow effects, on the order of 0.946 were calculated for a 60 wt percent gel, assuming a fragmentation ratio of 5.

Apparatus for reading two-dimensional electrophoretograms containing. beta. -ray-emitting labeled compounds

DOEpatents

Anderson, H.L.; Kinnison, W.W.; Lillberg, J.W.

1985-04-30

An apparatus and method for electronically reading planar two-dimensional ..beta..-ray emitter-labeled gel electrophoretograms. A single, flat rectangular multiwire proportional chamber is placed in close proximity to the gel and the assembly placed in an intense uniform magnetic field disposed in a perpendicular manner to the rectangular face of the proportional chamber. Beta rays emitted in the direction of the proportional chamber are caused to execute helical motions which substantially preserve knowledge the coordinates of their origin in the gel. Perpendicularly oriented, parallel wire, parallel plane cathodes electronically sense the location of the ..beta..-rays from ionization generated thereby in a detection gas coupled with an electron avalanche effect resulting from the action of a parallel wire anode located therebetween. A scintillator permits the present apparatus to be rendered insensitive when signals are generated from cosmic rays incident on the proportional chamber. Resolution for concentrations of radioactive compounds in the gel exceeds 700-..mu..m. The apparatus and method of the present invention represent a significant improvement over conventional autoradiographic techniques in dynamic range, linearity and sensitivity of data collection. A concentration and position map for gel electrophoretograms having significant concentrations of labeled compounds and/or highly radioactive labeling nuclides can generally be obtained in less than one hour.

Apparatus and method for reading two-dimensional electrophoretograms containing .beta.-ray-emitting labeled compounds

DOEpatents

Anderson, Herbert L.; Kinnison, W. Wayne; Lillberg, John W.

1987-01-01

Apparatus and method for electronically reading planar two dimensional .beta.-ray emitter-labeled gel electrophoretograms. A single, flat rectangular multiwire proportional chamber is placed in close proximity to the gel and the assembly placed in an intense uniform magnetic field disposed in a perpendicular manner to the rectangular face of the proportional chamber. Beta rays emitted in the direction of the proportional chamber are caused to execute helical motions which substantially preserve knowledge of the coordinates of their origin in the gel. Perpendicularly oriented, parallel wire, parallel plane cathodes electronically sense the location of the .beta.-rays from ionization generated thereby in a detection gas coupled with an electron avalanche effect resulting from the action of a parallel wire anode located therebetween. A scintillator permits the present apparatus to be rendered insensitive when signals are generated from cosmic rays incident on the proportional chamber. Resolution for concentrations of radioactive compounds in the gel exceeds 700 .mu.m. The apparatus and method of the present invention represent a significant improvement over conventional autoradiographic techniques in dynamic range, linearity and sensitivity of data collection. A concentration and position map for gel electrophoretograms having significant concentrations of labeled compounds and/or highly radioactive labeling nuclides can generally be obtained in less than one hour.

Biochemical analysis of secretory proteins synthesized by normal rat pancreas and by pancreatic acinar tumor cells

PubMed Central

1982-01-01

We have examined the secretogogue responsiveness and the pattern of secretory proteins produced by a transplantable rat pancreatic acinar cell tumor. Dispersed tumor cells were found to discharge secretory proteins in vitro when incubated with hormones that act on four different classes of receptors: carbamylcholine, caerulein, secretin- vasoactive intestinal peptide, and bombesin. With all hormones tested, maximal discharge from tumor cells was only about one-half that of control pancreatic lobules, but occurred at the same dose optima except for secretin, whose dose optimum was 10-fold higher. Biochemical analysis of secretory proteins discharged by the tumor cells was carried out by crossed immunoelectrophoresis and by two-dimensional isoelectric focusing-SDS polyacrylamide gel electrophoresis. To establish a baseline for comparison, secretory proteins from normal rat pancreas were identified according to enzymatic activity and correlated with migration position on two-dimensional gels. Our results indicate that a group of basic polypeptides including proelastase, basic trypsinogen, basic chymotrypsinogen, and ribonuclease, two out of three forms of procarboxypeptidase B, and the major lipase species were greatly reduced or absent in tumor cell secretion. In contrast, the amount of acidic chymotrypsinogen was notably increased compared with normal acinar cells. Although the acinar tumor cells are highly differentiated cytologically and express functional receptors for several classes of pancreatic secretagogues, they show quantitative and qualitative differences when compared with normal pancreas with regard to their production of secretory proteins. PMID:6185502

Calibration of mass spectrometric peptide mass fingerprint data without specific external or internal calibrants

PubMed Central

Wolski, Witold E; Lalowski, Maciej; Jungblut, Peter; Reinert, Knut

2005-01-01

Background Peptide Mass Fingerprinting (PMF) is a widely used mass spectrometry (MS) method of analysis of proteins and peptides. It relies on the comparison between experimentally determined and theoretical mass spectra. The PMF process requires calibration, usually performed with external or internal calibrants of known molecular masses. Results We have introduced two novel MS calibration methods. The first method utilises the local similarity of peptide maps generated after separation of complex protein samples by two-dimensional gel electrophoresis. It computes a multiple peak-list alignment of the data set using a modified Minimum Spanning Tree (MST) algorithm. The second method exploits the idea that hundreds of MS samples are measured in parallel on one sample support. It improves the calibration coefficients by applying a two-dimensional Thin Plate Splines (TPS) smoothing algorithm. We studied the novel calibration methods utilising data generated by three different MALDI-TOF-MS instruments. We demonstrate that a PMF data set can be calibrated without resorting to external or relying on widely occurring internal calibrants. The methods developed here were implemented in R and are part of the BioConductor package mscalib available from . Conclusion The MST calibration algorithm is well suited to calibrate MS spectra of protein samples resulting from two-dimensional gel electrophoretic separation. The TPS based calibration algorithm might be used to correct systematic mass measurement errors observed for large MS sample supports. As compared to other methods, our combined MS spectra calibration strategy increases the peptide/protein identification rate by an additional 5 – 15%. PMID:16102175

Unravelling the differences: comparative proteomic analysis of a clonal virulent and an attenuated Histomonas meleagridis strain.

PubMed

Monoyios, Andreas; Patzl, Martina; Schlosser, Sarah; Hess, Michael; Bilic, Ivana

2018-02-01

The current study focused on Histomonas meleagridis, a unicellular protozoan, responsible for histomonosis in poultry. Recently, the occurrence of the disease increased due to the ban of effective chemotherapeutic drugs. Basic questions regarding the molecular biology, virulence mechanisms or even life cycle of the flagellate are still puzzling. In order to address some of these issues, we conducted a comparative proteomic analysis of a virulent and an attenuated H. meleagridis strain traced back to a single cell and propagated in vitro as monoxenic mono-eukaryotic cultures. Using two-dimensional electrophoresis (2-DE) for proteome visualization with computational 2-DE gel image and statistical analysis, upregulated proteins in either of the two H. meleagridis strains were detected. Statistical analysis fulfilling two criteria (≥threefold upregulation and P < 0.05) revealed 119 differentially expressed protein spots out of which 62 spots were noticed in gels with proteins from the virulent and 57 spots in gels with proteins from the attenuated culture. Mass spectrometric analysis of 32 protein spots upregulated in gels of the virulent strain identified 17 as H. meleagridis-specific. The identification revealed that these spots belonged to eight different proteins, with the majority related to cellular stress management. Two ubiquitous cellular proteins, actin and enolase, were upregulated in multiple gel positions in this strain, indicating either post-translational modification or truncation, or even both. Additionally, a known virulence factor named legumain cysteine peptidase was also detected. In contrast to this, mass spectrometric analysis of 49 protein spots, upregulated in gels of the attenuated strain, singled out 32 spots as specific for the flagellate. These spots were shown to correspond to 24 different proteins that reflect the increased metabolism, in vitro adaptation of the parasite, and amoeboid morphology. In addition to H. meleagridis proteins, the analysis identified differential expression of Escherichia coli DH5α proteins that could have been influenced by the co-cultivated H. meleagridis strain, indicating a reciprocal interaction of these two organisms during monoxenic cultivation. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Photonic devices based on patterning by two photon induced polymerization techniques

NASA Astrophysics Data System (ADS)

Fortunati, I.; Dainese, T.; Signorini, R.; Bozio, R.; Tagliazucca, V.; Dirè, S.; Lemercier, G.; Mulatier, J.-C.; Andraud, C.; Schiavuta, P.; Rinaldi, A.; Licoccia, S.; Bottazzo, J.; Franco Perez, A.; Guglielmi, M.; Brusatin, G.

2008-04-01

Two and three dimensional structures with micron and submicron resolution have been achieved in commercial resists, polymeric materials and sol-gel materials by several lithographic techniques. In this context, silicon-based sol-gel materials are particularly interesting because of their versatility, chemical and thermal stability, amount of embeddable active compounds. Compared with other micro- and nano-fabrication schemes, the Two Photon Induced Polymerization is unique in its 3D processing capability. The photopolymerization is performed with laser beam in the near-IR region, where samples show less absorption and less scattering, giving rise to a deeper penetration of the light. The use of ultrashort laser pulses allows the starting of nonlinear processes like multiphoton absorption at relatively low average power without thermally damaging the samples. In this work we report results on the photopolymerization process in hybrid organic-inorganic films based photopolymerizable methacrylate-containing Si-nanobuilding blocks. Films, obtained through sol-gel synthesis, are doped with a photo-initiator allowing a radical polymerization of methacrylic groups. The photo-initiator is activated by femtosecond laser source, at different input energies. The development of the unexposed regions is performed with a suitable solvent and the photopolymerized structures are characterized by microscopy techniques.

NopB, a type III secreted protein of Rhizobium sp. strain NGR234, is associated with pilus-like surface appendages.

PubMed

Saad, Maged M; Kobayashi, Hajime; Marie, Corinne; Brown, Ian R; Mansfield, John W; Broughton, William J; Deakin, William J

2005-02-01

Rhizobium sp. strain NGR234 possesses a functional type three secretion system (TTSS), through which a number of proteins, called nodulation outer proteins (Nops), are delivered to the outside of the cell. A major constraint to the identification of Nops is their low abundance in the supernatants of NGR234 strains grown in culture. To overcome this limitation, a more sensitive proteomics-based strategy was developed. Secreted proteins from wild-type NGR234 were separated by two-dimensional gel electrophoresis, and the gel was compared to similar gels containing the proteins from a TTSS mutant (NGROmegarhcN). To identify the proteins, spots unique to the NGR234 gels were analyzed by matrix-assisted laser desorption ionization-time of flight mass spectrometry and the data were compared to the sequence of the symbiotic plasmid of NGR234. A nonpolar mutant of one of these proteins was generated called NopB. NopB is required for Nop secretion but inhibits the interaction with Pachyrhizus tuberosus and augments nodulation of Tephrosia vogelii. Flavonoids and a functional TTSS are required for the formation of some surface appendages on NGR234. In situ immunogold labeling and isolation of these pili showed that they contain NopB.

NopB, a Type III Secreted Protein of Rhizobium sp. Strain NGR234, Is Associated with Pilus-Like Surface Appendages

PubMed Central

Saad, Maged M.; Kobayashi, Hajime; Marie, Corinne; Brown, Ian R.; Mansfield, John W.; Broughton, William J.; Deakin, William J.

2005-01-01

Rhizobium sp. strain NGR234 possesses a functional type three secretion system (TTSS), through which a number of proteins, called nodulation outer proteins (Nops), are delivered to the outside of the cell. A major constraint to the identification of Nops is their low abundance in the supernatants of NGR234 strains grown in culture. To overcome this limitation, a more sensitive proteomics-based strategy was developed. Secreted proteins from wild-type NGR234 were separated by two-dimensional gel electrophoresis, and the gel was compared to similar gels containing the proteins from a TTSS mutant (NGRΩrhcN). To identify the proteins, spots unique to the NGR234 gels were analyzed by matrix-assisted laser desorption ionization-time of flight mass spectrometry and the data were compared to the sequence of the symbiotic plasmid of NGR234. A nonpolar mutant of one of these proteins was generated called NopB. NopB is required for Nop secretion but inhibits the interaction with Pachyrhizus tuberosus and augments nodulation of Tephrosia vogelii. Flavonoids and a functional TTSS are required for the formation of some surface appendages on NGR234. In situ immunogold labeling and isolation of these pili showed that they contain NopB. PMID:15659692

Development of a highly sensitive three-dimensional gel electrophoresis method for characterization of monoclonal protein heterogeneity.

PubMed

Nakano, Keiichi; Tamura, Shogo; Otuka, Kohei; Niizeki, Noriyasu; Shigemura, Masahiko; Shimizu, Chikara; Matsuno, Kazuhiko; Kobayashi, Seiichi; Moriyama, Takanori

2013-07-15

Three-dimensional gel electrophoresis (3-DE), which combines agarose gel electrophoresis and isoelectric focusing/SDS-PAGE, was developed to characterize monoclonal proteins (M-proteins). However, the original 3-DE method has not been optimized and its specificity has not been demonstrated. The main goal of this study was to optimize the 3-DE procedure and then compare it with 2-DE. We developed a highly sensitive 3-DE method in which M-proteins are extracted from a first-dimension agarose gel, by diffusing into 150 mM NaCl, and the recovery of M-proteins was 90.6%. To validate the utility of the highly sensitive 3-DE, we compared it with the original 3-DE method. We found that highly sensitive 3-DE provided for greater M-protein recovery and was more effective in terms of detecting spots on SDS-PAGE gels than the original 3-DE. Moreover, highly sensitive 3-DE separates residual normal IgG from M-proteins, which could not be done by 2-DE. Applying the highly sensitive 3-DE to clinical samples, we found that the characteristics of M-proteins vary tremendously between individuals. We believe that our highly sensitive 3-DE method described here will prove useful in further studies of the heterogeneity of M-proteins. Copyright © 2013 Elsevier Inc. All rights reserved.

TM4SF5 promotes metastatic behavior of cells in 3D extracellular matrix gels by reducing dependency on environmental cues

PubMed Central

Nam, Seo Hee; Cheong, Jin-Gyu; Jeong, Doyoung; Lee, Seo-Jin; Pan, Cheol-Ho; Jung, Jae Woo; Kim, Hye-Jin; Ryu, Jihye; Kim, Ji Eon; Kim, Somi; Cho, Chang Yun; Kang, Min-Kyung; Lee, Kyung-Min; Lee, Jung Weon

2017-01-01

Transmembrane 4 L six family member 5 (TM4SF5) is highly expressed in hepatocellular carcinoma tissues and enhances migration in two-dimensional environments. Here, we investigated how TM4SF5 is involved in diverse pro-metastatic phenotypes in in vivo-like three-dimensional (3D) extracellular matrix gels. TM4SF5-positive cells aggressively formed invasive foci in 3D Matrigel, depending on TM4SF5-mediated signaling activity, cytoskeletal organization, and matrix metallopeptidase (MMP) 2-mediated extracellular remodeling, whereas TM4SF5-null cells did not. The TM4SF5-null cells did, however, form invasive foci in 3D Matrigel following inhibition of Rho-associated protein kinase or addition of collagen I, suggesting that collagen I compensated for TM4SF5 expression. Similarly, TM4SF5-positive cells expressing vascular endothelial-cadherin formed network-like vasculogenic mimicry in 3D Matrigel and collagen I mixture gels, whereas TM4SF5-negative cells in the mixture gels displayed the network structures only upon further treatment with epidermal growth factor. The foci formation also required MMP2-mediated remodeling of the extracellular matrix. Co-cultures exhibited TM4SF5-positive or cancer-associated fibroblasts at the outward edges of TM4SF5-null cell clusters. Compared with TM4SF5-null cells, TM4SF5-positive cells in 3D collagen gels showed a more invasive outgrowth with dramatic invadopodia. These observations suggest that TM4SF5 plays roles in the promotion of diverse metastatic properties with fewer environmental requirements than TM4SF5-negative cells. PMID:29137358

Isoforms of a cuticular protein from larvae of the meal beetle, Tenebrio molitor, studied by mass spectrometry in combination with Edman degradation and two-dimensional polyacrylamide gel electrophoresis.

PubMed Central

Haebel, S.; Jensen, C.; Andersen, S. O.; Roepstorff, P.

1995-01-01

Simultaneous sequencing, using a combination of mass spectrometry and Edman degradation, of three approximately 15-kDa variants of a cuticular protein extracted from the meal beetle Tenebrio molitor larva is demonstrated. The information obtained by matrix-assisted laser desorption ionization mass spectrometry (MALDI MS) time-course monitoring of enzymatic digests was found essential to identify the differences among the three variants and for alignment of the peptides in the sequence. To determine whether each individual insect larva contains all three protein variants, proteins extracted from single animals were separated by two-dimensional gel electrophoresis, electroeluted from the gel spots, and analyzed by MALDI MS. Molecular weights of the proteins present in each sample could be obtained, and mass spectrometric mapping of the peptides after digestion with trypsin gave additional information. The protein isoforms were found to be allelic variants. PMID:7795523

Isoforms of a cuticular protein from larvae of the meal beetle, Tenebrio molitor, studied by mass spectrometry in combination with Edman degradation and two-dimensional polyacrylamide gel electrophoresis.

PubMed

Haebel, S; Jensen, C; Andersen, S O; Roepstorff, P

1995-03-01

Simultaneous sequencing, using a combination of mass spectrometry and Edman degradation, of three approximately 15-kDa variants of a cuticular protein extracted from the meal beetle Tenebrio molitor larva is demonstrated. The information obtained by matrix-assisted laser desorption ionization mass spectrometry (MALDI MS) time-course monitoring of enzymatic digests was found essential to identify the differences among the three variants and for alignment of the peptides in the sequence. To determine whether each individual insect larva contains all three protein variants, proteins extracted from single animals were separated by two-dimensional gel electrophoresis, electroeluted from the gel spots, and analyzed by MALDI MS. Molecular weights of the proteins present in each sample could be obtained, and mass spectrometric mapping of the peptides after digestion with trypsin gave additional information. The protein isoforms were found to be allelic variants.

MAGIC polymer gel for dosimetric verification in boron neutron capture therapy

PubMed Central

Heikkinen, Sami; Kotiluoto, Petri; Serén, Tom; Seppälä, Tiina; Auterinen, Iiro; Savolainen, Sauli

2007-01-01

Radiation‐sensitive polymer gels are among the most promising three‐dimensional dose verification tools developed to date. We tested the normoxic polymer gel dosimeter known by the acronym MAGIC (methacrylic and ascorbic acid in gelatin initiated by copper) to evaluate its use in boron neutron capture therapy (BNCT) dosimetry. We irradiated a large cylindrical gel phantom (diameter: 10 cm; length: 20 cm) in the epithermal neutron beam of the Finnish BNCT facility at the FiR 1 nuclear reactor. Neutron irradiation was simulated with a Monte Carlo radiation transport code MCNP. To compare dose–response, gel samples from the same production batch were also irradiated with 6 MV photons from a medical linear accelerator. Irradiated gel phantoms then underwent magnetic resonance imaging to determine their R2 relaxation rate maps. The measured and normalized dose distribution in the epithermal neutron beam was compared with the dose distribution calculated by computer simulation. The results support the feasibility of using MAGIC gel in BNCT dosimetry. PACS numbers: 87.53.Qc, 87.53.Wz, 87.66.Ff PMID:17592463

MAGIC-f Gel in Nuclear Medicine Dosimetry: study in an external beam of Iodine-131

NASA Astrophysics Data System (ADS)

Schwarcke, M.; Marques, T.; Garrido, C.; Nicolucci, P.; Baffa, O.

2010-11-01

MAGIC-f gel applicability in Nuclear Medicine dosimetry was investigated by exposure to a 131I source. Calibration was made to provide known absorbed doses in different positions around the source. The absorbed dose in gel was compared with a Monte Carlo Simulation using PENELOPE code and a thermoluminescent dosimetry (TLD). Using MRI analysis for the gel a R2-dose sensitivity of 0.23 s-1Gy-1was obtained. The agreement between dose-distance curves obtained with Monte Carlo simulation and TLD was better than 97% and for MAGIC-f and TLD was better than 98%. The results show the potential of polymer gel for application in nuclear medicine where three dimensional dose distribution is demanded.

Hindered diffusion of high molecular weight compounds in brain extracellular microenvironment measured with integrative optical imaging.

PubMed

Nicholson, C; Tao, L

1993-12-01

This paper describes the theory of an integrative optical imaging system and its application to the analysis of the diffusion of 3-, 10-, 40-, and 70-kDa fluorescent dextran molecules in agarose gel and brain extracellular microenvironment. The method uses a precisely defined source of fluorescent molecules pressure ejected from a micropipette, and a detailed theory of the intensity contributions from out-of-focus molecules in a three-dimensional medium to a two-dimensional image. Dextrans tagged with either tetramethylrhodamine or Texas Red were ejected into 0.3% agarose gel or rat cortical slices maintained in a perfused chamber at 34 degrees C and imaged using a compound epifluorescent microscope with a 10 x water-immersion objective. About 20 images were taken at 2-10-s intervals, recorded with a cooled CCD camera, then transferred to a 486 PC for quantitative analysis. The diffusion coefficient in agarose gel, D, and the apparent diffusion coefficient, D*, in brain tissue were determined by fitting an integral expression relating the measured two-dimensional image intensity to the theoretical three-dimensional dextran concentration. The measurements in dilute agarose gel provided a reference value of D and validated the method. Values of the tortuosity, lambda = (D/D*)1/2, for the 3- and 10-kDa dextrans were 1.70 and 1.63, respectively, which were consistent with previous values derived from tetramethylammonium measurements in cortex. Tortuosities for the 40- and 70-kDa dextrans had significantly larger values of 2.16 and 2.25, respectively. This suggests that the extracellular space may have local constrictions that hinder the diffusion of molecules above a critical size that lies in the range of many neurotrophic compounds.

Comparative proteomic expression profile in all-trans retinoic acid differentiated neuroblastoma cell line.

PubMed

Cimmino, Flora; Spano, Daniela; Capasso, Mario; Zambrano, Nicola; Russo, Roberta; Zollo, Massimo; Iolascon, Achille

2007-07-01

Neuroblastoma (NB) is an infant tumor which frequently differentiates into neurons. We used two-dimensional differential in-gel electrophoresis (2D-DIGE) to analyze the cytosolic and nuclear protein expression patterns of LAN-5 cells following neuronal differentiating agent all-trans-retinoic acid treatment. We identified several candidate proteins, from which G beta2 and Prefoldin 3 may have a role on NB development. These results strength the use of proteomics to discover new putative protein targets in cancer.

Mechanotransduction mechanisms in growing spherically structured tissues

NASA Astrophysics Data System (ADS)

Littlejohns, Euan; Dunlop, Carina M.

2018-04-01

There is increasing experimental interest in mechanotransduction in multi-cellular tissues as opposed to single cells. This is driven by a growing awareness of the importance of physiologically relevant three-dimensional culture and of cell–cell and cell–gel interactions in directing growth and development. The paradigm biophysical technique for investigating tissue level mechanobiology in this context is to grow model tissues in artificial gels with well-defined mechanical properties. These studies often indicate that the stiffness of the encapsulating gel can significantly alter cellular behaviours. We demonstrate here potential mechanisms linking tissue growth with stiffness-mediated mechanotransduction. We show how tissue growth in gel systems generates points at which there is a significant qualitative change in the cellular stress and strain experienced. We show analytically how these potential switching points depend on the mechanical properties of the constraining gel and predict when they will occur. Significantly, we identify distinct mechanisms that act separately in each of the stress and strain fields at different times. These observations suggest growth as a potential physical mechanism coupling gel stiffness with cellular mechanotransduction in three-dimensional tissues. We additionally show that non-proliferating areas, in the case that the constraining gel is soft compared with the tissue, will expand and contract passively as a result of growth. Central compartment size is thus seen to not be a reliable indicator on its own for growth initiation or active behaviour.

SU-E-T-799: Verification of a Simultaneous Treatment of Multiple Brain Metastases Using VMAT Technique by a Composite Alanine-Gel Dosimeter Phantom

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pavoni, J; Silveira, M; Filho, O Baffa

Purpose: This work presents an end-to-end test using a Gel-Alanine phantom to validate the three-dimensional (3D) dose distribution (DD) delivered by a single isocenter VMAT technique on the simultaneous treatment of multiple brain metastases. Methods: Three cylindrical phantons containing MAGIC-f gel dosimeter were used to measure the 3D DD of a VMAT treatment, the first two were filled with the gel dosimeter (Gel 1 and 2) and the third one was filled with gel and 12 alanine dosimeters distributed along it (Gel 3). Gels 1 and 3 were irradiated and gel 2 was used to map the magnetic resonance imagemore » (MRI) scanner field inomogeneities. A CT scan of gel 3 was used for the VMAT treatment planning and 5 alanine pellets were chosen as lesions, around them a PTV was grown and different dose prescriptions were assigned for each one, varying from 5 to 9Gy. Before treatment, the plan was approved in a QA based on an ionization chamber absolute dose measurement, a radiochromic film planar dose measurement and a portal dosimetry per field verification; and also the phantons positioning were verified by ExacTrac 6D correction and OBI kV Cone Beam CT. The gels were irradiated, the MRIs were acquired 24 hours after irradiation and finally, the alanine dosimeters were analysed in a X-band Electron Spin Resonance spectrometer. Results: The association of the two detectors enabled the 3D dose evaluation by gel and punctually inside target volumes by alanine. In the gamma analyses (3%/3mm) comparing the 5 PTVs’ central images DD with TPS expected DD more than 95% of the points were approved. The alanine absolute dose measurements were in agreement with TPS by less than 5%. Conclusion: The gel-alanine phantom enabled the dosimetric validation of multiple brain metastases treatment using VMAT, being an almost ideal tool for this application. This work is partially supported by FAPESP.« less

FRET Imaging in Three-dimensional Hydrogels

PubMed Central

Taboas, Juan M.

2016-01-01

Imaging of Förster resonance energy transfer (FRET) is a powerful tool for examining cell biology in real-time. Studies utilizing FRET commonly employ two-dimensional (2D) culture, which does not mimic the three-dimensional (3D) cellular microenvironment. A method to perform quenched emission FRET imaging using conventional widefield epifluorescence microscopy of cells within a 3D hydrogel environment is presented. Here an analysis method for ratiometric FRET probes that yields linear ratios over the probe activation range is described. Measurement of intracellular cyclic adenosine monophosphate (cAMP) levels is demonstrated in chondrocytes under forskolin stimulation using a probe for EPAC1 activation (ICUE1) and the ability to detect differences in cAMP signaling dependent on hydrogel material type, herein a photocrosslinking hydrogel (PC-gel, polyethylene glycol dimethacrylate) and a thermoresponsive hydrogel (TR-gel). Compared with 2D FRET methods, this method requires little additional work. Laboratories already utilizing FRET imaging in 2D can easily adopt this method to perform cellular studies in a 3D microenvironment. It can further be applied to high throughput drug screening in engineered 3D microtissues. Additionally, it is compatible with other forms of FRET imaging, such as anisotropy measurement and fluorescence lifetime imaging (FLIM), and with advanced microscopy platforms using confocal, pulsed, or modulated illumination. PMID:27500354

Microstructure of β-Sitosterol:γ-Oryzanol Edible Organogels.

PubMed

Matheson, Andrew B; Koutsos, Vasileios; Dalkas, Georgios; Euston, Stephen; Clegg, Paul

2017-05-09

Rheology and atomic force microscopy (AFM) were employed to examine the microstructure of β-sitosterol:γ-oryzanol organogels in sunflower oil. Using time-resolved rheology, we followed gel formation, paying specific attention to the fibril aggregation process, which had not been studied in detail previously for this system. Using AFM, we observed gel structures directly and obtained detailed information on the gel structure, far exceeding previous studies. Our analysis suggests that though gels are formed by the self-assembly and aggregation of one-dimensional fibrils, the manner in which these fibrils aggregate into ribbons results in complex structures of higher dimensionality. We emphasize that it is a surprise to find ribbons and not twisted strands. Comparing AFM images of 10% w/w and 20% w/w gelator systems, we observed differences in the degree of branching which are consistent with the rheology. We also observed the individual self-assembled fibrils which make up these gels with much greater clarity than in previous microscopy studies, and the fibril diameters of ∼9.8 nm we measured agree excellently with those obtained from existing small-angle neutron scattering data. These results provide new insight into the structure and formation kinetics of this important organogel system.

Three-Dimensional Reflectance Traction Microscopy

PubMed Central

Jones, Christopher A. R.; Groves, Nicholas Scott; Sun, Bo

2016-01-01

Cells in three-dimensional (3D) environments exhibit very different biochemical and biophysical phenotypes compared to the behavior of cells in two-dimensional (2D) environments. As an important biomechanical measurement, 2D traction force microscopy can not be directly extended into 3D cases. In order to quantitatively characterize the contraction field, we have developed 3D reflectance traction microscopy which combines confocal reflection imaging and partial volume correlation postprocessing. We have measured the deformation field of collagen gel under controlled mechanical stress. We have also characterized the deformation field generated by invasive breast cancer cells of different morphologies in 3D collagen matrix. In contrast to employ dispersed tracing particles or fluorescently-tagged matrix proteins, our methods provide a label-free, computationally effective strategy to study the cell mechanics in native 3D extracellular matrix. PMID:27304456

Fabrication of three dimensional patterns of wide dimensional range using microbes and their applications

NASA Astrophysics Data System (ADS)

Mehta, Sunita; Murugeson, Saravanan; Prakash, Balaji; Deepak

2015-10-01

Inspired by the wound healing property of certain trees, we report a novel microbes based additive process for producing three dimensional patterns, which has a potential of engineering applications in a variety of fields. Imposing a two dimensional pattern of microbes on a gel media and allowing them to grow in the third dimension is known from its use in biological studies. Instead, we have introduced an intermediate porous substrate between the gel media and the microbial growth, which enables three dimensional patterns in specific forms that can be lifted off and used in engineering applications. In order to demonstrate the applicability of this idea in a diverse set of areas, two applications are selected. In one, using this method of microbial growth, we have fabricated microlenses for enhanced light extraction in organic light emitting diodes, where densely packed microlenses of the diameters of hundreds of microns lead to luminance increase by a factor of 1.24X. In another entirely different type of application, braille text patterns are prepared on a normal office paper where the grown microbial colonies serve as braille tactile dots. Braille dot patterns thus prepared meet the standard specifications (size and spacing) for braille books.

Fabrication of 3D-culture platform with sandwich architecture for preserving liver-specific functions of hepatocytes using 3D bioprinter.

PubMed

Arai, Kenichi; Yoshida, Toshiko; Okabe, Motonori; Goto, Mitsuaki; Mir, Tanveer Ahmad; Soko, Chika; Tsukamoto, Yoshinari; Akaike, Toshihiro; Nikaido, Toshio; Zhou, Kaixuan; Nakamura, Makoto

2017-06-01

The development of new three-dimensional (3D) cell culture system that maintains the physiologically relevant signals of hepatocytes is essential in drug discovery and tissue engineering research. Conventional two-dimensional (2D) culture yields cell growth, proliferation, and differentiation. However, gene expression and signaling profiles can be different from in vivo environment. Here, we report the fabrication of a 3D culture system using an artificial scaffold and our custom-made inkjet 3D bioprinter as a new strategy for studying liver-specific functions of hepatocytes. We built a 3D culture platform for hepatocytes-attachment and formation of cell monolayer by interacting the galactose chain of galactosylated alginate gel (GA-gel) with asialoglycoprotein receptor (ASGPR) of hepatocytes. The 3D geometrical arrangement of cells was controlled by using 3D bioprinter, and cell polarity was controlled with the galactosylated hydrogels. The fabricated GA-gel was able to successfully promote adhesion of hepatocytes. To observe liver-specific functions and to mimic hepatic cord, an additional parallel layer of hepatocytes was generated using two gel sheets. These results indicated that GA-gel biomimetic matrices can be used as a 3D culture system that could be effective for the engineering of liver tissues. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1583-1592, 2017. © 2017 Wiley Periodicals, Inc.

Introducing Proteomics in the Undergraduate Curriculum: A Simple 2D Gel Electrophoresis Exercise with Serum Proteins

ERIC Educational Resources Information Center

Kim, Thomas D.; Craig, Paul A.

2010-01-01

Two-dimensional gel electrophoresis (2DGE) remains an important tool in the study of biological systems by proteomics. While the use of 2DGE is commonplace in research publications, there are few instructional laboratories that address the use of 2DGE for analyzing complex protein samples. One reason for this lack is the fact that the preparation…

Altered Protein Expression of Streptococcus oralis Cultured at Low pH Revealed by Two-Dimensional Gel Electrophoresis

PubMed Central

Wilkins, Joanna C.; Homer, Karen A.; Beighton, David

2001-01-01

Streptococcus oralis is the predominant aciduric nonmutans streptococcus isolated from the human dentition, but the role of this organism in the initiation and progression of dental caries has yet to be established. To identify proteins that are differentially expressed by S. oralis growing under conditions of low pH, soluble cellular proteins extracted from bacteria grown in batch culture at pH 5.2 or 7.0 were analyzed by two-dimensional (2-D) gel electrophoresis. Thirty-nine proteins had altered expression at low pH; these were excised, digested with trypsin using an in-gel protocol, and further analyzed by peptide mass fingerprinting using matrix-assisted laser desorption ionization mass spectrometry. The resulting fingerprints were compared with the genomic database for Streptococcus pneumoniae, an organism that is phylogenetically closely related to S. oralis, and putative functions for the majority of these proteins were determined on the basis of functional homology. Twenty-eight proteins were up-regulated following growth at pH 5.2; these included enzymes of the glycolytic pathway (glyceraldehyde-3-phosphate dehydrogenase and lactate dehydrogenase), the polypeptide chains comprising ATP synthase, and proteins that are considered to play a role in the general stress response of bacteria, including the 60-kDa chaperone, Hsp33, and superoxide dismutase, and three distinct ABC transporters. These data identify, for the first time, gene products that may be important in the survival and proliferation of nonmutans aciduric S. oralis under conditions of low pH that are likely to be encountered by this organism in vivo. PMID:11472910

A three-dimensionally chitin nanofiber/carbon nanotube hydrogel network for foldable conductive paper.

PubMed

Chen, Chuchu; Yang, Chuang; Li, Suiyi; Li, Dagang

2015-12-10

We reported a highly conductive nanocomposite made with multiwalled carbon nanotubes (MWCNTs) and chitin nanofibers (ChNFs). The MWCNTs were dispersed into ChNFs by the simple process of vacuum-filtration, forming a three-dimensional network structure. In this approach, MWCNT acted as a filler to introduce electron channel paths throughout the ChNF skeleton. And then, a hybrid hydrogel system (20 wt.% NaOH, -18 °C) was applied to prepare the ChNF/MWCNT gel-film followed with drying process. It is found that the resultant ChNF/MWCNT gel-film exposed much more MWCNT areas forming denser structure due to the shrinking of ChNFs after the gelation treatment. Compared with ChNF/MWCNT film, the one treated under hydrogel system (ChNF/MWCNT gel-film) exhibited almost twice higher conductivity (9.3S/cm for 50 wt.% MWCNTs in gel-film; whereas 4.7S/cm for 50 wt.% MWCNTs in film). Moreover, the facile and low-cost of this conductive paper may have great potential in development of foldable electronic devices. Copyright © 2015 Elsevier Ltd. All rights reserved.

Proteomic Analysis Reveals Differences in Tolerance to Acid Rain in Two Broad-Leaf Tree Species, Liquidambar formosana and Schima superba

PubMed Central

Wang, Chao; Liu, Ting-Wu; Chalifour, Annie; Chen, Juan; Shen, Zhi-Jun; Liu, Xiang; Wang, Wen-Hua; Zheng, Hai-Lei

2014-01-01

Acid rain (AR) is a serious environmental issue inducing harmful impacts on plant growth and development. It has been reported that Liquidambar formosana, considered as an AR-sensitive tree species, was largely injured by AR, compared with Schima superba, an AR-tolerant tree species. To clarify the different responses of these two species to AR, a comparative proteomic analysis was conducted in this study. More than 1000 protein spots were reproducibly detected on two-dimensional electrophoresis gels. Among them, 74 protein spots from L. formosana gels and 34 protein spots from S. superba gels showed significant changes in their abundances under AR stress. In both L. formosana and S. superba, the majority proteins with more than 2 fold changes were involved in photosynthesis and energy production, followed by material metabolism, stress and defense, transcription, post-translational and modification, and signal transduction. In contrast with L. formosana, no hormone response-related protein was found in S. superba. Moreover, the changes of proteins involved in photosynthesis, starch synthesis, and translation were distinctly different between L. formosana and S. superba. Protein expression analysis of three proteins (ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit, ascorbate peroxidase and glutathione-S-transferase) by Western blot was well correlated with the results of proteomics. In conclusion, our study provides new insights into AR stress responses in woody plants and clarifies the differences in strategies to cope with AR between L. formosana and S. superba. PMID:25025692

A sensitive gel-based global O-glycomics approach reveals high levels of mannosyl glycans in the high mass region of the mouse brain proteome.

PubMed

Breloy, Isabelle; Pacharra, Sandra; Aust, Christina; Hanisch, Franz-Georg

2012-08-01

We developed a gel-based global O-glycomics method applicable for highly complex protein mixtures entrapped in discontinuous gradient gel layers. The protocol is based on in-gel proteolysis with pronase followed by (glyco)peptide elution and off-gel reductive β-elimination. The protocol offers robust performance with sensitivity in the low picomolar range, is compatible with gel-based proteomics, and shows superior performance in global applications in comparison with workflows eliminating glycans in-gel or from electroblotted glycoproteins. By applying this method, we analyzed the O-glycome of human myoblasts and of the mouse brain O-glycoproteome. After semipreparative separation of mouse brain proteins by one-dimensional SDS gel electrophoresis, the O-glycans from proteins in different mass ranges were characterized with a focus on O-mannose-based glycans. The relative proportion of the latter, which generally represent a rare modification, increases to comparatively high levels in the mouse brain proteome in dependence of increasing protein masses.

Automated apparatus for producing gradient gels

DOEpatents

Anderson, N.L.

1983-11-10

Apparatus for producing a gradient gel which serves as a standard medium for a two-dimensional analysis of proteins, the gel having a density gradient along its height formed by a variation in gel composition, with the apparatus including first and second pumping means each including a plurality of pumps on a common shaft and driven by a stepping motor capable of providing small incremental changes in pump outputs for the gel ingredients, the motors being controlled, by digital signals from a digital computer, a hollow form or cassette for receiving the gel composition, means for transferring the gel composition including a filler tube extending near the bottom of the cassette, adjustable horizontal and vertical arms for automatically removing and relocating the filler tube in the next cassette, and a digital computer programmed to automatically control the stepping motors, arm movements, and associated sensing operations involving the filling operation.

Automated apparatus for producing gradient gels

DOEpatents

Anderson, Norman L.

1986-01-01

Apparatus for producing a gradient gel which serves as a standard medium for a two-dimensional analysis of proteins, the gel having a density gradient along its height formed by a variation in gel composition, with the apparatus including first and second pumping means each including a plurality of pumps on a common shaft and driven by a stepping motor capable of providing small incremental changes in pump outputs for the gel ingredients, the motors being controlled, by digital signals from a digital computer, a hollow form or cassette for receiving the gel composition, means for transferring the gel composition including a filler tube extending near the bottom of the cassette, adjustable horizontal and vertical arms for automatically removing and relocating the filler tube in the next cassette, and a digital computer programmed to automatically control the stepping motors, arm movements, and associated sensing operations involving the filling operation.

Upon Generating (2+1)-dimensional Dynamical Systems

NASA Astrophysics Data System (ADS)

Zhang, Yufeng; Bai, Yang; Wu, Lixin

2016-06-01

Under the framework of the Adler-Gel'fand-Dikii(AGD) scheme, we first propose two Hamiltonian operator pairs over a noncommutative ring so that we construct a new dynamical system in 2+1 dimensions, then we get a generalized special Novikov-Veselov (NV) equation via the Manakov triple. Then with the aid of a special symmetric Lie algebra of a reductive homogeneous group G, we adopt the Tu-Andrushkiw-Huang (TAH) scheme to generate a new integrable (2+1)-dimensional dynamical system and its Hamiltonian structure, which can reduce to the well-known (2+1)-dimensional Davey-Stewartson (DS) hierarchy. Finally, we extend the binormial residue representation (briefly BRR) scheme to the super higher dimensional integrable hierarchies with the help of a super subalgebra of the super Lie algebra sl(2/1), which is also a kind of symmetric Lie algebra of the reductive homogeneous group G. As applications, we obtain a super 2+1 dimensional MKdV hierarchy which can be reduced to a super 2+1 dimensional generalized AKNS equation. Finally, we compare the advantages and the shortcomings for the three schemes to generate integrable dynamical systems.

Initial proteome analysis of caffeine-induced proteins in Aspergillus tamarii using two-dimensional fluorescence difference gel electrophoresis.

PubMed

Gutiérrez-Sánchez, Gerardo; Atwood, James; Kolli, V S Kumar; Roussos, Sévastianos; Augur, Christopher

2012-04-01

Caffeine is toxic to most microorganisms. However, some filamentous fungi, such as Aspergillus tamarii, are able to metabolize this alkaloid when fed caffeine as the sole nitrogen source. The aim of the present work was to identify intracellular A. tamarii proteins, regulated by caffeine, using fluorescence difference two-dimensional gel electrophoresis. Specific proteins from two culture media of A. tamarii grown either on ammonium sulfate or caffeine as the sole nitrogen source were analysed by mass spectrometry. Thirteen out of a total of 85 differentially expressed spots were identified after database search. Identified up-regulated proteins include phosphoglycerate kinase, malate dehydrogenase, dyp-type peroxidase family protein, heat shock protein, Cu, Zn superoxidase dismutase and xanthine dehydrogenase. Some of the proteins identified in this study are involved in the caffeine degradation pathway as well as in stress response, suggesting that stress proteins could be involved in caffeine metabolism in filamentous fungi.

Preprocessing of 2-Dimensional Gel Electrophoresis Images Applied to Proteomic Analysis: A Review.

PubMed

Goez, Manuel Mauricio; Torres-Madroñero, Maria Constanza; Röthlisberger, Sarah; Delgado-Trejos, Edilson

2018-02-01

Various methods and specialized software programs are available for processing two-dimensional gel electrophoresis (2-DGE) images. However, due to the anomalies present in these images, a reliable, automated, and highly reproducible system for 2-DGE image analysis has still not been achieved. The most common anomalies found in 2-DGE images include vertical and horizontal streaking, fuzzy spots, and background noise, which greatly complicate computational analysis. In this paper, we review the preprocessing techniques applied to 2-DGE images for noise reduction, intensity normalization, and background correction. We also present a quantitative comparison of non-linear filtering techniques applied to synthetic gel images, through analyzing the performance of the filters under specific conditions. Synthetic proteins were modeled into a two-dimensional Gaussian distribution with adjustable parameters for changing the size, intensity, and degradation. Three types of noise were added to the images: Gaussian, Rayleigh, and exponential, with signal-to-noise ratios (SNRs) ranging 8-20 decibels (dB). We compared the performance of wavelet, contourlet, total variation (TV), and wavelet-total variation (WTTV) techniques using parameters SNR and spot efficiency. In terms of spot efficiency, contourlet and TV were more sensitive to noise than wavelet and WTTV. Wavelet worked the best for images with SNR ranging 10-20 dB, whereas WTTV performed better with high noise levels. Wavelet also presented the best performance with any level of Gaussian noise and low levels (20-14 dB) of Rayleigh and exponential noise in terms of SNR. Finally, the performance of the non-linear filtering techniques was evaluated using a real 2-DGE image with previously identified proteins marked. Wavelet achieved the best detection rate for the real image. Copyright © 2018 Beijing Institute of Genomics, Chinese Academy of Sciences and Genetics Society of China. Production and hosting by Elsevier B.V. All rights reserved.

Three-dimensional dosimetry of small megavoltage radiation fields using radiochromic gels and optical CT scanning

NASA Astrophysics Data System (ADS)

Babic, Steven; McNiven, Andrea; Battista, Jerry; Jordan, Kevin

2009-04-01

The dosimetry of small fields as used in stereotactic radiotherapy, radiosurgery and intensity-modulated radiation therapy can be challenging and inaccurate due to partial volume averaging effects and possible disruption of charged particle equilibrium. Consequently, there exists a need for an integrating, tissue equivalent dosimeter with high spatial resolution to avoid perturbing the radiation beam and artificially broadening the measured beam penumbra. In this work, radiochromic ferrous xylenol-orange (FX) and leuco crystal violet (LCV) micelle gels were used to measure relative dose factors (RDFs), percent depth dose profiles and relative lateral beam profiles of 6 MV x-ray pencil beams of diameter 28.1, 9.8 and 4.9 mm. The pencil beams were produced via stereotactic collimators mounted on a Varian 2100 EX linear accelerator. The gels were read using optical computed tomography (CT). Data sets were compared quantitatively with dosimetric measurements made with radiographic (Kodak EDR2) and radiochromic (GAFChromic® EBT) film, respectively. Using a fast cone-beam optical CT scanner (Vista™), corrections for diffusion in the FX gel data yielded RDFs that were comparable to those obtained by minimally diffusing LCV gels. Considering EBT film-measured RDF data as reference, cone-beam CT-scanned LCV gel data, corrected for scattered stray light, were found to be in agreement within 0.5% and -0.6% for the 9.8 and 4.9 mm diameter fields, respectively. The validity of the scattered stray light correction was confirmed by general agreement with RDF data obtained from the same LCV gel read out with a laser CT scanner that is less prone to the acceptance of scattered stray light. Percent depth dose profiles and lateral beam profiles were found to agree within experimental error for the FX gel (corrected for diffusion), LCV gel (corrected for scattered stray light), and EBT and EDR2 films. The results from this study reveal that a three-dimensional dosimetry method utilizing optical CT-scanned radiochromic gels allows for the acquisition of a self-consistent volumetric data set in a single exposure, with sufficient spatial resolution to accurately characterize small fields.

Simplification and improvement of protein detection in two-dimensional electrophoresis gels with SERVA HPE™ lightning red.

PubMed

Griebel, Anja; Obermaier, Christian; Westermeier, Reiner; Moche, Martin; Büttner, Knut

2013-07-01

A new fluorescent amino-reactive dye has been tested for both labelling proteins prior to electrophoretic separations and between the two steps of two-dimensional electrophoresis. A series of experiments showed, that the labelling of lysines with this dye is compatible with all standard additives used for sample preparation, including reducing substances and carrier ampholytes. Using this dye for pre-labelling considerably simplifies the electrophoresis and detection workflow and provides highly sensitive and quantitative visualisation of proteins.

Exposures of Sus scrofa to a TASER(®) conducted electrical weapon: no effects on 2-dimensional gel electrophoresis patterns of plasma proteins.

PubMed

Jauchem, James R; Cerna, Cesario Z; Lim, Tiffany Y; Seaman, Ronald L

2014-12-01

In an earlier study, we found significant changes in red-blood-cell, leukocyte, and platelet counts, and in red-blood-cell membrane proteins, following exposures of anesthetized pigs to a conducted electrical weapon. In the current study, we examined potential changes in plasma proteins [analyzed via two-dimensional gel electrophoresis (2-DGE)] following two 30 s exposures of anesthetized pigs (Sus scrofa) to a TASER (®) C2 conducted electrical weapon. Patterns of proteins, separated by 2-DGE, were consistent and reproducible between animals and between times of sampling. We determined that the blood plasma collection, handling, storage, and processing techniques we used are suitable for swine blood. There were no statistically significant changes in plasma proteins following the conducted-electrical-weapon exposures. Overall gel patterns of fibrinogen were similar to results of other studies of both pigs and humans (in control settings, not exposed to conducted electrical weapons). The lack of significant changes in plasma proteins may be added to the body of evidence regarding relative safety of TASER C2 device exposures.

Techniques for characterization and eradication of potato cyst nematode: a review.

PubMed

Bairwa, Aarti; Venkatasalam, E P; Sudha, R; Umamaheswari, R; Singh, B P

2017-09-01

Correct identification of species and pathotypes is must for eradication of potato cyst nematodes (PCN). The identification of PCN species after completing the life cycle is very difficult because it is based on morphological and morphometrical characteristics. Genetically different populations of PCN are morphologically same and differentiated based on the host differential study. Later on these traditional techniques have been replaced by biochemical techniques viz, one and two dimensional gel electrophoresis, capillary gel electrophoresis, isozymes, dot blot hybridization and isoelectric focusing etc. to distinguish both the species. One and two dimensional gel electrophoresis has used to examine inter- and intra-specific differences in proteins of Globodera rostochiensis and G. pallida . Now application of PCR and DNA based characterization techniques like RAPD, AFLP and RFLP are the important tools for differentiating inter- and intra specific variation in PCN and has given opportunities to accurate identification of PCN. For managing the PCN, till now we are following integrated pest management (IPM) strategies, however these strategies are not effective to eradicate the PCN. Therefore to eradicate the PCN we need noval management practices like RNAi (RNA interference) or Gene silencing.

Water equivalence of NIPAM based polymer gel dosimeters with enhanced sensitivity for x-ray CT

NASA Astrophysics Data System (ADS)

Gorjiara, Tina; Hill, Robin; Bosi, Stephen; Kuncic, Zdenka; Baldock, Clive

2013-10-01

Two new formulations of N-isopropylacrylamide (NIPAM) based three dimensional (3D) gel dosimeters have recently been developed with improved sensitivity to x-ray CT readout, one without any co-solvent and the other one with isopropanol co-solvent. The water equivalence of the NIPAM gel dosimeters was investigated using different methods to calculate their radiological properties including: density, electron density, number of electrons per grams, effective atomic number, photon interaction probabilities, mass attenuation and energy absorption coefficients, electron collisional, radiative and total mass stopping powers and electron mass scattering power. Monte Carlo modelling was also used to compare the dose response of these gel dosimeters with water for kilovoltage and megavoltage x-ray beams and for megavoltage electron beams. We found that the density and electron density of the co-solvent free gel dosimeter are more water equivalent with less than a 2.6% difference compared to a 5.7% difference for the isopropanol gel dosimeter. Both the co-solvent free and isopropanol solvent gel dosimeters have lower effective atomic numbers than water, differing by 2.2% and 6.5%, respectively. As a result, their photoelectric absorption interaction probabilities are up to 6% and 19% different from water, respectively. Compton scattering and pair production interaction probabilities of NIPAM gel with isopropanol differ by up to 10% from water while for the co-solvent free gel, the differences are 3%. Mass attenuation and energy absorption coefficients of the co-solvent free gel dosimeter and the isopropanol gel dosimeter are up to 7% and 19% lower than water, respectively. Collisional and total mass stopping powers of both gel dosimeters differ by less than 2% from those of water. The dose response of the co-solvent free gel dosimeter is water equivalent (with <1% discrepancy) for dosimetry of x-rays with energies <100 keV while the discrepancy increases (up to 5%) for the isopropanol gel dosimeter over the same energy range. For x-ray beams over the energy range 180 keV-18 MV, both gel dosimeters have less than 2% discrepancy with water. For megavoltage electron beams, the dose differences with water reach 7% and 14% for the co-solvent free gel dosimeter and the isopropanol gel dosimeter, respectively. Our results demonstrate that for x-ray beam dosimetry with photon energies higher than 100 keV and megavoltage electron beams, correction factors are needed for both NIPAM gels to be used as water equivalent dosimeters.

Space Science

NASA Image and Video Library

2003-10-28

These gels were obtained by two-dimensional (2D) electrophoresis, in which proteins move different substances through a polyacrylamide gel matrix based on their molecular weight and total charge in an electric field. The gels illustrate principal investigator David Niesel’s findings that exposure to modeled microgravity results in some Streptoccoccus Pneumonia’s proteins being upregulated and others being downregulated. In 2D protein profiles of whole cell lysates of Streptoccoccus Pneumonia, 6,304 cultured under normal gravity (left), appear to be expressed at higher levels indicated with black circles. Red circles (right) indicate proteins that were grown under modeled microgravity in a high aspect ratio vessel HARV).

Increased expression of annexin I and thioredoxin detected by two-dimensional gel electrophoresis of drug resistant human stomach cancer cells.

PubMed

Sinha, P; Hütter, G; Köttgen, E; Dietel, M; Schadendorf, D; Lage, H

1998-11-18

The therapy of advanced cancer using chemotherapy alone or in combination with radiation or hyperthermia yields an overall response rate of about 20-50%. This success is often marred by the development of resistance to cytostatic drugs. Our aim was to study the global analysis of protein expression in the development of chemoresistance in vitro. We therefore used a cell culture model derived from the gastric carcinoma cell line EPG 85-257P. A classical multidrug-resistant subline EPG85-257RDB selected to daunorubicin and an atypical multidrug-resistant cell variant EPG85-257RNOV selected to mitoxantrone, were analysed using two-dimensional electrophoresis in immobilized pH-gradients (pH 4.0-8.0) in the first dimension and linear polyacrylamide gels (12%) in the second dimension. After staining with coomassie brilliant blue, image analysis was performed using the PDQuest system. Spots of interest were isolated using preparative two-dimensional electrophoresis and subjected to microsequencing. A total of 241 spots from the EPG85-257RDB-standard and 289 spots from the EPG85-257RNOV-standard could be matched to the EPG85-257P-standard. Microsequencing after enzymatic hydrolysis in gel, mass spectrometric data and sequencing of the peptides after their fractionation using microbore HPLC identified that two proteins annexin I and thioredoxin were overexpressed in chemoresistant cell lines. Annexin I was present in both the classical and the atypical multidrug-resistant cells. Thioredoxin was found to be overexpressed only in the atypical multidrug-resistant cell line.

Fabrication of three dimensional patterns of wide dimensional range using microbes and their applications

PubMed Central

Mehta, Sunita; Murugeson, Saravanan; Prakash, Balaji; Deepak

2015-01-01

Inspired by the wound healing property of certain trees, we report a novel microbes based additive process for producing three dimensional patterns, which has a potential of engineering applications in a variety of fields. Imposing a two dimensional pattern of microbes on a gel media and allowing them to grow in the third dimension is known from its use in biological studies. Instead, we have introduced an intermediate porous substrate between the gel media and the microbial growth, which enables three dimensional patterns in specific forms that can be lifted off and used in engineering applications. In order to demonstrate the applicability of this idea in a diverse set of areas, two applications are selected. In one, using this method of microbial growth, we have fabricated microlenses for enhanced light extraction in organic light emitting diodes, where densely packed microlenses of the diameters of hundreds of microns lead to luminance increase by a factor of 1.24X. In another entirely different type of application, braille text patterns are prepared on a normal office paper where the grown microbial colonies serve as braille tactile dots. Braille dot patterns thus prepared meet the standard specifications (size and spacing) for braille books. PMID:26486847

Gel properties and interactions of Mesona blumes polysaccharide-soy protein isolates mixed gel: The effect of salt addition.

PubMed

Wang, Wenjie; Shen, Mingyue; Liu, Suchen; Jiang, Lian; Song, Qianqian; Xie, Jianhua

2018-07-15

Effect of different salt ions on the gel properties and microstructure of Mesona blumes polysaccharide (MBP)-soy protein isolates (SPI) mixed gels were investigated. Sodium and calcium ions were chosen to explore their effects on the rheological behavior and gel properties of MBP-SPI mixed gels were evaluated by using rheological, X-ray diffraction, protein solubility determination, and microstructure analysis. Results showed that the addition of salt ions change the crystalline state of gels system, the crystal of gel was enhanced at low ion concentrations (0.005-0.01 M). The two peaks of gel characteristic at 8.9° and 19.9° almost disappeared at high salt ions concentrations (0.015-0.02 M), and new crystallization peaks appeared at around 30° and 45°. The elasticity, viscosity, gel strength, water holding capacity, and thermal stability of gel were increased at low ion concentration. Results showed that the main interactions which promoted gel formation and maintain the three-dimensional structure of the gel were electrostatic interactions, hydrophobic interactions, and disulfide interactions. Copyright © 2018 Elsevier Ltd. All rights reserved.

Apparatus for electrophoresis separation

DOEpatents

Anderson, Norman L.

1978-01-01

An apparatus is disclosed for simultaneously performing electrophoresis separations on a plurality of slab gels containing samples of protein, protein subunits or nucleic acids. A reservoir of buffer solution is divided into three compartments by two parallel partitions having vertical slots spaced along their length. A sheet of flexible, electrically insulative material is attached to each partition and is provided with vertical slits aligned with the slots. Slab-gel holders are received within the slots with the flexible material folded outwardly as flaps from the slits to overlay portions of the holder surfaces and thereby act as electrical and liquid seals. An elongated, spaghetti-like gel containing a sample of specimen that was previously separated by isoelectric focusing techniques is vertically positioned along a marginal edge portion of the slab gel. On application of an electrical potential between the two outer chambers of buffer solution, a second dimensional electrophoresis separation in accordance with molecular weight occurs as the specimen molecules migrate across the slab gel.

Two-dimensional polyacrylamide gel electrophoresis of equine seminal plasma proteins and their relation with semen freezability.

PubMed

Jobim, M I M; Trein, C; Zirkler, H; Gregory, R M; Sieme, H; Mattos, R C

2011-09-01

The objective was to evaluate protein profiles of equine seminal plasma using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and to determine whether any of these proteins were related to semen freezability. Seminal plasma was collected from 10 stallions, of high and low semen freezability, housed at the State Stud of Lower Saxony, and routinely used in AI programs. Twenty-five protein spots were identified from the two-dimensional gel (12%), seven of which were present in all samples (all proteins were identified by MALDI-MS). Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) has been used to generate ion images of samples in one or more mass-to-charge (m/z) values, providing the capability of mapping specific molecules to two-dimensional coordinates of the original sample. Of the 25 proteins identified, two spots had greater relative content (P < 0.05) in seminal plasma samples collected from stallions with high semen freezability: spot 5 (80-85 kDa, isoelectric point [pI] 7.54), identified as CRISP-3; and spot 45 (18.2 kDa, pI 5.0-5.2), identified as HSP-2. Conversely, protein content was greater (P < 0.05) in seminal plasma samples from stallions with low semen freezability: spot 7 (75.4 kDa, pI 6.9-7.4), identified as lactoferrin; spot 15 (26.7 kDa, pI 5.51), identified as kallikrein; spot 25 (25 kDa, pI 7.54), identified as CRISP-3; and spot 35 (13.9 kDa, pI 3.8-4.2), identified as HSP-1. In conclusion, there were differences in the seminal plasma protein profile from stallions with high and low semen freezability. Furthermore, CRISP-3 and HSP-2 were potential seminal plasma markers of high semen freezability. Copyright © 2011 Elsevier Inc. All rights reserved.

Fast Two-Dimensional Bubble Analysis of Biopolymer Filamentous Networks Pore Size from Confocal Microscopy Thin Data Stacks

PubMed Central

Molteni, Matteo; Magatti, Davide; Cardinali, Barbara; Rocco, Mattia; Ferri, Fabio

2013-01-01

The average pore size ξ0 of filamentous networks assembled from biological macromolecules is one of the most important physical parameters affecting their biological functions. Modern optical methods, such as confocal microscopy, can noninvasively image such networks, but extracting a quantitative estimate of ξ0 is a nontrivial task. We present here a fast and simple method based on a two-dimensional bubble approach, which works by analyzing one by one the (thresholded) images of a series of three-dimensional thin data stacks. No skeletonization or reconstruction of the full geometry of the entire network is required. The method was validated by using many isotropic in silico generated networks of different structures, morphologies, and concentrations. For each type of network, the method provides accurate estimates (a few percent) of the average and the standard deviation of the three-dimensional distribution of the pore sizes, defined as the diameters of the largest spheres that can be fit into the pore zones of the entire gel volume. When applied to the analysis of real confocal microscopy images taken on fibrin gels, the method provides an estimate of ξ0 consistent with results from elastic light scattering data. PMID:23473499

Two-dimensional electrophoretic profiling of normal human kidney glomerulus proteome and construction of an extensible markup language (XML)-based database.

PubMed

Yoshida, Yutaka; Miyazaki, Kenji; Kamiie, Junichi; Sato, Masao; Okuizumi, Seiji; Kenmochi, Akihisa; Kamijo, Ken'ichi; Nabetani, Takuji; Tsugita, Akira; Xu, Bo; Zhang, Ying; Yaoita, Eishin; Osawa, Tetsuo; Yamamoto, Tadashi

2005-03-01

To contribute to physiology and pathophysiology of the glomerulus of human kidney, we have launched a proteomic study of human glomerulus, and compiled a profile of proteins expressed in the glomerulus of normal human kidney by two-dimensional gel electrophoresis (2-DE) and identification with matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) and/or liquid chromatography-tandem mass spectrometry (LC-MS/MS). Kidney cortices with normal appearance were obtained from patients under surgical nephrectomy due to renal tumor, and glomeruli were highly purified by a standard sieving method followed by picking-up under a phase-contrast microscope. The glomerular proteins were separated by 2-DE with 24 cm immobilized pH gradient strips in the 3-10 range in the first dimension and 26 x 20 cm sodium dodecyl sulfate polyacrylamide electrophoresis gels of 12.5% in the second dimension. Gels were silver-stained, and valid spots were processed for identification through an integrated robotic system that consisted of a spot picker, an in-gel digester, and a MALDI-TOF MS and / or a LC-MS/MS. From 2-DE gel images of glomeruli of four subjects with no apparent pathologic manifestations, a synthetic gel image of normal glomerular proteins was created. The synthetic gel image contained 1713 valid spots, of which 1559 spots were commonly observed in the respective 2-DE gels. Among the 1559 spots, 347 protein spots, representing 212 proteins, have so far been identified, and used for the construction of an extensible markup language (XML)-based database. The database is deposited on a web site (http://www.sw.nec.co.jp/bio/rd/hgldb/index.html) in a form accessible to researchers to contribute to proteomic studies of human glomerulus in health and disease.

A Study of the Effects of High Power Pulsed 2450 MHz Microwaves, ELF modulated Microwaves, and ELF Fields on Human Lymphocytes and Selected Cell Lines

DTIC Science & Technology

1993-01-27

Considerable effect was expended in investigating shifts in intercellular calcium of one particular cell line, Jurket, using flow cytometry methods. No...culture. The following analysis were used to characterize the immortalized cell lines: flow cytometry , electron microscopy, two-dimensional protein gel...further characterized by flow cytometry , electron microscopy, two dimensional protein electrophoresis and nuclear run-off assay. Flow cytometric analysis of

Quantitative phase analysis and microstructure characterization of magnetite nanocrystals obtained by microwave assisted non-hydrolytic sol–gel synthesis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sciancalepore, Corrado, E-mail: corrado.sciancalepore@unimore.it; Bondioli, Federica; INSTM Consortium, Via G. Giusti 9, 51121 Firenze

2015-02-15

An innovative preparation procedure, based on microwave assisted non-hydrolytic sol–gel synthesis, to obtain spherical magnetite nanoparticles was reported together with a detailed quantitative phase analysis and microstructure characterization of the synthetic products. The nanoparticle growth was analyzed as a function of the synthesis time and was described in terms of crystallization degree employing the Rietveld method on the magnetic nanostructured system for the determination of the amorphous content using hematite as internal standard. Product crystallinity increases as the microwave thermal treatment is increased and reaches very high percentages for synthesis times longer than 1 h. Microstructural evolution of nanocrystals wasmore » followed by the integral breadth methods to obtain information on the crystallite size-strain distribution. The results of diffraction line profile analysis were compared with nanoparticle grain distribution estimated by dimensional analysis of the transmission electron microscopy (TEM) images. A variation both in the average grain size and in the distribution of the coherently diffraction domains is evidenced, allowing to suppose a relationship between the two quantities. The traditional integral breadth methods have proven to be valid for a rapid assessment of the diffraction line broadening effects in the above-mentioned nanostructured systems and the basic assumption for the correct use of these methods are discussed as well. - Highlights: • Fe{sub 3}O{sub 4} nanocrystals were obtained by MW-assisted non-hydrolytic sol–gel synthesis. • Quantitative phase analysis revealed that crystallinity up to 95% was reached. • The strategy of Rietveld refinements was discussed in details. • Dimensional analysis showed nanoparticles ranging from 4 to 8 nm. • Results of integral breadth methods were compared with microscopic analysis.« less

Enhancement of the Gelation Properties of Surimi from Yellowtail Seabream (Parargyrops edita, Sparidae) with Chinese Oak Silkworm Pupa, Antheraea pernyi.

PubMed

Zhu, Jialin; Fan, Daming; Zhao, Jianxin; Zhang, Hao; Huang, Jianlian; Zhou, Wenguo; Zhang, Wenhai; Chen, Wei

2016-02-01

In this study, the textural properties and micromechanism of yellowtail seabream (Parargyrops edita, Sparidae) surimi, with and without Chinese oak silkworm pupa homogenate (SPH), were investigated at different levels. The fresh, freeze-dried, and oven-dried SPH all showed a gel-enhancing ability in suwari (40/90 °C) and modori (67/90 °C) gels, in a concentration-dependent manner. Though the drying treatments can improve the storability of SPH, compared with fresh, the effect of the active substance was weakened. Suwari and modori gels added with 5%(w/w, whole product) fresh SPH had the increase in breaking force and deformation by 37.39% and 47.98%, and 85.14% and 78.49%, respectively, compared with the control gel (without SPH addition). The major myofibrillar protein, especially myosin heavy chain (MHC), was better retained by the addition of SPH. Compared the control group, a finer, denser, and more ordered 3-dimensional gel network microstructure was obtained, and different Df (Fractal dimension) was analyzed by using the box count method. This was found in all samples from 2.838 to 2.864 for suwari gels and 2.795 to 2.857 for modori gels, respectively. Therefore, the modori of yellowtail seabream surimi, linked with endogenous proteases, could be retarded in the presence of SPH, leading to an increase in gel strength. © 2015 Institute of Food Technologists®

Antibody response to a sterile filtered PPD tuberculin in M. bovis infected and M. bovis sensitized cattle.

PubMed

Rennie, Bryan; Filion, Lionel G; Smart, Nonie

2010-11-09

Bovine tuberculosis, caused by Mycobacterium bovis, afflicts approximately 50 million cattle worldwide and is detected by the tuberculin skin test (TST). While it has long been recognized that purified protein derivative (PPD) tuberculin is composed of a mixture of M. bovis derived protein components, little is known about the quality, relative quantity and identity of the proteins that make up PPD tuberculin. We manufactured a sterile filtered PPD tuberculin (SF-PPD) from a nine-week-old M. bovis culture supernatant in order to characterise the culture filtrate proteins (CFP) which make up M. bovis PPD tuberculin and to compare the antibody response of M. bovis infected versus M. bovis sensitized cattle. SF-PPD resolved into approximately 200 discrete spots using two-dimensional polyacrylamide gel electrophoresis (2-DE) while fewer than 65 spots could be discerned from 2-DE gels of tuberculin derived from autoclaved culture supernatant. Two dimensional Western blot analyses indicated that sera from M. bovis sensitized cattle recognized additional SF-PPD antigens as compared to M. bovis infected cattle at seven weeks post infection/sensitization. However, application of a comparative tuberculin skin test resulted in an antibody boosting response to the same set of M. bovis CFPs in both the M. bovis infected and M. bovis sensitized cattle. We concluded that it is the heat sterilization of the M. bovis CFPs that causes severe structural changes to the M. bovis proteins. This work suggests that M. bovis infected cattle and cattle artificially sensitized to M. bovis with an injection of heat killed cells exhibit similar antibody responses to M. bovis antigens.

Application of 2-dimensional difference gel electrophoresis (2D-DIGE) to the study of thrombin-activated human platelet secretome.

PubMed

Della Corte, Anna; Maugeri, Norma; Pampuch, Agnieszka; Cerletti, Chiara; de Gaetano, Giovanni; Rotilio, Domenico

2008-02-01

Thrombin is an agonist inducing platelet activation. We combined two-dimensional difference gel electrophoresis (2D-DIGE) and mass spectrometry (MALDI-TOF MS) to analyse differentially expressed proteins secreted from thrombin-stimulated platelets. Human washed platelets, from healthy volunteers, were stimulated with thrombin 0.5 U/ml at 37 degrees C without stirring and the secreted proteins were resolved by 2D-DIGE. By image analysis, 1094 spots were detected in the 2D gel. The spots whose mean intensity showed at least a five-fold change intensity increase or decrease in the thrombin-activated platelet gel in comparison with unstimulated control were digested by trypsin and subjected to MALDI-TOF MS analysis. Peptides from mass spectra of in-gel digest samples were matched against available databases, using the Mascot search engine (Matrix Science) for peptide mass fingerprint. In the activated platelet secretome, transferrin, glutathione-transferase, WD repeat protein, ER-60, thrombospondin-1 precursor and thrombospondin were the most abundant. Also lamin A, a nuclear protein, not previously identified in platelets, appeared to be released. The novel strategy to combine 2D-DIGE with MALDI-TOF MS is a useful approach for a quantitative analysis of the effect of thrombin on the secretome profile of human platelets.

Identifying sites of replication initiation in yeast chromosomes: looking for origins in all the right places.

PubMed

van Brabant, A J; Hunt, S Y; Fangman, W L; Brewer, B J

1998-06-01

DNA fragments that contain an active origin of replication generate bubble-shaped replication intermediates with diverging forks. We describe two methods that use two-dimensional (2-D) agarose gel electrophoresis along with DNA sequence information to identify replication origins in natural and artificial Saccharomyces cerevisiae chromosomes. The first method uses 2-D gels of overlapping DNA fragments to locate an active chromosomal replication origin within a region known to confer autonomous replication on a plasmid. A variant form of 2-D gels can be used to determine the direction of fork movement, and the second method uses this technique to find restriction fragments that are replicated by diverging forks, indicating that a bidirectional replication origin is located between the two fragments. Either of these two methods can be applied to the analysis of any genomic region for which there is DNA sequence information or an adequate restriction map.

Two- and three-dimensional transvaginal ultrasound with power Doppler angiography and gel infusion sonography for diagnosis of endometrial malignancy.

PubMed

Dueholm, M; Christensen, J W; Rydbjerg, S; Hansen, E S; Ørtoft, G

2015-06-01

To evaluate the diagnostic efficiency of two-dimensional (2D) and three-dimensional (3D) transvaginal ultrasonography, power Doppler angiography (PDA) and gel infusion sonography (GIS) at offline analysis for recognition of malignant endometrium compared with real-time evaluation during scanning, and to determine optimal image parameters at 3D analysis. One hundred and sixty-nine consecutive women with postmenopausal bleeding and endometrial thickness ≥ 5 mm underwent systematic evaluation of endometrial pattern on 2D imaging, and 2D videoclips and 3D volumes were later analyzed offline. Histopathological findings at hysteroscopy or hysterectomy were used as the reference standard. The efficiency of the different techniques for diagnosis of malignancy was calculated and compared. 3D image parameters, endometrial volume and 3D vascular indices were assessed. Optimal 3D image parameters were transformed by logistic regression into a risk of endometrial cancer (REC) score, including scores for body mass index, endometrial thickness and endometrial morphology at gray-scale and PDA and GIS. Offline 2D and 3D analysis were equivalent, but had lower diagnostic performance compared with real-time evaluation during scanning. Their diagnostic performance was not markedly improved by the addition of PDA or GIS, but their efficiency was comparable with that of real-time 2D-GIS in offline examinations of good image quality. On logistic regression, the 3D parameters from the REC-score system had the highest diagnostic efficiency. The area under the curve of the REC-score system at 3D-GIS (0.89) was not improved by inclusion of vascular indices or endometrial volume calculations. Real-time evaluation during scanning is most efficient, but offline 2D and 3D analysis is useful for prediction of endometrial cancer when good image quality can be obtained. The diagnostic efficiency at 3D analysis may be improved by use of REC-scoring systems, without the need for calculation of vascular indices or endometrial volume. The optimal imaging modality appears to be real-time 2D-GIS. Copyright © 2014 ISUOG. Published by John Wiley & Sons Ltd.

A gel as an array of channels.

PubMed

Zimm, B H

1996-06-01

We consider the theory of charged point molecules ('probes') being pulled by an electric field through a two-dimensional net of channels that represents a piece of gel. Associated with the position in the net is a free energy of interaction between the probe and the net; this free energy fluctuates randomly with the position of the probe in the net. The free energy is intended to represent weak interactions between the probe and the gel, such as entropy associated with the restriction of the freedom of motion of the probe by the gel, or electrostatic interactions between the probe and charges fixed to the gel. The free energy can be thought of as a surface with the appearance of a rough, hilly landscape spread over the net; the roughness is measured by the standard deviation of the free-energy distribution. Two variations of the model are examined: (1) the net is assumed to have all channels open, or (2) only channels parallel to the electric field are open and all the cross-connecting channels are closed. Model (1) is more realistic but presents a two-dimensional mathematical problem which can only be solved by slow iteration methods, while model (2) is less realistic but presents a one-dimensional problem that can be reduced to simple quadratures and is easy to solve by numerical integration. In both models the mobility of the probe decreases as the roughness parameter is increased, but the effect is larger in the less realistic model (2) if the same free-energy surface is used in both. The mobility in model (2) is reduced both by high points in the rough surface ('bumps') and by low points ('traps'), while in model (1) only the traps are effective, since the probes can flow around the bumps through the cross channels. The mobility in model (2) can be made to agree with model (1) simply by cutting off the bumps of the surface. Thus the simple model (2) can be used in place of the more realistic model (1) that is more difficult to compute.

Evaluation of several two-dimensional gel electrophoresis techniques in cardiac proteomics.

PubMed

Li, Zhao Bo; Flint, Paul W; Boluyt, Marvin O

2005-09-01

Two-dimensional gel electrophoresis (2-DE) is currently the best method for separating complex mixtures of proteins, and its use is gradually becoming more common in cardiac proteome analysis. A number of variations in basic 2-DE have emerged, but their usefulness in analyzing cardiac tissue has not been evaluated. The purpose of the present study was to systematically evaluate the capabilities and limitations of several 2-DE techniques for separating proteins from rat heart tissue. Immobilized pH gradient strips of various pH ranges, parameters of protein loading and staining, subcellular fractionation, and detection of phosphorylated proteins were studied. The results provide guidance for proteome analysis of cardiac and other tissues in terms of selection of the isoelectric point separating window for cardiac proteins, accurate quantitation of cardiac protein abundance, stabilization of technical variation, reduction of sample complexity, enrichment of low-abundant proteins, and detection of phosphorylated proteins.

Separation and identification of Musa acuminate Colla (banana) leaf proteins by two-dimensional gel electrophoresis and mass spectrometry.

PubMed

Lu, Y; Qi, Y X; Zhang, H; Zhang, H Q; Pu, J J; Xie, Y X

2013-12-19

To establish a proteomic reference map of Musa acuminate Colla (banana) leaf, we separated and identified leaf proteins using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and mass spectrometry (MS). Tryptic digests of 44 spots were subjected to peptide mass fingerprinting (PMF) by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS. Three spots that were not identified by MALDI-TOF MS analysis were identified by searching against the NCBInr, SwissProt, and expressed sequence tag (EST) databases. We identified 41 unique proteins. The majority of the identified leaf proteins were found to be involved in energy metabolism. The results indicate that 2D-PAGE is a sensitive and powerful technique for the separation and identification of Musa leaf proteins. A summary of the identified proteins and their putative functions is discussed.

A cytotoxic and apoptosis-inducing sesquiterpenoid isolated from the aerial parts of Artemisia princeps PAMPANINI (Sajabalssuk).

PubMed

Bang, Myun-Ho; Han, Min-Woo; Song, Myoung-Chong; Cho, Jin-Gyeong; Chung, Hae-Gon; Jeong, Tae-Sook; Lee, Kyung-Tae; Choi, Myung-Sook; Kim, Se-Young; Baek, Nam-In

2008-08-01

Repeated silica gel and octadecyl silica gel (ODS) column chromatography of the aerial parts of Artemisia princeps PAMPANINI (Sajabalssuk) led to the isolation of a new sesquiterpenoid, 3-((S)-2-methylbutyryloxy)-costu-1(10),4(5)-dien-12,6 alpha-olide (2), along with two previously reported sesquiterpenoids: 8 alpha-angeloyloxy-3beta,4 beta-epoxy-6 beta H,7 alpha H,8 beta H-guaia-1(10),11(13)-dien-12,6 alpha-olide (1, carlaolide B) and 3beta,4 beta-epoxy-8 alpha-isobutyryloxy-6 beta H,7 alpha H,8 beta H-guaia-1(10),11(13)-dien-12,6 alpha-olide (3, carlaolide A). The structure of compound 2 was elucidated by spectroscopic data analysis, including one dimensional (1D) and two dimensional (2D) nuclear magnetic resonance (NMR) experiments. Of the isolates, compound 2 exhibited potent cytotoxicity against human cervix adenocarcinoma cells and induced apoptosis.

Application of a proteomic approach to identify proteins associated with primary graft non-function after liver transplantation.

PubMed

Kornasiewicz, Oskar; Bojarczuk, Kamil; Bugajski, Marek; Golab, Jakub; Krawczyk, Marek

2012-10-01

Primary graft non-function (PNF) is a rare, life-threatening complication of liver transplantation. Increasing use of extended criteria donor pools and high-risk recipients seem to influence the incidence of PNF. Primary failure is associated with high patient morbidity and inferior graft survival. The only available treatment for PNF is emergency hepatic retransplantation, which is also correlated with significant morbidity and mortality. Therefore, researchers are working to identify risk factors of diagnostic value to prevent PNF. The current study attempted to explore liver proteomic patterns in patients with PNF. Using two-dimensional gel electrophoresis and liquid chromatography-mass spectrometry (LC-MS), we compared liver protein homogenates from 3 patients with PNF to those obtained from 6 healthy liver samples to identify potential new biomarkers of PNF. Our comparisons revealed 21 proteins with differential expression (13 upregulated and 8 downregulated). Most of these proteins are involved in energy metabolism, lipid metabolism, peptide cleavage, cell differentiation, and apoptosis. Although none of these proteins appeared more than once in separate analyses, this preliminary study shows that two-dimensional gel electrophoresis and LC-MS may allow identification of characteristic proteins to be used as biomarkers of a life-threatening complication of liver transplantation. Larger-scale analyses could improve patient care by finding suitable prognostic and therapeutic options. These data represent the first global proteomic approach to study PNF.

Gangliosides During Tumor Progression in Patients With Prostate Cancer

DTIC Science & Technology

2004-07-01

plates (10xlO cm) precoated with Silica Gel 60 ( glass or aluminium backing) (E. Merck, Darmstadt, Germany) were used. Two dimensional HPTLC was...Investigation, 15 (1997) 491-499. 16 16. T. Shiraishi, M. T. Kinter, S. E. Mills, M. C. Lippert , G. S. Bova, W. W. Jr. Young, The glycosphingolipids of human...described earlier (21, 22). HPTLC plates (1 Ox1 0 cm) precoated with silica gel 60 ( glass or aluminum backing) (E. Merck, Darmstadt, Germany) were

Analysis of the Liver Soluble Proteome from Bull Terriers Affected with Inherited Lethal Acrodermatitis

PubMed Central

Mouat, Michael F.; Mauldin, Elizabeth A.; Casal, Margret L.

2012-01-01

Lethal acrodermatitis (LAD) is a genetic disease affecting bull terrier dogs. The phenotype is similar to that for acrodermatitis enteropathica in humans, but is currently without treatment. The purpose of the research presented here is to determine the biochemical defects associated with LAD using proteomic methodologies. Two affected (male and female) and one unaffected (male) bull terrier pups were euthanized at 14 weeks of age, their livers dissected and prepared for two-dimensional gel electrophoresis (2DE) and densitometry. Approximately 200 protein spots were observed. The density of the spots within each gel was normalized to the total spot volume of the gel; only those soluble liver protein spots that were consistently different in both of the livers of the affected pups compared to the unaffected pup were excised manually and submitted for MALDI mass spectrometry. Thirteen proteins were identified as differentially expressed in the affected, compared to the unaffected, pups. The proteins were involved in numerous cellular physiological functions, including chaperones, calcium binding, and energy metabolism, as well as being associated with the inflammatory response. Of note were haptoglobin, glutamine synthetase, prohibitin and keratin 10 which exhibited at least a 4-fold level of differential expression. These data represent the first proteomic analysis of this mutation. The differentially expressed proteins that were identified may be key in understanding the etiology of LAD, and may lead to diagnostic tools for its identification within the bull terrier population. PMID:17693109

Modeling the Dynamics of Gel Electrophorresis in the High School Classroom

NASA Astrophysics Data System (ADS)

Saucedo, Skyler R.

2013-01-01

Gel electrophoresis, used by geneticists and forensic experts alike, is an immensely popular technique that utilizes an electric field to separate molecules and proteins by size and charge. At the microscopic level, a dye or complex protein like DNA is passed through agarose, a gelatinous three-dimensional matrix of pores and nano-sized tunnels. When forced through a maze of holes, the molecule unravels, forming a long chain, slithering through the field of pores in a process colloquially coined "reputation." As a result, the smaller molecules travel farther through the gel when compared to molecules of larger molecular weight. This highly effective "molecular sieve" provides consistent data and allows scientists to compare similar sequences of DNA base pairs in a routine fashion.2 When performed at the high school level, gel electrophoresis provides students the opportunity to learn about a contemporary lab technique of great scientific relevance. Doing real science certainly excites students and motivates them to learn more.

Quantitative ultra-fast MRI of HPMC swelling and dissolution.

PubMed

Chen, Ya Ying; Hughes, L P; Gladden, L F; Mantle, M D

2010-08-01

For the first time quantitative Rapid Acquisition with Relaxation Enhancement (RARE) based ultra-fast two-dimensional magnetic resonance imaging has been used to follow the dissolution of hydroxypropylmethyl cellulose (HPMC) in water. Quantitative maps of absolute water concentration, spin-spin relaxation times and water self-diffusion coefficient are obtained at a spatial resolution of 469 microm in less than 3 min each. These maps allow the dynamic development of the medium release rate HPMC/water system to be followed. It is demonstrated that the evolution of the gel layer and, in particular, the gradient in water concentration across it, is significantly different when comparing the quantitative RARE sequence with a standard (nonquantitative) implementation of RARE. The total gel thickness in the axial direction grows faster than that in the radial direction and that the dry core initially expands anisotropically. Additionally, while HPMC absorbs a large amount of water during the dissolution process, the concentration gradient of water within the gel layer is relatively small. For the first time MRI evidence is presented for a transition swollen glassy layer which resides between the outer edge of the dry tablet core and the inner edge of the gel layer. (c) 2010 Wiley-Liss, Inc. and the American Pharmacists Association

Three-dimensional ultrasound and magnetic resonance imaging assessment of cervix and vagina in women with uterine malformations.

PubMed

Bermejo, C; Martínez-Ten, P; Recio, M; Ruiz-López, L; Díaz, D; Illescas, T

2014-03-01

To investigate the accuracy of three-dimensional ultrasound (3D-US) with respect to magnetic resonance imaging (MRI), and compared to clinical examination, in the assessment of cervix and vagina in women with uterine malformations. In this prospective study, 16 patients diagnosed with uterine malformation with cervical involvement underwent 3D-US examination. The acquisition of cervical volumes was transvaginal, with four cases repeated in the peri-ovulation period, while vaginal volumes were acquired by transperineal imaging following filling of the vagina with gel. MRI was performed in 13 patients using endovaginal gel. All cases underwent clinical examination, comprising bimanual gynecological examination and speculoscopy. Diagnostic concordance of each of the methods with the gold standard was calculated. 3D-US cervical examinations revealed 12 cases of duplicate cervix, two of complete septate cervix and two of incomplete septate cervix. Images of the cervical canal in the peri-ovulation period were judged subjectively to be better in quality, but did not lead us to change any diagnosis. 3D-US vaginal examinations revealed four cases with a vaginal dividing wall and two with a blind hemivagina. None of the 3D-US findings contradicted the clinical findings of the cervix; however, clinically we observed two cases with vaginal dividing wall that had not been diagnosed with 3D-US. MRI diagnosed nine cases of duplicate cervix, three of complete septate cervix, one of incomplete septate cervix, five of vaginal dividing wall and two of blind hemivagina. One case diagnosed as complete septate cervix was in fact a duplicate cervix on 3D-US and on clinical examination. Compared with the gold standard, both 3D-US and MRI were highly efficient in the diagnosis of anomalies of the cervix and vagina. The overall diagnostic concordance of 3D-US with clinical examination (kappa, 0.84; 95% CI, 0.62-1) was slightly inferior to that of MRI with clinical examination (kappa, 0.9; 95% CI, 0.72-1), but this difference was not statistically significant. The acquisition of isolated cervical volumes, without including the uterus, defines the extent of the ectocervix and the limits of the cervical canal in uterine malformations. The use of endovaginal gel makes possible the diagnosis of associated vaginal anomalies with 3D-US. Copyright © 2013 ISUOG. Published by John Wiley & Sons Ltd.

Effects of Microgravity on Streptoccoccus Pneumonia

NASA Technical Reports Server (NTRS)

2003-01-01

These gels were obtained by two-dimensional (2D) electrophoresis, in which proteins move different substances through a polyacrylamide gel matrix based on their molecular weight and total charge in an electric field. The gels illustrate principal investigator David Niesel's findings that exposure to modeled microgravity results in some Streptoccoccus Pneumonia's proteins being upregulated and others being downregulated. In 2D protein profiles of whole cell lysates of Streptoccoccus Pneumonia, 6,304 cultured under normal gravity (left), appear to be expressed at higher levels indicated with black circles. Red circles (right) indicate proteins that were grown under modeled microgravity in a high aspect ratio vessel HARV).

Proteomic analysis of albumin and globulin fractions of pea (Pisum sativum L.) seeds.

PubMed

Dziuba, Jerzy; Szerszunowicz, Iwona; Nałęcz, Dorota; Dziuba, Marta

2014-01-01

Proteomic analysis is emerging as a highly useful tool in food research, including studies of food allergies. Two-dimensional gel electrophoresis involving isoelectric focusing and sodium dodecyl sulfate polyacrylamide gel electrophoresis is the most effective method of separating hundreds or even thousands of proteins. In this study, albumin and globulin tractions of pea seeds cv. Ramrod were subjected to proteomic analysis. Selected potentially alergenic proteins were identified based on their molecular weights and isoelectric points. Pea seeds (Pisum sativum L.) cv. Ramrod harvested over a period of two years (Plant Breeding Station in Piaski-Szelejewo) were used in the experiment. The isolated albumins, globulins and legumin and vicilin fractions of globulins were separated by two-dimensional gel electrophoresis. Proteomic images were analysed in the ImageMaster 2D Platinum program with the use of algorithms from the Melanie application. The relative content, isoelectric points and molecular weights were computed for all identified proteins. Electrophoregrams were analysed by matching spot positions from three independent replications. The proteomes of albumins, globulins and legumin and vicilin fractions of globulins produced up to several hundred spots (proteins). Spots most characteristic of a given fraction were identified by computer analysis and spot matching. The albumin proteome accumulated spots of relatively high intensity over a broad range of pi values of ~4.2-8.1 in 3 molecular weight (MW) ranges: I - high molecular-weight albumins with MW of ~50-110 kDa, II - average molecular-weight albumins with MW of ~20-35 kDa, and III - low molecular-weight albumins with MW of ~13-17 kDa. 2D gel electrophoregrams revealed the presence of 81 characteristic spots, including 24 characteristic of legumin and 14 - of vicilin. Two-dimensional gel electrophoresis proved to be a useful tool for identifying pea proteins. Patterns of spots with similar isoelectric points and different molecular weights or spots with different isoelectric points and similar molecular weights play an important role in proteome analysis. The regions characteristic of albumin, globulin and legumin and vicilin fractions of globulin with typical MW and pi values were identified as the results of performed 2D electrophoretic separations of pea proteins. 2D gel electrophoresis of albumins and the vicilin fraction of globulins revealed the presence of 4 and 2 spots, respectively, representing potentially allergenic proteins. They probably corresponded to vicilin fragments synthesized during post-translational modification of the analysed protein.

Two-dimensional electrophoretic analysis of nuclear matrix proteins in human colon adenocarcinoma.

PubMed

Toumpanaki, A; Baltatzis, G E; Gaitanarou, E; Seretis, E; Toumpanakis, C; Aroni, K; Kittas, Christos; Voloudakis-Baltatzis, I E

2009-01-01

The aim of the present study was to observe possible qualitative and quantitative expression differences between nuclear matrix proteins (NMPs) of human colon adenocarcinoma and their mirror biopsies, using the technique of two-dimensional gel electrophoresis, in order to identify the existence of specific NMP fingerprints for colon cancer. Colon tissues were examined ultrastructurally and NMPs were isolated biochemically, by serial extraction of lipids, soluble proteins, DNA, RNA, and intermediate filaments and were separated according to their isoelectric point (pI) and their molecular weight (MW) by high-resolution two-dimensional electrophoresis (2D). By comparing the 2D electropherograms of colon cancer tissues and mirror biopsy tissues we observed qualitative and quantitative expression differences between their NMPs but also a differentiation of NMP composition between the stages of malignancy. Moreover, despite the similarities between mirror biopsy samples, a highlight percentage of exception was observed. Electrophoretic results provided in this study demonstrated that the examined NMPs could be further investigated as potential markers for detection of colorectal cancer in an early stage, for the assessment of the disease progression, as well as useful tools for individual therapy and for preventing a possible recurrence of cancer and metastasis.

Small Angle Neutron Scattering (SANS) Studies on the Structural Evolution of Pyromellitamide Self-assembled Gels

DOE PAGES

Scott, Jamieson; Tong, Katie; William, Hamilton; ...

2014-10-31

The kinetics of aggregation of two pyromellitamide gelators; tetrabutyl- (C4) and tetrahexylpyromellitamide (C6), in deuterated cyclohexane has been investigated by small angle neutron scattering (SANS) for up to six days. The purpose of this study was to improve our understanding of how self-assembled gels are formed. Short-term (< 3 hour) time scales revealed multiple phases with the data for the tetrabutylpyromellitamide C4 indicating one dimensional stacking and aggregation corresponding to a multi-fiber braided cluster arrangement that is about 35 Å in diameter. The corresponding tetrahexylpyromellitamide C6 data suggests that the C6 also forms one-dimensional stacks but that these aggregate tomore » a thicker multi-fiber braided cluster that have a diameter of 61.8 Å. Over a longer period of time, the radius, persistence length and contour length all continue to increase in 6 days after cooling. This data suggests that structural changes in self-assembled gels occur over a period exceeding several days and that fairly subtle changes in the structure (e.g. tail-length) can influence the packing of molecules in self-assembled gels on the single-to-few fiber bundle stage.« less

Small angle neutron scattering (SANS) studies on the structural evolution of pyromellitamide self-assembled gels.

PubMed

Jamieson, Scott A; Tong, Katie W K; Hamilton, William A; He, Lilin; James, Michael; Thordarson, Pall

2014-11-25

The kinetics of aggregation of two pyromellitamide gelators, tetrabutyl- (C4) and tetrahexyl-pyromellitamide (C6), in deuterated cyclohexane has been investigated by small angle neutron scattering (SANS) for up to 6 days. The purpose of this study was to improve our understanding of how self-assembled gels are formed. Short-term (< 3 h) time scales revealed multiple phases with the data for the tetrabutylpyromellitamide C4, indicating one-dimensional stacking and aggregation corresponding to a multifiber braided cluster arrangement that is about 35 Å in diameter. The corresponding tetrahexylpyromellitamide C6 data suggest that the C6 also forms one-dimensional stacks but that these aggregate to a thicker multifiber braided cluster that has a diameter of about 62 Å. Over a longer period of time, the radius, persistence length, and contour length all continue to increase in 6 days after cooling. These data suggest that structural changes in self-assembled gels occur over a period exceeding several days and that fairly subtle changes in the structure (e.g., tail-length) can influence the packing of molecules in self-assembled gels on the single-to-few fiber bundle stage.

Small Angle Neutron Scattering (SANS) Studies on the Structural Evolution of Pyromellitamide Self-assembled Gels

DOE Office of Scientific and Technical Information (OSTI.GOV)

Scott, Jamieson; Tong, Katie; William, Hamilton

The kinetics of aggregation of two pyromellitamide gelators; tetrabutyl- (C4) and tetrahexylpyromellitamide (C6), in deuterated cyclohexane has been investigated by small angle neutron scattering (SANS) for up to six days. The purpose of this study was to improve our understanding of how self-assembled gels are formed. Short-term (< 3 hour) time scales revealed multiple phases with the data for the tetrabutylpyromellitamide C4 indicating one dimensional stacking and aggregation corresponding to a multi-fiber braided cluster arrangement that is about 35 Å in diameter. The corresponding tetrahexylpyromellitamide C6 data suggests that the C6 also forms one-dimensional stacks but that these aggregate tomore » a thicker multi-fiber braided cluster that have a diameter of 61.8 Å. Over a longer period of time, the radius, persistence length and contour length all continue to increase in 6 days after cooling. This data suggests that structural changes in self-assembled gels occur over a period exceeding several days and that fairly subtle changes in the structure (e.g. tail-length) can influence the packing of molecules in self-assembled gels on the single-to-few fiber bundle stage.« less

Therapeutic Potential of Human Adipose-Derived Stem/Stromal Cell Microspheroids Prepared by Three-Dimensional Culture in Non-Cross-Linked Hyaluronic Acid Gel.

PubMed

Mineda, Kazuhide; Feng, Jingwei; Ishimine, Hisako; Takada, Hitomi; Doi, Kentaro; Kuno, Shinichiro; Kinoshita, Kahori; Kanayama, Koji; Kato, Harunosuke; Mashiko, Takanobu; Hashimoto, Ichiro; Nakanishi, Hideki; Kurisaki, Akira; Yoshimura, Kotaro

2015-12-01

Three-dimensional culture of mesenchymal stem/stromal cells for spheroid formation is known to enhance their therapeutic potential for regenerative medicine. Spheroids were prepared by culturing human adipose-derived stem/stromal cells (hASCs) in a non-cross-linked hyaluronic acid (HA) gel and compared with dissociated hASCs and hASC spheroids prepared using a nonadherent dish. Preliminary experiments indicated that a 4% HA gel was the most appropriate for forming hASC spheroids with a relatively consistent size (20-50 µm) within 48 hours. Prepared spheroids were positive for pluripotency markers (NANOG, OCT3/4, and SOX-2), and 40% of the cells were SSEA-3-positive, a marker of the multilineage differentiating stress enduring or Muse cell. In contrast with dissociated ASCs, increased secretion of cytokines such as hepatocyte growth factor was detected in ASC spheroids cultured under hypoxia. On microarray ASC spheroids showed upregulation of some pluripotency markers and downregulation of genes related to the mitotic cell cycle. After ischemia-reperfusion injury to the fat pad in SCID mice, local injection of hASC spheroids promoted tissue repair and reduced the final atrophy (1.6%) compared with that of dissociated hASCs (14.3%) or phosphate-buffered saline (20.3%). Part of the administered hASCs differentiated into vascular endothelial cells. ASC spheroids prepared in a HA gel contain undifferentiated cells with therapeutic potential to promote angiogenesis and tissue regeneration after damage. This study shows the therapeutic value of human adipose-derived stem cell spheroids prepared in hyarulonic acid gel. The spheroids have various benefits as an injectable cellular product and show therapeutic potential to the stem cell-depleted conditions such as diabetic chronic skin ulcer. ©AlphaMed Press.

Identification of Methanococcus Jannaschii Proteins in 2-D Gel Electrophoresis Patterns by Mass Spectrometry

DOE R&D Accomplishments Database

Liang, X.

1998-06-10

The genome of Methanococcus jannaschii has been sequenced completely and has been found to contain approximately 1,770 predicted protein-coding regions. When these coding regions are expressed and how their expression is regulated, however, remain open questions. In this work, mass spectrometry was combined with two-dimensional gel electrophoresis to identify which proteins the genes produce under different growth conditions, and thus investigate the regulation of genes responsible for functions characteristic of this thermophilic representative of the methanogenic Archaea.

Efficient method of protein extraction from Theobroma cacao L. roots for two-dimensional gel electrophoresis and mass spectrometry analyses.

PubMed

Bertolde, F Z; Almeida, A-A F; Silva, F A C; Oliveira, T M; Pirovani, C P

2014-07-04

Theobroma cacao is a woody and recalcitrant plant with a very high level of interfering compounds. Standard protocols for protein extraction were proposed for various types of samples, but the presence of interfering compounds in many samples prevented the isolation of proteins suitable for two-dimensional gel electrophoresis (2-DE). An efficient method to extract root proteins for 2-DE was established to overcome these problems. The main features of this protocol are: i) precipitation with trichloroacetic acid/acetone overnight to prepare the acetone dry powder (ADP), ii) several additional steps of sonication in the ADP preparation and extractions with dense sodium dodecyl sulfate and phenol, and iii) adding two stages of phenol extractions. Proteins were extracted from roots using this new protocol (Method B) and a protocol described in the literature for T. cacao leaves and meristems (Method A). Using these methods, we obtained a protein yield of about 0.7 and 2.5 mg per 1.0 g lyophilized root, and a total of 60 and 400 spots could be separated, respectively. Through Method B, it was possible to isolate high-quality protein and a high yield of roots from T. cacao for high-quality 2-DE gels. To demonstrate the quality of the extracted proteins from roots of T. cacao using Method B, several protein spots were cut from the 2-DE gels, analyzed by tandem mass spectrometry, and identified. Method B was further tested on Citrus roots, with a protein yield of about 2.7 mg per 1.0 g lyophilized root and 800 detected spots.

Optimization of Large Gel 2D Electrophoresis for Proteomic Studies of Skeletal Muscle

PubMed Central

Reed, Patrick W.; Densmore, Allison; Bloch, Robert J.

2013-01-01

We describe improved methods for large format, 2-dimensional gel electrophoresis (2-DE) that improve protein solubility and recovery, minimize proteolysis, and reduce the loss of resolution due to contaminants and manipulations of the gels, and thus enhance quantitative analysis of protein spots. Key modifications are: (i) the use of 7M urea + 2 M thiourea, instead of 9M urea, in sample preparation and in the tops of the gel tubes; (ii) standardized deionization of all solutions containing urea with a mixed bed ion exchange resin and removal of urea from the electrode solutions; and (iii) use of a new gel tank and cooling device that eliminate the need to run two separating gels in the SDS dimension. These changes make 2D-GE analysis more reproducible and sensitive, with minimal artifacts. Application of this method to the soluble fraction of muscle tissues reliably resolves ~1800 protein spots in adult human skeletal muscle and over 2800 spots in myotubes. PMID:22589104

X-Ray Reflectometry of DMPS Monolayers on a Water Substrate

NASA Astrophysics Data System (ADS)

Tikhonov, A. M.; Asadchikov, V. E.; Volkov, Yu. O.; Roshchin, B. S.; Ermakov, Yu. A.

2017-12-01

The molecular structure of dimyristoyl phosphatidylserine (DMPS) monolayers on a water substrate in different phase states has been investigated by X-ray reflectometry with a photon energy of 8 keV. According to the experimental data, the transition from a two-dimensional expanded liquid state to a solid gel state (liquid crystal) accompanied by the ordering of the hydrocarbon tails C14H27 of the DMPS molecule occurs in the monolayer as the surface pressure rises. The monolayer thickness is 20 ± 3 and 28 ± 2 Å in the liquid and solid phases, respectively, with the deflection angle of the molecular tail axis from the normal to the surface in the gel phase being 26° ± 8°. At least a twofold decrease in the degree of hydration of the polar lipid groups also occurs under two-dimensional monolayer compression. The reflectometry data have been analyzed using two approaches: under the assumption about the presence of two layers with different electron densities in the monolayer and without any assumptions about the transverse surface structure. Both approaches demonstrate satisfactory agreement between themselves in describing the experimental results.

Multiscale Mechanical Characterization of Biomimetic Physically Associating Gels

DTIC Science & Technology

2006-09-01

work, a ballistic gelatin and two styrene- isoprene triblock copolymer gels are tested and compared using both macroscale and microscale measurements... isoprene triblock copolymer gels are tested and compared using both macroscale and microscale measurements. A methodology is presented to conduct

Coordination polymer gels with important environmental and biological applications.

PubMed

Jung, Jong Hwa; Lee, Ji Ha; Silverman, Julian R; John, George

2013-02-07

Coordination Polymer Gels (CPGs) constitute a subset of solid-like metal ion and bridging organic ligand structures (similar to metal-organic frameworks) that form multi-dimensional networks through a trapped solvent as a result of non-covalent interactions. While physical properties of these gels are similar to conventional high molecular weight organic polymer gels, coordination polymer gel systems are often fully reversible and can be assembled and disassembled in the presence of additional energy (heat, sonication, shaking) to give a solution of solvated gelators. Compared to gels resulting from purely organic self-assembled low molecular weight gelators, metal ions incorporated into the fibrilar networks spanning the bulk solvent can impart CPGs with added functionalities. The solid/liquid nature of the gels allows for species to migrate through the gel system and interact with metals, ligands, and the solvent. Chemosensing, catalysis, fluorescence, and drug-delivery applications are some of the many potential uses for these dynamic systems, taking advantage of the metal ion's coordination, the organic polydentate ligand's orientation and functionality, or a combination of these properties. By fine tuning these systems through metal ion and ligand selection and by directing self-assembly with external stimuli the rational synthesis of practical systems can be envisaged.

Platelet-Rich Plasma Gel Combined with Bovine-Derived Xenograft for the Treatment of Dehiscence Around Immediately Placed Conventionally Loaded Dental Implants in Humans: Cone Beam Computed Tomography and Three-Dimensional Image Evaluation.

PubMed

ArRejaie, Aws; Al-Harbi, Fahad; Alagl, Adel S; Hassan, Khalid S

2016-01-01

This study clinically and radiographically investigated the potential of platelet-rich plasma (PRP) gel combined with bovine-derived xenograft to treat dehiscence defects around immediate dental implants. This study was performed on 32 sites from 16 patients who each received an immediate implant for a single tooth replacement at a maxillary anterior or premolar site. Patients were divided into two groups according to the augmented materials used. One group received an immediate implant and filling of defects using a PRP gel plus bovine-derived xenograft. The other group received an immediate implant and filling of defects with a bovine-derived xenograft without PRP gel. Cone beam computed tomography (CBCT) was taken before placement, and at 6 and 12 months postsurgery. Both treatment procedures resulted in significant improvements for the primary outcome regarding bone fill, as well as the marginal bone level. In addition, statistically significant differences were found in the bone density for the combined therapy compared with sites treated with bovine-derived xenografts alone (P ≤ .01). Autogenous PRP gel combined with bovine-derived xenograft demonstrated superiority to the bovine-derived xenograft alone, which suggested that it could be successfully applicable for the treatment of dehiscence around an immediate dental implant. Moreover, CBCT can be used to measure dehiscence and to assess bone thickness along the implant.

Detecting peroxiredoxin hyperoxidation by one-dimensional isoelectric focusing.

PubMed

Cao, Zhenbo; Bulleid, Neil J

The activity of typical 2-cys peroxiredoxin (Prxs) can be regulated by hyperoxidation with a consequent loss of redox activity. Here we developed a simple assay to monitor the level of hyperoxidation of different typical 2-cys prxs simultaneously. This assay only requires standard equipment and can compare different samples on the same gel. It requires much less time than conventional 2D gels and gives more information than Western blotting with an antibody specific for hyperoxidized peroxiredoxin. This method could also be used to monitor protein modification with a charge difference such as phosphorylation.

[Proteomic analysis of myocardial hypertrophy induced by left kidney artery coarctation in rats].

PubMed

Lv, Yuan-yuan; Sun, Biao; Ma, Ji-zheng

2009-05-01

To identify the expression of proteins in cardiomyocytes in rats with left kidney artery coarctation. 16 male SD rats were separated into 2 groups (n=8): 2 kidney 1 Clip group (2K1C) and sham operation group (SO). The postoperational 8th week, after examination by normal doppler and tissue doppler echocardiography, the extracted proteins from cardiomyocytes were isolated by two-dimensional gel electrophoresis with staining. The gel images were acquired by scanner and 2-DE analysis software. Different spots observed on two 2D gels were selected and identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Overall, 21 protein spots showed significant difference, and 14 out of which were identified. Kidney artery coactation-induced cardiac hypertrophy displays different expression of proteins in cardiomyocytes.

Evaluation of dose delivery accuracy of gamma knife using MRI polymer gel dosimeter in an inhomogeneous phantom

NASA Astrophysics Data System (ADS)

Pourfallah T, A.; Alam N, Riahi; M, Allahverdi; M, Ay; M, Zahmatkesh

2009-05-01

Polymer gel dosimetry is still the only dosimetry method for directly measuring three-dimensional dose distributions. MRI Polymer gel dosimeters are tissue equivalent and can act as a phantom material. Because of high dose response sensitivity, the MRI was chosen as readout device. In this study dose profiles calculated with treatment-planning software (LGP) and measurements with the MR polymer gel dosimeter for single-shot irradiations were compared. A custom-built 16 cm diameter spherical plexiglas head phantom was used in this study. Inside the phantom, there is a cubic cutout for insertion of gel phantoms and another cutout for inserting the inhomogeneities. The phantoms were scanned with a 1.5T MRI (Siemens syngo MR 2004A 4VA25A) scanner. The multiple spin-echo sequence with 32 echoes was used for the MRI scans. Calibration relations between the spin-spin relaxation rate and the absorbed dose were obtained by using small cylindrical vials, which were filled with the PAGAT polymer gel from the same batch as for the spherical phantom. 1D and 2D data obtained using gel dosimeter for homogeneous and inhomogeneous phantoms were compared with dose obtained using LGP calculation. The distance between relative isodose curves obtained for homogeneous phantom and heterogeneous phantoms exceed the accepted total positioning error (>±2mm). The findings of this study indicate that dose measurement using PAGAT gel dosimeter can be used for verifying dose delivering accuracy in GK unit in presence of inhomogeneities.

Identification of Fur, Aconitase, and Other Proteins Expressed by Mycobacterium tuberculosis under Conditions of Low and High Concentrations of Iron by Combined Two-Dimensional Gel Electrophoresis and Mass Spectrometry

PubMed Central

Wong, Diane K.; Lee, Bai-Yu; Horwitz, Marcus A.; Gibson, Bradford W.

1999-01-01

Iron plays a critical role in the pathophysiology of Mycobacterium tuberculosis. To gain a better understanding of iron regulation by this organism, we have used two-dimensional (2-D) gel electrophoresis, mass spectrometry, and database searching to study protein expression in M. tuberculosis under conditions of high and low iron concentration. Proteins in cellular extracts from M. tuberculosis Erdman strain grown under low-iron (1 μM) and high-iron (70 μM) conditions were separated by 2-D polyacrylamide gel electrophoresis, which allowed high-resolution separation of several hundred proteins, as visualized by Coomassie staining. The expression of at least 15 proteins was induced, and the expression of at least 12 proteins was decreased under low-iron conditions. In-gel trypsin digestion was performed on these differentially expressed proteins, and the digestion mixtures were analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry to determine the molecular masses of the resulting tryptic peptides. Partial sequence data on some of the peptides were obtained by using after source decay and/or collision-induced dissociation. The fragmentation data were used to search computerized peptide mass and protein sequence databases for known proteins. Ten iron-regulated proteins were identified, including Fur and aconitase proteins, both of which are known to be regulated by iron in other bacterial systems. Our study shows that, where large protein sequence databases are available from genomic studies, the combined use of 2-D gel electrophoresis, mass spectrometry, and database searching to analyze proteins expressed under defined environmental conditions is a powerful tool for identifying expressed proteins and their physiologic relevance. PMID:9864233

Digital-Micromirror-Device Projection Printing System for Meniscus Tissue Engineering

PubMed Central

Grogan, Shawn P; Chung, Peter H; Soman, Pranav; Chen, Peter; Lotz, Martin K; Chen, Shaochen; D’Lima, Darryl D

2013-01-01

Meniscus degeneration due to age or injury can lead to osteoarthritis. Though promising, current cell-based approaches show limited success. Here we present three-dimensional methacrylated gelatin (GelMA) scaffolds patterned via projection stereolithography to emulate the circumferential alignment of cells in native meniscus tissue. Cultured human avascular zone meniscus cells from normal meniscus were seeded on the scaffolds. Cell viability was monitored, and neo-tissue formation was assessed by gene expression analysis and histology after two weeks in serum free culture with TGFβ1 (10ng/ml). Light, confocal and scanning electron microscopy was used to observe cell/GelMA interactions. Tensile mechanical testing was performed on unseeded, fresh scaffolds and two-week old cell-seeded and unseeded scaffolds. Two-week old cell/GelMA constructs were implanted into surgically created meniscus defects in an explant organ culture model. No cytotoxic effects were observed three weeks after implantation, and cells grew and aligned to the patterned GelMA strands. Gene expression profiles and histology indicated promotion of a fibrocartilage-like meniscus phenotype, and scaffold integration with repair tissue was observed in the explant model. We show that micropatterned GelMA scaffolds are non-toxic, produce organized cellular alignment, and promote meniscus-like tissue formation. Prefabrication of GelMA scaffolds with architectures mimicking meniscus collagen bundle organization shows promise for meniscal repair. Furthermore, the technique presented may be scaled to repair larger defects. PMID:23523536

Comparative proteome analysis of monolayer and spheroid culture of canine osteosarcoma cells.

PubMed

Gebhard, Christiane; Miller, Ingrid; Hummel, Karin; Neschi Née Ondrovics, Martina; Schlosser, Sarah; Walter, Ingrid

2018-04-15

Osteosarcoma is an aggressive bone tumor with high metastasis rate in the lungs and affects both humans and dogs in a similar way. Three-dimensional tumor cell cultures mimic the in vivo situation of micro-tumors and metastases and are therefore better experimental in vitro models than the often applied two-dimensional monolayer cultures. The aim of the present study was to perform comparative proteomics of standard monolayer cultures of canine osteosarcoma cells (D17) and three-dimensional spheroid cultures, to better characterize the 3D model before starting with experiments like migration assays. Using DIGE in combination with MALDI-TOF/TOF we found 27 unique canine proteins differently represented between these two culture systems, most of them being part of a functional network including mainly chaperones, structural proteins, stress-related proteins, proteins of the glycolysis/gluconeogenesis pathway and oxidoreductases. In monolayer cells, a noticeable shift to more acidic pI values was noticed for several proteins of medium to high abundance; two proteins (protein disulfide isomerase A3, stress-induced-phosphoprotein 1) showed an increase of phosphorylated protein species. Protein distribution within the cells, as detected by immunohistochemistry, displayed a switch of stress-induced-phosphoprotein 1 from the cytoplasm (in monolayer cultures) to the nucleus (in spheroid cultures). Additionally, Western blot testing revealed upregulated concentrations of metastasin (S100A4), triosephosphate isomerase 1 and septin 2 in spheroid cultures, in contrast to decreased concentrations of CCT2, a subunit of the T-complex. Results indicate regulation of stress proteins in the process of three-dimensional organization characterized by a hypoxic and nutrient-deficient environment comparable to tumor micro-metastases. Osteosarcoma is an aggressive bone tumor that early spreads to the lungs. Three-dimensional tumor cell cultures represent the avascular stage of micro-tumors and metastases, and should therefore represent a better experimental in vitro model compared to two-dimensional monolayer cultures. Significant differences have been reported in response to drug and radiation treatment between these two culture systems. A gel-based proteomic investigation was performed to compare protein patterns of a canine osteosarcoma cell line cultivated under those two conditions, to learn more about altered cell composition and its impact on cell behaviour. Due to the fact that the canine osteosarcoma is an accepted model for the human disease, results will be relevant for the human species as well. Copyright © 2018 Elsevier B.V. All rights reserved.

Neural stem cell proliferation and differentiation in the conductive PEDOT-HA/Cs/Gel scaffold for neural tissue engineering.

PubMed

Wang, Shuping; Guan, Shui; Xu, Jianqiang; Li, Wenfang; Ge, Dan; Sun, Changkai; Liu, Tianqing; Ma, Xuehu

2017-09-26

Engineering scaffolds with excellent electro-activity is increasingly important in tissue engineering and regenerative medicine. Herein, conductive poly(3,4-ethylenedioxythiophene) doped with hyaluronic acid (PEDOT-HA) nanoparticles were firstly synthesized via chemical oxidant polymerization. A three-dimensional (3D) PEDOT-HA/Cs/Gel scaffold was then developed by introducing PEDOT-HA nanoparticles into a chitosan/gelatin (Cs/Gel) matrix. HA, as a bridge, not only was used as a dopant, but also combined PEDOT into the Cs/Gel via chemical crosslinking. The PEDOT-HA/Cs/Gel scaffold was used as a conductive substrate for neural stem cell (NSC) culture in vitro. The results demonstrated that the PEDOT-HA/Cs/Gel scaffold had excellent biocompatibility for NSC proliferation and differentiation. 3D confocal fluorescence images showed cells attached on the channel surface of Cs/Gel and PEDOT-HA/Cs/Gel scaffolds with a normal neuronal morphology. Compared to the Cs/Gel scaffold, the PEDOT-HA/Cs/Gel scaffold not only promoted NSC proliferation with up-regulated expression of Ki67, but also enhanced NSC differentiation into neurons and astrocytes with up-regulated expression of β tubulin-III and GFAP, respectively. It is expected that this electro-active and bio-active PEDOT-HA/Cs/Gel scaffold will be used as a conductive platform to regulate NSC behavior for neural tissue engineering.

Comparative efficacy and safety of two 0.025% tretinoin gels: results from a multicenter double-blind, parallel study.

PubMed

Lucky, A W; Cullen, S I; Jarratt, M T; Quigley, J W

1998-04-01

The addition of polyolprepolymer-2 in tretinoin formulations may reduce tretinoin-induced cutaneous irritation. This study compared the efficacy and safety of a new 0.025% tretinoin gel containing polyolprepolymer-2, its vehicle, and a commercially-available 0.025% tretinoin gel in patients with mild to moderate acne vulgaris. In this 12-week multicenter, double-blind, parallel group study, efficacy was evaluated by objective lesion counts and the investigators' global evaluations. Subjective assessment of cutaneous irritation by the investigators and patients evaluated safety. The efficacy of the two active treatments in this 215 patient study was comparable, and both treatments were statistically significantly more effective than vehicle. When compared with the commercially-available tretinoin gel, the formulation containing polyolprepolymer-2 demonstrated statistically significantly less peeling at days 28, 56, and 84, statistically significantly less dryness by day 84, and statistically significantly less itching at day 14. Irritation scores for the formulation containing polyolprepolymer-2 were numerically lower but not statistically different from those of the commercially-available gel for erythema and burning. The number of cutaneous and noncutaneous adverse events were similar for both active medications. The two 0.025% gels studied demonstrated comparable efficacy. However, the gel formulation containing polyolprepolymer-2 caused significantly less peeling and drying than the commercially-available formulation by day 84 of the study.

Drying Affects the Fiber Network in Low Molecular Weight Hydrogels

PubMed Central

2017-01-01

Low molecular weight gels are formed by the self-assembly of a suitable small molecule gelator into a three-dimensional network of fibrous structures. The gel properties are determined by the fiber structures, the number and type of cross-links and the distribution of the fibers and cross-links in space. Probing these structures and cross-links is difficult. Many reports rely on microscopy of dried gels (xerogels), where the solvent is removed prior to imaging. The assumption is made that this has little effect on the structures, but it is not clear that this assumption is always (or ever) valid. Here, we use small angle neutron scattering (SANS) to probe low molecular weight hydrogels formed by the self-assembly of dipeptides. We compare scattering data for wet and dried gels, as well as following the drying process. We show that the assumption that drying does not affect the network is not always correct. PMID:28631478

A modified method using the SonoPrep ultrasonic skin permeation system for sampling human interstitial fluid is compatible with proteomic techniques.

PubMed

Lecomte, Marie M J; Atkinson, Kelly R; Kay, Daniel P; Simons, Joanne L; Ingram, John R

2013-02-01

The use of biomarkers in skin is a novel diagnostic tool. Interstitial fluid (ISF) from skin provides a snapshot of proteins secreted at the time of sampling giving insights into the patient's health status. A minimally invasive technique for the transdermal collection of human ISF proteins. A low frequency ultrasonic skin permeation device (SonoPrep ultrasonic skin permeation system) was used to produce micropores in the stratum corneum through which ISF was extracted using a portable pulsed vacuum ISF collection device. On average, protein concentrations recovered ranged between 0.064 and 4.792 μg/μL (mean 1.258 μg/μL). Two-dimensional gel electrophoresis revealed that this sample type was amenable to this type of analysis. Gel images indicated that both highly abundant proteins and lower abundance proteins were isolated from the skin. Western blot analysis confirmed the presence of proteins commonly found in plasma and the epidermis. A minimally invasive method for the transdermal recovery of ISF proteins has been developed. We have demonstrated that ISF samples obtained using this approach can be analysed with proteomic techniques, such as two-dimensional gel electrophoresis and western blots, providing another tool for the identification of disease specific protein biomarkers. © 2012 John Wiley & Sons A/S.

Sandwiched polymer fibre in fibrin matrices for the dictation of endothelial cells undergoing angiogenesis

NASA Astrophysics Data System (ADS)

Sukmana, I.; Djuansjah, J. R. P.

2013-04-01

We present here a three-dimensional (3D) sandwich system made by poly(ethylene terephthalate) (PET) fibre and fibrin extracellular matrix (ECM) for endothelial cell dictation and angiogenesis guidance. In this three-dimensional system, Human Umbilical Vein Endothelial cells (HUVECs) were firstly cultured for 2 (two) days to cover the PET fibre before sandwiched in two layer fibrin gel containing HUVECs. After 4 (four) days of culture, cel-to-cel connection, tube-like structure and multi-cellular lumen formation were then assessed and validated. Phase contrast and fluorescence imaging using an inverted microscope were used to determine cell-to-cell and cell-ECM interactions. Laser scanning confocal microscopy and histological techniques were used to confirm the development of tube-like structure and multi-cellular lumen formation. This study shows that polymer fibres sandwiched in fibrin gel can be used to dictate endothelial cells undergoing angiogenesis with potential application in cancer and cardiovascular study and tissue engineering vascularisation.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Okita, T.W.; Nakata, P.A.; Anderson, J.M.

ADPglucose pyrophosphorylase has been extensively purified from potato (Solanum tuberosum L.) tuber tissue to study its structure. By employing a modified published procedure together with Mono Q chromatography, a near homogeneous enzyme preparation was obtained with substantial improvement in enzyme yield and specific activity. In single dimensional sodium dodecyl sulfate polyacrylamide gels, the enzyme migrated as a single polypeptide band with a mobility of about 50,000 daltons. Analysis by two-dimensional polyacrylamide gel electrophoresis, however, revealed the presence of two types of subunits which could be distinguished by their slight differences in net charge and molecular weight. The smaller potato tubermore » subunit was recognized by antiserum prepared against the smaller spinach leaf 51 kilodalton ADPglucose pyrophosphorylase subunit. In contrast, the anti-54 kilodalton raised against the spinach leaf subunit did not significantly react to the tuber enzyme subunits. The results are consistent with the hypothesis that the potato tuber ADPglucose pyrophosphorylase is not composed of a simple homotetramer as previously suggested, but is a product of two separate and distinct subunits as observed for the spinach leaf and maize enzymes.« less

Engineering fibrin hydrogels to promote the wound healing potential of mesenchymal stem cell spheroids.

PubMed

Murphy, Kaitlin C; Whitehead, Jacklyn; Zhou, Dejie; Ho, Steve S; Leach, J Kent

2017-12-01

Mesenchymal stem cells (MSCs) secrete endogenous factors such as vascular endothelial growth factor (VEGF) and prostaglandin E2 (PGE 2 ) that promote angiogenesis, modulate the inflammatory microenvironment, and stimulate wound repair, and MSC spheroids secrete more trophic factors than dissociated, individual MSCs. Compared to injection of cells alone, transplantation of MSCs in a biomaterial can enhance their wound healing potential by localizing cells at the defect site and upregulating trophic factor secretion. To capitalize on the therapeutic potential of spheroids, we engineered a fibrin gel delivery vehicle to simultaneously enhance the proangiogenic and anti-inflammatory potential of entrapped human MSC spheroids. We used multifactorial statistical analysis to determine the interaction between four input variables derived from fibrin gel synthesis on four output variables (gel stiffness, gel contraction, and secretion of VEGF and PGE 2 ). Manipulation of the four input variables tuned fibrin gel biophysical properties to promote the simultaneous secretion of VEGF and PGE 2 by entrapped MSC spheroids while maintaining overall gel integrity. MSC spheroids in stiffer gels secreted the most VEGF, while PGE 2 secretion was highest in more compliant gels. Simultaneous VEGF and PGE 2 secretion was greatest using hydrogels with intermediate mechanical properties, as small increases in stiffness increased VEGF secretion while maintaining PGE 2 secretion by entrapped spheroids. The fibrin gel formulation predicted to simultaneously increase VEGF and PGE 2 secretion stimulated endothelial cell proliferation, enhanced macrophage polarization, and promoted angiogenesis when used to treat a wounded three-dimensional human skin equivalent. These data demonstrate that a statistical approach is an effective strategy to formulate fibrin gel formulations that enhance the wound healing potential of human MSCs. Mesenchymal stem cells (MSCs) are under investigation for wound healing applications due to their secretion of bioactive factors that enhance granulation tissue formation, blood vessel ingrowth, and reduce inflammation. However, the effectiveness of cell-based therapies is reduced due to poor engraftment and high rates of cell death when transplanted into harsh environments characteristic of large wounds. Compared to dissociated cells, MSCs exhibit increased overall function when aggregated into three-dimensional spheroids, and transplantation of cells using biomaterials is one strategy for guiding cell function in the defect site. The present study demonstrates that the biophysical properties of fibrin hydrogels, designed for use as a cell carrier, can be engineered to dictate the secretion of bioactive factors by entrapped MSC spheroids. This strategy enables MSCs to contribute to wound healing by synergistically promoting neovascularization and modulating the inflammatory milieu. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

The Goat (Capra hircus) Mammary Gland Mitochondrial Proteome: A Study on the Effect of Weight Loss Using Blue-Native PAGE and Two-Dimensional Gel Electrophoresis.

PubMed

Cugno, Graziano; Parreira, José R; Ferlizza, Enea; Hernández-Castellano, Lorenzo E; Carneiro, Mariana; Renaut, Jenny; Castro, Noemí; Arguello, Anastasio; Capote, Juan; Campos, Alexandre M O; Almeida, André M

2016-01-01

Seasonal weight loss (SWL) is the most important limitation to animal production in the Tropical and Mediterranean regions, conditioning producer's incomes and the nutritional status of rural communities. It is of importance to produce strategies to oppose adverse effects of SWL. Breeds that have evolved in harsh climates have acquired tolerance to SWL through selection. Most of the factors determining such ability are related to changes in biochemical pathways as affected by SWL. In this study, a gel based proteomics strategy (BN: Blue-Native Page and 2DE: Two-dimensional gel electrophoresis) was used to characterize the mitochondrial proteome of the secretory tissue of the goat mammary gland. In addition, we have conducted an investigation of the effects of weight loss in two goat breeds with different levels of adaptation to nutritional stress: Majorera (tolerant) and Palmera (susceptible). The study used Majorera and Palmera dairy goats, divided in 4 sets, 2 for each breed: underfed group fed on wheat straw (restricted diet, so their body weight would be 15-20% reduced by the end of experiment), and a control group fed with an energy-balanced diet. At the end of the experimental period (22 days), mammary gland biopsies were obtained for all experimental groups. The proteomic analysis of the mitochondria enabled the resolution of a total of 277 proteins, and 148 (53%) were identified by MALDI-TOF/TOF mass spectrometry. Some of the proteins were identified as subunits of the glutamate dehydrogenase complex and the respiratory complexes I, II, IV, V from mitochondria, as well as numerous other proteins with functions in: metabolism, development, localization, cellular organization and biogenesis, biological regulation, response to stimulus, among others, that were mapped in both BN and 2DE gels. The comparative proteomics analysis enabled the identification of several proteins: NADH-ubiquinone oxidoreductase 75 kDa subunit and lamin B1 mitochondrial (up-regulated in the Palmera breed), Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-2 (up-regulated in the Majorera breed) and cytochrome b-c1 complex subunit 1, mitochondrial and Chain D, Bovine F1-C8 Sub-Complex Of Atp Synthase (down-regulated in the Majorera breed) as a consequence of weight loss.

The Goat (Capra hircus) Mammary Gland Mitochondrial Proteome: A Study on the Effect of Weight Loss Using Blue-Native PAGE and Two-Dimensional Gel Electrophoresis

PubMed Central

Cugno, Graziano; Parreira, José R.; Ferlizza, Enea; Hernández-Castellano, Lorenzo E.; Carneiro, Mariana; Renaut, Jenny; Castro, Noemí; Arguello, Anastasio; Capote, Juan

2016-01-01

Seasonal weight loss (SWL) is the most important limitation to animal production in the Tropical and Mediterranean regions, conditioning producer’s incomes and the nutritional status of rural communities. It is of importance to produce strategies to oppose adverse effects of SWL. Breeds that have evolved in harsh climates have acquired tolerance to SWL through selection. Most of the factors determining such ability are related to changes in biochemical pathways as affected by SWL. In this study, a gel based proteomics strategy (BN: Blue-Native Page and 2DE: Two-dimensional gel electrophoresis) was used to characterize the mitochondrial proteome of the secretory tissue of the goat mammary gland. In addition, we have conducted an investigation of the effects of weight loss in two goat breeds with different levels of adaptation to nutritional stress: Majorera (tolerant) and Palmera (susceptible). The study used Majorera and Palmera dairy goats, divided in 4 sets, 2 for each breed: underfed group fed on wheat straw (restricted diet, so their body weight would be 15–20% reduced by the end of experiment), and a control group fed with an energy-balanced diet. At the end of the experimental period (22 days), mammary gland biopsies were obtained for all experimental groups. The proteomic analysis of the mitochondria enabled the resolution of a total of 277 proteins, and 148 (53%) were identified by MALDI-TOF/TOF mass spectrometry. Some of the proteins were identified as subunits of the glutamate dehydrogenase complex and the respiratory complexes I, II, IV, V from mitochondria, as well as numerous other proteins with functions in: metabolism, development, localization, cellular organization and biogenesis, biological regulation, response to stimulus, among others, that were mapped in both BN and 2DE gels. The comparative proteomics analysis enabled the identification of several proteins: NADH-ubiquinone oxidoreductase 75 kDa subunit and lamin B1 mitochondrial (up-regulated in the Palmera breed), Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-2 (up-regulated in the Majorera breed) and cytochrome b-c1 complex subunit 1, mitochondrial and Chain D, Bovine F1-C8 Sub-Complex Of Atp Synthase (down-regulated in the Majorera breed) as a consequence of weight loss. PMID:27031334

Fabrication and characterization of hexagonally patterned quasi-1D ZnO nanowire arrays

PubMed Central

2014-01-01

Quasi-one-dimensional (quasi-1D) ZnO nanowire arrays with hexagonal pattern have been successfully synthesized via the vapor transport process without any metal catalyst. By utilizing polystyrene microsphere self-assembled monolayer, sol–gel-derived ZnO thin films were used as the periodic nucleation sites for the growth of ZnO nanowires. High-quality quasi-1D ZnO nanowires were grown from nucleation sites, and the original hexagonal periodicity is well-preserved. According to the experimental results, the vapor transport solid condensation mechanism was proposed, in which the sol–gel-derived ZnO film acting as a seed layer for nucleation. This simple method provides a favorable way to form quasi-1D ZnO nanostructures applicable to diverse fields such as two-dimensional photonic crystal, nanolaser, sensor arrays, and other optoelectronic devices. PMID:24521308

Gel compression considerations for chromatography scale-up for protein C purification.

PubMed

He, W; Bruley, D F; Drohan, W N

1998-01-01

This work is to establish theoretical and experimental relationships for the scale-up of Immobilized Metal Affinity Chromatography (IMAC) and Immuno Affinity Chromatography for the low cost production of large quantities of Protein C. The external customer requirements for this project have been established for Protein C deficient people with the goal of providing prophylactic patient treatment. Deep vein thrombosis is the major symptom for protein C deficiency creating the potential problem of embolism transport to important organs, such as, lung and brain. Gel matrices for protein C separation are being analyzed to determine the relationship between the material properties of the gel and the column collapse characteristics. The fluid flow rate and pressure drop is being examined to see how they influence column stability. Gel packing analysis includes two considerations; one is bulk compression due to flow rate, and the second is gel particle deformation due to fluid flow and pressure drop. Based on the assumption of creeping flow, Darcy's law is being applied to characterize the flow through the gel particles. Biot's mathematical description of three-dimensional consolidation in porous media is being used to develop a set of system equations. Finite difference methods are being utilized to obtain the equation solutions. In addition, special programs such as finite element approaches, ABAQUS, will be studied to determine their application to this particular problem. Experimental studies are being performed to determine flow rate and pressure drop correlation for the chromatographic columns with appropriate gels. Void fraction is being measured using pulse testing to allow Reynolds number calculations. Experimental yield stress is being measured to compare with the theoretical calculations. Total Quality Management (TQM) tools have been utilized to optimize this work. For instance, the "Scatter Diagram" has been used to evaluate and select the appropriate gels and operating conditions via Taguchi techniques. Targeting customer requirements under the structure of TQM represents a novel approach to graduate student research in an academic institution which is designed to simulate an industrial environment.

Comparison of camel tear proteins between summer and winter

PubMed Central

Chen, Ziyan; Shamsi, Farrukh A.; Li, Kaijun; Huang, Qiang; Al-Rajhi, Ali A.; Chaudhry, Imtiaz A.

2011-01-01

Purpose Proteins in the tear fluid have positive effects on maintaining the integrity and stabilization of the tear film, which is affected by several environmental factors. The aim of this study is to investigate seasonal variation of protein patterns in camel tears collected during the summer and winter season. Methods Tears from both eyes of 50 clinically normal camels (Camelus dromedarius) were collected in the summer (June – July) and in the winter (December – January) respectively. Pooled tear protein samples from two seasons were separated by SDS–PAGE and two-dimensional electrophoresis (2-DE). Protein spots of differential expression in two season gels were excised and subjected to in-gel digestion and identification by matrix assisted laser desorption/ionization-time of flight/time of flight-mass spectrum (MALDI-TOF/TOF-MS) analysis. Two differentially expressed proteins, lactoferrin (LF) and vitelline membrane outer layer protein 1 homolog (VMO1 homolog), were validated by western blotting. Results Thirteen well resolved bands were detected in SDS–PAGE gels of both summer and winter camel tears. By band densitometry, significantly higher intensities of band 6, 7, 11, and lower intensity of band 13 were observed in the summer group compared to the winter group. In 2-DE profiles of camel tears, four protein spots were found expressed differentially in two seasons. Further protein identification by MALDI-TOF/TOF-MS and confirmation by western blotting indicated that there was a significant decrease in LF (p=0.002) and an increase in VMO1 homolog (p=0.042) in tears in the summer compared to the winter. Conclusions The seasonal variation of camel tear fluids has been found in the composition of proteins, including LF and VMO1 homolog. This result will expand our knowledge of physiologic characteristics of tear fluids and establish a foundation for the mechanistic studies and clinical practices on ocular surface disorders. PMID:21293736

Statistical Methods for Proteomic Biomarker Discovery based on Feature Extraction or Functional Modeling Approaches.

PubMed

Morris, Jeffrey S

2012-01-01

In recent years, developments in molecular biotechnology have led to the increased promise of detecting and validating biomarkers, or molecular markers that relate to various biological or medical outcomes. Proteomics, the direct study of proteins in biological samples, plays an important role in the biomarker discovery process. These technologies produce complex, high dimensional functional and image data that present many analytical challenges that must be addressed properly for effective comparative proteomics studies that can yield potential biomarkers. Specific challenges include experimental design, preprocessing, feature extraction, and statistical analysis accounting for the inherent multiple testing issues. This paper reviews various computational aspects of comparative proteomic studies, and summarizes contributions I along with numerous collaborators have made. First, there is an overview of comparative proteomics technologies, followed by a discussion of important experimental design and preprocessing issues that must be considered before statistical analysis can be done. Next, the two key approaches to analyzing proteomics data, feature extraction and functional modeling, are described. Feature extraction involves detection and quantification of discrete features like peaks or spots that theoretically correspond to different proteins in the sample. After an overview of the feature extraction approach, specific methods for mass spectrometry ( Cromwell ) and 2D gel electrophoresis ( Pinnacle ) are described. The functional modeling approach involves modeling the proteomic data in their entirety as functions or images. A general discussion of the approach is followed by the presentation of a specific method that can be applied, wavelet-based functional mixed models, and its extensions. All methods are illustrated by application to two example proteomic data sets, one from mass spectrometry and one from 2D gel electrophoresis. While the specific methods presented are applied to two specific proteomic technologies, MALDI-TOF and 2D gel electrophoresis, these methods and the other principles discussed in the paper apply much more broadly to other expression proteomics technologies.

Effect of 5% benzocaine gel on relieving pain caused by fixed orthodontic appliance activation. A double-blind randomized controlled trial.

PubMed

Eslamian, L; Gholami, H; Mortazavi, S A R; Soheilifar, S

2016-11-01

To compare the effectiveness of 5% benzocaine gel and placebo gel on reducing pain caused by fixed orthodontic appliance activation. Thirty subjects (15-25 years) undergoing fixed orthodontics. A randomized, double-blind, placebo-controlled and cross-over clinical trial study was conducted. Subjects were asked to apply a placebo gel and 5% benzocaine gel, exchangeable in two consecutive appointments, twice a day for 3 days and mark their level of pain on a VAS scale. The pain severity was evaluated by means of Mann-Whitney U-test for comparing two gel groups, Kruskal-Wallis nonparametric test for overall differences and post hoc test of Dunnett for paired multiple comparisons. p-value was assigned <0.05. The overall mean value of pain intensity for benzocaine and placebo gels was 0.89 and 1.15, respectively. The Mann-Whitney U-test indicated that there was no significant difference between overall pain in both groups (mean difference = 0.258 p ˂ 0.21). For both groups, pain intensity was significantly lower at 2, 6 and 24 h compared with pain experienced at days 2, 3 and 7. Benzocaine gel caused a decrease in pain perception at 2 h compared with placebo gel. Peak pain intensity was at 2 h for placebo gel and at 6 h for benzocaine gel, followed by a decline in pain perception from that point to day 7 for both gels. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Peritoneal Culture Alters Streptococcus pneumoniae Protein Profiles and Virulence Properties

PubMed Central

Orihuela, Carlos J.; Janssen, Rob; Robb, Christopher W.; Watson, David A.; Niesel, David W.

2000-01-01

We have examined the properties of Streptococcus pneumoniae cultured in the murine peritoneal cavity and compared its virulence-associated characteristics to those of cultures grown in vitro. Analysis of mRNA levels for specific virulence factors demonstrated a 2.8-fold increase in ply expression and a 2.2-fold increase in capA3 expression during murine peritoneal culture (MPC). Two-dimensional gels and immunoblots using convalescent-phase patient sera and murine sera revealed distinct differences in protein production in vivo (MPC). MPC-grown pneumococci adhered to A549 epithelial cell lines at levels 10-fold greater than those cultured in vitro. PMID:10992528

Peritoneal culture alters Streptococcus pneumoniae protein profiles and virulence properties

NASA Technical Reports Server (NTRS)

Orihuela, C. J.; Janssen, R.; Robb, C. W.; Watson, D. A.; Niesel, D. W.

2000-01-01

We have examined the properties of Streptococcus pneumoniae cultured in the murine peritoneal cavity and compared its virulence-associated characteristics to those of cultures grown in vitro. Analysis of mRNA levels for specific virulence factors demonstrated a 2.8-fold increase in ply expression and a 2.2-fold increase in capA3 expression during murine peritoneal culture (MPC). Two-dimensional gels and immunoblots using convalescent-phase patient sera and murine sera revealed distinct differences in protein production in vivo (MPC). MPC-grown pneumococci adhered to A549 epithelial cell lines at levels 10-fold greater than those cultured in vitro.

Investigation of Preparation and Mechanisms of a Dispersed Particle Gel Formed from a Polymer Gel at Room Temperature

PubMed Central

Zhao, Guang; Dai, Caili; Zhao, Mingwei; You, Qing; Chen, Ang

2013-01-01

A dispersed particle gel (DPG) was successfully prepared from a polymer gel at room temperature. The polymer gel system, morphology, viscosity changes, size distribution, and zeta potential of DPG particles were investigated. The results showed that zirconium gel systems with different strengths can be cross-linked within 2.5 h at low temperature. Scanning electron microscopy (SEM), transmission electron microscopy (TEM), and atomic force microscopy (AFM) results showed that the particles were polygonal particles with nano-size distribution. According to the viscosity changes, the whole preparation process can be divided into two major stages: the bulk gel cross-linking reaction period and the DPG particle preparation period. A polymer gel with a 3-dimensional network was formed in the bulk gel cross-linking reaction period whereas shearing force and frictional force were the main driving forces for the preparation of DPG particles, and thus affected the morphology of DPG particles. High shearing force and frictional force reduced the particle size distribution, and then decreased the zeta potential (absolute value). The whole preparation process could be completed within 3 h at room temperature. It could be an efficient and energy-saving technology for preparation of DPG particles. PMID:24324817

Heat- and light-induced thiol-ene oligomerization of soybean oil-based polymercaptan

USDA-ARS?s Scientific Manuscript database

Polymercaptanized soybean oil (PMSO), the product of a thiol-ene reaction between soybean oil and hydrogen sulfide, is a material of interest as a lubricant additive and polymer precursor. We investigated with gel permeation chromatography, nuclear magnetic resonance (one-dimensional and two-dimensi...

Analysis of proteins using DIGE and MALDI mass spectrometry

EPA Science Inventory

In this work the sensitivity of the quantitative proteomics approach 2D-DIGE/MS (twoDimensional Difference Gel Electrophoresis / Mass Spectrometry) was tested by detecting decreasing amounts of a specific protein at the low picomole and sub-picomole range. Sensitivity of the 2D-D...

Equilibrium polymerization of cyclic carbonate oligomers. III. Chain branching and the gel transition

NASA Astrophysics Data System (ADS)

Ballone, P.; Jones, R. O.

2002-10-01

Ring-opening polymerization of cyclic polycarbonate oligomers, where monofunctional active sites act on difunctional monomers to produce an equilibrium distribution of rings and chains, leads to a "living polymer." Monte Carlo simulations [two-dimensional (2D) and three-dimensional (3D)] of the effects of single [J. Chem. Phys. 115, 3895 (2001)] and multiple active sites [J. Chem. Phys. 116, 7724 (2002)] are extended here to trifunctional active sites that lead to branching. Low concentrations of trifunctional particles c3 reduce the degree of polymerization significantly in 2D, and higher concentrations (up to 32%) lead to further large changes in the phase diagram. Gel formation is observed at high total density and sizable c3 as a continuous transition similar to percolation. Polymer and gel are much more stable in 3D than in 2D, and both the total density and the value of c3 required to produce high molecular weight aggregates are reduced significantly. The degree of polymerization in high-density 3D systems is increased by the addition of trifunctional monomers and reduced slightly at low densities and low c3. The presence of branching makes equilibrium states more sensitive (in 2D and 3D) to changes in temperature T. The stabilities of polymer and gel are enhanced by increasing T, and—for sufficiently high values of c3—there is a reversible polymer-gel transformation at a density-dependent floor temperature.

Comparative evaluation of the antimicrobial efficacy of aloe vera tooth gel and two popular commercial toothpastes: an in vitro study.

PubMed

George, Dilip; Bhat, Sham S; Antony, Beena

2009-01-01

Aloe vera (Aloe barbadensis Miller) has been suggested for a wide variety of ailments but its use in dentistry is limited. This article reviews the uses of the plant and describes an in vitro investigation that compared the antimicrobial effectiveness of aloe vera tooth gel with two popular, commercially available dentifrices. The preliminary results showed that aloe vera tooth gel and the toothpastes were equally effective against Candida albicans, Streptococcus mutans, Lactobacillus acidophilus, Enterococcus faecalis, Prevotella intermedia, and Peptostreptococcus anaerobius. Aloe vera tooth gel demonstrated enhanced antibacterial effect against S. mitis.

Fabrication and characterization of sol-gel derived 45S5 Bioglass®-ceramic scaffolds.

PubMed

Chen, Qi-Zhi; Thouas, George A

2011-10-01

Although Bioglass® has existed for nearly half a century its ability to trigger bone formation and tuneable degradability is vastly superior to other bioceramics, such as SiO(2)-CaO bioactive glasses. The sol-gel process of producing glass foams is well established for SiO(2)-CaO compositions, but not yet established for 45S5 composites containing Na(2)O. In this work the sol-gel derived 45S5 Bioglass® has for the first time been foamed into highly porous three-dimensional scaffolds using a surfactant, combined with vigorous mechanical stirring and subsequent sintering at 1000°C for 2 h. It was found that the mechanical strength of the sintered sol-gel derived Bioglass® scaffolds was significantly improved, attributable to the small fraction of material on the pore walls. More importantly, the compressive strength of the three-dimensional scaffolds produced by this surfactant foaming method could be predicted using Gibson and Ashby's closed cell model of porous networks. A comparative experiment revealed that ion release from the sol-gel derived Bioglass® foams was faster than that of counterparts produced by the replication technique. In vitro evaluation using osteoblast-like cells demonstrated that the sol-gel derived 45S5 Bioglass foams supported the proliferation of viable cell populations on the surface of the scaffolds, although few cells were observed to migrate into the virtually closed pores within the foams. Further work should be focused on modifications of the reaction conditions or alternative foaming techniques to improve pore interconnection. Copyright © 2011 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Investigation of water mobility and diffusivity in hydrating micronized low-substituted hydroxypropyl cellulose, hydroxypropylmethyl cellulose, and hydroxypropyl cellulose matrix tablets by magnetic resonance imaging (MRI).

PubMed

Kojima, Masazumi; Nakagami, Hiroaki

2002-12-01

The water mobility and diffusivity in the gel-layer of hydrating low-substituted hydroxypropyl cellulose (LH41) tablets with or without a drug were investigated by magnetic resonance imaging (MRI) and compared with those properties in the gel-layer of hydroxypropylmethyl cellulose (HPMC) and hydroxypropyl cellulose (HPC) tablets. For this purpose, a localized image-analysis method was newly developed, and the spin-spin relaxation time (T(2)) and apparent self-diffusion coefficient (ADC) of water in the gel-layer were visualized in one-dimensional maps. Those maps showed that the extent of gel-layer growth in the tablets was in the order of HPC>HPMC>LH41, and there was a water mobility gradient across the gel-layers of all three tablet formulations. The T(2) and ADC in the outer parts of the gel-layers were close to those of free water. In contrast, these values in the inner parts of the gel-layer decreased progressively; suggesting that the water mobility and diffusivity around the core interface were highly restricted. Furthermore, the correlation between the T(2) of (1)H proton in the gel-layer of the tablets and the drug release rate from the tablets was observed.

Development in electrophoresis: instrumentation for two-dimensional gel electrophoresis of protein separation and application of capillary electrophoresis in micro-bioanalysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Aoshuang

2008-01-01

This dissertation begins with a general introduction of topics related to this work. The following chapters contain three scientific manuscripts, each presented in a separate chapter with accompanying tables, figures, and literature citations. The final chapter summarizes the work and provides some prospective on this work. This introduction starts with a brief treatment of the basic principles of electrophoresis separation, followed by a discussion of gel electrophoresis and particularly polyacrylamide gel electrophoresis for protein separation, a summary of common capillary electrophoresis separation modes, and a brief treatment of micro-bioanalysis application of capillary electrophoresis, and ends with an overview of proteinmore » conformation and dynamics.« less

Osteochondral Biopsy Analysis Demonstrates That BST-CarGel Treatment Improves Structural and Cellular Characteristics of Cartilage Repair Tissue Compared With Microfracture

PubMed Central

Méthot, Stéphane; Changoor, Adele; Tran-Khanh, Nicolas; Hoemann, Caroline D.; Stanish, William D.; Restrepo, Alberto; Shive, Matthew S.; Buschmann, Michael D.

2016-01-01

Objective The efficacy and safety of BST-CarGel, a chitosan-based medical device for cartilage repair, was compared with microfracture alone at 1 year during a multicenter randomized controlled trial (RCT) in the knee. The quality of repair tissue of osteochondral biopsies collected from a subset of patients was compared using blinded histological assessments. Methods The international RCT evaluated repair tissue quantity and quality by 3-dimensional quantitative magnetic resonance imaging as co-primary endpoints at 12 months. At an average of 13 months posttreatment, 21/41 BST-CarGel and 17/39 microfracture patients underwent elective second look arthroscopies as a tertiary endpoint, during which ICRS (International Cartilage Repair Society) macroscopic scoring was carried out, and osteochondral biopsies were collected. Stained histological sections were evaluated by blinded readers using ICRS I and II histological scoring systems. Collagen organization was evaluated using a polarized light microscopy score. Results BST-CarGel treatment resulted in significantly better ICRS macroscopic scores (P = 0.0002) compared with microfracture alone, indicating better filling, integration, and tissue appearance. Histologically, BST-CarGel resulted in a significant improvement of structural parameters—Surface Architecture (P = 0.007) and Surface/Superficial Assessment (P = 0.042)—as well as cellular parameters—Cell Viability (P = 0.006) and Cell Distribution (P = 0.032). No histological parameters were significantly better for the microfracture group. BST-CarGel treatment also resulted in a more organized repair tissue with collagen stratification more similar to native hyaline cartilage, as measured by polarized light microscopy scoring (P = 0.0003). Conclusion Multiple and independent analyses in this biopsy substudy demonstrated that BST-CarGel treatment results in improved structural and cellular characteristics of repair tissue at 1 year posttreatment compared with microfracture alone, supporting previously reported results by quantitative magnetic resonance imaging. PMID:26958314

Analysis of soybean tissue culture protein dynamics using difference gel electrophoresis.

PubMed

Miernyk, Ján A; Jett, Alissa A; Johnston, Mark L

2016-01-01

Excised hypocotyls from developing soybean (Glycine max (L.) merr. cv. Jack) were cultivated on agar-solidified medium until callus formed. The calli were then propagated in liquid medium until stable, relatively uniform, finely-divided suspension cultures were obtained. Cells were typically transferred to fresh medium at 7-day intervals. Cultures were harvested by filtration five days (early log phase) or eight days (late log phase) after transfer. In order to evaluate dynamic changes, both intracellular and extracellular proteins were analyzed by 2-dimensional difference gel electrophoresis. Selected spots were subjected to in-gel tryptic-digestion and the resultant peptides were analyzed by nLC-MS/MS. In follow-up studies gel-free shot-gun analyses led to identification of 367 intracellular proteins and 188 extracellular proteins. The significance of the described research is two-fold. First a gel-based proteomics method was applied to the study of the dynamics of the secretome (extracellular proteins). Second, results of a shot-gun non-gel based proteomic survey of both cellular and extracellular proteins are presented. Published by Elsevier B.V.

A novel collagen hydrogel cross-linked by gamma-ray irradiation in acidic pH conditions.

PubMed

Inoue, Naoki; Bessho, Masahiko; Furuta, Masakazu; Kojima, Takao; Okuda, Shuichi; Hara, Masayuki

2006-01-01

We made a new type of collagen gel by gamma-ray irradiation of an acidic solution of type-I collagen, and performed comparative studies on a conventional gel and the new type of gel. The neutral gel, a conventional 0.3% (w/v) collagen gel, was formed at neutral pH and then irradiated by gamma-rays. The acidic gel, a 0.3% (w/v) collagen gel, was formed directly from the acidic solution of collagen by y-ray irradiation. Both types of gel were prepared, swollen in water and then dried for the measurement of specific water content. The neutral gel showed a relatively high specific water content and shrunk moderately, depending on the dose, while the acidic gel showed lower specific water content and shrunk clearly by y-ray irradiation. A three-dimensional tangled network of microfibrils was clearly observed in the neutral gels by scanning electron microscopy, but not in the acidic gels. From these results, we concluded that the acidic gel was quite different from a conventional collagen gel. Sodium dodecylsulfate-polyacrylamide gel electrophoresis showed that the alpha1 subunit and alpha2 subunit of the collagen molecule were cross-linked. The triple-helical structure of collagen was only partially perturbed, but not denatured completely, because the circular dichroism spectrum of the collagen solution irradiated at 1.3 kGy was similar to that of native collagen solution. Amino-acid analysis revealed that tyrosine, phenylalanine and histidine decreased by irradiation in the neutral gel. In the case of the acidic gel, these three amino acids and methionine decreased. We considered that these amino acids were cross-linking points between the collagen subunits during the gamma-ray irradiation.

Pilot randomized trial for treatment of bacterial vaginosis using in situ forming metronidazole vaginal gel.

PubMed

Shaaban, Omar M; Fetih, Gihan N; Abdellah, Noura H; Ismail, Saeyd; Ibrahim, Maggie A; Ibrahim, El-sayed A

2011-07-01

To compare the efficacy of a novel vaginal delivery system for metronidazole (0.8% MTZ in situ gel) versus a conventional MTZ vaginal gel product in the treatment of bacterial vaginosis (BV). All consecutive patients who presented to a tertiary care hospital with symptoms suggestive of BV were approached to participate in the study. Forty-two eligible participants were randomly assigned to either MTZ in situ gel or a conventional vaginal gel product twice daily for 5 days. All participants were re-examined after one and 4 weeks of the beginning of treatment to ensure cure of infection and any side-effects. Demographic criteria of the participants were comparable in the two treatment groups. The cure rate after one week from the treatment was 85% in the in situ gel group and 71.4% in the conventional vaginal gel group (P = 0.294), while after 4 weeks, the cure rate showed significant difference in the in situ gel group as compared to the conventional vaginal gel group (16/20 [80%]) and (9/19 [47.4%]), respectively (P = 0.034). Pilot testing showed that in situ MTZ vaginal gel is more effective than the conventional vaginal gel for long-term cure of BV. These findings suggest a novel and efficient long-term treatment of BV. © 2011 The Authors. Journal of Obstetrics and Gynaecology Research © 2011 Japan Society of Obstetrics and Gynecology.

Comparative proteomics of human endothelial cell caveolae and rafts using two-dimensional gel electrophoresis and mass spectrometry.

PubMed

Sprenger, Richard R; Speijer, Dave; Back, Jaap Willem; De Koster, Chris G; Pannekoek, Hans; Horrevoets, Anton J G

2004-01-01

The human endothelial cell plasma membrane harbors two subdomains of similar lipid composition, caveolae and rafts, both crucially involved in various essential cellular processes like transcytosis, signal transduction and cholesterol homeostasis. Caveolin-enriched membranes, isolated by either cationic silica or buoyant density methods, were explored by comparing large series of two-dimensional (2-D) maps and subsequent identification of over 100 protein spots by matrix-assisted laser desorption/ionization (MALDI) peptide mass fingerprinting. Improved representation and identification of membrane proteins and valuable information on various post-translational modifications was achieved by the presented optimized procedures for solubilization, destaining and database searching/computing. Whereas the cationic silica purification yielded predominantly known endoplasmic reticulum residents, the cold-detergent method yielded a large number of known caveolae residents, including caveolin-1. Thus, a large part of this subproteome was established, including known (trans-)membrane, signal transduction and glycosyl phosphatidylinositol (GPI)-anchored proteins. Several predicted proteins from the human genome were isolated for the first time from biological samples, including SGRP58, SLP-2, C8ORF2, and XRP-2. These findings and various optimized procedures can serve as a reference to study the differential composition of endothelial cell caveolae and rafts, known to be involved in pathologies like cancer and cardiovascular disease.

Swelling-induced and controlled curving in layered gel beams

PubMed Central

Lucantonio, A.; Nardinocchi, P.; Pezzulla, M.

2014-01-01

We describe swelling-driven curving in originally straight and non-homogeneous beams. We present and verify a structural model of swollen beams, based on a new point of view adopted to describe swelling-induced deformation processes in bilayered gel beams, that is based on the split of the swelling-induced deformation of the beam at equilibrium into two components, both depending on the elastic properties of the gel. The method allows us to: (i) determine beam stretching and curving, once assigned the characteristics of the solvent bath and of the non-homogeneous beam, and (ii) estimate the characteristics of non-homogeneous flat gel beams in such a way as to obtain, under free-swelling conditions, three-dimensional shapes. The study was pursued by means of analytical, semi-analytical and numerical tools; excellent agreement of the outcomes of the different techniques was found, thus confirming the strength of the method. PMID:25383031

Root responses of Medicago truncatula plants grown in two different iron deficiency conditions: changes in root protein profile and riboflavin biosynthesis

USDA-ARS?s Scientific Manuscript database

Iron deficiency is a yield-limiting factor with major implications for field crop production in one-third of the world's agricultural areas, especially those with high soil CaCO3. A two-dimensional gel electrophoresis proteomic approach was combined with a study on the riboflavin synthesis pathway, ...

DOE Office of Scientific and Technical Information (OSTI.GOV)

Horie, Masafumi; Saito, Akira, E-mail: asaitou-tky@umin.ac.jp; Mikami, Yu

Highlights: Black-Right-Pointing-Pointer We established three patient-paired sets of CAFs and NFs. Black-Right-Pointing-Pointer CAFs and NFs were analyzed using three-dimensional co-culture experiments. Black-Right-Pointing-Pointer CAFs clearly enhanced collagen gel contraction. Black-Right-Pointing-Pointer CAFs showed higher {alpha}-SMA expression than NFs. Black-Right-Pointing-Pointer CAFs were implicated in invasion and differentiation of lung cancer cells. -- Abstract: Lung cancer is the most common cause of cancer-related death worldwide. Stromal cancer-associated fibroblasts (CAFs) play crucial roles in carcinogenesis, proliferation, invasion, and metastasis of non-small cell lung carcinoma, and targeting of CAFs could be a novel strategy for cancer treatment. However, the characteristics of human CAFs still remain tomore » be better defined. In this study, we established patient-matched CAFs and normal fibroblasts (NFs), from tumoral and non-tumoral portions of resected lung tissue from lung cancer patients. CAFs showed higher {alpha}-smooth muscle actin ({alpha}-SMA) expression than NFs, and CAFs clearly enhanced collagen gel contraction. Furthermore, we employed three-dimensional co-culture assay with A549 lung cancer cells, where CAFs were more potent in inducing collagen gel contraction. Hematoxylin and eosin staining of co-cultured collagen gel revealed that CAFs had the potential to increase invasion of A549 cells compared to NFs. These observations provide evidence that lung CAFs have the tumor-promoting capacity distinct from NFs.« less

Relative quantitative comparisons of the extracellular protein profiles of Staphylococcus aureus UAMS-1 and its sarA, agr, and sarA agr regulatory mutants using one-dimensional polyacrylamide gel electrophoresis and nanocapillary liquid chromatography coupled with tandem mass spectrometry.

PubMed

Jones, Richard C; Deck, Joanna; Edmondson, Ricky D; Hart, Mark E

2008-08-01

One-dimensional polyacrylamide gel electrophoresis followed by nanocapillary liquid chromatography coupled with mass spectrometry was used to analyze proteins isolated from Staphylococcus aureus UAMS-1 after 3, 6, 12, and 24 h of in vitro growth. Protein abundance was determined using a quantitative value termed normalized peptide number, and overall, proteins known to be associated with the cell wall were more abundant early on in growth, while proteins known to be secreted into the surrounding milieu were more abundant late in growth. In addition, proteins from spent media and cell lysates of strain UAMS-1 and its isogenic sarA, agr, and sarA agr regulatory mutant strains during exponential growth were identified, and their relative abundances were compared. Extracellular proteins known to be regulated by the global regulators sarA and agr displayed protein levels in accordance with what is known regarding the effects of these regulators. For example, cysteine protease (SspB), endopeptidase (SspA), staphopain (ScpA), and aureolysin (Aur) were higher in abundance in the sarA and sarA agr mutants than in strain UAMS-1. The immunoglobulin G (IgG)-binding protein (Sbi), immunodominant staphylococcal antigen A (IsaA), IgG-binding protein A (Spa), and the heme-iron-binding protein (IsdA) were most abundant in the agr mutant background. Proteins whose abundance was decreased in the sarA mutant included fibrinogen-binding protein (Fib [Efb]), IsaA, lipase 1 and 2, and two proteins identified as putative leukocidin F and S subunits of the two-component leukotoxin family. Collectively, this approach identified 1,263 proteins (matches of two peptides or more) and provided a convenient and reliable way of identifying proteins and comparing their relative abundances.

Comparison of Grain Proteome Profiles of Four Brazilian Common Bean (Phaseolus vulgaris L.) Cultivars.

PubMed

Rossi, Gabriela Barbosa; Valentim-Neto, Pedro Alexandre; Blank, Martina; Faria, Josias Correa de; Arisi, Ana Carolina Maisonnave

2017-08-30

Common bean (Phaseolus vulgaris L.) is a source of proteins for about one billion people worldwide. In Brazil, 'BRS Sublime', 'BRS Vereda', 'BRS Esteio', and 'BRS Estilo' cultivars were developed by Embrapa to offer high yield to farmers and excellent quality to final consumers. In this work, grain proteomes of these common bean cultivars were compared based on two-dimensional gel electrophoresis (2-DE) and tandem mass spectrometry (MS/MS). Principal component analysis (PCA) was applied to compare 349 matched spots in these cultivars proteomes, and all cultivars were clearly separated in PCA plot. Thirty-two differentially accumulated proteins were identified by MS. Storage proteins such as phaseolins, legumins, and lectins were the most abundant, and novel proteins were also identified. We have built a useful platform that could be used to analyze other Brazilian cultivars and genotypes of common beans.

The Tolerability Profile of Clindamycin 1%/Benzoyl Peroxide 5% Gel vs. Adapalene 0.1%/Benzoyl Peroxide 2.5% Gel for Facial Acne

PubMed Central

Cirigliano, Marcela; Gwazdauskas, Jennifer A; Gonzalez, Pablo

2012-01-01

Objective: To compare the first two weeks of tolerability of clindamycin/benzoyl peroxide gel versus adapalene/benzoyl peroxide gel followed by six weeks of open-label clindamycin/benzoyl peroxide gel therapy in subjects with mild-to-moderate acne who participated in two eight-week, identically designed, clinical studies. Methods: Using a split-face method, patients received both clindamycin/benzoyl peroxide gel and adapalene/benzoyl peroxide gel once daily for two weeks (allocation to the right or left side of the face was randomized) in an investigator-blinded fashion. Patients then went on to receive a further six weeks of open-label, full-face clindamycin/benzoyl peroxide gel. The primary outcome was to compare signs and symptoms of tolerability during the first two weeks of treatment using an investigator-assessed 4-point rating scale. Secondary endpoints included assessment of acne severity (Investigator Static Global Assessment and lesion counts), quality of life, product acceptability/preference, and patient assessments of tolerability and safety. Results: Of the 76 subjects enrolled in the two studies, 72 completed them. Overall both products were well tolerated, but mean scores for erythema, dryness, and peeling were significantly higher with adapalene/benzoyl peroxide gel than with clindamycin/benzoyl peroxide gel at both Weeks 1 and 2 (p<0.03). Patients also rated clindamycin/benzoyl peroxide gel significantly more tolerable than adapalene/benzoyl peroxide gel for redness, dryness, burning, itching, and scaling at Weeks 1 and 2 (p 0.0073). Mean Investigator Static Global Assessment score improved with both products during the first two weeks of treatment and continued to show significant improvement versus baseline when treatment with clindamycin/benzoyl peroxide gel was continued for a further six weeks (p<0.001 at Week 8). Lesion counts improved throughout the study with significant reductions from baseline occurring at Weeks 5 and 8 (p<0.0001 for both time points for total lesion counts). Clindamycin/benzoyl peroxide gel and adapalene/benzoyl peroxide gel were well tolerated, with most adverse events of mild-to-moderate severity. Conclusion: Clindamycin/benzoyl peroxide gel had better tolerability with regard to erythema, dryness, and peeling than adapalene/benzoyl peroxide gel during the first two weeks of treatment. PMID:22808305

[Azelaic acid 15% gel in the treatment of acne vulgaris. Combined results of two double-blind clinical comparative studies].

PubMed

Gollnick, Harald P M; Graupe, Klaus; Zaumseil, Rolf-Peter

2004-10-01

Topical measures are still the mainstay in the therapy of mild-to-moderate acne vulgaris. Azelaic acid 20% in a cream formulation has been established as an efficacious and safe topical drug for 15 years. A new non-alcoholic hydrogel formulation containing 15% azelaic acid was clinically tested against two standard drugs--5% benzoyl peroxide (BPO) and 1% clindamycin. In two independent, randomized, blinded comparative trials 15% azelaic acid gel was clinically tested against 5% benzoyl peroxide (BPO) gel in 351 patients and against 1% clindamycin gel in 229 patients. The drugs were applied b.i.d. for 4 months. Azelaic acid 15% gel proved to be as effective as BPO and clindamycin with median % reduction of the inflamed lesion (papules and pustules) of 70%, and 71% respectively. The azelaic acid gel was well-tolerated, the side effects (local burning and irritation) were distinctly less than with BPO but more pronounced than with clindamycin. Despite these side effects, the treatment was well-accepted by the majority of patients. Azelaic acid gel is an effective topical monotherapy for mild-to-moderate acne vulgaris; its new gel form is an enrichment of acne therapy.

Electrophoresis and isoelectric focusing of whole cell and membrane proteins from the extremely halophilic archaebacteria

NASA Technical Reports Server (NTRS)

Stan-Lotter, Helga; Lang, Frank J., Jr.; Hochstein, Lawrence I.

1989-01-01

The subunits from two purified halobacterial membrane enzymes (ATPase and nitrate reductase) behaved differently with respect to isoelectric focusing, silver staining and interaction with ampholytes. Differential behavior was also observed in whole cell proteins from Halobacterium saccharovorum regarding resolution in two-dimensional gels and silver staining. It is proposed that these differences reflect the existence of two classes of halobacterial proteins.

Quasi-one-dimensional nanostructured cobalt (Co) intercalated vanadium oxide (V{sub 2}O{sub 5}): Peroxovanadate sol gel synthesis and structural study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Langie da Silva, Douglas, E-mail: douglas.langie@ufpel.edu.br; Moreira, Eduardo Ceretta; Dias, Fábio Teixeira

2015-01-15

Nanostructured cobalt vanadium oxide (V{sub 2}O{sub 5}) xerogels spread onto crystalline Si substrates were synthesized via peroxovanadate sol gel route. The resulting products were characterized by distinct experimental techniques. The surface morphology and the nanostructure of xerogels correlate with Co concentration. The decrease of the structural coherence length is followed by the formation of a loose network of nanopores when the concentration of intercalated species was greater than 4 at% of Co. The efficiency of the synthesis route also drops with the increase of Co concentration. The interaction between the Co(OH{sub 2}){sub 6}{sup 2+} cations and the (H{sub 2}V{sub 10}O{submore » 28}){sup 4−} anions during the synthesis was suggested as a possible explanation for the incomplete condensation of the V{sub 2}O{sub 5} gel. Finally the experimental results points for the intercalation of Co between the bilayers of the V{sub 2}O{sub 5}. In this scenario two possible preferential occupation sites for the metallic atoms in the framework of the xerogel were proposed. - Graphical abstract: Quasi-one-dimensional nanostructured cobalt (Co) intercalated vanadium oxide (V{sub 2}O{sub 5}) nanoribbons synthesized by peroxovanadate sol gel route. - Highlights: • Nanostructured cobalt V{sub 2}O{sub 5} gel spread onto c{sub S}i were synthesized via peroxovanadate sol gel route. • The micro and nanostructure correlates with the cobalt content. • The efficiency of the synthesis route shows to be also dependent of Co content. • The experimental results points for the intercalation of Co between the bilayers of the V{sub 2}O{sub 5} xerogel.« less

Comparative effect of topical silicone gel and topical tretinoin cream for the prevention of hypertrophic scar and keloid formation and the improvement of scars.

PubMed

Kwon, S Y; Park, S D; Park, K

2014-08-01

Numerous modalities have been used to treat keloids and hypertrophic scars; however, optimal treatment has not yet been established. Therefore, prevention is the mainstay. Recently, silicone gel and tretinoin cream have been shown to be useful for the prevention of hypertrophic scars and keloids. However, there has been no comparative study of the two topical agents thus far. To determine and compare the effectiveness of silicone gel and tretinoin cream for the prevention of hypertrophic scars and keloids resulting from postoperative wounds and for scar improvement. This study included 26 patients with 44 different wounds. The postoperative wounds were divided into two treatment groups and one control group. The patients in the first and second treatment group applied silicone gel and tretinoin cream, respectively, twice a day on their wounds after their stitches were removed. In contrast, the control group patients did not apply anything. We used the Modified Vancouver Scar Scale to quantitatively examine the effectiveness of silicone gel and tretinoin cream just after stitches removal, and at 4, 8, 12 and 24 weeks after removal of the stitches. The silicone gel and tretinoin cream effectively prevented hypertrophic scars and keloids and improved scar effects in the two treatment groups compared with those in the control group. However, no significant difference was noted between the two treatment groups. To prevent hypertrophic scars and keloids and improve scars after surgery, application of a silicone gel or a tretinoin cream to the wounds is needed. © 2013 European Academy of Dermatology and Venereology.

Thermal stability and structural characterization of organic/inorganic hybrid nonlinear optical material containing a two-dimensional chromophore.

PubMed

Chang, Po-Hsun; Tsai, Hsieh-Chih; Chen, Yu-Ren; Chen, Jian-Yu; Hsiue, Ging-Ho

2008-10-21

In this study, two nonlinear optic hybrid materials with different dimensional alkoxysilane dyes were prepared and characterized. One NLO silane (Cz2PhSO 2OH- TES), a two-dimensional structure based on carbazole, had a larger rotational volume than the other (DR19-TES). Second harmonic ( d 33) analysis verified there is an optimum heating process for the best poling efficiency. The maximum d 33 value of NLO hybrid film containing Cz2PhSO 2OH was obtained for 10.7 pm/V after precuring at 150 degrees C for 3 h and poling at 210 degrees C for 60 min. The solid-state (29)Si NMR spectrum shows that the main factor influencing poling efficiency and thermal stability was cross-linking degree of NLO silane, but not that of TMOS. In particular, the two-dimensional sol-gel system has a greater dynamic and temporary stability than the one-dimensional system due to Cz2PhSO 2OH-TES requiring a larger volume to rotate in the hybrid matrix after cross-linking.

Rifaximin-mediated changes to the epithelial cell proteome: 2-D gel analysis.

PubMed

Schrodt, Caroline; McHugh, Erin E; Gawinowicz, Mary Ann; Dupont, Herbert L; Brown, Eric L

2013-01-01

Rifaximin is a semi-synthetic rifamycin derivative that is used to treat different conditions including bacterial diarrhea and hepatic encephalopathy. Rifaximin is of particular interest because it is poorly adsorbed in the intestines and has minimal effect on colonic microflora. We previously demonstrated that rifaximin affected epithelial cell physiology by altering infectivity by enteric pathogens and baseline inflammation suggesting that rifaximin conferred cytoprotection against colonization and infection. Effects of rifaximin on epithelial cells were further examined by comparing the protein expression profile of cells pretreated with rifaximin, rifampin (control antibiotic), or media (untreated). Two-dimensional (2-D) gel electrophoresis identified 36 protein spots that were up- or down-regulated by over 1.7-fold in rifaximin treated cells compared to controls. 15 of these spots were down-regulated, including annexin A5, intestinal-type alkaline phosphatase, histone H4, and histone-binding protein RbbP4. 21 spots were up-regulated, including heat shock protein (HSP) 90α and fascin. Many of the identified proteins are associated with cell structure and cytoskeleton, transcription and translation, and cellular metabolism. These data suggested that in addition to its antimicrobial properties, rifaximin may alter host cell physiology that provides cytoprotective effects against bacterial pathogens.

Rifaximin-Mediated Changes to the Epithelial Cell Proteome: 2-D Gel Analysis

PubMed Central

Schrodt, Caroline; McHugh, Erin E.; Gawinowicz, Mary Ann; DuPont, Herbert L.; Brown, Eric L.

2013-01-01

Rifaximin is a semi-synthetic rifamycin derivative that is used to treat different conditions including bacterial diarrhea and hepatic encephalopathy. Rifaximin is of particular interest because it is poorly adsorbed in the intestines and has minimal effect on colonic microflora. We previously demonstrated that rifaximin affected epithelial cell physiology by altering infectivity by enteric pathogens and baseline inflammation suggesting that rifaximin conferred cytoprotection against colonization and infection. Effects of rifaximin on epithelial cells were further examined by comparing the protein expression profile of cells pretreated with rifaximin, rifampin (control antibiotic), or media (untreated). Two-dimensional (2-D) gel electrophoresis identified 36 protein spots that were up- or down-regulated by over 1.7-fold in rifaximin treated cells compared to controls. 15 of these spots were down-regulated, including annexin A5, intestinal-type alkaline phosphatase, histone H4, and histone-binding protein RbbP4. 21 spots were up-regulated, including heat shock protein (HSP) 90α and fascin. Many of the identified proteins are associated with cell structure and cytoskeleton, transcription and translation, and cellular metabolism. These data suggested that in addition to its antimicrobial properties, rifaximin may alter host cell physiology that provides cytoprotective effects against bacterial pathogens. PMID:23922656

Effects of calorie restriction on the zebrafish liver proteome

PubMed Central

Jury, David R.; Kaveti, Suma; Duan, Zhong-Hui; Willard, Belinda; Kinter, Michael; Londraville, Richard

2012-01-01

A proteomic approach was taken to study how fish respond to changes in calorie availability, with the longer-term goal of understanding the evolution of lipid metabolism in vertebrates. Zebrafish (Danio rerio) were fed either high (3 rations/day) or low (1 ration/7 days) calorie diets for 5 weeks and liver proteins extracted for proteomic analyses. Proteins were separated on two-dimensional electrophoresis gels and homologous spots compared between treatments to determine which proteins were up-regulated with high-calorie diet. Fifty-five spots were excised from the gel and analyzed via LC–ESI MS/MS, which resulted in the identification of 69 unique proteins (via multiple peptides). Twenty-nine of these proteins were differentially expressed between treatments. Differentially expressed proteins were mapped to Gene Ontology (GO) terms, and these terms compared to the entire zebrafish GO annotation set by Fisher's exact test. The most significant GO terms associated with high-calorie diet are related to a decrease in oxygen-binding activity in the high-calorie treatment. This response is consistent with a well-characterized response in obese humans, indicating there may be a link between lipid storage and hypoxia sensitivity in vertebrates. PMID:20494847

Proteomic differential display analysis for TS-1-resistant and -sensitive pancreatic cancer cells using two-dimensional gel electrophoresis and mass spectrometry.

PubMed

Yoshida, Kanako; Kuramitsu, Yasuhiro; Murakami, Kohei; Ryozawa, Shomei; Taba, Kumiko; Kaino, Seiji; Zhang, Xiulian; Sakaida, Isao; Nakamura, Kazuyuki

2011-06-01

TS-1 is an oral anticancer agent containing two biochemical modulators for 5-fluorouracil (5-FU) and tegafur (FT), a metabolically activated prodrug of 5-FU. TS-1 has been recognized as an effective anticancer drug using standard therapies for patients with advanced pancreatic cancer along with gemcitabine. However, a high level of inherent and acquired tumor resistance to TS-1 induces difficulty in the treatment. To identify proteins linked to the TS-1-resistance of pancreatic cancer, we profiled protein expression levels in samples of TS-1-resistant and -sensitive pancreatic cancer cell lines by using two-dimensional gel electrophoresis (2-DE) and liquid chromatography-tandem mass spectrometry (LC-MS/MS). The cytotoxicity of a 5-FU/5-chloro-2,4-dihydroxypyridine (CDHP) combination towards pancreatic cancer cell lines was evaluated by MTS assay. Panc-1, BxPC-3, MiaPaCa-2 and PK59 showed high sensitivity to the 5-FU/CDHP combination (TS-1-sensitive), whereas PK45p and KLM-1 were much less sensitive (TS-1-resistant). Proteomic analysis showed that eleven spots, including T-complex protein 1 subunit beta, ribonuclease inhibitor, elongation factor 1-delta, peroxiredoxin-2 and superoxide dismutase (Cu-Zn), appeared to be down-regulated, and 29 spots, including hypoxia up-regulated protein 1, lamin-A/C, endoplasmin, fascin and annexin A1, appeared to be up-regulated in TS-1-resistant cells compared with -sensitive cells. These results suggest that the identified proteins showing different expression between TS-1-sensitive and -resistant pancreatic cancer cells possibly relate to TS-1-sensitivity. These findings could be useful to overcome the TS-1-resistance of pancreatic cancer cells.

MAGIC with formaldehyde applied to dosimetry of HDR brachytherapy source

NASA Astrophysics Data System (ADS)

Marques; T; Fernandes; J; Barbi; G; Nicolucci; P; Baffa; O

2009-05-01

The use of polymer gel dosimeters in brachytherapy can allow the determination of three-dimensional dose distributions in large volumes and with high spatial resolution if an adequate calibration process is performed. One of the major issues in these experiments is the polymer gel response dependence on dose rate when high dose rate sources are used and the doses in the vicinity of the sources are to be determinated. In this study, the response of a modified MAGIC polymer gel with formaldehyde around an Iridium-192 HDR brachytherapy source is presented. Experimental results obtained with this polymer gel were compared with ionization chamber measurements and with Monte Carlo simulation with PENELOPE. A maximum difference of 3.10% was found between gel dose measurements and Monte Carlo simulation at a radial distance of 18 mm from the source. The results obtained show that the gel's response is strongly influenced by dose rate and that a different calibration should be used for the vicinity of the source and for regions of lower dose rates. The results obtained in this study show that, provided the proper calibration is performed, MAGIC with formaldehyde can be successfully used to accurate determinate dose distributions form high dose rate brachytherapy sources.

How many proteins can be identified in a 2DE gel spot within an analysis of a complex human cancer tissue proteome?

PubMed

Zhan, Xianquan; Yang, Haiyan; Peng, Fang; Li, Jianglin; Mu, Yun; Long, Ying; Cheng, Tingting; Huang, Yuda; Li, Zhao; Lu, Miaolong; Li, Na; Li, Maoyu; Liu, Jianping; Jungblut, Peter R

2018-04-01

Two-dimensional gel electrophoresis (2DE) in proteomics is traditionally assumed to contain only one or two proteins in each 2DE spot. However, 2DE resolution is being complemented by the rapid development of high sensitivity mass spectrometers. Here we compared MALDI-MS, LC-Q-TOF MS and LC-Orbitrap Velos MS for the identification of proteins within one spot. With LC-Orbitrap Velos MS each Coomassie Blue-stained 2DE spot contained an average of at least 42 and 63 proteins/spot in an analysis of a human glioblastoma proteome and a human pituitary adenoma proteome, respectively, if a single gel spot was analyzed. If a pool of three matched gel spots was analyzed this number further increased up to an average of 230 and 118 proteins/spot for glioblastoma and pituitary adenoma proteome, respectively. Multiple proteins per spot confirm the necessity of isotopic labeling in large-scale quantification of different protein species in a proteome. Furthermore, a protein abundance analysis revealed that most of the identified proteins in each analyzed 2DE spot were low-abundance proteins. Many proteins were present in several of the analyzed spots showing the ability of 2DE-MS to separate at the protein species level. Therefore, 2DE coupled with high-sensitivity LC-MS has a clearly higher sensitivity as expected until now to detect, identify and quantify low abundance proteins in a complex human proteome with an estimated resolution of about 500 000 protein species. This clearly exceeds the resolution power of bottom-up LC-MS investigations. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Silk-fibrin/hyaluronic acid composite gels for nucleus pulposus tissue regeneration.

PubMed

Park, Sang-Hyug; Cho, Hongsik; Gil, Eun Seok; Mandal, Biman B; Min, Byoung-Hyun; Kaplan, David L

2011-12-01

Scaffold designs are critical for in vitro culture of tissue-engineered cartilage in three-dimensional environments to enhance cellular differentiation for tissue engineering and regenerative medicine. In the present study we demonstrated silk and fibrin/hyaluronic acid (HA) composite gels as scaffolds for nucleus pulposus (NP) cartilage formation, providing both biochemical support for NP outcomes as well as fostering the retention of size of the scaffold during culture due to the combined features of the two proteins. Passage two (P2) human chondrocytes cultured in 10% serum were encapsulated within silk-fibrin/HA gels. Five study groups with fibrin/HA gel culture (F/H) along with varying silk concentrations (2% silk gel only, fibrin/HA gel culture with 1% silk [F/H+1S], 1.5% silk [F/H+1.5S], and 2% silk [F/H+2S]) were cultured in serum-free chondrogenic defined media (CDM) for 4 weeks. Histological examination with alcian blue showed a defined chondrogenic area at 1 week in all groups that widened homogenously until 4 weeks. In particular, chondrogenic differentiation observed in the F/H+1.5S had no reduction in size throughout the culture period. The results of biochemical and molecular biological evaluations supported observations made during histological examination. Mechanical strength measurements showed that the silk mixed gels provided stronger mechanical properties for NP tissue than fibrin/HA composite gels in CDM. This effect could potentially be useful in the study of in vitro NP tissue engineering as well as for clinical implications for NP tissue regeneration.

Silk-Fibrin/Hyaluronic Acid Composite Gels for Nucleus Pulposus Tissue Regeneration

PubMed Central

Park, Sang-Hyug; Cho, Hongsik; Gil, Eun Seok; Mandal, Biman B.; Min, Byoung-Hyun

2011-01-01

Scaffold designs are critical for in vitro culture of tissue-engineered cartilage in three-dimensional environments to enhance cellular differentiation for tissue engineering and regenerative medicine. In the present study we demonstrated silk and fibrin/hyaluronic acid (HA) composite gels as scaffolds for nucleus pulposus (NP) cartilage formation, providing both biochemical support for NP outcomes as well as fostering the retention of size of the scaffold during culture due to the combined features of the two proteins. Passage two (P2) human chondrocytes cultured in 10% serum were encapsulated within silk-fibrin/HA gels. Five study groups with fibrin/HA gel culture (F/H) along with varying silk concentrations (2% silk gel only, fibrin/HA gel culture with 1% silk [F/H+1S], 1.5% silk [F/H+1.5S], and 2% silk [F/H+2S]) were cultured in serum-free chondrogenic defined media (CDM) for 4 weeks. Histological examination with alcian blue showed a defined chondrogenic area at 1 week in all groups that widened homogenously until 4 weeks. In particular, chondrogenic differentiation observed in the F/H+1.5S had no reduction in size throughout the culture period. The results of biochemical and molecular biological evaluations supported observations made during histological examination. Mechanical strength measurements showed that the silk mixed gels provided stronger mechanical properties for NP tissue than fibrin/HA composite gels in CDM. This effect could potentially be useful in the study of in vitro NP tissue engineering as well as for clinical implications for NP tissue regeneration. PMID:21736446

Mannan-binding lectin of the sea urchin Strongylocentrotus nudus.

PubMed

Bulgakov, Aleksandr A; Eliseikina, Marina G; Kovalchuk, Svetlana N; Petrova, Irina Yu; Likhatskaya, Galina N; Shamshurina, Ekaterina V; Rasskazov, Valery A

2013-02-01

A novel lectin specific to low-branched mannans (MBL-SN) was isolated from coelomic plasma of the sea urchin Strongylocentrotus nudus by combining anion-exchange liquid chromatography on DEAE Toyopearl 650 M, affinity chromatography on mannan-Sepharose and gel filtration on the Sephacryl S-200. The molecular mass of MBL-SN was estimated by sodium dodecyl sulphate polyacrylamide gel electrophoresis under non-reducing conditions to be about 34 kDa. MBL-SN was shown to be a dimer with two identical subunits of about 17 kDa. The native MBL-SN exists as a tetramer. The physico-chemical properties of MBL-SN indicate that it belongs to C-type mannan-binding lectins. The cDNA encoding MBL-SN was cloned from the total cDNA of S. nudus coelomocytes and encodes a 17-kDa protein of 144 amino acid residues that contains a single carbohydrate-recognition domain of C-type lectins. Prediction of the MBL-SN tertiary structure using comparative modelling revealed that MBL-SN is an α/β-protein with eight β-strands and two α-helices. Comparison of the MBL-SN model with available three-dimensional structures of C-type lectins revealed that they share a common fold pattern.

Comparison of the longissimus muscle proteome between obese and lean pigs at 180 days.

PubMed

Li, Anning; Mo, Delin; Zhao, Xiao; Jiang, Wei; Cong, Peiqing; He, Zuyong; Xiao, Shuqi; Liu, Xiaohong; Chen, Yaosheng

2013-02-01

Production of high-quality meat is important to satisfy the consumer and make the pig industry competitive. Obese and lean breeds of pig show clear differences in adipogenic capacity and meat quality, but the underlying molecular mechanism remains unclear. We have compared protein expression of the longissimus muscle between Lantang (LT, obese) and Landrace (LR, lean) pigs at the age of 180 days using two-dimensional fluorescence difference gel electrophoresis. Of the 1,400 protein spots detected per gel, 18 were differentially expressed between the two breeds. Using peptide mass fingerprint and tandem mass spectrometry, 17 protein spots were identified, corresponding to ten different proteins that could be divided into four groups: metabolism-related, structure-related, stress-related, and other (unclassified). Among the metabolism-related proteins, COX5A and ATP5B, which participate in oxidative phosphorylation, were highly expressed in LT, whereas ENO3, which is involved in glycolysis, was highly expressed in LR. These results may contribute valuable information to our understanding of the molecular mechanism responsible for differences between obese and lean pigs, such as growth rate and meat quality.

Induction of labor using prostaglandin E2 (PGE2) vaginal gel in triacetin base. An efficacy study comparing two dosage regimens.

PubMed

Seeras, R C; Olatunbosun, O A; Pierson, R A; Turnell, R W

1995-01-01

To compare two dosage regimens for the administration of vaginal prostaglandin gel in triacetin base for induction of labor. Seventy subjects planned for elective induction of labor at term were randomized to treatment with PGE2 vaginal gel every 6 or 12 hours. The 6-hourly group received an initial dose of 1 mg, followed by 2 mg at 6 hour intervals for a maximum of two additional doses if not in active labor. The 12-hourly group had an initial dose of 2 mg followed by two additional doses at 12 hour intervals if not in active labor. Successful induction rate was higher in the 12-hourly as compared to 6-hourly gel regimen (100% vs. 91%, P > 0.05). Twelve hours after the initial dose, delivery occurred in 34% delivery had occurred in 57% and 37% respectively (P < 0.01). We found no difference in the induction-active labor interval (P > 0.05), and the induction-delivery interval (P > 0.05) between the two groups. Active labor followed a single dose of gel in 66% of the 12-hourly group compared to 40% of the 6-hourly group (P < 0.01). Syntocinon augmentation was needed in 6% of subjects in the 12-hourly group as compared to 26% in the 6-hourly group (P < 0.01). The cesarean section rate was similar in both groups. Uterine hyperstimulation occurred less frequently in the 12-hourly group (P < 0.05). The perinatal outcome was similar in both groups. The 12-hourly regimen was more effective than the 6-hourly regimen in initiating labor. The majority of the subjects in the 12 hourly group achieved labor following a single dose of gel. Induction delivery interval, however, was similar in both groups.

Modification of vital wheat gluten with phosphoric acid to produce high free-solution capacity

USDA-ARS?s Scientific Manuscript database

Wheat gluten reacts with phosphoric acid to produce natural superabsorbent gels. The gel properties are defined by Fourier Transform Infra-red (FTIR) spectroscopy, 2-dimensional gel electrophoresis (2DE), and uptake of water, salt solutions, and aqueous ethanol. Temperatures above 120'C and dry cond...

Distinguishing between respiratory syncytial virus subgroups by protein profile analysis.

PubMed Central

Walpita, P; Mufson, M A; Stanek, R J; Pfeifer, D; Connor, J D

1992-01-01

We subgrouped 75 strains of respiratory syncytial virus by a protein profile method (PPM) which relies on different mobilities of the phosphoprotein in one-dimensional polyacrylamide gel electrophoresis and does not require monoclonal antibodies. When compared with enzyme immunoassay, PPM correctly subgrouped 54 of 56 subgroup A and all 19 subgroup B strains. Images PMID:1572961

Comparison between Monte Carlo simulation and measurement with a 3D polymer gel dosimeter for dose distributions in biological samples

NASA Astrophysics Data System (ADS)

Furuta, T.; Maeyama, T.; Ishikawa, K. L.; Fukunishi, N.; Fukasaku, K.; Takagi, S.; Noda, S.; Himeno, R.; Hayashi, S.

2015-08-01

In this research, we used a 135 MeV/nucleon carbon-ion beam to irradiate a biological sample composed of fresh chicken meat and bones, which was placed in front of a PAGAT gel dosimeter, and compared the measured and simulated transverse-relaxation-rate (R2) distributions in the gel dosimeter. We experimentally measured the three-dimensional R2 distribution, which records the dose induced by particles penetrating the sample, by using magnetic resonance imaging. The obtained R2 distribution reflected the heterogeneity of the biological sample. We also conducted Monte Carlo simulations using the PHITS code by reconstructing the elemental composition of the biological sample from its computed tomography images while taking into account the dependence of the gel response on the linear energy transfer. The simulation reproduced the experimental distal edge structure of the R2 distribution with an accuracy under about 2 mm, which is approximately the same as the voxel size currently used in treatment planning.

Comparison between Monte Carlo simulation and measurement with a 3D polymer gel dosimeter for dose distributions in biological samples.

PubMed

Furuta, T; Maeyama, T; Ishikawa, K L; Fukunishi, N; Fukasaku, K; Takagi, S; Noda, S; Himeno, R; Hayashi, S

2015-08-21

In this research, we used a 135 MeV/nucleon carbon-ion beam to irradiate a biological sample composed of fresh chicken meat and bones, which was placed in front of a PAGAT gel dosimeter, and compared the measured and simulated transverse-relaxation-rate (R2) distributions in the gel dosimeter. We experimentally measured the three-dimensional R2 distribution, which records the dose induced by particles penetrating the sample, by using magnetic resonance imaging. The obtained R2 distribution reflected the heterogeneity of the biological sample. We also conducted Monte Carlo simulations using the PHITS code by reconstructing the elemental composition of the biological sample from its computed tomography images while taking into account the dependence of the gel response on the linear energy transfer. The simulation reproduced the experimental distal edge structure of the R2 distribution with an accuracy under about 2 mm, which is approximately the same as the voxel size currently used in treatment planning.

Oxygen diffusion and consumption in extracellular matrix gels: implications for designing three-dimensional cultures.

PubMed

Colom, Adai; Galgoczy, Roland; Almendros, Isaac; Xaubet, Antonio; Farré, Ramon; Alcaraz, Jordi

2014-08-01

Three-dimensional (3D) cultures are increasingly used as tissue surrogates to study many physiopathological processes. However, to what extent current 3D culture protocols provide physiologic oxygen tension conditions remains ill defined. To address this limitation, oxygen tension was measured in a panel of acellular or cellularized extracellular matrix (ECM) gels with A549 cells, and analyzed in terms of oxygen diffusion and consumption. Gels included reconstituted basement membrane, fibrin and collagen. Oxygen diffusivity in acellular gels was up to 40% smaller than that of water, and the lower values were observed in the denser gels. In 3D cultures, physiologic oxygen tension was achieved after 2 days in dense (≥3 mg/mL) but not sparse gels, revealing that the latter gels are not suitable tissue surrogates in terms of oxygen distribution. In dense gels, we observed a dominant effect of ECM composition over density in oxygen consumption. All diffusion and consumption data were used in a simple model to estimate ranges for gel thickness, seeding density and time-window that may support physiologic oxygen tension. Thus, we identified critical variables for oxygen tension in ECM gels, and introduced a model to assess initial values of these variables, which may short-cut the optimization step of 3D culture studies. © 2013 Wiley Periodicals, Inc.

Comparative proteomic analyses of macular and peripheral retina of cynomolgus monkeys (Macaca fascicularis).

PubMed

Okamoto, Haru; Umeda, Shinsuke; Nozawa, Takehiro; Suzuki, Michihiro T; Yoshikawa, Yasuhiro; Matsuura, Etsuko T; Iwata, Takeshi

2010-01-01

The central region of the primate retina is called the macula. The fovea is located at the center of the macula, where the photoreceptors are concentrated to create a neural network adapted for high visual acuity. Damage to the fovea, e.g., by macular dystrophies and age-related macular degeneration, can reduce central visual acuity. The molecular mechanisms leading to these diseases are most likely dependent on the proteins in the macula which differ from those in the peripheral retina in expression level. To investigate whether the distribution of proteins in the macula is different from the peripheral retina, proteomic analyses of tissues from these two regions of cynomolgus monkeys were compared. Two-dimensional gel electrophoresis and mass spectrometry identified 26 proteins that were present only in the macular gel spots. The expression levels of five proteins, cone photoreceptor specific arrestin-C, gamma-synuclein, epidermal fatty acid binding protein, tropomyosin 1alpha chain, and heterogeneous nuclear ribonucleoproteins A2/B1, were significantly higher in the macula than in the peripheral retina. Immunostaining of macula sections by antibodies to each identified protein revealed unique localization in the retina, retinal pigment epithelial cells and the choroidal layer. Some of these proteins were located in cells with higher densities in the macula. We suggest that it will be important to study these proteins to determine their contribution to the pathogenesis and progression of macula diseases.

Proteomic analysis of common bean (Phaseolus vulgaris) by two-dimensional gel electrophoresis and mass spectrometry

USDA-ARS?s Scientific Manuscript database

The modern cultivated common bean (Phaseolus vulgaris) has evolved from wild common beans distributed in Central America, Mexico and the Andean region of South America. It has been reported that wild common bean accessions have higher levels of protein content than the domesticated dry bean cultiva...

Flexible and transparent polyimide films containing two-dimensional alumina nanosheets templated by graphene oxide for improved barrier property.

PubMed

Tseng, I-Hsiang; Tsai, Mei-Hui; Chung, Chi-Wei

2014-08-13

Unique two-dimensional alumina nanosheets (Alns) using graphene oxide (GO) as templates are fabricated and successfully incorporated with organo-soluble polyimide (PI) to obtain highly transparent PI nanocomposite films with improved moisture barrier property. The effects of filler types and contents on water vapor transmission rate (WVTR) and transparency of PI are systematically studied. The hydroxyl groups on GO react with aluminum isopropoxide via sol-gel process to obtain alumina coverd-GO (Al-GO), and then thermal decomposition is applied to obtain Alns. Alns are the most efficient fillers among others to restrict the diffusion of water vapor within PI matrix and simultaneously maintain the transparency of PI. XRD pattern, TEM, and AFM images confirm the sheet-like morphology of Alns with ultrahigh aspect ratio. With only 0.01 wt % of Alns, the PI nanocomposite film exhibits the most significant reduction of 95% in WVTR as compared to that of pure PI film. Most importantly, the resultant PI/Alns-0.01 film exhibits excellent optical transparency and high mechanical strength and great thermal stability.

Evaluation of different protein extraction methods for banana (Musa spp.) root proteome analysis by two-dimensional electrophoresis.

PubMed

Vaganan, M Mayil; Sarumathi, S; Nandakumar, A; Ravi, I; Mustaffa, M M

2015-02-01

Four protocols viz., the trichloroacetic acid-acetone (TCA), phenol-ammonium acetate (PAA), phenol/SDS-ammonium acetate (PSA) and trisbase-acetone (TBA) were evaluated with modifications for protein extraction from banana (Grand Naine) roots, considered as recalcitrant tissues for proteomic analysis. The two-dimensional electrophoresis (2-DE) separated proteins were compared based on protein yield, number of resolved proteins, sum of spot quantity, average spot intensity and proteins resolved in 4-7 pI range. The PAA protocol yielded more proteins (0.89 mg/g of tissues) and protein spots (584) in 2-DE gel than TCA and other protocols. Also, the PAA protocol was superior in terms of sum of total spot quantity and average spot intensity than TCA and other protocols, suggesting phenol as extractant and ammonium acetate as precipitant of proteins were the most suitable for banana rooteomics analysis by 2-DE. In addition, 1:3 ratios of root tissue to extraction buffer and overnight protein precipitation were most efficient to obtain maximum protein yield.

Identification of novel proteins in culture filtrates of Mycobacterium bovis bacillus Calmette-Guérin in the isoelectric point range 6-11.

PubMed

Florio, Walter; Batoni, Giovanna; Esin, Semih; Bottai, Daria; Maisetta, Giuseppantonio; Pardini, Manuela; Campa, Mario

2003-05-01

Two-dimensional gel electrophoresis and mass spectrometry were used to identify proteins in the isoelectric point range 6-11 in culture filtrates of Mycobacterium bovis bacillus Calmette-Guérin (BCG). Twelve proteins were identified, three of which had not been described previously. The expression of the identified proteins was comparatively analyzed in culture filtrates of BCG in different growth phases and culture conditions. For some of these proteins, the relative protein abundance in the different culture filtrate preparations was significantly different. The differential expression of the identified proteins is discussed in relation to their putative localization and/or biological function.

Full evaporation dynamic headspace in combination with selectable one-dimensional/two-dimensional gas chromatography-mass spectrometry for the determination of suspected fragrance allergens in cosmetic products.

PubMed

Devos, Christophe; Ochiai, Nobuo; Sasamoto, Kikuo; Sandra, Pat; David, Frank

2012-09-14

Suspected fragrance allergens were determined in cosmetic products using a combination of full evaporation-dynamic headspace (FEDHS) with selectable one-dimensional/two-dimensional GC-MS. The full evaporation dynamic headspace approach allows the non-discriminating extraction and injection of both apolar and polar fragrance compounds, without contamination of the analytical system by high molecular weight non-volatile matrix compounds. The method can be applied to all classes of cosmetic samples, including water containing matrices such as shower gels or body creams. In combination with selectable (1)D/(2)D GC-MS, consisting of a dedicated heart-cutting GC-MS configuration using capillary flow technology (CFT) and low thermal mass GC (LTM-GC), a highly flexible and easy-to-use analytical solution is offered. Depending on the complexity of the perfume fraction, analyses can be performed in one-dimensional GC-MS mode or in heart-cutting two-dimensional GC-MS mode, without the need of hardware reconfiguration. The two-dimensional mode with independent temperature control of the first and second dimension column is especially useful to confirm the presence of detected allergen compounds when mass spectral deconvolution is not possible. Copyright © 2012 Elsevier B.V. All rights reserved.

Human Mesenchymal Stem Cell Spheroids in Fibrin Hydrogels Exhibit Improved Cell Survival and Potential for Bone Healing

PubMed Central

Murphy, Kaitlin C.; Fang, Sophia Y.; Leach, J. Kent

2014-01-01

Mesenchymal stem cells (MSC) have great therapeutic potential for the repair of nonhealing bone defects due to their proliferative capacity, multilineage potential, trophic factor secretion, and lack of immunogenicity. However, a major barrier to the translation of cell-based therapies into clinical practice is ensuring their survival and function upon implantation into the defect site. We hypothesized that forming MSC into more physiologic 3-dimensional spheroids, rather than employing dissociated cells from 2-dimensional monolayer culture, would enhance their survival when exposed to a harsh microenvironment while maintaining their osteogenic potential. MSC spheroids were formed using the hanging drop method with increasing cell numbers. Compared to larger spheroids, the smallest spheroids which contained 15,000 cells exhibited increased metabolic activity, reduced apoptosis, and the most uniform distribution of proliferating cells. Spheroids were then entrapped in fibrin gels and cultured in serum-free media and 1% oxygen. Compared to identical numbers of dissociated MSC in fibrin gels, spheroids exhibited significantly reduced apoptosis and secreted up to 100-fold more VEGF. We also observed that fibrin gels containing spheroids and those containing an equivalent number of dissociated cells exhibited similar expression levels of early and late markers of osteogenic differentiation. These data demonstrate that MSC spheroids exhibit greater resistance to apoptosis and enhanced proangiogenic potential, while maintaining similar osteogenic potential to dissociated MSC entrapped in a clinically relevant biomaterial, supporting the use of MSC spheroids in cell-based approaches to bone repair. PMID:24781147

Investigation of dose characteristics in three-dimensional MAGAT-type polymer gel dosimetry with MSE MR imaging

NASA Astrophysics Data System (ADS)

Lee, Jason J. S.; Tsai, Chia-Jung; Lo, Man-Kuok; Huang, Yung-Hui; Chen, Chien-Chuan; Wu, Jay; Tyan, Yeu-Sheng; Wu, Tung-Hsin

2008-05-01

A new type of normoxic polymer gel dosimeter, named MAGAT responses well to absorbed dose even when manufacturing in the presence of normal levels of oxygen. The aim of this study was to evaluate dose response, diffusion effect and cumulated dose response under multiple fractional irradiations of the MAGAT gel dosimeter using Multiple Spin-Echo (MSE) Magnetic Resonance (MR) sequence. Dose response was performed by irradiating MAGAT-gel-filled testing vials with a 6 MV linear accelerator and a linear relationship was present with doses from 0 to 6 Gy, but gradually, a bi-exponential function result was obtained with given doses up to 20 Gy. No significant difference in dose response was present between single and cumulated doses (p > 0.05). For study of diffusion effect, edge sharpness of the R2 map imaging between two split doses was smaller than 1 cm of dose profile penumbra between 20% and 80%. In conclusion, the MAGAT polymer gel dosimeter with MSE MR imaging is a promising method for dose verification in clinical radiation therapy practice.

Swimming micro-robot powered by stimuli-sensitive gel

NASA Astrophysics Data System (ADS)

Masoud, Hassan; Alexeev, Alexander

2012-11-01

Using three-dimensional computer simulations, we design a simple maneuverable micro-swimmer that can self-propel and navigate in highly viscous (low Reynolds-number) environments. Our simple swimmer consists of a cubic gel body which periodically changes volume in response to external stimuli, two rigid rectangular flaps attached to the opposite sides of the gel body, and a flexible steering flap at the front end of the swimmer. The stimuli-sensitive body undergoes periodic expansions (swelling) and contractions (deswelling) leading to a time-irreversible beating motion of the propulsive flaps that propel the micro-swimmer. Thus, the responsive gel body acts as an ``engine'' actuating the motion of the swimmer. We examine how the swimming speed depends on the gel and flap properties. We also probe how the swimmer trajectory can be changed using a responsive steering flap whose curvature is controlled by an external stimulus. We show that the turning occurs due to steering flap bending and periodic beating. Furthermore, our simulations reveal that the turning direction can be regulated by changing the intensity of external stimulus.

Analysis of proteomic differences between liquefied after-cataracts and normal lenses using two-dimensional gel electrophoresis and mass spectrometry

PubMed Central

Ge, Jia-Jia; Huang, Yu-Sen

2017-01-01

AIM To analyze and identify the proteomic differences between liquefied after-cataracts and normal lenses by means of liquefied chromatography-tandem mass spectrometry (LC-MS/MS). METHODS Three normal lenses and three liquefied after-cataracts were exposed to depolymerizing reagents to extract the total proteins. Protein concentrations were separated using two-dimensional gel electrophoresis (2-DE). The digitized images obtained with a GS-800 scanner were then analyzed with PDQuest7.0 software to detect the differentially-expressed protein spots. These protein spots were cut from the gel using a proteome work spot cutter and subjected to in-gel digestion with trypsin. The digested peptide separation was conducted by LC-MS/MS. RESULTS The 2-DE maps showed that lens proteins were in a pH range of 3-10 with a relative molecular weight of 21-70 kD. The relative molecular weight of the more abundant proteins was localized at 25-50 kD, and the isoelectric points were found to lie between PI 4-9. The maps also showed that the protein level within the liquefied after-cataracts was at 29 points and significantly lower than in normal lenses. The 29 points were identified by LC-MS/MS, and ten of these proteins were identified by mass spectrometry and database queries: beta-crystallin B1, glyceraldehyde-3-phosphate dehydrogenase, carbonyl reductase (NADPH) 1, cDNA FLJ55253, gamma-crystallin D, GAS2-like protein 3, sorbitol dehydrogenase, DNA FLJ60282, phosphoglycerate kinase, and filensin. CONCLUSION The level of the ten proteins may play an important role in the development of liquefied after-cataracts. PMID:28944190

Combining high-throughput MALDI-TOF mass spectrometry and isoelectric focusing gel electrophoresis for virtual 2D gel-based proteomics.

PubMed

Lohnes, Karen; Quebbemann, Neil R; Liu, Kate; Kobzeff, Fred; Loo, Joseph A; Ogorzalek Loo, Rachel R

2016-07-15

The virtual two-dimensional gel electrophoresis/mass spectrometry (virtual 2D gel/MS) technology combines the premier, high-resolution capabilities of 2D gel electrophoresis with the sensitivity and high mass accuracy of mass spectrometry (MS). Intact proteins separated by isoelectric focusing (IEF) gel electrophoresis are imaged from immobilized pH gradient (IPG) polyacrylamide gels (the first dimension of classic 2D-PAGE) by matrix-assisted laser desorption/ionization (MALDI) MS. Obtaining accurate intact masses from sub-picomole-level proteins embedded in 2D-PAGE gels or in IPG strips is desirable to elucidate how the protein of one spot identified as protein 'A' on a 2D gel differs from the protein of another spot identified as the same protein, whenever tryptic peptide maps fail to resolve the issue. This task, however, has been extremely challenging. Virtual 2D gel/MS provides access to these intact masses. Modifications to our matrix deposition procedure improve the reliability with which IPG gels can be prepared; the new procedure is described. Development of this MALDI MS imaging (MSI) method for high-throughput MS with integrated 'top-down' MS to elucidate protein isoforms from complex biological samples is described and it is demonstrated that a 4-cm IPG gel segment can now be imaged in approximately 5min. Gel-wide chemical and enzymatic methods with further interrogation by MALDI MS/MS provide identifications, sequence-related information, and post-translational/transcriptional modification information. The MSI-based virtual 2D gel/MS platform may potentially link the benefits of 'top-down' and 'bottom-up' proteomics. Copyright © 2016 Elsevier Inc. All rights reserved.

Generating multiple independent retention index data in dual-secondary column comprehensive two-dimensional gas chromatography.

PubMed

Bieri, Stefan; Marriott, Philip J

2006-12-01

A method producing simultaneously three retention indexes for compounds has been developed for comprehensive two-dimensional gas chromatography by using a dual secondary column approach (GC x 2GC). For this purpose, the primary flow of the first dimension column was equally diverted into two secondary microbore columns of identical geometry by means of a three-way flow splitter positioned after the longitudinally modulated cryogenic system. This configuration produced a pair of comprehensive two-dimensional chromatograms and generated retention data on three different stationary phases in a single run. First dimension retention indexes were determined on a polar SolGel-Wax column under linear programmed-temperature conditions according to the van den Dool approach using primary alcohol homologues as the reference scale. Calculation of pseudoisothermal retention indexes in both second dimensions was performed on low-polarity 5% phenyl equivalent polysilphenylene/siloxane (BPX5) and 14% cyanopropylphenyl/86% dimethylpolysiloxane (BP10) columns. To construct a retention correlation map in the second dimension separation space upon which KovAts indexes can be derived, two methods exploiting "isovolatility" relationships of alkanes were developed. The first involved 15 sequential headspace samplings of selected n-alkanes by solid-phase microextraction (SPME), with each sampling followed by their injection into the GC at predetermined times during the chromatographic run. The second method extended the second dimension retention map and consisted of repetitive introduction of SPME-sampled alkane mixtures at various isothermal conditions incremented over the temperature program range. Calculated second dimension retention indexes were compared with experimental values obtained in conventional one-dimensional GC. A case study mixture including 24 suspected allergens (i.e., fragrance ingredients) was used to demonstrate the feasibility and potential of retention index information in comprehensive 2D-GC.

Direct visualization of in vitro drug mobilization from Lescol XL tablets using two-dimensional (19)F and (1)H magnetic resonance imaging.

PubMed

Chen, Chen; Gladden, Lynn F; Mantle, Michael D

2014-02-03

This article reports the application of in vitro multinuclear ((19)F and (1)H) two-dimensional magnetic resonance imaging (MRI) to study both dissolution media ingress and drug egress from a commercial Lescol XL extended release tablet in a United States Pharmacopeia Type IV (USP-IV) dissolution cell under pharmacopoeial conditions. Noninvasive spatial maps of tablet swelling and dissolution, as well as the mobilization and distribution of the drug are quantified and visualized. Two-dimensional active pharmaceutical ingredient (API) mobilization and distribution maps were obtained via (19)F MRI. (19)F API maps were coregistered with (1)H T2-relaxation time maps enabling the simultaneous visualization of drug distribution and gel layer dynamics within the swollen tablet. The behavior of the MRI data is also discussed in terms of its relationship to the UV drug release behavior.

Aluminum/hydrocarbon gel propellants: An experimental and theoretical investigation of secondary atomization and predicted rocket engine performance

NASA Astrophysics Data System (ADS)

Mueller, Donn Christopher

1997-12-01

Experimental and theoretical investigations of aluminum/hydrocarbon gel propellant secondary atomization and its potential effects on rocket engine performance were conducted. In the experimental efforts, a dilute, polydisperse, gel droplet spray was injected into the postflame region of a burner and droplet size distributions was measured as a function of position above the burner using a laser-based sizing/velocimetry technique. The sizing/velocimetry technique was developed to measure droplets in the 10-125 mum size range and avoids size-biased detection through the use of a uniformly illuminated probe volume. The technique was used to determine particle size distributions and velocities at various axial locations above the burner for JP-10, and 50 and 60 wt% aluminum gels. Droplet shell formation models were applied to aluminum/hydrocarbon gels to examine particle size and mass loading effects on the minimum droplet diameter that will permit secondary atomization. This diameter was predicted to be 38.1 and 34.7 mum for the 50 and 60 wt% gels, which is somewhat greater than the experimentally measured 30 and 25 mum diameters. In the theoretical efforts, three models were developed and an existing rocket code was exercised to gain insights into secondary atomization. The first model was designed to predict gel droplet properties and shell stresses after rigid shell formation, while the second, a one-dimensional gel spray combustion model was created to quantify the secondary atomization process. Experimental and numerical comparisons verify that secondary atomization occurs in 10-125 mum diameter particles although an exact model could not be derived. The third model, a one-dimensional gel-fueled rocket combustion chamber, was developed to evaluate secondary atomization effects on various engine performance parameters. Results show that only modest secondary atomization may be required to reduce propellant burnout distance and radiation losses. A solid propellant engine code was employed to estimate nozzle two-phase flow losses and engine performance for upper-stage and booster missions (3-6% and 2-3%, respectively). Given these losses and other difficulties, metallized gel propellants may be impractical in high-expansion ratio engines. Although uncertainties remain, it appears that performance gains will be minimal in gross-weight limited missions, but that significant gains may arise in volume-limited missions.

From the Macro to the Micro: Gel Mapping to Differentiate between Sporozoites of Two Immunologically Distinct Strains of Eimeria maxima (Strains M6 and Guelph)

PubMed Central

Liu, Hongbin; Al Nasr, Ibrahim; Liu, Xianyong; Suo, Xun; Barta, John

2015-01-01

Two immunologically distinct strains of E. maxima were examined in this study: the M6 strain and the Guelph strain. The differential expression between the sporozoites of the two strains of E. maxima was determined by image analysis of 100 μg of protein from each strain separated by standard one- and conventional two-dimensional polyacrylamide gel electrophoresis. In addition to differences in both molecular weight and the electrophoretic mobility, differences in the intensity of polypeptide bands for example, GS 136.4 and M6 169 were explored. Pooled gels were prepared from each strain. A representative 2D-PAGE gel spanning a non-linear pH range of 3–10 of E. maxima strain M6 consisted of approximately 694 polypeptide spots with about 67 (9.6%) of the polypeptide spots being unique relative to the other strain. E. maxima strain GS had about 696 discernable polypeptide spots with 69 spots (9.9%) that differed from those of the M6 strain. In-depth characterization of the variable polypeptide spots; unique polypeptide spots (absence or presence) and shared polypeptide spots with modifications may lead to novel vaccine target in the form of multi-component, multi-stage, multi-immunovariant strains, multi-species subunit vaccine, and diagnostic probe for E. maxima. PMID:26641262

From the Macro to the Micro: Gel Mapping to Differentiate between Sporozoites of Two Immunologically Distinct Strains of Eimeria maxima (Strains M6 and Guelph).

PubMed

El-Ashram, Saeed; Yin, Qing; Liu, Hongbin; Al Nasr, Ibrahim; Liu, Xianyong; Suo, Xun; Barta, John

2015-01-01

Two immunologically distinct strains of E. maxima were examined in this study: the M6 strain and the Guelph strain. The differential expression between the sporozoites of the two strains of E. maxima was determined by image analysis of 100 μg of protein from each strain separated by standard one- and conventional two-dimensional polyacrylamide gel electrophoresis. In addition to differences in both molecular weight and the electrophoretic mobility, differences in the intensity of polypeptide bands for example, GS 136.4 and M6 169 were explored. Pooled gels were prepared from each strain. A representative 2D-PAGE gel spanning a non-linear pH range of 3-10 of E. maxima strain M6 consisted of approximately 694 polypeptide spots with about 67 (9.6%) of the polypeptide spots being unique relative to the other strain. E. maxima strain GS had about 696 discernable polypeptide spots with 69 spots (9.9%) that differed from those of the M6 strain. In-depth characterization of the variable polypeptide spots; unique polypeptide spots (absence or presence) and shared polypeptide spots with modifications may lead to novel vaccine target in the form of multi-component, multi-stage, multi-immunovariant strains, multi-species subunit vaccine, and diagnostic probe for E. maxima.

Proteomic Analysis of Synovial Fluid Obtained From a Dog Diagnosed With Idiopathic Immune-Mediated Polyarthritis.

PubMed

Tan, Wei Miao; Lau, Seng Fong; Ajat, Mokrish; Mansor, Rozaihan; Abd Rani, Puteri Azaziah Megat; Rahmad, Norasfaliza Binti

2017-03-01

This case study is to report the proteins detected by proteomic analysis of synovial fluid from a dog diagnosed with idiopathic immune-mediated polyarthritis, and to compare it with healthy dogs. Synovial fluid was collected via arthrocentesis from a dog diagnosed with immune-mediated polyarthritis. Protein precipitation was performed on the synovial fluid, followed by isoelectric focusing and 2-dimensional gel electrophoresis. The spots on the 2-dimensional gels were analyzed using MALDI-TOF/MS. The results were then analyzed against the MASCOT database. The results from the proteomic analysis revealed an abundance of several types of immunoglobulins together with the presence of complement C4b-binding protein alpha chain. Actin and keratin were also among the proteins detected. Proteomic studies, facilitate a better understanding of the different levels of proteins expressed during disease activity. Potential disease biomarkers can aid in the diagnosis of disease, as well as help in monitoring treatment efficacy and providing prognosis for the patient. Copyright © 2017 Elsevier Inc. All rights reserved.

The characterisation of novel secreted Ly-6 proteins from rat urine by the combined use of two-dimensional gel electrophoresis, microbore high performance liquid chromatography and expressed sequence tag data.

PubMed

Southan, Christopher; Cutler, Paul; Birrell, Helen; Connell, John; Fantom, Kenneth G M; Sims, Matthew; Shaikh, Narjis; Schneider, Klaus

2002-02-01

A proteomic study of rat urine was undertaken using two-dimensional gel electrophoresis, microbore high performance liquid chromatography, mass spectrometry and N-terminal sequencing. Five known urinary proteins were identified but two novel peptide fragments matched a large number of rat expressed sequence tags (ESTs) from a liver library. By combining protein chemical and nucleotide data, two 101-residue open reading frames with 90% amino acid identity were determined, rat urinary protein 1 (RUP-1) and RUP-2. The data established signal peptide removal and provided evidence for N-glycosylation. A third related sequence, rat spleen protein (RSP-1) was confirmed from EST searches. These three proteins have been submitted to SWISS-PROT as P81827, P81828 and Q9QXN2, respectively. A fourth novel homologue was found in porcine and bovine ESTs from embryo libraries. Alignment with known homologues showed conserved cysteine positions characteristic of a secreted subfamily of Ly-6 proteins. In two cases, antineoplastic urinary protein and caltrin, these homologues have unverified functional annotations. The RUP sequences showed high scoring matches to three unrelated rat mRNAs subsequently established to be chimeric. Two of these share extended sectional identity to RUP-1 but the third may represent another novel Ly-6 homologue. These chimeras have caused serious annotation errors in secondary databases.

Studies on a new class of organogelator containing 2-anthracenecarboxylic acid: influence of gelator and solvent on stereochemistry of the photodimers.

PubMed

Dawn, Arnab; Fujita, Norifumi; Haraguchi, Shuichi; Sada, Kazuki; Tamaru, Shun-ichi; Shinkai, Seiji

2009-11-07

A new class of binary organogelator (G1, G2 and G3) based on 2-anthracenecarboxylic acid (2Ac), attached noncovalently with the gelator counterpart containing a 3,4,5-tris(n-dodecyloxy)benzoylamide backbone has been developed. Among the three gelators, two (G2 and G3) are chiral containing D-alanine or L-2-phenylglycine moieties, respectively. They can act as efficient gelators of organic solvents with varying polarity depending upon the gelator systems. Gelator G1 even gelates chiral solvents. The photoirradiation of the gel samples produces photocyclodimers having different degrees of stereoselectivity for different systems. Gels with G1 and G2 produce head-to-head (h-h) photodimers as major products, whereas the stereoselectivity is reversed for the gels with G3 producing head-to-tail (h-t) photodimers as major products. Among those, G2/cyclohexane gel shows the highest degree of stereoselectivity, producing only h-h photodimers with some significant amount of chiral induction. Other chiral systems exhibit low to moderate chiral inductions. The gelator G1 can differentiate between the racemic and enantiomerically pure varieties of a solvent by exhibiting different gel melting temperatures (T(gel)). For different gel systems, T(gel) increases in all the cases as a consequence of photoreaction, except for the G2/cyclohexane gel, where a prominent gel-to-sol phase transition can be observed during the photoreaction. Hydrogen-bonding and pi-pi stacking interactions play the principal roles in constructing the gel structure. The morphologies of the gel systems vary between one-dimensional fibrils and a fibrillar network structure. In addition, the influences of the gelator and solvent polarity on the rate of photoreactions, photoproduct distributions as well as gel structures are investigated.

Quantitative analysis of fragrance in selectable one dimensional or two dimensional gas chromatography-mass spectrometry with simultaneous detection of multiple detectors in single injection.

PubMed

Tan, Hui Peng; Wan, Tow Shi; Min, Christina Liew Shu; Osborne, Murray; Ng, Khim Hui

2014-03-14

A selectable one-dimensional ((1)D) or two-dimensional ((2)D) gas chromatography-mass spectrometry (GC-MS) system coupled with flame ionization detector (FID) and olfactory detection port (ODP) was employed in this study to analyze perfume oil and fragrance in shower gel. A split/splitless (SSL) injector and a programmable temperature vaporization (PTV) injector are connected via a 2-way splitter of capillary flow technology (CFT) in this selectable (1)D/(2)D GC-MS/FID/ODP system to facilitate liquid sample injections and thermal desorption (TD) for stir bar sorptive extraction (SBSE) technique, respectively. The dual-linked injectors set-up enable the use of two different injector ports (one at a time) in single sequence run without having to relocate the (1)D capillary column from one inlet to another. Target analytes were separated in (1)D GC-MS/FID/ODP and followed by further separation of co-elution mixture from (1)D in (2)D GC-MS/FID/ODP in single injection without any instrumental reconfiguration. A (1)D/(2)D quantitative analysis method was developed and validated for its repeatability - tR; calculated linear retention indices (LRI); response ratio in both MS and FID signal, limit of detection (LOD), limit of quantitation (LOQ), as well as linearity over a concentration range. The method was successfully applied in quantitative analysis of perfume solution at different concentration level (RSD≤0.01%, n=5) and shower gel spiked with perfume at different dosages (RSD≤0.04%, n=5) with good recovery (96-103% for SSL injection; 94-107% for stir bar sorptive extraction-thermal desorption (SBSE-TD). Copyright © 2014 Elsevier B.V. All rights reserved.

Serum proteomic profiling in granumomatosis with polyangiitis using two-dimensional gel electrophoresis along with matrix assisted laser desorption ionization time of flight mass spectrometry.

PubMed

Rani, Lekha; Minz, Ranjana W; Arora, Amit; Kannan, Monica; Sharma, Aman; Anand, Shashi; Gupta, Dheeraj; Panda, Naresh K; Sakhuja, Vinay K

2014-11-01

The present study is a proteomic approach to find differentially expressed proteins in sera of limited and systemic subsets of active disease versus their remitting state in patients with granulomatosis with polyangiitis (GPA) and their correlation with disease activity. Eighteen patients with GPA in active as well as in remitting state and four healthy controls (HC) were included in the study. For proteomics analysis, two-dimensional gel electrophoresis along with matrix-assisted laser desorption ionization time-of-flight mass spectrometry were performed. A total of 14 gels were run from pooled patients' sera from active GPA and remission as well as pooled HC serum. There was significant differential expression of proteins in limited versus systemic GPA and between active systemic versus remitting patients of systemic disease. We identified nine maximally differentially expressed and five proteins which were not detected in HC. Among nine proteins, one (Prolow density lipoprotein receptor-related protein 1) was downregulated and four proteins (haptoglobin Hp, Hp2, vitamin D binding protein, killer cell lectin-like receptor subfamily F member 2), were up-regulated in both limited and systemic active disease, two proteins like Ig gamma-4 chain C region protein and serum albumin were up-regulated in limited active GPA and two proteins, that is, cysteine rich secretory protein LCCL domain-containing 2 precursor and serine-threonine-protein kinase A-Raf were up-regulated in systemic active disease. Levels of interleukin-17 and vitamin-D binding protein (VDBP) by enzyme-linked immunosorbent assay could distinctly demarcate active disease versus remission. Our study provides potential protein markers of active disease versus remission in GPA. © 2014 Asia Pacific League of Associations for Rheumatology and Wiley Publishing Asia Pty Ltd.

Development of a patient-specific 3D dose evaluation program for QA in radiation therapy

NASA Astrophysics Data System (ADS)

Lee, Suk; Chang, Kyung Hwan; Cao, Yuan Jie; Shim, Jang Bo; Yang, Dae Sik; Park, Young Je; Yoon, Won Sup; Kim, Chul Yong

2015-03-01

We present preliminary results for a 3-dimensional dose evaluation software system ( P DRESS, patient-specific 3-dimensional dose real evaluation system). Scanned computed tomography (CT) images obtained by using dosimetry were transferred to the radiation treatment planning system (ECLIPSE, VARIAN, Palo Alto, CA) where the intensity modulated radiation therapy (IMRT) nasopharynx plan was designed. We used a 10 MV photon beam (CLiX, VARIAN, Palo Alto, CA) to deliver the nasopharynx treatment plan. After irradiation, the TENOMAG dosimeter was scanned using a VISTA ™ scanner. The scanned data were reconstructed using VistaRecon software to obtain a 3D dose distribution of the optical density. An optical-CT scanner was used to readout the dose distribution in the gel dosimeter. Moreover, we developed the P DRESS by using Flatform, which were developed by our group, to display the 3D dose distribution by loading the DICOM RT data which are exported from the radiotherapy treatment plan (RTP) and the optical-CT reconstructed VFF file, into the independent P DRESS with an ioniz ation chamber and EBT film was used to compare the dose distribution calculated from the RTP with that measured by using a gel dosimeter. The agreement between the normalized EBT, the gel dosimeter and RTP data was evaluated using both qualitative and quantitative methods, such as the isodose distribution, dose difference, point value, and profile. The profiles showed good agreement between the RTP data and the gel dosimeter data, and the precision of the dose distribution was within ±3%. The results from this study showed significantly discrepancies between the dose distribution calculated from the treatment plan and the dose distribution measured by a TENOMAG gel and by scanning with an optical CT scanner. The 3D dose evaluation software system ( P DRESS, patient specific dose real evaluation system), which were developed in this study evaluates the accuracies of the three-dimensional dose distributions. Further applications of the system utility are expected to result from future studies.

Polymer gel dosimeters for pretreatment radiotherapy verification using the three-dimensional gamma evaluation and pass rate maps.

PubMed

Hsieh, Ling-Ling; Shieh, Jiunn-I; Wei, Li-Ju; Wang, Yi-Chun; Cheng, Kai-Yuan; Shih, Cheng-Ting

2017-05-01

Polymer gel dosimeters (PGDs) have been widely studied for use in the pretreatment verification of clinical radiation therapy. However, the readability of PGDs in three-dimensional (3D) dosimetry remain unclear. In this study, the pretreatment verifications of clinical radiation therapy were performed using an N-isopropyl-acrylamide (NIPAM) PGD, and the results were used to evaluate the performance of the NIPAM PGD on 3D dose measurement. A gel phantom was used to measure the dose distribution of a clinical case of intensity-modulated radiation therapy. Magnetic resonance imaging scans were performed for dose readouts. The measured dose volumes were compared with the planned dose volume. The relative volume histograms showed that relative volumes with a negative percent dose difference decreased as time elapsed. Furthermore, the histograms revealed few changes after 24h postirradiation. For the 3%/3mm and 2%/2mm criteria, the pass rates of the 12- and 24-h dose volumes were higher than 95%, respectively. This study thus concludes that the pass rate map can be used to evaluate the dose-temporal readability of PGDs and that the NIPAM PGD can be used for clinical pretreatment verifications. Copyright © 2017 Associazione Italiana di Fisica Medica. Published by Elsevier Ltd. All rights reserved.

Quantitative molecular characterization of bovine vitreous and lens with non-invasive dynamic light scattering

NASA Technical Reports Server (NTRS)

Ansari, R. R.; Suh, K. I.; Dunker, S.; Kitaya, N.; Sebag, J.

2001-01-01

The non-invasive technique of dynamic light scattering (DLS) was used to quantitatively characterize vitreous and lens structure on a molecular level by measuring the sizes of the predominant particles and mapping the three-dimensional topographic distribution of these structural macromolecules in three spatial dimensions. The results of DLS measurements in five fresh adult bovine eyes were compared to DLS measurements in model solutions of hyaluronan (HA) and collagen (Coll). In the bovine eyes DLS measurements were obtained from excised samples of gel and liquid vitreous and compared to the model solutions. Measurements in whole vitreous were obtained at multiple points posterior to the lens to generate a three-dimensional 'map' of molecular structure. The macromolecule distribution in bovine lens was similarly characterized.In each bovine vitreous (Bo Vit) specimen, DLS predominantly detected two distinct particles, which differed in diffusion properties and hence size. Comparisons with model vitreous solutions demonstrated that these most likely corresponded to the Coll and HA components of vitreous. Three-dimensional mapping of Bo Vit found heterogeneity throughout the vitreous body, with different particle size distributions for Coll and HA at different loci. In contrast, the three-dimensional distribution of lens macromolecules was more homogeneous. Thus, the non-invasive DLS technique can quantitate the average sizes of vitreous and lens macromolecules and map their three-dimensional distribution. This method to assess quantitatively the macromolecular structure of vitreous and lens should be useful for clinical as well as experimental applications in health and disease. Copyright 2001 Academic Press.

Proteomic analysis of differentially expressed proteins in kidneys of brain dead rabbits

PubMed Central

Li, Ling; Li, Ning; He, Chongxiang; Huang, Wei; Fan, Xiaoli; Zhong, Zibiao; Wang, Yanfeng; Ye, Qifa

2017-01-01

A large number of previous clinical studies have reported a delayed graft function for brain dead donors, when compared with living relatives or cadaveric organ transplantations. However, there is no accurate method for the quality evaluation of kidneys from brain-dead donors. In the present study, two-dimensional gel electrophoresis and MALDI-TOF MS-based comparative proteomic analysis were conducted to profile the differentially-expressed proteins between brain death and the control group renal tissues. A total of 40 age- and sex-matched rabbits were randomly divided into donation following brain death (DBD) and control groups. Following the induction of brain death via intracranial progressive pressure, the renal function and the morphological alterations were measured 2, 6 and 8 h afterwards. The differentially expressed proteins were detected from renal histological evidence at 6 h following brain death. Although 904±19 protein spots in control groups and 916±25 in DBD groups were identified in the two-dimensional gel electrophoresis, >2-fold alterations were identified by MALDI-TOF MS and searched by NCBI database. The authors successfully acquired five downregulated proteins, these were: Prohibitin (isoform CRA_b), beta-1,3-N-acetylgalactosaminyltransferase 1, Annexin A5, superoxide dismutase (mitochondrial) and cytochrome b-c1 complex subunit 1 (mitochondrial precursor). Conversely, the other five upregulated proteins were: PRP38 pre-mRNA processing factor 38 (yeast) domain containing A, calcineurin subunit B type 1, V-type proton ATPase subunit G 1, NADH dehydrogenase [ubiquinone] 1 beta subcomplex subunit 10 and peroxiredoxin-3 (mitochondrial). Immunohistochemical results revealed that the expressions of prohibitin (PHB) were gradually increased in a time-dependent manner. The results indicated that there were alterations in levels of several proteins in the kidneys of those with brain death, even if the primary function and the morphological changes were not obvious. PHB may therefore be a novel biomarker for primary quality evaluation of kidneys from brain-dead donors. PMID:28534953

Neurite outgrowth at the interface of 2D and 3D growth environments

NASA Astrophysics Data System (ADS)

Kofron, Celinda M.; Fong, Vivian J.; Hoffman-Kim, Diane

2009-02-01

Growing neurons navigate complex environments, but in vitro systems for studying neuronal growth typically limit the cues to flat surfaces or a single type of cue, thereby limiting the resulting growth. Here we examined the growth of neurons presented with two-dimensional (2D) substrate-bound cues when these cues were presented in conjunction with a more complex three-dimensional (3D) architecture. Dorsal root ganglia (DRG) explants were cultured at the interface between a collagen I matrix and a glass coverslip. Laminin (LN) or chondroitin sulfate proteoglycans (CSPG) were uniformly coated on the surface of the glass coverslip or patterned in 50 µm tracks by microcontact printing. Quantitative analysis of neurite outgrowth with a novel grid system at multiple depths in the gel revealed several interesting trends. Most of the neurites extended at the surface of the gel when LN was presented whereas more neurites extended into the gel when CSPG was presented. Patterning of cues did not affect neurite density or depth of growth. However, neurite outgrowth near the surface of the gel aligned with LN patterns, and these extensions were significantly longer than neurites extended in other cultures. In interface cultures, DRG growth patterns varied with the type of cue where neurite density was higher in cultures presenting LN than in cultures presenting CSPG. These results represent an important step toward understanding how neurons integrate local structural and chemical cues to make net growth decisions.

PLMA vs. I-gel: A Comparative Evaluation of Respiratory Mechanics in Laparoscopic Cholecystectomy

PubMed Central

Sharma, Bimla; Sehgal, Raminder; Sahai, Chand; Sood, Jayashree

2010-01-01

Background: Supraglottic airway devices (SADs), such as ProSealTM laryngeal mask airway (PLMA), which produce high oropharyngeal seal pressure (OSP) and have the facility for gastric decompression have been used in laparoscopic procedures. i-gel is a new SAD which shares these features with the PLMA. This study was designed to compare the respiratory mechanics of these two devices during positive pressure ventilation in anaesthetised adult patients undergoing laparoscopic cholecystectomy. Patients & Methods: The study included 60 ASA I-II adult patients scheduled for laparoscopic cholecystectomy. The patients were randomized to two groups of 30 each, with either PLMA or i-gel as their airway device. Anaesthesia and premedication were standardized for both the groups. In addition to routine monitoring, neuromuscular monitoring with TOF ratio, OSP and respiratory mechanics monitoring (dynamic compliance, resistance, work of breathing, measured minute ventilation and peak airway pressures) were employed. Fibreoptic evaluation of positioning of the devices and adverse events related to them were also compared. Results: The OSP (cm H2O) were higher for PLMA (38.9 vs. 35.6, P=0.007). The respiratory mechanics parameters using the two devices were comparable apart from the dynamic compliance, which was significantly higher with i-gel (P < 0.05). Malrotation was higher with i-gel than with PLMA (15 vs. 5, P = 0.006). Conclusion: The PLMA formed a better seal while the dynamic compliance was higher with the i-gel. Both devices provided optimal ventilation and oxygenation and the adverse events were also comparable. PMID:21547168

Comparative Proteomic Analysis of Two Varieties of Genetically Modified (GM) Embrapa 5.1 Common Bean (Phaseolus vulgaris L.) and Their Non-GM Counterparts.

PubMed

Balsamo, Geisi M; Valentim-Neto, Pedro A; Mello, Carla S; Arisi, Ana C M

2015-12-09

The genetically modified (GM) common bean event Embrapa 5.1 was commercially approved in Brazil in 2011; it is resistant to golden mosaic virus infection. In the present work grain proteome profiles of two Embrapa 5.1 common bean varieties, Pérola and Pontal, and their non-GM counterparts were compared by two-dimensional gel electrophoresis (2-DE) followed by mass spectrometry (MS). Analyses detected 23 spots differentially accumulated between GM Pérola and non-GM Pérola and 21 spots between GM Pontal and non-GM Pontal, although they were not the same proteins in Pérola and Pontal varieties, indicating that the variability observed may not be due to the genetic transformation. Among them, eight proteins were identified in Pérola varieties, and four proteins were identified in Pontal. Moreover, we applied principal component analysis (PCA) on 2-DE data, and variation between varieties was explained in the first two principal components. This work provides a first 2-DE-MS/MS-based analysis of Embrapa 5.1 common bean grains.

Aqueous clay suspensions stabilized by alginate fluid gels for coal spontaneous combustion prevention and control.

PubMed

Qin, Botao; Ma, Dong; Li, Fanglei; Li, Yong

2017-11-01

We have developed aqueous clay suspensions stabilized by alginate fluid gels (AFG) for coal spontaneous combustion prevention and control. Specially, this study aimed to characterize the effect of AFG on the microstructure, static and dynamic stability, and coal fire inhibition performances of the prepared AFG-stabilized clay suspensions. Compared with aqueous clay suspensions, the AFG-stabilized clay suspensions manifest high static and dynamic stability, which can be ascribed to the formation of a robust three-dimensional gel network by AFG. The coal acceleration oxidation experimental results show that the prepared AFG-stabilized clay suspensions can improve the coal thermal stability and effectively inhibit the coal spontaneous oxidation process by increasing crossing point temperature (CPT) and reducing CO emission. The prepared low-cost and nontoxic AFG-stabilized clay suspensions, exhibiting excellent coal fire extinguishing performances, indicate great application potentials in coal spontaneous combustion prevention and control.

Multiple forms of ADP-glucose pyrophosphorylase from tomato fruit

NASA Technical Reports Server (NTRS)

Chen, B. Y.; Janes, H. W.

1997-01-01

ADP-glucose pyrophosphorylase (AGP) was purified from tomato (Lycopersicon esculentum Mill.) fruit to apparent homogeneity. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis the enzyme migrated as two close bands with molecular weights of 50,000 and 51,000. Two-dimensional polyacrylamide gel electrophoresis analysis of the purified enzyme, however, revealed at least five major protein spots that could be distinguished by their slight differences in net charge and molecular weight. Whereas all of the spots were recognized by the antiserum raised against tomato fruit AGP holoenzyme, only three of them reacted strongly with antiserum raised against the potato tuber AGP large subunit, and the other two spots (with lower molecular weights) reacted specifically with antisera raised against spinach leaf AGP holoenzyme and the potato tuber AGP small subunit. The results suggest the existence of at least three isoforms of the AGP large subunit and two isoforms of the small subunit in tomato fruit in vivo. The native molecular mass of the enzyme determined by gel filtration was 220 +/- 10 kD, indicating a tetrameric structure for AGP from tomato fruit. The purified enzyme is very sensitive to 3-phosphoglycerate/inorganic phosphate regulation.

The proteins of Fusobacterium spp. involved in hydrogen sulfide production from L-cysteine.

PubMed

Basic, Amina; Blomqvist, Madeleine; Dahlén, Gunnar; Svensäter, Gunnel

2017-03-14

Hydrogen sulfide (H 2 S) is a toxic foul-smelling gas produced by subgingival biofilms in patients with periodontal disease and is suggested to be part of the pathogenesis of the disease. We studied the H 2 S-producing protein expression of bacterial strains associated with periodontal disease. Further, we examined the effect of a cysteine-rich growth environment on the synthesis of intracellular enzymes in F. nucleatum polymorphum ATCC 10953. The proteins were subjected to one-dimensional (1DE) and two-dimensional (2DE) gel electrophoresis An in-gel activity assay was used to detect the H 2 S-producing enzymes; Sulfide from H 2 S, produced by the enzymes in the gel, reacted with bismuth forming bismuth sulfide, illustrated as brown bands (1D) or spots (2D) in the gel. The discovered proteins were identified with liquid chromatography - tandem mass spectrometry (LC-MS/MS). Cysteine synthase and proteins involved in the production of the coenzyme pyridoxal 5'phosphate (that catalyzes the production of H 2 S) were frequently found among the discovered enzymes. Interestingly, a higher expression of H 2 S-producing enzymes was detected from bacteria incubated without cysteine prior to the experiment. Numerous enzymes, identified as cysteine synthase, were involved in the production of H 2 S from cysteine and the expression varied among Fusobacterium spp. and strains. No enzymes were detected with the in-gel activity assay among the other periodontitis-associated bacteria tested. The expression of the H 2 S-producing enzymes was dependent on environmental conditions such as cysteine concentration and pH but less dependent on the presence of serum and hemin.

Bacterial community structure analysis of sediment in the Sagami River, Japan using a rapid approach based on two-dimensional DNA gel electrophoresis mapping with selective primer pairs.

PubMed

Liu, Guo-hua; Rajendran, Narasimmalu; Amemiya, Takashi; Itoh, Kiminori

2011-11-01

A rapid approach based on two-dimensional DNA gel electrophroesis (2-DGE) mapping with selective primer pairs was employed to analyze bacterial community structure in sediments from upstream, midstream and downstream of Sagami River in Japan. The 2-DGE maps indicated that Alpha- and Delta-proteobacteria were major bacterial populations in the upstream and midstream sediments. Further bacterial community structure analysis showed that richness proportion of Alpha- and Delta-proteobacterial groups reflected a trend toward decreasing from the upstream to downstream sediments. The biomass proportion of bacterial populations in the midstream sediment showed a significantly difference from that in the other sediments, suggesting that there may be an environmental pressure on the midstream bacterial community. Lorenz curves, together with Gini coefficients were successfully applied to the 2-DGE mapping data for resolving evenness of bacterial populations, and showed that the plotted curve from high-resolution 2-DGE mapping became less linear and more an exponential function than that of the 1-DGE methods such as chain length analysis and denaturing gradient gel electrophoresis, suggesting that the 2-DGE mapping may achieve a more detailed evaluation of bacterial community. In conclusion, the 2-DGE mapping combined with the selective primer pairs enables bacterial community structure analysis in river sediment and thus it can also monitor sediment pollution based on the change of bacterial community structure.

Cellulose nanocrystal-based composite electrolyte with superior dimensional stability for alkaline fuel cell membranes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lu, Yuan; Artmentrout, Aaron A.; Li, Juchuan

2015-05-13

Cellulose nanocrystal (CNC)-based composite films were prepared as a solid electrolyte for alkaline fuel cells. Poly (vinyl alcohol) (PVA) and silica gel hybrid was used to bind the CNCs to form a robust composite film. The mass ratio (i.e., 1 : 1, 1 : 2) of PVA and silica gel was tuned to control the hydrophobicity of the resulting films. Composite films with a range of CNC content (i.e., 20 to 60%) were prepared to demonstrate the impact of CNC on the performance of these materials as a solid electrolyte for alkaline fuel cells. Different from previously reported cross-linked polymermore » films, CNC-based composite films with 40% hydrophobic binder (i.e., PVA : silica gel=1 : 2) exhibited simultaneous low water swelling (e.g., ~5%) and high water uptake (e.g., ~80%) due to the hydrophilicity and extraordinary dimensional stability of CNC. It also showed a conductivity of 0.044 and 0.065 S/cm at 20 and 60 oC, respectively. To the best of our knowledge, the film with 60% CNC and 40% binder is characterized by the lowest hydroxide conductivity-normalized swelling ratio. Decreased CNC content (i.e., 40 and 20%) resulted in comparable hydroxide conductivity but a greater swelling ratio. Finally, these results demonstrate the advantage of CNC as a key component for a solid electrolyte for alkaline fuel cells over conventional polymers, suggesting the great potential of CNCs in improving the dimensional stability while maintaining the conductivity of existing anion exchange membranes.« less

Tuning three-dimensional collagen matrix stiffness independently of collagen concentration modulates endothelial cell behavior.

PubMed

Mason, Brooke N; Starchenko, Alina; Williams, Rebecca M; Bonassar, Lawrence J; Reinhart-King, Cynthia A

2013-01-01

Numerous studies have described the effects of matrix stiffening on cell behavior using two-dimensional synthetic surfaces; however, less is known about the effects of matrix stiffening on cells embedded in three-dimensional in vivo-like matrices. A primary limitation in investigating the effects of matrix stiffness in three dimensions is the lack of materials that can be tuned to control stiffness independently of matrix density. Here, we use collagen-based scaffolds where the mechanical properties are tuned using non-enzymatic glycation of the collagen in solution, prior to polymerization. Collagen solutions glycated prior to polymerization result in collagen gels with a threefold increase in compressive modulus without significant changes to the collagen architecture. Using these scaffolds, we show that endothelial cell spreading increases with matrix stiffness, as does the number and length of angiogenic sprouts and the overall spheroid outgrowth. Differences in sprout length are maintained even when the receptor for advanced glycation end products is inhibited. Our results demonstrate the ability to de-couple matrix stiffness from matrix density and structure in collagen gels, and that increased matrix stiffness results in increased sprouting and outgrowth. Copyright © 2012 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Evaluation of polymer gels and MRI as a 3-D dosimeter for intensity-modulated radiation therapy.

PubMed

Low, D A; Dempsey, J F; Venkatesan, R; Mutic, S; Markman, J; Mark Haacke, E; Purdy, J A

1999-08-01

BANG gel (MGS Research, Inc., Guilford, CT) has been evaluated for measuring intensity-modulated radiation therapy (IMRT) dose distributions. Treatment plans with target doses of 1500 cGy were generated by the Peacock IMRT system (NOMOS Corp., Sewickley, PA) using test target volumes. The gels were enclosed in 13 cm outer diameter cylindrical glass vessels. Dose calibration was conducted using seven smaller (4 cm diameter) cylindrical glass vessels irradiated to 0-1800 cGy in 300 cGy increments. Three-dimensional maps of the proton relaxation rate R2 were obtained using a 1.5 T magnetic resonance imaging (MRI) system (Siemens Medical Systems, Erlangen, Germany) and correlated with dose. A Hahn spin echo sequence was used with TR = 3 s, TE = 20 and 100 ms, NEX = 1, using 1 x 1 x 3 mm3 voxels. The MRI measurements were repeated weekly to identify the gel-aging characteristics. Ionization chamber, thermoluminescent dosimetry (TLD), and film dosimetry measurements of the IMRT dose distributions were obtained to compare against the gel results. The other dosimeters were used in a phantom with the same external cross-section as the gel phantom. The irradiated R2 values of the large vessels did not precisely track the smaller vessels, so the ionization chamber measurements were used to normalize the gel dose distributions. The point-to-point standard deviation of the gel dose measurements was 7.0 cGy. When compared with the ionization chamber measurements averaged over the chamber volume, 1% agreement was obtained. Comparisons against radiographic film dose distribution measurements and the treatment planning dose distribution calculation were used to determine the spatial localization accuracy of the gel and MRI. Spatial localization was better than 2 mm, and the dose was accurately determined by the gel both within and outside the target. The TLD chips were placed throughout the phantom to determine gel measurement precision in high- and low-dose regions. A multidimensional dose comparison tool that simultaneously examines the dose-difference and distance-to-agreement was used to evaluate the gel in both low-and high-dose gradient regions. When 3% and 3 mm criteria were used for the comparisons, more than 90% of the TLD measurements agreed with the gel, with the worst of 309 TLD chip measurements disagreeing by 40% of the criteria. All four MRI measurement session gel-measured dose distributions were compared to evaluate the time behavior of the gel. The low-dose regions were evaluated by comparison with TLD measurements at selected points, while high-dose regions were evaluated by directly comparing measured dose distributions. Tests using the multidimensional comparison tool showed detectable degradation beyond one week postirradiation, but all low-dose measurements passed relative to the test criteria and the dose distributions showed few regions that failed.

Comparative Study of Early Cold-Regulated Proteins by Two-Dimensional Difference Gel Electrophoresis Reveals a Key Role for Phospholipase Dα1 in Mediating Cold Acclimation Signaling Pathway in Rice.

PubMed

Huo, Chenmin; Zhang, Baowen; Wang, Hui; Wang, Fawei; Liu, Meng; Gao, Yingjie; Zhang, Wenhua; Deng, Zhiping; Sun, Daye; Tang, Wenqiang

2016-04-01

To understand the early signaling steps that regulate cold responses in rice, two-dimensional difference gel electrophoresis (2-D DIGE)(1)was used to study early cold-regulated proteins in rice seedlings. Using mass spectrometry, 32 spots, which represent 26 unique proteins that showed an altered expression level within 5 min of cold treatment were identified. Among these proteins, Western blot analyses confirmed that the cellular phospholipase D α1 (OsPLDα1) protein level was increased as early as 1 min after cold treatment. Genetic studies showed that reducing the expression ofOsPLDα1makes rice plants more sensitive to chilling stress as well as cold acclimation increased freezing tolerance. Correspondingly, cold-regulated proteomic changes and the expression of the cold-responsive C repeat/dehydration-responsive element binding 1 (OsDREB1) family of transcription factors were inhibited in thepldα1mutant. We also found that the expression ofOsPLDα1is directly regulated by OsDREB1A. This transcriptional regulation ofOsPLDα1could provide positive feedback regulation of the cold signal transduction pathway in rice. OsPLDα1 hydrolyzes phosphatidylcholine to produce the signal molecule phosphatidic acid (PA). By lipid-overlay assay, we demonstrated that the rice cold signaling proteins, MAP kinase 6 (OsMPK6) and OsSIZ1, bind directly to PA. Taken together, our results suggest that OsPLDα1 plays a key role in transducing cold signaling in rice by producing PA and regulatingOsDREB1s' expression by OsMPK6, OsSIZ1, and possibly other PA-binding proteins. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Identification and In Silico Analysis of Major Redox Modulated Proteins from Brassica juncea Seedlings Using 2D Redox SDS PAGE (2-Dimensional Diagonal Redox Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis).

PubMed

Chaurasia, Satya Prakash; Deswal, Renu

2017-02-01

The thiol-disulphide exchange regulates the activity of proteins by redox modulation. Many studies to analyze reactive oxygen species (ROS), particularly, hydrogen peroxide (H 2 O 2 ) induced changes in the gene expression have been reported, but efforts to detect H 2 O 2 modified proteins are comparatively few. Two-dimensional diagonal redox sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE) was used to detect polypeptides which undergo thiol-disulphide exchange in Brassica juncea seedlings following H 2 O 2 (10 mM) treatment for 30 min. Eleven redox responsive polypeptides were identified which included cruciferin, NLI [Nuclear LIM (Lin11, Isl-1 & Mec-3 domains)] interacting protein phosphatase, RuBisCO (ribulose-1,5-bisphosphate carboxylase/oxygenase) large subunit, and myrosinase. Redox modulation of RuBisCO large subunit was further confirmed by western blotting. However, the small subunit of RuBisCO was not affected by these redox changes. All redox modulated targets except NLI interacting protein (although it contains two cysteines) showed oxidation sensitive cysteines by in silico analysis. Interestingly, interactome of cruciferin and myrosinase indicated that they may have additional function(s) beside their well-known roles in the seedling development and abiotic stress respectively. Cruciferin showed interactions with stress associated proteins like defensing-like protein 192 and 2-cys peroxiredoxin. Similarly, myrosinase showed interactions with nitrilase and cytochrome p450 which are involved in nitrogen metabolism and/or hormone biosynthesis. This simple procedure can be used to detect major stress mediated redox changes in other plants.

Evaluation of proteome alterations induced by cadmium stress in sunflower (Helianthus annuus L.) cultures.

PubMed

Lopes Júnior, Cícero Alves; Barbosa, Herbert de Sousa; Moretto Galazzi, Rodrigo; Ferreira Koolen, Hector Henrique; Gozzo, Fábio Cesar; Arruda, Marco Aurélio Zezzi

2015-09-01

The present study evaluates, at a proteomic level, changes in protein abundance in sunflower leaves in the absence or presence (at 50 or 700mg) of cadmium (as CdCl2). At the end of the cultivation period (45 days), proteins are extracted from leaves with phenol, separated by two-dimensional difference gel electrophoresis (2-D DIGE), and excised from the gels. The differential protein abundances (for proteins differing by more than 1.8 fold, which corresponds to 90% variation) are characterized using nESI-LC-MS/MS. The protein content decreases by approximately 41% in plants treated with 700mg Cd compared with control plants. By comparing all groups of plants evaluated in this study (Control vs. Cd-lower, Control vs. Cd-higher and Cd-lower vs. Cd-higher), 39 proteins are found differential and 18 accurately identified; the control vs. Cd-higher treatment is that presenting the most differential proteins. From identified proteins, those involved in energy and disease/defense (including stress), are the ribulose bisphosphate carboxylase large chain, transketolase, and heat shock proteins are the most differential abundant proteins. Thus, at the present study, photosynthesis is the main process affected by Cd in sunflowers, although these plants are highly tolerant to Cd. Copyright © 2015 Elsevier Inc. All rights reserved.

[Comparison of acetonitrile, ethanol and chromatographic column to eliminate high-abundance proteins in human serum].

PubMed

Li, Yin; Liao, Ming; He, Xiao; Zhou, Yi; Luo, Rong; Li, Hongtao; Wang, Yun; He, Min

2012-11-01

To compare the effects of acetonitrile precipitation, ethanol precipitation and multiple affinity chromatography column Human 14 removal to eliminate high-abundance proteins in human serum. Elimination of serum high-abundance proteins performed with acetonitrile precipitation, ethanol precipitation and multiple affinity chromatography column Human 14 removal. Bis-Tris Mini Gels electrophoresis and two-dimensional gel electrophoresis to detect the effect. Grey value analysis from 1-DE figure showed that after serum processed by acetonitrile method, multiple affinity chromatography column Human 14 removal method and ethanol method, the grey value of albumin changed into 157.2, 40.8 and 8.2 respectively from the original value of 19. 2-DE analysis results indicated that using multiple affinity chromatography column Human 14 method, the protein points noticeable increased by 137 compared to the original serum. After processed by acetonitrile method and ethanol method, the protein point reduced, but the low abundance protein point emerged. The acetonitrile precipitation could eliminate the vast majority of high abundance proteins in serum and gain more proteins of low molecular weight, ethanol precipitation could eliminate part of high abundance proteins in serum, but low abundance proteins less harvested, and multiple affinity chromatography column Human 14 method could effectively removed the high abundance proteins, and keep a large number of low abundance proteins.

2-DE Compared with iTRAQ-based Proteomic Analysis of the Functional Regulation of Proteins in Rhodococcus sp. BAP-1 Response to Fluoranthene

NASA Astrophysics Data System (ADS)

Xu, Jie; Wang, Hongqi; Kong, Dekang

2018-01-01

Although the degradation pathways of Polycyclic aromatic hydrocarbons (PAHs) have been extensively studied in many bacteria, the variations in the expression levels of the key functional regulation of proteins during catabolism are still not quantitatively understood. In this study, we compared two proteomic methods, that one is two-dimensional gel electrophoresis (2-DE), a traditional widely used way and the other is isobaric tags for relative and absolute quantization (iTRAQ), an innovative approach, in order to analyze the functional regulation at the protein level in high effective fluoranthene-degrading bacteria named Rhodococcus sp. BAP-1. The number of differentially expressed proteins identified using iTRAQ is much larger than employing 2-DE. Response to fluoranthene, the key over expressed proteins in BAP-1 were NADPH-dependent FMN reductase, 30S ribosomal protein S2, S-ribosylhomocysteinase, etc.; the significant down-regulated proteins were cytochrome ubiquinol oxidase subunit, NAD(P) transhydrogenase subunit alpha, 5-methyltetrahydropteroyltriglutamate-homocysteine methyltransferase, et al.

Layer-by-Layer Self-Assembling Gold Nanorods and Glucose Oxidase onto Carbon Nanotubes Functionalized Sol-Gel Matrix for an Amperometric Glucose Biosensor.

PubMed

Wu, Baoyan; Hou, Shihua; Miao, Zhiying; Zhang, Cong; Ji, Yanhong

2015-09-18

A novel amperometric glucose biosensor was fabricated by layer-by-layer self-assembly of gold nanorods (AuNRs) and glucose oxidase (GOD) onto single-walled carbon nanotubes (SWCNTs)-functionalized three-dimensional sol-gel matrix. A thiolated aqueous silica sol containing SWCNTs was first assembled on the surface of a cleaned Au electrode, and then the alternate self-assembly of AuNRs and GOD were repeated to assemble multilayer films of AuNRs-GOD onto SWCNTs-functionalized silica gel for optimizing the biosensor. Among the resulting glucose biosensors, the four layers of AuNRs-GOD-modified electrode showed the best performance. The sol-SWCNTs-(AuNRs- GOD)₄/Au biosensor exhibited a good linear range of 0.01-8 mM glucose, high sensitivity of 1.08 μA/mM, and fast amperometric response within 4 s. The good performance of the proposed glucose biosensor could be mainly attributed to the advantages of the three-dimensional sol-gel matrix and stereo self-assembly films, and the natural features of one-dimensional nanostructure SWCNTs and AuNRs. This study may provide a new facile way to fabricate the enzyme-based biosensor with high performance.

A pilot clinical trial comparing an acid-buffering formulation (ACIDFORM gel) with metronidazole gel for the treatment of symptomatic bacterial vaginosis

PubMed Central

Simoes, Jose A; Bahamondes, Luis G; Camargo, Rodrigo P S; Alves, Valeria M N; Zaneveld, Lourens J D; Waller, Donald P; Schwartz, Jill; Callahan, Marianne M; Mauck, Christine K

2006-01-01

Aim To compare the effectiveness of an acid-buffering formulation gel (ACIDFORM) with metronidazole gel in the treatment of symptomatic bacterial vaginosis (BV). Methods After a confirmed diagnosis of BV according to the criteria established by Nugent and Amsel, 30 nonpregnant women were enrolled in a randomized, double-blind clinical study. The women were randomly assigned to receive either 5 g ACIDFORM gel (n = 13) or 10% metronidazole gel (n = 17) intravaginally once daily for five consecutive days. Participants were evaluated in two follow-up visits (7–12 days and 28–35 days after treatment). Therapeutic success was defined as the presence of less than three of Amsel's criteria. If three or more criteria were present at first or second follow-up visit, the woman was excluded from the study and treated orally with metronidazole. Nugent scores were recorded at each visit but these were not used to define cure. Results At the first follow-up visit, 15 (88%) of the women in the metronidazole group were cured compared with only three (23%) in the ACIDFORM group (P < 0.001). The remaining 12 women (10 of the ACIDFORM group and two of the metronidazole group) were considered as failure and were treated orally with metronidazole. At the second follow-up visit, two of the ACIDFORM-treated women and six of the metronidazole-treated women presented recurrent BV. Four women in the ACIDFORM group and one in the metronidazole group reported occasional burning and itching during product use. Conclusion ACIDFORM gel was significantly less effective than high-dose metronidazole gel for the treatment of symptomatic BV. PMID:16433875

Oral dextrose gel for the treatment of hypoglycaemia in newborn infants.

PubMed

Weston, Philip J; Harris, Deborah L; Battin, Malcolm; Brown, Julie; Hegarty, Joanne E; Harding, Jane E

2016-05-04

Neonatal hypoglycaemia, a common condition, can be associated with brain injury. It is frequently managed by providing infants with an alternative source of glucose, given enterally with formula or intravenously with dextrose solution. This often requires that mother and baby are cared for in separate environments and may inhibit breast feeding. Dextrose gel is simple and inexpensive and can be administered directly to the buccal mucosa for rapid correction of hypoglycaemia, in association with continued breast feeding and maternal care. To assess the effectiveness of dextrose gel in correcting hypoglycaemia and in reducing long-term neurodevelopmental impairment. We searched MEDLINE, EMBASE, the Cochrane Central Register of Controlled Trials (CENTRAL), the Cumulative Index to Nursing and Allied Health Literature (CINAHL) and Web of Science from inception of the database to February 2016. We also searched international clinical trials networks and handsearched proceedings of specific scientific meetings. Randomised and quasi-randomised studies comparing dextrose gel versus placebo, no treatment or other therapies for treatment of neonatal hypoglycaemia. Two review authors independently assessed trial quality and extracted data and did not assess publications for which they themselves were study authors. We included two trials involving 312 infants. No data were available for correction of hypoglycaemia for each hypoglycaemic event. We found no evidence of a difference between dextrose gel and placebo gel for major neurosensory disability at two-year follow-up (risk ratio (RR) 6.27, 95% confidence interval (CI) 0.77 to 51.03; one trial, n = 184; quality of evidence very low). Dextrose gel compared with placebo gel or no gel did not alter the need for intravenous treatment for hypoglycaemia (typical RR 0.78, 95% CI 0.46 to 1.32; two trials, 312 infants; quality of evidence very low). Infants treated with dextrose gel were less likely to be separated from their mothers for treatment of hypoglycaemia (RR 0.54, 95% CI 0.31 to 0.93; one trial, 237 infants; quality of evidence moderate) and were more likely to be exclusively breast fed after discharge (RR 1.10, 95% CI 1.01 to 1.18; one trial, 237 infants; quality of evidence moderate). Estimated rise in blood glucose concentration following dextrose gel was 0.4 mmol/L (95% CI -0.14 to 0.94; one trial, 75 infants). Investigators in one trial reported no adverse outcomes (n = 237 infants). Treatment of infants with neonatal hypoglycaemia with 40% dextrose gel reduces the incidence of mother-infant separation for treatment and increases the likelihood of full breast feeding after discharge compared with placebo gel. No evidence suggests occurrence of adverse effects during the neonatal period or at two years' corrected age. Oral dextrose gel should be considered first-line treatment for infants with neonatal hypoglycaemia.

Effect of manuka honey on the expression of universal stress protein A in meticillin-resistant Staphylococcus aureus.

PubMed

Jenkins, Rowena; Burton, Neil; Cooper, Rose

2011-04-01

Staphylococcus aureus is an important pathogen that can cause many problems, from impetigo to endocarditis. With its continued resistance to multiple antibiotics, S. aureus remains a serious health threat. Honey has been used to eradicate meticillin-resistant S. aureus (MRSA) strains from wounds, but its mode of action is not yet understood. Proteomics provides a potent group of techniques that can be used to analyse differences in protein expression between untreated bacterial cells and those treated with inhibitory concentrations of manuka honey. In this study, two-dimensional (2D) electrophoresis was combined with matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) to determine the identities of proteins whose levels of expression were changed at least two-fold following treatment with manuka honey. Protein extracts were obtained from cells grown in tryptone soy broth (with or without manuka honey) by mechanical disruption and were separated on 2D polyacrylamide gels. A protein was isolated in gels prepared from untreated cell extract that was absent from gels made using honey-treated cell extract. Using MALDI-TOF MS, the protein was identified as universal stress protein A (UspA). Downregulation of this protein was confirmed by real-time polymerase chain reaction (PCR), which showed a 16-fold downregulation in honey-treated cells compared with untreated samples. This protein is involved in the stress stamina response and its downregulation could help to explain the inhibition of MRSA by manuka honey. Copyright © 2011 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

Nuclear Heat Shock Response and Novel Nuclear Domain 10 Reorganization in Respiratory Syncytial Virus-Infected A549 Cells Identified by High-Resolution Two-Dimensional Gel Electrophoresis

PubMed Central

Brasier, Allan R.; Spratt, Heidi; Wu, Zheng; Boldogh, Istvan; Zhang, Yuhong; Garofalo, Roberto P.; Casola, Antonella; Pashmi, Jawad; Haag, Anthony; Luxon, Bruce; Kurosky, Alexander

2004-01-01

The pneumovirus respiratory syncytial virus (RSV) is a leading cause of epidemic respiratory tract infection. Upon entry, RSV replicates in the epithelial cytoplasm, initiating compensatory changes in cellular gene expression. In this study, we have investigated RSV-induced changes in the nuclear proteome of A549 alveolar type II-like epithelial cells by high-resolution two-dimensional gel electrophoresis (2DE). Replicate 2D gels from uninfected and RSV-infected nuclei were compared for changes in protein expression. We identified 24 different proteins by peptide mass fingerprinting after matrix-assisted laser desorption ionization-time of flight mass spectrometry (MS), whose average normalized spot intensity was statistically significant and differed by ±2-fold. Notable among the proteins identified were the cytoskeletal cytokeratins, RNA helicases, oxidant-antioxidant enzymes, the TAR DNA binding protein (a protein that associates with nuclear domain 10 [ND10] structures), and heat shock protein 70- and 60-kDa isoforms (Hsp70 and Hsp60, respectively). The identification of Hsp70 was also validated by liquid chromatography quadropole-TOF tandem MS (LC-MS/MS). Separate experiments using immunofluorescence microscopy revealed that RSV induced cytoplasmic Hsp70 aggregation and nuclear accumulation. Data mining of a genomic database showed that RSV replication induced coordinate changes in Hsp family proteins, including the 70, 70-2, 90, 40, and 40-3 isoforms. Because the TAR DNA binding protein associates with ND10s, we examined the effect of RSV infection on ND10 organization. RSV induced a striking dissolution of ND10 structures with redistribution of the component promyelocytic leukemia (PML) and speckled 100-kDa (Sp100) proteins into the cytoplasm, as well as inducing their synthesis. Our findings suggest that cytoplasmic RSV replication induces a nuclear heat shock response, causes ND10 disruption, and redistributes PML and Sp100 to the cytoplasm. Thus, a high-resolution proteomics approach, combined with immunofluorescence localization and coupled with genomic response data, yielded unexpected novel insights into compensatory nuclear responses to RSV infection. PMID:15479789

Computer-based image analysis of one-dimensional electrophoretic gels used for the separation of DNA restriction fragments.

PubMed Central

Gray, A J; Beecher, D E; Olson, M V

1984-01-01

A stand-alone, interactive computer system has been developed that automates the analysis of ethidium bromide-stained agarose and acrylamide gels on which DNA restriction fragments have been separated by size. High-resolution digital images of the gels are obtained using a camera that contains a one-dimensional, 2048-pixel photodiode array that is mechanically translated through 2048 discrete steps in a direction perpendicular to the gel lanes. An automatic band-detection algorithm is used to establish the positions of the gel bands. A color-video graphics system, on which both the gel image and a variety of operator-controlled overlays are displayed, allows the operator to visualize and interact with critical stages of the analysis. The principal interactive steps involve defining the regions of the image that are to be analyzed and editing the results of the band-detection process. The system produces a machine-readable output file that contains the positions, intensities, and descriptive classifications of all the bands, as well as documentary information about the experiment. This file is normally further processed on a larger computer to obtain fragment-size assignments. Images PMID:6320097

Topical adapalene gel 0.1% vs. isotretinoin gel 0.05% in the treatment of acne vulgaris: a randomized open-label clinical trial.

PubMed

Ioannides, D; Rigopoulos, D; Katsambas, A

2002-09-01

Topical application of isotretinoin and adapalene has proved effective in treating acne vulgaris. Both drugs demonstrate therapeutic advantages and less irritancy over tretinoin, the most widely used treatment for acne. They both act as retinoid agonists, but differ in their affinity profile for nuclear and cytosolic retinoic acid receptors. To compare the efficacy and tolerability of adapalene gel 0.1% and isotretinoin gel 0.05% in the treatment of acne vulgaris of the face, in a randomized open-label clinical trial. Eighty patients were enrolled and were instructed to apply adapalene gel 0.1% or isotretinoin gel 0.05% once daily over a 12-week treatment period. Efficacy determination included noninflammatory and inflammatory lesion counts by the investigator and global evaluation of improvement. Cutaneous tolerance was assessed by determining erythema, scaling and burning with pruritus. Adapalene and isotretinoin gels were highly effective in treating facial acne. Adapalene gel produced greater reductions in noninflammatory and inflammatory lesion counts than did isotretinoin gel, but differences between treatments were not statistically significant. Adapalene gel was significantly better tolerated than isotretinoin gel during the whole treatment period. The two gels studied demonstrated comparable efficacy. When adapalene and isotretinoin were compared, significantly lower skin irritation was noted with adapalene, indicating that adapalene may begin a new era of treatment with low-irritant retinoids.

Statistical Analysis of Variation in the Human Plasma Proteome

DOE PAGES

Corzett, Todd H.; Fodor, Imola K.; Choi, Megan W.; ...

2010-01-01

Quantifying the variation in the human plasma proteome is an essential prerequisite for disease-specific biomarker detection. We report here on the longitudinal and individual variation in human plasma characterized by two-dimensional difference gel electrophoresis (2-D DIGE) using plasma samples from eleven healthy subjects collected three times over a two week period. Fixed-effects modeling was used to remove dye and gel variability. Mixed-effects modeling was then used to quantitate the sources of proteomic variation. The subject-to-subject variation represented the largest variance component, while the time-within-subject variation was comparable to the experimental variation found in a previous technical variability study where onemore » human plasma sample was processed eight times in parallel and each was then analyzed by 2-D DIGE in triplicate. Here, 21 protein spots had larger than 50% CV, suggesting that these proteins may not be appropriate as biomarkers and should be carefully scrutinized in future studies. Seventy-eight protein spots showing differential protein levels between different individuals or individual collections were identified by mass spectrometry and further characterized using hierarchical clustering. The results present a first step toward understanding the complexity of longitudinal and individual variation in the human plasma proteome, and provide a baseline for improved biomarker discovery.« less

Statistical analysis of variation in the human plasma proteome.

PubMed

Corzett, Todd H; Fodor, Imola K; Choi, Megan W; Walsworth, Vicki L; Turteltaub, Kenneth W; McCutchen-Maloney, Sandra L; Chromy, Brett A

2010-01-01

Quantifying the variation in the human plasma proteome is an essential prerequisite for disease-specific biomarker detection. We report here on the longitudinal and individual variation in human plasma characterized by two-dimensional difference gel electrophoresis (2-D DIGE) using plasma samples from eleven healthy subjects collected three times over a two week period. Fixed-effects modeling was used to remove dye and gel variability. Mixed-effects modeling was then used to quantitate the sources of proteomic variation. The subject-to-subject variation represented the largest variance component, while the time-within-subject variation was comparable to the experimental variation found in a previous technical variability study where one human plasma sample was processed eight times in parallel and each was then analyzed by 2-D DIGE in triplicate. Here, 21 protein spots had larger than 50% CV, suggesting that these proteins may not be appropriate as biomarkers and should be carefully scrutinized in future studies. Seventy-eight protein spots showing differential protein levels between different individuals or individual collections were identified by mass spectrometry and further characterized using hierarchical clustering. The results present a first step toward understanding the complexity of longitudinal and individual variation in the human plasma proteome, and provide a baseline for improved biomarker discovery.

Meso-Decorated Switching-Knot Gels

NASA Astrophysics Data System (ADS)

Gong, Jin; Sawamura, Kensuke; Makino, Masato; Kabir, M. H.; Furukawa, Hidemitsu

Gels are a new material having three-dimensional network structures of macromolecules. They possess excellent properties as swellability, high permeability and biocompatibility, and have been applied in various fields of daily life, food, medicine, architecture, and chemistry .In this study, we tried to prepare new multi-functional and high-strength gels by using Meso-Decoration (Meso-Deco), one new method of structure design at intermediate mesoscale. High-performance rigid-rod aromatic polymorphic crystals. The strengthening of gels can be realized by meso-decorating the gels' structure using high-performance polymorphic crystals. New gels with good mechanical properties, novel optical properties and thermal properties are expected to be developed.

Neurotrophins differentially stimulate the growth of cochlear neurites on collagen surfaces and in gels☆

PubMed Central

Xie, Joanna; Pak, Kwang; Evans, Amaretta; Kamgar-Parsi, Andy; Fausti, Stephen; Mullen, Lina; Ryan, Allen Frederic

2013-01-01

The electrodes of a cochlear implant are located far from the surviving neurons of the spiral ganglion, which results in decreased precision of neural activation compared to the normal ear. If the neurons could be induced to extend neurites toward the implant, it might be possible to stimulate more discrete subpopulations of neurons, and to increase the resolution of the device. However, a major barrier to neurite growth toward a cochlear implant is the fluid filling the scala tympani, which separates the neurons from the electrodes. The goal of this study was to evaluate the growth of cochlear neurites in three-dimensional extracellular matrix molecule gels, and to increase biocompatibility by using fibroblasts stably transfected to produce neurotrophin-3 and brain-derived neurotrophic factor. Spiral ganglion explants from neonatal rats were evaluated in cultures. They were exposed to soluble neurotrophins, cells transfected to secrete neurotrophins, and/or collagen gels. We found that cochlear neurites grew readily on collagen surfaces and in three-dimensional collagen gels. Co-culture with cells producing neurotrophin-3 resulted in increased numbers of neurites, and neurites that were longer than when explants were cultured with control fibroblasts stably transfected with green fluorescent protein. Brain-derived neurotrophic factor-producing cells resulted in a more dramatic increase in the number of neurites, but there was no significant effect on neurite length. It is suggested that extracellular matrix molecule gels and cells transfected to produce neurotrophins offer an opportunity to attract spiral ganglion neurites toward a cochlear implant. PMID:24459465

Dose evaluation of an NIPAM polymer gel dosimeter using gamma index

NASA Astrophysics Data System (ADS)

Chang, Yuan-Jen; Lin, Jing-Quan; Hsieh, Bor-Tsung; Yao, Chun-Hsu; Chen, Chin-Hsing

2014-11-01

An N-isopropylacrylamide (NIPAM) polymer gel dosimeter has great potential in clinical applications. However, its three-dimensional dose distribution must be assessed. In this work, a quantitative evaluation of dose distributions was performed to evaluate the NIPAM polymer gel dosimeter using gamma analysis. A cylindrical acrylic phantom filled with NIPAM gel measuring 10 cm (diameter) by 10 cm (height) by 3 mm (thickness) was irradiated by a 4×4 cm2 square light field. The irradiated gel phantom was scanned using an optical computed tomography (optical CT) scanner (OCTOPUS™, MGS Research, Inc., Madison, CT, USA) at 1 mm resolution. The projection data were transferred to an image reconstruction program, which was written using MATLAB (The MathWorks, Natick, MA, USA). The program reconstructed the image of the optical density distribution using the algorithm of a filter back-projection. Three batches of replicated gel phantoms were independently measured. The average uncertainty of the measurements was less than 1%. The gel was found to have a high degree of spatial uniformity throughout the dosimeter and good temporal stability. A comparison of the line profiles of the treatment planning system and of the data measured by optical CT showed that the dose was overestimated in the penumbra region because of two factors. The first is light scattering due to changes in the refractive index at the edge of the irradiated field. The second is the edge enhancement caused by free radical diffusion. However, the effect of edge enhancement on the NIPAM gel dosimeter is not as significant as that on the BANG gel dosimeter. Moreover, the dose uncertainty is affected by the inaccuracy of the gel container positioning process. To reduce the uncertainty of 3D dose distribution, improvements in the gel container holder must be developed.

Phytochemical Evaluation of Moth Bean (Vigna aconitifolia L.) Seeds and Their Divergence

PubMed Central

Gupta, Neha; Shrivastava, Nidhi; Singh, Pramod Kumar; Bhagyawant, Sameer S.

2016-01-01

In the present study, phytochemical contents of 25 moth bean (Vigna aconitifolia) seed accessions were evaluated. This includes protease inhibitors, phytic acid, radical scavenging activity, and tannins. The studies revealed significant variation in the contents of theses phytochemicals. Presence of photochemical composition was correlated with seed storage proteins like albumin and globulin. Qualitative identification of total seed storage protein abundance across two related moth bean accessions using two-dimensional gel electrophoresis (2D-GE) was performed. Over 20 individual protein fractions were distributed over the gel as a series of spots in two moth bean accessions. Seed proteome accumulated spots of high intensity over a broad range of pI values of 3–10 in a molecular weight range of 11–170 kDa. In both seed accessions maximum protein spots are seen in the pI range of 6–8. PMID:27239343

Phytochemical Evaluation of Moth Bean (Vigna aconitifolia L.) Seeds and Their Divergence.

PubMed

Gupta, Neha; Shrivastava, Nidhi; Singh, Pramod Kumar; Bhagyawant, Sameer S

2016-01-01

In the present study, phytochemical contents of 25 moth bean (Vigna aconitifolia) seed accessions were evaluated. This includes protease inhibitors, phytic acid, radical scavenging activity, and tannins. The studies revealed significant variation in the contents of theses phytochemicals. Presence of photochemical composition was correlated with seed storage proteins like albumin and globulin. Qualitative identification of total seed storage protein abundance across two related moth bean accessions using two-dimensional gel electrophoresis (2D-GE) was performed. Over 20 individual protein fractions were distributed over the gel as a series of spots in two moth bean accessions. Seed proteome accumulated spots of high intensity over a broad range of pI values of 3-10 in a molecular weight range of 11-170 kDa. In both seed accessions maximum protein spots are seen in the pI range of 6-8.

Multilineage differentiation of rhesus monkey embryonic stem cells in three-dimensional culture systems

NASA Technical Reports Server (NTRS)

Chen, Silvia S.; Revoltella, Roberto P.; Papini, Sandra; Michelini, Monica; Fitzgerald, Wendy; Zimmerberg, Joshua; Margolis, Leonid

2003-01-01

In the course of normal embryogenesis, embryonic stem (ES) cells differentiate along different lineages in the context of complex three-dimensional (3D) tissue structures. In order to study this phenomenon in vitro under controlled conditions, 3D culture systems are necessary. Here, we studied in vitro differentiation of rhesus monkey ES cells in 3D collagen matrixes (collagen gels and porous collagen sponges). Differentiation of ES cells in these 3D systems was different from that in monolayers. ES cells differentiated in collagen matrixes into neural, epithelial, and endothelial lineages. The abilities of ES cells to form various structures in two chemically similar but topologically different matrixes were different. In particular, in collagen gels ES cells formed gland-like circular structures, whereas in collagen sponges ES cells were scattered through the matrix or formed aggregates. Soluble factors produced by feeder cells or added to the culture medium facilitated ES cell differentiation into particular lineages. Coculture with fibroblasts in collagen gel facilitated ES cell differentiation into cells of a neural lineage expressing nestin, neural cell adhesion molecule, and class III beta-tubulin. In collagen sponges, keratinocytes facilitated ES cell differentiation into cells of an endothelial lineage expressing factor VIII. Exogenous granulocyte-macrophage colony-stimulating factor further enhanced endothelial differentiation. Thus, both soluble factors and the type of extracellular matrix seem to be critical in directing differentiation of ES cells and the formation of tissue-like structures. Three-dimensional culture systems are a valuable tool for studying the mechanisms of these phenomena.

The PA influenza virus polymerase subunit is a phosphorylated protein.

PubMed

Sanz-Ezquerro, J J; Fernández Santarén, J; Sierra, T; Aragón, T; Ortega, J; Ortín, J; Smith, G L; Nieto, A

1998-03-01

The induction of proteolysis by expression of the influenza virus PA polymerase subunit is the only biochemical activity ascribed to this protein. In the course of studying viral protein synthesis by two-dimensional gel electrophoresis, we observed the existence of several PA isoforms with different isoelectric points. These isoforms were also present when the PA gene was singly expressed in three different expression systems, indicating that a cellular activity is responsible for its post-translational modification. In vivo labelling with [32P]orthophosphate, followed by two-dimensional gel electrophoresis, clearly demonstrated the incorporation of phosphate into the PA molecule. Phosphoserine and phosphothreonine epitopes were present in PA, while phosphotyrosine residues were absent, as tested by immunoblotting with specific antibodies. These facts, as well as the presence of multiple consensus sites for casein kinase II (CKII) phosphorylation, prompted us to test the involvement of this kinase in PA covalent modification. PA protein purified by immunoprecipitation could be specifically labelled by the catalytic alpha subunit of human CKII, which was expressed and purified from bacteria. Collectively, these data demonstrate that the PA subunit of the influenza virus RNA polymerase is a phosphoprotein.

Retrospective study of radiotherapy-induced skin reactions in breast cancer patients: reduced incidence of moist desquamation with a hydroactive colloid gel versus dexpanthenol.

PubMed

Censabella, Sandrine; Claes, Stefan; Orlandini, Marc; Braekers, Roel; Thijs, Herbert; Bulens, Paul

2014-10-01

Dermatitis is a very frequent and distressing side effect of radiation therapy that may necessitate a treatment interruption when evolving towards more severe forms such as moist desquamation (MD). The aim of this study was to compare the efficacy of two topical agents, a dexpanthenol cream vs a hydroactive colloid gel combining absorbing and moisturising properties, in preventing MD in breast cancer patients. This retrospective study compared two successive groups of breast cancer patients undergoing radiotherapy after breast-sparing surgery between 2008 and 2012. A group of 267 patients applied a 5% dexpanthenol cream on the irradiated zone throughout the course of their radiotherapy. Another group of 216 patients applied first the dexpanthenol cream then replaced it by the hydroactive colloid gel after 11-14 days of radiotherapy. Radiation treatment (total dose, technique, and equipment) was the same for the two groups. The clinical outcomes were the occurrence and time to onset of moist desquamation. The overall incidence of MD was significantly lower in patients who applied the hydroactive colloid gel (16%) than in those who applied the dexpanthenol cream (32%, odds-ratio = 0.35). Also, MD occurred significantly later with the hydroactive colloid gel than with the dexpanthenol cream (hazard ratio = 0.39). Compared with the dexpanthenol cream, the hydroactive colloid gel significantly reduced the risk of developing MD in patients undergoing radiotherapy for breast cancer. These promising results warrant further research on the efficacy of hydroactive colloid gels in managing radiation dermatitis. Copyright © 2014 Elsevier Ltd. All rights reserved.

Experimental study of sector and linear array ultrasound accuracy and the influence of navigated 3D-reconstruction as compared to MRI in a brain tumor model.

PubMed

Siekmann, Max; Lothes, Thomas; König, Ralph; Wirtz, Christian Rainer; Coburger, Jan

2018-03-01

Currently, intraoperative ultrasound in brain tumor surgery is a rapidly propagating option in imaging technology. We examined the accuracy and resolution limits of different ultrasound probes and the influence of 3D-reconstruction in a phantom and compared these results to MRI in an intraoperative setting (iMRI). An agarose gel phantom with predefined gel targets was examined with iMRI, a sector (SUS) and a linear (LUS) array probe with two-dimensional images. Additionally, 3D-reconstructed sweeps in perpendicular directions were made of every target with both probes, resulting in 392 measurements. Statistical calculations were performed, and comparative boxplots were generated. Every measurement of iMRI and LUS was more precise than SUS, while there was no apparent difference in height of iMRI and 3D-reconstructed LUS. Measurements with 3D-reconstructed LUS were always more accurate than in 2D-LUS, while 3D-reconstruction of SUS showed nearly no differences to 2D-SUS in some measurements. We found correlations of 3D-reconstructed SUS and LUS length and width measurements with 2D results in the same image orientation. LUS provides an accuracy and resolution comparable to iMRI, while SUS is less exact than LUS and iMRI. 3D-reconstruction showed the potential to distinctly improve accuracy and resolution of ultrasound images, although there is a strong correlation with the sweep direction during data acquisition.

Detection and three-dimensional visualization of lesion models using sonoelastography

NASA Astrophysics Data System (ADS)

Taylor, Lawrence S.; Gaborski, Thomas R.; Strang, John G.; Rubens, Deborah; Parker, Kevin J.

2002-04-01

Sonoelastography is a vibration Doppler technique for imaging the relative elasticity of tissues. Detectability of hard lesions of various sizes has previously been demonstrated in tissue phantoms by our group. Because real tissue differs from phantom material, the injection of formaldehyde in fresh liver tissue is being used as an in-vitro lesion model. Pieces of fresh calf liver were embedded in an agar gel then injected with a bolus of 37% formaldehyde to create a stiff lesion. Two and three-dimensional sonoelastography and b-mode images were acquired. The lesions were visible in each sonoelastography image as a region of reduced vibration. After imaging, lesions were dissected and measured for size and volume. One 0.4 cc bolus injection of formaldehyde created a lesion with a volume of 10.3 cc in the sonoelastography image compared to 9.3 cc using fluid displacement of the dissected lesion. A 0.33 cc injection of formaldehyde lesion created a volume of 5 cc in the sonoelastography image compared to 4.4 cc using fluid displacement. Sonoelastography imaging techniques for imaging hard lesions in phantoms can be successfully extended to imaging formaldehyde induced lesions in real tissue.

Biogelx: Cell Culture on Self-Assembling Peptide Gels.

PubMed

Harper, Mhairi M; Connolly, Michael L; Goldie, Laura; Irvine, Eleanore J; Shaw, Joshua E; Jayawarna, Vineetha; Richardson, Stephen M; Dalby, Matthew J; Lightbody, David; Ulijn, Rein V

2018-01-01

Aromatic peptide amphiphiles can form self-supporting nanostructured hydrogels with tunable mechanical properties and chemical compositions. These hydrogels are increasingly applied in two-dimensional (2D) and three-dimensional (3D) cell culture, where there is a rapidly growing need to store, grow, proliferate, and manipulate naturally derived cells within a hydrated, 3D matrix. Biogelx Limited is a biomaterials company, created to commercialize these bio-inspired hydrogels to cell biologists for a range of cell culture applications. This chapter describes methods of various characterization and cell culture techniques specifically optimized for compatibility with Biogelx products.

Granular gel support-enabled extrusion of three-dimensional alginate and cellular structures.

PubMed

Jin, Yifei; Compaan, Ashley; Bhattacharjee, Tapomoy; Huang, Yong

2016-06-03

Freeform fabrication of soft structures has been of great interest in recent years. In particular, it is viewed as a critical step toward the grand vision of organ printing--the on-demand design and fabrication of three-dimensional (3D) human organ constructs for implantation and regenerative medicine. The objective of this study is to develop a novel granular gel support material-enabled, two-step gelation-based 'printing-then-gelation' approach to fabricate 3D alginate structures using filament extrusion. Specifically, a granular Carbopol microgel bath holds the ungelled alginate structure being extruded, avoiding the instantaneous gelation of each printed layer as well as resultant surface tension-induced nozzle clogging. Since Carbopol microgels react with multivalent cations, which are needed for alginate crosslinking, gelatin is introduced as a sacrificial material to make an alginate and gelatin bioink for extrusion, which gels thermally (step-one gelation) to initially stabilize the printed structure for removal from Carbopol. Then gelatin is melted and diffused away while alginate is ionically crosslinked in a 37 °C calcium chloride bath (step-two gelation), resulting in an alginate structure. The proposed 'printing-then-gelation' approach works for alginate structure fabrication, and it is also applicable for the printing of cellular constructs and other similar homogeneous soft structures using a two-step or even multi-step approach. The main conclusions are: (1) 0.8% (w/v) Carbopol bath with a neutral pH value may be most suitable for soft structure printing; (2) it is most effective to use a 0.9% (w/v) NaCl solution to facilitate the removal of residual Carbopol; and (3) alginate structures fabricated using the proposed approach demonstrate better mechanical properties than those fabricated using the conventional 'gelation-while-printing' approach.

Synthesis and Structural Studies of Calcium and Magnesium Phosphinate and Phosphonate Compounds

NASA Astrophysics Data System (ADS)

Bampoh, Victoria Naa Kwale

The work presented herein describes synthetic methodologies leading to the design of a wide array of magnesium and calcium based phosphinate and phosphonates with possible applications as bone scaffolding materials or additives to bone cements. The challenge to the chemistry of the alkaline earth phosphonate target compounds includes poor solubility of compounds, and poorly understood details on the control of the metal's coordination environment. Hence, less is known on phosphonate based alkaline earth metal organic frameworks as compared to transition metal phosphonates. Factors governing the challenges in obtaining crystalline, well-defined magnesium and calcium solids lie in the large metal diameters, the absence of energetically available d-orbitals to direct metal geometry, as well as the overall weakness of the metal-ligand bonds. A significant part of this project was concerned with the development of suitable reaction conditions to obtain X-ray quality crystals of the reaction products to allow for structural elucidation of the novel compounds. Various methodologies to aid in crystal growth including hydrothermal methods and gel crystallization were employed. We have used phosphinate and phosphonate ligands with different number of phosphorus oxygen atoms as well as diphosphonates with different linker lengths to determine their effects on the overall structural features. An interesting correlation is observed between the dimensionality of products and the increasing number of donor oxygen atoms in the ligands as we progress from phosphinic acid to the phosphorous acids. As an example, monophosphinate ligand only yielded one-dimensional compounds, whereas the phosphonates crystallize as one and two-dimensional compounds, and the di- and triphosphonate based compounds display two or three-dimensional geometries. This thesis provides a selection of calcium and magnesium compounds with one-dimensional geometry, as represented in a calcium phosphinate to novel two-dimensional sheets of magnesium and pillared calcium phosphonates. The preparation of these novel compounds has led to the establishment of synthetic protocols that allow for the direct preparation of compounds with defined structural features.

Comparative Proteome Analysis of Brucella melitensis Vaccine Strain Rev 1 and a Virulent Strain, 16M

PubMed Central

Eschenbrenner, Michel; Wagner, Mary Ann; Horn, Troy A.; Kraycer, Jo Ann; Mujer, Cesar V.; Hagius, Sue; Elzer, Philip; DelVecchio, Vito G.

2002-01-01

The genus Brucella consists of bacterial pathogens that cause brucellosis, a major zoonotic disease characterized by undulant fever and neurological disorders in humans. Among the different Brucella species, Brucella melitensis is considered the most virulent. Despite successful use in animals, the vaccine strains remain infectious for humans. To understand the mechanism of virulence in B. melitensis, the proteome of vaccine strain Rev 1 was analyzed by two-dimensional gel electrophoresis and compared to that of virulent strain 16M. The two strains were grown under identical laboratory conditions. Computer-assisted analysis of the two B. melitensis proteomes revealed proteins expressed in either 16M or Rev 1, as well as up- or down-regulation of proteins specific for each of these strains. These proteins were identified by peptide mass fingerprinting. It was found that certain metabolic pathways may be deregulated in Rev 1. Expression of an immunogenic 31-kDa outer membrane protein, proteins utilized for iron acquisition, and those that play a role in sugar binding, lipid degradation, and amino acid binding was altered in Rev 1. PMID:12193611

Comparative proteome analysis of Brucella melitensis vaccine strain Rev 1 and a virulent strain, 16M.

PubMed

Eschenbrenner, Michel; Wagner, Mary Ann; Horn, Troy A; Kraycer, Jo Ann; Mujer, Cesar V; Hagius, Sue; Elzer, Philip; DelVecchio, Vito G

2002-09-01

The genus Brucella consists of bacterial pathogens that cause brucellosis, a major zoonotic disease characterized by undulant fever and neurological disorders in humans. Among the different Brucella species, Brucella melitensis is considered the most virulent. Despite successful use in animals, the vaccine strains remain infectious for humans. To understand the mechanism of virulence in B. melitensis, the proteome of vaccine strain Rev 1 was analyzed by two-dimensional gel electrophoresis and compared to that of virulent strain 16M. The two strains were grown under identical laboratory conditions. Computer-assisted analysis of the two B. melitensis proteomes revealed proteins expressed in either 16M or Rev 1, as well as up- or down-regulation of proteins specific for each of these strains. These proteins were identified by peptide mass fingerprinting. It was found that certain metabolic pathways may be deregulated in Rev 1. Expression of an immunogenic 31-kDa outer membrane protein, proteins utilized for iron acquisition, and those that play a role in sugar binding, lipid degradation, and amino acid binding was altered in Rev 1.

Structure and morphology evolution of silica-modified pseudoboehmite aerogels during heat treatment

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pakharukova, V.P., E-mail: verapakh@catalysis.ru; Novosibirsk State University, Pirogova Street 2, 630090 Novosibirsk; Research and Educational Center for Energy Efficient Catalysis, Novosibirsk State University, Novosibirsk 630090

Silica-modified pseudoboehmite aerogels (0, 10, 20 at% of Si) were prepared by sol–gel method followed by supercritical drying. The phase transformations, changes in structure and morphology upon calcination were thoroughly investigated by advanced X-Ray diffraction (XRD) techniques and high-resolution transmission electron microscopy (HRTEM). Obtained pseudoboehmite samples had specific nanostructure: ultrathin two-dimensional (2D) crystallites were loosely packed. The silica dopant drastically enhanced the crystallite anisotropy. Thus, the aerogel with Al:Si atomic ratio of 9:1 consisted of the pseudoboehmite nanosheets with thickness of one unit cell (average dimensions of 14.0×1.2×14.5 nm). The specific nanostructure caused remarkable features of experimental XRD patterns, includingmore » anisotropic peak broadening and appearance of forbidden reflection. Direct simulation of XRD patterns with using the Debye Scattering Equation allowed the size and morphology of pseudoboehmite crystallites to be determined. The silica addition strongly delayed formation of γ-alumina and further phase transformations upon calcinaton. Thermal stability of alumina was suggested to be affected by the particle morphology inherited from the pseudoboehmite precursor. - Graphical abstract: Pseudoboehmite samples had specific nanostructure: ultrathin two-dimensional (2D) crystallites were loosely packed. - Highlights: • Silica-doped boehmites were prepared by sol–gel method with supercritical drying. • Ultrathin two-dimensional crystallites of pseudoboehmite were obtained. • Changes in structure and morphology upon calcination were studied. • Simulation of XRD patterns was performed with use of the Debye Scattering Equation. • Thermal stability of alumina depended on morphology inherited from pseudoboehmite.« less

Imaging of Selenium by Laser Ablation Inductively Coupled Plasma Mass Spectrometry (LA-ICP-MS) in 2-D Electrophoresis Gels and Biological Tissues.

PubMed

Cruz, Elisa Castañeda Santa; Susanne Becker, J; Sabine Becker, J; Sussulini, Alessandra

2018-01-01

Selenium and selenoproteins are important components of living organisms that play a role in different biological processes. Laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) is a powerful analytical technique that has been employed to obtain distribution maps of selenium in biological tissues in a direct manner, as well as in selenoproteins, previously separated by their molecular masses and isoelectric points using two-dimensional polyacrylamide gel electrophoresis (2-D PAGE). In this chapter, we present the protocols to perform LA-ICP-MS imaging experiments, allowing the distribution visualization and determination of selenium and/or selenoproteins in biological systems.

A spot-matching method using cumulative frequency matrix in 2D gel images

PubMed Central

Han, Chan-Myeong; Park, Joon-Ho; Chang, Chu-Seok; Ryoo, Myung-Chun

2014-01-01

A new method for spot matching in two-dimensional gel electrophoresis images using a cumulative frequency matrix is proposed. The method improves on the weak points of the previous method called ‘spot matching by topological patterns of neighbour spots’. It accumulates the frequencies of neighbour spot pairs produced through the entire matching process and determines spot pairs one by one in order of higher frequency. Spot matching by frequencies of neighbour spot pairs shows a fairly better performance. However, it can give researchers a hint for whether the matching results can be trustworthy or not, which can save researchers a lot of effort for verification of the results. PMID:26019609

Cytokeratin expression of engrafted three-dimensional culture tissues using epithelial cells derived from porcine periodontal ligaments.

PubMed

Yamada, Rie; Kitajima, Kayoko; Arai, Kyoko; Igarashi, Masaru

2014-09-01

This study investigated the differentiation and proliferation of epithelial cells derived from periodontal ligaments after three-dimensional culture using collagen gel with fibroblasts in vitro and in vivo. Epithelial cells and fibroblasts were derived from porcine periodontal ligaments. Epithelial cells were labeled using a fluorescent red membrane marker (PKH-26GL) and were seeded onto collagen gel with fibroblasts, followed by incubation in an air-liquid interface for 7 days. Three-dimensional cultures were grafted onto the backs of nude mice and removed at 1, 7, and 14 days after surgery (in vivo model). Unfixed sections (5 μm) were used to detect the presence of red fluorescent cells. Paraffin sections were analyzed histologically and immunohistochemically. Specimens were compared with three-dimensional culture tissues at 8, 14 and 21 days (in vitro model). Grafted three-dimensional cultures formed a stratified epithelial structure similar to skin in vivo. Epithelial cells were sequenced in basal-layer-like structures at 14 days in vivo. Immunohistochemical findings showed that the expression of cytokeratin was detected in the epithelial layer in in vitro and in vivo models. Ck8 + 18 + 19 was expressed in the upper epithelial layer in the in vitro model at 14 and 21 days, but not in vivo. Involucrin was expressed in the certified layers in vitro at 14 days, but not in vivo. Laminin was detected at the dermo-epidermal junction in vivo at 7 and 14 days, but not in vitro. These results suggest that differentiation of three-dimensional culture tissues differs in vivo and in vitro. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Gel performance in rheology and profile control under low-frequency vibration: coupling application of physical and chemical EOR techniques.

PubMed

Zheng, Li Ming; Pu, Chun Sheng; Liu, Jing; Ma, Bo; Khan, Nasir

2017-01-01

Flowing gel plugging and low-frequency vibration oil extraction technology have been widely applied in low-permeability formation. High probability of overlapping in action spheres of two technologies might lead to poor operating efficiency during gel injection. Study on flowing gel rheological properties under low-frequency vibration was essential, which was carried out indoor with viscosity measurement. Potential dynamic mechanisms were analyzed for the rheological variation. Under low-frequency vibration, gel rheological properties were found to be obviously influenced, with vibration delaying gel cross-linking in induction period, causing a two-stage gel viscosity change in acceleration period, and decreasing gel strength in stable period. Surface of gel system under vibration presented different fluctuating phenomenon from initial harmonic vibrating to heterogeneous fluctuating (droplet separation might appear) to final harmonic vibrating again. Dynamic displacement in unconsolidated sand pack revealed that low-frequency vibration during gel injection might be a measure to achieve deep profile control, with the gel injection depth increased by 65.8 % compared with the vibration-free sample. At last, suggestions for field test were given in the paper to achieve lower injection friction and better gel plugging efficiency.

Small-Field Measurements of 3D Polymer Gel Dosimeters through Optical Computed Tomography.

PubMed

Shih, Tian-Yu; Wu, Jay; Shih, Cheng-Ting; Lee, Yao-Ting; Wu, Shin-Hua; Yao, Chun-Hsu; Hsieh, Bor-Tsung

2016-01-01

With advances in therapeutic instruments and techniques, three-dimensional dose delivery has been widely used in radiotherapy. The verification of dose distribution in a small field becomes critical because of the obvious dose gradient within the field. The study investigates the dose distributions of various field sizes by using NIPAM polymer gel dosimeter. The dosimeter consists of 5% gelatin, 5% monomers, 3% cross linkers, and 5 mM THPC. After irradiation, a 24 to 96 hour delay was applied, and the gel dosimeters were read by a cone beam optical computed tomography (optical CT) scanner. The dose distributions measured by the NIPAM gel dosimeter were compared to the outputs of the treatment planning system using gamma evaluation. For the criteria of 3%/3 mm, the pass rates for 5 × 5, 3 × 3, 2 × 2, 1 × 1, and 0.5 × 0.5 cm2 were as high as 91.7%, 90.7%, 88.2%, 74.8%, and 37.3%, respectively. For the criteria of 5%/5 mm, the gamma pass rates of the 5 × 5, 3 × 3, and 2 × 2 cm2 fields were over 99%. The NIPAM gel dosimeter provides high chemical stability. With cone-beam optical CT readouts, the NIPAM polymer gel dosimeter has potential for clinical dose verification of small-field irradiation.

Enhanced biocontrol activity of Rhodotorula mucilaginosa cultured in media containing chitosan against postharvest diseases in strawberries: possible mechanisms underlying the effect.

PubMed

Zhang, Hongyin; Ge, Lingling; Chen, Keping; Zhao, Lina; Zhang, Xiaoyun

2014-05-07

The effect of Rhodotorula mucilaginosa cultured in media containing chitosan on its antogonistic activity against postharvest diseases of strawberries and the possible mechanisms involved are discussed. Two-dimensional gel electrophoresis were applied in the analysis of the proteins of R. mucilaginosa in response to chitosan. Compared with the application of R. mucilaginosa alone, the biocontrol efficacy of the yeast combined with 0.5% chitosan was enhanced greatly, with significant increase in chitinase activity of antagonistic yeast, polyphenoloxidase, peroxidase, phenylalanine ammonia lyase, chitinase and β-1,3-glucanase activity, and with an inhibition of lipid peroxidation of strawberries. The population of R. mucilaginosa harvested from NYDB amended with chitosan at 0.5% increased rapidly in strawberry wounds compared with those harvested from NYDB without chitosan. In the cellular proteome, several differentially expressed proteins were identified, most of which were related to basic metabolism.

Comparative Proteome Analysis of the Tuberous Roots of Six Cassava (Manihot esculenta) Varieties Reveals Proteins Related to Phenotypic Traits.

PubMed

Schmitz, Gabriela Justamante Händel; de Magalhães Andrade, Jonathan; Valle, Teresa Losada; Labate, Carlos Alberto; do Nascimento, João Roberto Oliveira

2016-04-27

Cassava (Manihot esculenta Crantz) is a staple food and an important source of starch, and the attributes of its tuberous root largely depend on the variety. The proteome of cassava has been investigated; however, to date, no study has focused on varieties that reveal the molecular basis of phenotypical characteristics. Therefore, we aimed to compare the proteome of the tuberous roots of six cassava varieties that differed in carbohydrates, carotenoids, and resistance to diseases, among other attributes. Two-dimensional gels showed 146 differential spots between the varieties, and the functional roles of some differential proteins were correlated to phenotypic characteristics of the varieties, such as the amount of carbohydrates or carotenoids and the resistance to biotic or abiotic stresses. The results obtained here highlight elements that might help to direct the improvement of new cultivars of cassava, which is an economically and socially relevant crop worldwide.

An anthropomorphic abdominal phantom for deformable image registration accuracy validation in adaptive radiation therapy.

PubMed

Liao, Yuliang; Wang, Linjing; Xu, Xiangdong; Chen, Haibin; Chen, Jiawei; Zhang, Guoqian; Lei, Huaiyu; Wang, Ruihao; Zhang, Shuxu; Gu, Xuejun; Zhen, Xin; Zhou, Linghong

2017-06-01

To design and construct a three-dimensional (3D) anthropomorphic abdominal phantom for geometric accuracy and dose summation accuracy evaluations of deformable image registration (DIR) algorithms for adaptive radiation therapy (ART). Organ molds, including liver, kidney, spleen, stomach, vertebra, and two metastasis tumors, were 3D printed using contours from an ovarian cancer patient. The organ molds were molded with deformable gels made of different mixtures of polyvinyl chloride (PVC) and the softener dioctyl terephthalate. Gels with different densities were obtained by a polynomial fitting curve that described the relation between the Hounsfield unit (HU) and PVC-softener blending ratio. The rigid vertebras were constructed by molding of white cement and cellulose pulp. The final abdominal phantom was assembled by arranging all the fabricated organs inside a hollow dummy according to their anatomies, and sealed by deformable gel with averaged HU of muscle and fat. Fiducial landmarks were embedded inside the phantom for spatial accuracy and dose accumulation accuracy studies. Two channels were excavated to facilitate ionization chamber insertion for dosimetric measurements. Phantom properties such as deformable gel elasticity and HU stability were studied. The dosimetric measurement accuracy in the phantom was performed, and the DIR accuracies of three DIR algorithms available in the open source DIR toolkit-DIRART were also validated. The constructed deformable gel showed elastic behavior and was stable in HU values over times, proving to be a practical material for the deformable phantom. The constructed abdominal phantom consisted of realistic anatomies in terms of both anatomical shapes and densities when compared with its reference patient. The dosimetric measurements showed a good agreement with the calculated doses from the treatment planning system. Fiducial-based accuracy analysis conducted on the constructed phantom demonstrated the feasibility of applying the phantom for organ-wise DIR accuracy assessment. We have designed and constructed an anthropomorphic abdominal deformable phantom with satisfactory elastic property, realistic organ density, and anatomy. This physical phantom can be used for routine validations of DIR geometric accuracy and dose accumulation accuracy in ART. © 2017 American Association of Physicists in Medicine.

Protein Separation by Capillary Gel Electrophoresis: A Review

PubMed Central

Zhu, Zaifang; Lu, Joann J.; Liu, Shaorong

2011-01-01

Capillary gel electrophoresis (CGE) has been used for protein separation for more than two decades. Due to the technology advancement, current CGE methods are becoming more and more robust and reliable for protein analysis, and some of the methods have been routinely used for the analysis of protein-based pharmaceuticals and quality controls. In light of this progress, we survey 147 papers related to CGE separations of proteins and present an overview of this technology. We first introduce briefly the early development of CGE. We then review the methodology, in which we specifically describe the matrices, coatings, and detection strategies used in CGE. CGE using microfabricated channels and incorporation of CGE with two-dimensional protein separations are also discussed in this section. We finally present a few representative applications of CGE for separating proteins in real-world samples. PMID:22122927

Intraoperative use of a reflective blanket (Sirius rescue sheet) for temperature management in dogs less than 10 kg.

PubMed

Tünsmeyer, J; Bojarski, I; Nolte, I; Kramer, S

2009-07-01

To compare the effects of the Sirius rescue sheet with gel pads versus gel pads alone on intraoperative body temperature in dogs less than 10 kg. Forty small breed dogs undergoing elective surgical procedures were randomly assigned to two groups. One group was intraoperatively laid on warmed gel pads, and the other group was additionally wrapped in a Sirius rescue sheet. Oesophageal body temperature was determined every 10 minutes and compared between groups. Temperature of gel pads was measured preoperatively and postoperatively to compare heat loss of the gel pads between groups. The body temperature of dogs wrapped with the Sirius rescue sheet increased intraoperatively. In dogs just lying on warmed gel pads, a decrease in mean body temperature was revealed and mean body temperatures differed between groups after 40 minutes. Extent of heat loss from the gel pads did not differ between the groups. The Sirius rescue sheet, used in addition to warmed gel pads, led to higher intraoperative body temperatures in small breed dogs undergoing surgical procedures to the extremities and the head. The cost-effectiveness and ease of handling make this a useful addition to clinical practice.

Bioprinting 3D cell-laden hydrogel microarray for screening human periodontal ligament stem cell response to extracellular matrix.

PubMed

Ma, Yufei; Ji, Yuan; Huang, Guoyou; Ling, Kai; Zhang, Xiaohui; Xu, Feng

2015-12-22

Periodontitis is an inflammatory disease negatively affecting up to 15% of adults worldwide. Periodontal ligament stem cells (PDLSCs) hold great promises for periodontal tissue regeneration, where it is necessary to find proper extracellular matrix (ECM) materials (e.g., composition, concentration). In this study, we proposed a bioprinting-based approach to generate nano-liter sized three-dimensional (3D) cell-laden hydrogel array with gradient of ECM components, through controlling the volume ratio of two hydrogels, such as gelatin methacrylate (GelMA) and poly(ethylene glycol) (PEG) dimethacrylate. The resulting cell-laden array with a gradient of GelMA/PEG composition was used to screen human PDLSC response to ECM. The behavior (e.g., cell viability, spreading) of human PDLSCs in GelMA/PEG array were found to be depended on the volume ratios of GelMA/PEG, with cell viability and spreading area decreased along with increasing the ratio of PEG. The developed approach would be useful for screening cell-biomaterial interaction in 3D and promoting regeneration of functional tissue.

Measuring Dilution of Microbicide Gels with Optical Imaging

PubMed Central

Drake, Tyler K.; Shah, Tejen; Peters, Jennifer J.; Wax, Adam; Katz, David F.

2013-01-01

We present a novel approach for measuring topical microbicide gel dilution using optical imaging. The approach compares gel thickness measurements from fluorimetry and multiplexed low coherence interferometry in order to calculate dilution of a gel. As a microbicide gel becomes diluted at fixed thickness, its mLCI thickness measurement remains constant, while the fluorimetry signal decreases in intensity. The difference between the two measurements is related to the extent of gel dilution. These two optical modalities are implemented in a single endoscopic instrument that enables simultaneous data collection. A preliminary validation study was performed with in vitro placebo gel measurements taken in a controlled test socket. It was found that change in slope of the regression line between fluorimetry and mLCI based measurements indicates dilution. A dilution calibration curve was then generated by repeating the test socket measurements with serial dilutions of placebo gel with vaginal fluid simulant. This methodology can provide valuable dilution information on candidate microbicide products, which could substantially enhance our understanding of their in vivo functioning. PMID:24340006

One plunge or two?--hand disinfection with alcohol gel.

PubMed

Macdonald, Duncan J M; Mckillop, Elisabeth C A; Trotter, Sylvia; Gray, Alastair J R

2006-04-01

To compare health care workers' hand surface coverage using two different volumes of alcohol gel for hand disinfection. and methods. A total of 84 members of staff in our hospital were studied. Subjects were asked to disinfect their hands with alcohol gel containing a clear fluorescent substance. Performance was assessed by using UV light to identify areas which had been missed, and the total surface area missed was calculated. A total of 42 subjects received 3.5 ml of alcohol gel, and 42 age-, sex-, and job-matched subjects received 1.75 ml of alcohol gel. Significantly less area was missed when hand disinfecting with double the volume of alcohol gel; 1.23 versus 6.35% surface area was missed (P < 0.001). Doubling the volume of alcohol gel used for hand disinfection significantly improves the efficiency of coverage of the hands with alcohol gel. This may result in lower bacterial count on the hands and may reduce the spread of nosocomial infections including that of methicillin-resistant Staphylococcus aureus.

Two-dimensional blue native/SDS-PAGE analysis of whole cell lysate protein complexes of rice in response to salt stress.

PubMed

Hashemi, Amenehsadat; Gharechahi, Javad; Nematzadeh, Ghorbanali; Shekari, Faezeh; Hosseini, Seyed Abdollah; Salekdeh, Ghasem Hosseini

2016-08-01

To understand the biology of a plant in response to stress, insight into protein-protein interactions, which almost define cell behavior, is thought to be crucial. Here, we provide a comparative complexomics analysis of leaf whole cell lysate of two rice genotypes with contrasting responses to salt using two-dimensional blue native/SDS-PAGE (2D-BN/SDS-PAGE). We aimed to identify changes in subunit composition and stoichiometry of protein complexes elicited by salt. Using mild detergent for protein complex solubilization, we were able to identify 9 protein assemblies as hetero-oligomeric and 30 as homo-oligomeric complexes. A total of 20 proteins were identified as monomers in the 2D-BN/SDS-PAGE gels. In addition to identifying known protein complexes that confirm the technical validity of our analysis, we were also able to discover novel protein-protein interactions. Interestingly, an interaction was detected for glycolytic enzymes enolase (ENO1) and triosephosphate isomerase (TPI) and also for a chlorophyll a-b binding protein and RuBisCo small subunit. To show changes in subunit composition and stoichiometry of protein assemblies during salt stress, the differential abundance of interacting proteins was compared between salt-treated and control plants. A detailed exploration of some of the protein complexes provided novel insight into the function, composition, stoichiometry and dynamics of known and previously uncharacterized protein complexes in response to salt stress. Copyright © 2016 Elsevier GmbH. All rights reserved.

Synthesis and evaluation of a maltose-bonded silica gel stationary phase for hydrophilic interaction chromatography and its application in Ginkgo Biloba extract separation in two-dimensional systems.

PubMed

Sheng, Qianying; Yang, Kaiya; Ke, Yanxiong; Liang, Xinmiao; Lan, Minbo

2016-09-01

Maltose covalently bonded to silica was prepared by using carbonyl diimidazole as a cross-linker and employed as a stationary phase for hydrophilic interaction liquid chromatography. The column efficiency and the effect of water content, buffer concentration, and pH value influenced on retention were investigated. The separation or enrichment selectivity was also studied with nucleosides, saccharides, amino acids, peptides, and glycopeptides. The results indicated that the stationary phase processed good separation efficiency and separation selectivity in hydrophilic interaction liquid chromatography mode. Moreover, a two-dimensional hydrophilic interaction liquid chromatography× reversed-phase liquid chromatography method with high orthogonality was developed to analyze the Ginkgo Biloba extract fractions. The development of this two-dimensional chromatographic system would be an effective tool for the separation of complex samples of different polarities and contents. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Nanoparticulate delivery of agents for induced elastogenesis in three-dimensional collagenous matrices.

PubMed

Venkataraman, Lavanya; Sivaraman, Balakrishnan; Vaidya, Pratik; Ramamurthi, Anand

2016-12-01

The degradation of elastic matrix in the infrarenal aortic wall is a critical parameter underlying the formation and progression of abdominal aortic aneurysms. It is mediated by the chronic overexpression of matrix metalloprotease (MMP)-2 and MMP-9, leading to a progressive loss of elasticity and weakening of the aortic wall. Delivery of therapeutic agents to inhibit MMPs, while concurrently coaxing cell-based regenerative repair of the elastic matrix represents a potential strategy for slowing or arresting abdominal aortic aneurysm growth. Previous studies have demonstrated elastogenic induction of healthy and aneurysmal aortic smooth muscle cells and inhibition of MMPs, following exogenous delivery of elastogenic factors such as transforming growth factor (TGF)-β1, as well as MMP-inhibitors such as doxycycline (DOX) in two-dimensional culture. Based on these findings, and others that demonstrated elastogenic benefits of nanoparticulate delivery of these agents in two-dimensional culture, poly(lactide-co-glycolide) nanoparticles were developed for localized, controlled and sustained delivery of DOX and TGF-β1 to human aortic smooth muscle cells within a three-dimensional gels of type I collagen, which closely simulate the arterial tissue microenvironment. DOX and TGF-β1 released from these nanoparticles influenced elastogenic outcomes positively within the collagen constructs over 21 days of culture, which were comparable to that induced by exogenous supplementation of DOX and TGF-β1 within the culture medium. However, this was accomplished at doses ~20-fold lower than the exogenous dosages of the agents, illustrating that their localized, controlled and sustained delivery from nanoparticles embedded within a three-dimensional scaffold is an efficient strategy for directed elastogenesis. Copyright © 2014 John Wiley & Sons, Ltd. Copyright © 2014 John Wiley & Sons, Ltd.

Monoclonal antibodies against trophectoderm-specific markers during mouse blastocyst formation.

PubMed Central

Brûlet, P; Babinet, C; Kemler, R; Jacob, F

1980-01-01

Two-dimensional gel electrophoresis has allowed the detection of proteins characteristic of inner cell mass and trophectoderm in mouse blastocyst. Certain of the proteins characterizing trophectoderm copurify with intermediate filaments from trophectoderm and a trophoblastoma cell line. A monoclonal antibody prepared against proteins of these intermediate filaments labels a filament network in trophectoderm but not in inner cell mass cells. Images PMID:6933460

Layer-by-Layer Self-Assembling Gold Nanorods and Glucose Oxidase onto Carbon Nanotubes Functionalized Sol-Gel Matrix for an Amperometric Glucose Biosensor

PubMed Central

Wu, Baoyan; Hou, Shihua; Miao, Zhiying; Zhang, Cong; Ji, Yanhong

2015-01-01

A novel amperometric glucose biosensor was fabricated by layer-by-layer self-assembly of gold nanorods (AuNRs) and glucose oxidase (GOD) onto single-walled carbon nanotubes (SWCNTs)-functionalized three-dimensional sol-gel matrix. A thiolated aqueous silica sol containing SWCNTs was first assembled on the surface of a cleaned Au electrode, and then the alternate self-assembly of AuNRs and GOD were repeated to assemble multilayer films of AuNRs-GOD onto SWCNTs-functionalized silica gel for optimizing the biosensor. Among the resulting glucose biosensors, the four layers of AuNRs-GOD-modified electrode showed the best performance. The sol-SWCNTs-(AuNRs-GOD)4/Au biosensor exhibited a good linear range of 0.01–8 mM glucose, high sensitivity of 1.08 μA/mM, and fast amperometric response within 4 s. The good performance of the proposed glucose biosensor could be mainly attributed to the advantages of the three-dimensional sol-gel matrix and stereo self-assembly films, and the natural features of one-dimensional nanostructure SWCNTs and AuNRs. This study may provide a new facile way to fabricate the enzyme-based biosensor with high performance. PMID:28347080

A supramolecular gel electrolyte formed from amide based co-gelator for quasi-solid-state dye-sensitized solar cell with boosted electron kinetic processes

NASA Astrophysics Data System (ADS)

Huo, Zhipeng; Wang, Lu; Tao, Li; Ding, Yong; Yi, Jinxin; Alsaedi, Ahmed; Hayat, Tasawar; Dai, Songyuan

2017-08-01

A supramolecular gel electrolyte (Tgel > 100 °C) is formed from N,N‧-1,8-octanediylbis-dodecanamide and iodoacetamide as two-component co-gelator, and introduced into the quasi-solid-state dye-sensitized solar cells (QS-DSSCs). The different morphologies of microscopic network between two-component and single-component gel electrolytes have influence on the diffusion of redox couple in gel electrolytes and further affect the electron kinetic processes in QS-DSSCs. Compared with the single-component gel electrolyte, the two-component gel electrolyte has less compact gel network and weaker steric hindrance effect, which provides more effective charge transport channel for the diffusion of I3/I- redox couple. Meanwhile, the sbnd NH2 groups of iodoacetamide molecules interact with Li+ and I3-, which also accelerate the transport of I3-/I- and decrease in the I3- concentration in the TiO2/electrolyte interface. As a result, nearly a 12% improvement in short-circuit photocurrent density (Jsc) and much higher open circuit potential (Voc) are found in the two-component gel electrolyte based QS-DSSC. Consequently, the QS-DSSC based on the supramolecular gel electrolyte obtains a 17% enhancement in the photoelectric conversion efficiency (7.32%) in comparison with the QS-DSSC based on the single-component gel electrolyte (6.24%). Furthermore, the degradations of these QS-DSSCs are negligible after one sun light soaking with UV cutoff filter at 50 °C for 1000 h.

Architectural design, interior decoration, and three-dimensional plumbing en route to multifunctional nanoarchitectures.

PubMed

Long, Jeffrey W

2007-09-01

Ultraporous aperiodic solids, such as aerogels and ambigels, are sol-gel-derived equivalents of architectures. The walls are defined by the nanoscopic, covalently bonded solid network of the gel. The vast open, interconnected space characteristic of a building is represented by the three-dimensionally continuous nanoscopic pore network. We discuss how an architectural construct serves as a powerful metaphor that guides the chemist in the design of aerogel-like nanoarchitectures and in their physical and chemical transformation into multifunctional objects that yield high performance for rate-critical applications.

Enzymatically Cross-linked Alginic-Hyaluronic acid Composite Hydrogels As Cell Delivery Vehicles

PubMed Central

Ganesh, Nitya; Hanna, Craig; Nair, Shantikumar V.; Nair, Lakshmi S.

2013-01-01

An injectable composite gel was developed from alginic and hyaluronic acid. The ezymatically cross-linked injectable gels were prepared via the oxidative coupling of tyramine modified sodium algiante and sodium hyaluronate in the presence of horse radish peroxidase (HRP) and hydrogen peroxide (H2O2). The composite gels were prepared by mixing equal parts of the two tryaminated polymer solutions in 10U HRP and treating with 1.0% H2O2. The properties of the alginate gels were significanly affected by the addition of hyaluronic acid. The percentage water absorption and storage modulus of the composite gels were found to be lower than the alginate gels. The alginate and composite gels showed lower protein release compared to hyaluronate gels in the absence of hyaluronidase. Even hyaluronate gels showed only approximately 10% protein release after 14 days incubation in phosphate buffer solution. ATDC-5 cells encapsulated in the injectable gels showed high cell viability. The composite gels showed the presence of enlarged spherical cells with significantly higher metabolic activity compared to cells in hyaluronic and alginic acid gels. The results suggest the potential of the composite approach to develop covalently cross-linked hydrogels with tuneable physical, mechanical, and biological properties. PMID:23357799

Characterization of Two Cysteine Transfer RNA Genes from Xenopus Laevis

DTIC Science & Technology

1984-07-12

containing amino acids glycine, alanine and serine, are produced by the posterior silk gland of Bombyx mori and therefore high level of tRNAgly, tRNA^Ia...1979) Studies on tRNA adaptation, tRNA turnover, precursor tRNA and tRNA gene distribution in Bombyx mori by using two-dimensional polyacrylamlde gel...Nucleic Acids Research, 1^, 8537-8546. 26. Garber, R.L. and Gage, L.P. (1979) Transcription of a cloned Bombyx mori tRNA^2 gene: Nucleotide sequence of

Electrolyte diodes with weak acids and bases. I. Theory and an approximate analytical solution.

PubMed

Iván, Kristóf; Simon, Péter L; Wittmann, Mária; Noszticzius, Zoltán

2005-10-22

Until now acid-base diodes and transistors applied strong mineral acids and bases exclusively. In this work properties of electrolyte diodes with weak electrolytes are studied and compared with those of diodes with strong ones to show the advantages of weak acids and bases in these applications. The theoretical model is a one dimensional piece of gel containing fixed ionizable groups and connecting reservoirs of an acid and a base. The electric current flowing through the gel is measured as a function of the applied voltage. The steady-state current-voltage characteristic (CVC) of such a gel looks like that of a diode under these conditions. Results of our theoretical, numerical, and experimental investigations are reported in two parts. In this first, theoretical part governing equations necessary to calculate the steady-state CVC of a reverse-biased electrolyte diode are presented together with an approximate analytical solution of this reaction-diffusion-ionic migration problem. The applied approximations are quasielectroneutrality and quasiequilibrium. It is shown that the gel can be divided into an alkaline and an acidic zone separated by a middle weakly acidic region. As a further approximation it is assumed that the ionization of the fixed acidic groups is complete in the alkaline zone and that it is completely suppressed in the acidic one. The general solution given here describes the CVC and the potential and ionic concentration profiles of diodes applying either strong or weak electrolytes. It is proven that previous formulas valid for a strong acid-strong base diode can be regarded as a special case of the more general formulas presented here.

High transparent shape memory gel

NASA Astrophysics Data System (ADS)

Gong, Jin; Arai, Masanori; Kabir, M. H.; Makino, Masato; Furukawa, Hidemitsu

2014-03-01

Gels are a new material having three-dimensional network structures of macromolecules. They possess excellent properties as swellability, high permeability and biocompatibility, and have been applied in various fields of daily life, food, medicine, architecture, and chemistry. In this study, we tried to prepare new multi-functional and high-strength gels by using Meso-Decoration (Meso-Deco), one new method of structure design at intermediate mesoscale. High-performance rigid-rod aromatic polymorphic crystals, and the functional groups of thermoreversible Diels-Alder reaction were introduced into soft gels as crosslinkable pendent chains. The functionalization and strengthening of gels can be realized by meso-decorating the gels' structure using high-performance polymorphic crystals and thermoreversible pendent chains. New gels with good mechanical properties, novel optical properties and thermal properties are expected to be developed.

Meso-decorated self-healing gels: network structure and properties

NASA Astrophysics Data System (ADS)

Gong, Jin; Sawamura, Kensuke; Igarashi, Susumu; Furukawa, Hidemitsu

2013-04-01

Gels are a new material having three-dimensional network structures of macromolecules. They possess excellent properties as swellability, high permeability and biocompatibility, and have been applied in various fields of daily life, food, medicine, architecture, and chemistry. In this study, we tried to prepare new multi-functional and high-strength gels by using Meso-Decoration (Meso-Deco), one new method of structure design at intermediate mesoscale. High-performance rigid-rod aromatic polymorphic crystals, and the functional groups of thermoreversible Diels-Alder reaction were introduced into soft gels as crosslinkable pendent chains. The functionalization and strengthening of gels can be realized by meso-decorating the gels' structure using high-performance polymorphic crystals and thermoreversible pendent chains. New gels with good mechanical properties, novel optical properties and thermal properties are expected to be developed.

Human dermal and gingival fibroblasts in a three-dimensional culture: a comparative study on matrix remodeling.

PubMed

Chaussain Miller, C; Septier, D; Bonnefoix, M; Lecolle, S; Lebreton-Decoster, C; Coulomb, B; Pellat, B; Godeau, G

2002-03-01

Free-floating collagen lattice is considered a useful tool for assessing wound healing in vitro. This work compared extracellular matrix remodeling in collagen lattices populated by gingival or dermal fibroblasts. For 21 days we followed gel contraction and changes in cell number of collagen lattices seeded with l.5 x 10(5) fibroblasts of each tissue. We also used indirect immunodetection to study extracellular matrix components, metalloproteinases (MMPs), and their tissues inhibitors (TIMPs). In addition, the presence of MMPs and TIMPs in the culture media was analyzed by zymography and western blotting. No significant difference was found concerning gel contraction and changes in cell number. We observed the early expression of fibrillin I and collagen type III, apparently codistributed and at the end of the gel contraction their disappearance. Concomitantly we demonstrated the expression of MMPs and TIMPs, initially localized in cellular cytoplasm, then spreading in the extracellular compartment, and even found in the culture medium. This remodeling was more rapid and intense with gingival fibroblasts than dermal fibroblasts. In conclusion, gingival fibroblasts seem more efficient at remodeling the connective tissue than dermal fibroblasts and could lead to the better wound healing observed in vivo.

Novel royal jelly proteins identified by gel-based and gel-free proteomics.

PubMed

Han, Bin; Li, Chenxi; Zhang, Lan; Fang, Yu; Feng, Mao; Li, Jianke

2011-09-28

Royal jelly (RJ) plays an important role in caste determination of the honeybee; the genetically same female egg develops into either a queen or worker bee depending on the time and amount of RJ fed to the larvae. RJ also has numerous health-promoting properties for humans. Gel-based and gel-free proteomics approaches and high-performance liquid chromatography-chip quadruple time-of-flight tandem mass spectrometry were applied to comprehensively investigate the protein components of RJ. Overall, 37 and 22 nonredundant proteins were identified by one-dimensional gel electrophoresis and gel-free analysis, respectively, and 19 new proteins were found by these two proteomics approaches. Major royal jelly proteins (MRJPs) were identified as the principal protein components of RJ, and proteins related to carbohydrate metabolism such as glucose oxidase, α-glucosidase precursor, and glucose dehydrogenase were also successfully identified. Importantly, the 19 newly identified proteins were mainly classified into three functional categories: oxidation-reduction (ergic53 CG6822-PA isoform A isoform 1, Sec61 CG9539-PA, and ADP/ATP translocase), protein binding (regucalcin and translationally controlled tumor protein CG4800-PA isoform 1), and lipid transport (apolipophorin-III-like protein). These new findings not only significantly increase the RJ proteome coverage but also help to provide new knowledge of RJ for honeybee biology and potential use for human health promotion.

Effectiveness of Aloe Vera gel compared with 1% silver sulphadiazine cream as burn wound dressing in second degree burns.

PubMed

Shahzad, Muhammad Naveed; Ahmed, Naheed

2013-02-01

To assess the efficacy of Aloe Vera gel compared with 1% silver sulfadiazine cream as a burn dressing for the treatment of superficial and partial thickness burns. This Interventional Comparative study was carried out at the Burn unit and Plastic surgery department, Nishtar Hospital Multan, Pakistan from July 2008 to December 2010. A total of 50 patients with superficial and partial thickness burns were divided into two equal groups randomly by consecutive sampling method, one group was dressed with Aloe Vera gel while the other was treated with 1% silversulphadiazine cream, and the results regarding duration of wound epithelialization, pain relief and cost of treatment were compared. In patients treated with Aloe Vera gel, healing of burn wounds were remarkably early than those patients treated with 1% silver sulfadiazine. All the patients of Aloe Vera group were relieved of pain earlier than those patients who were treated with SSD. Thermal burns patients dressed with Aloe Vera gel showed advantage compared to those dressed with SSD regarding early wound epithelialization, earlier pain relief and cost-effectiveness.

Investigation of endogenous soybean food allergens by using a 2-dimensional gel electrophoresis approach.

PubMed

Rouquié, David; Capt, Annabelle; Eby, William H; Sekar, Vaithilingam; Hérouet-Guicheney, Corinne

2010-12-01

As part of the safety assessment of genetically modified (GM) soybean, 2-dimensional gel electrophoresis analyses were performed with the isoxaflutole and glyphosate tolerant soybean FG72, its non-GM near-isogenic counterpart (Jack) and three commercial non-GM soybean lines. The objective was to compare the known endogenous human food allergens in seeds in the five different soybean lines in order to evaluate any potential unintended effect(s) of the genetic modification. In total, 37 protein spots representing five well known soybean food allergen groups were quantified in each genotype. Qualitatively, all the allergenic proteins were detected in the different genetic backgrounds. Quantitatively, among 37 protein spots, the levels of accumulation of three allergens were slightly lower in the GM soybean than in the non-GM counterparts. Specifically, while the levels of two of these three allergens fell within the normal range of variation observed in the four non-GM varieties, the level of the third allergen was slightly below the normal range. Overall, there was no significant increase in the level of allergens in FG72 soybean seeds. Therefore, the FG72 soybean can be considered as safe as its non-GM counterpart with regards to endogenous allergenicity. Additional research is needed to evaluate the biological variability in the levels of endogenous soybean allergens and the correlation between level of allergens and allergenic potential in order to improve the interpretation of these data in the safety assessment of GM soybean context. Copyright © 2010 Elsevier Inc. All rights reserved.

Kinetics of swelling of polyelectrolyte gels: Fixed degree of ionization

NASA Astrophysics Data System (ADS)

Sen, Swati; Kundagrami, Arindam

2015-12-01

The swelling kinetics of uncharged and charged polymer (polyelectrolyte) gels in salt-free conditions is studied in one dimension by solving the constitutive equation of motion (Newton's law for the elementary gel volume) of the displacement variable by two theoretical methods: one in which the classical definition of stress is used with the bulk modulus taken as a parameter, and the other in which a phenomenological expression of the osmotic stress as a function of polymer density and degree of ionization is taken as an input to the dynamics. The time-evolution profiles for spatially varying polymer density and stress, along with the location of the gel-solvent interface, are obtained from the two methods. We show that both the polymer density (volume fraction) and stress inside the gel follow expected behaviours of being maximum for the uniformly shrunken gel, and relaxing slowly to the lowest values as the gel approaches equilibrium. We further show that, by comparing the temporal profiles of the gel-solvent interface and other variables between the two methods, one may attempt to assign an effective bulk modulus to the polyelectrolyte gel as a function of the degree of ionization and other parameters of the gel such as hydrophobicity, cross-link density, and the temperature. The major result we get is that the effective bulk modulus of a polyelectrolyte gel increases monotonically with its degree of ionization. In the process of identifying the parameters for a monotonic swelling, we calculated using a well-known expression of the free energy the equilibrium results of two-phase co-existence and the critical point of a polyelectrolyte gel with a fixed degree of ionization.

Kinetics of swelling of polyelectrolyte gels: Fixed degree of ionization.

PubMed

Sen, Swati; Kundagrami, Arindam

2015-12-14

The swelling kinetics of uncharged and charged polymer (polyelectrolyte) gels in salt-free conditions is studied in one dimension by solving the constitutive equation of motion (Newton's law for the elementary gel volume) of the displacement variable by two theoretical methods: one in which the classical definition of stress is used with the bulk modulus taken as a parameter, and the other in which a phenomenological expression of the osmotic stress as a function of polymer density and degree of ionization is taken as an input to the dynamics. The time-evolution profiles for spatially varying polymer density and stress, along with the location of the gel-solvent interface, are obtained from the two methods. We show that both the polymer density (volume fraction) and stress inside the gel follow expected behaviours of being maximum for the uniformly shrunken gel, and relaxing slowly to the lowest values as the gel approaches equilibrium. We further show that, by comparing the temporal profiles of the gel-solvent interface and other variables between the two methods, one may attempt to assign an effective bulk modulus to the polyelectrolyte gel as a function of the degree of ionization and other parameters of the gel such as hydrophobicity, cross-link density, and the temperature. The major result we get is that the effective bulk modulus of a polyelectrolyte gel increases monotonically with its degree of ionization. In the process of identifying the parameters for a monotonic swelling, we calculated using a well-known expression of the free energy the equilibrium results of two-phase co-existence and the critical point of a polyelectrolyte gel with a fixed degree of ionization.

Development of an in situ evaluation system for neural cells using extracellular matrix-modeled gel culture.

PubMed

Nagai, Takayuki; Ikegami, Yasuhiro; Mizumachi, Hideyuki; Shirakigawa, Nana; Ijima, Hiroyuki

2017-10-01

Two-dimensional monolayer culture is the most popular cell culture method. However, the cells may not respond as they do in vivo because the culture conditions are different from in vivo conditions. However, hydrogel-embedding culture, which cultures cells in a biocompatible culture substrate, can produce in vivo-like cell responses, but in situ evaluation of cells in a gel is difficult. In this study, we realized an in vivo-like environment in vitro to produce cell responses similar to those in vivo and established an in situ evaluation system for hydrogel-embedded cell responses. The extracellular matrix (ECM)-modeled gel consisted of collagen and heparin (Hep-col) to mimic an in vivo-like environment. The Hep-col gel could immobilize growth factors, which is important for ECM functions. Neural stem/progenitor cells cultured in the Hep-col gel grew and differentiated more actively than in collagen, indicating an in vivo-like environment in the Hep-col gel. Second, a thin-layered gel culture system was developed to realize in situ evaluation of the gel-embedded cells. Cells in a 200-μm-thick gel could be evaluated clearly by a phase-contrast microscope and immunofluorescence staining through reduced optical and diffusional effects. Finally, we found that the neural cells cultured in this system had synaptic connections and neuronal action potentials by immunofluorescence staining and Ca 2+ imaging. In conclusion, this culture method may be a valuable evaluation system for neurotoxicity testing. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Self-Assembly of a Strong Polyhedral Oligomeric Silsesquioxane Core-Based Aspartate Derivative Dendrimer Supramolecular Gelator in Different Polarity Solvents.

PubMed

He, Huiwen; Chen, Si; Tong, Xiaoqian; An, Zhihang; Ma, Meng; Wang, Xiaosong; Wang, Xu

2017-11-21

Aromatic groups are introduced into the end peripherals of polyhedral oligomeric silsesquioxane (POSS) core-based organic/inorganic hybrid supramolecules to get a novel dendrimer gelator POSS-Z-Asp(OBzl) (POSS-ASP), which have eight aspartate derivative arms to make full use of strong π-π stacking forces to get strong supramolecular gels in addition to multiple hydrogen bindings and van der Waals interactions. POSS-ASP can self-assemble into three-dimensional nanoscale gel networks to provide hybrid physical gels especially with strong mechanical properties and fast-recovery behaviors. Two totally different morphologies of the connected spherical particle structures and banded ultralong fibers are observed owing to the polarity of solvents confirmed by the scanning electron microscopy, polarized optical microscopy, and transmission electron microscopy techniques, expecting the existing various self-assembly models and illustrating the peripherals of the dendrimer and the polarity of solvents having huge influences in the supramolecular self-assembly mechanism. What is more, the thermal stability, rheological properties, and network architecture information have also been investigated via tube-inversion, rotational rheometer, and powder X-ray diffraction methods, the results of which confirm the two different gel formation mechanisms that make POSS-ASP to exhibit two totally different thermal and mechanical properties. Such a study reports a new gelation system in organic or organic/aqueous mixed solvents, which can be helpful for investigating the relationship of dendritic supramolecular gelation and different polarity solvents during the supramolecular self-assembly process of gelators.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bradley, B.; Brown, D.

Biological endpoints can complement chemical analyses in monitoring environmental remediation. In some cases the levels of chemical detection are so low that the costs of clean-up to no detection would be prohibitive. And chemical tests do not indicate the availability of the contaminants to the biota. On the other hand many if not most biological tests lack specificity. The authors have investigated a protein expression assay to establish an endpoint for clean-up of sulfur mustard and breakdown products. Earthworms (Lumbricus terrestris) were exposed to sulfur mustard (SM), a breakdown product thiodiethanol (TDE), and ethylene glycol, the solvent for the twomore » chemicals. Tissue from the lining of the coelomic cavity was taken from each of 6 worms in each treatment class. Soluble proteins were extracted and separated on one and two-dimensional (1D and 2D) gels. The 1 D gels showed no difference by eye but the patterns from control and solvent control worms on 2D gels differed from those of worms exposed to TDE and SM. The 1D gel data were digitized and analyzed by pattern recognition using artificial neural networks. The protein patterns under the two treatments and the two controls were learned in one set of data and successfully recognized in a second. This indicated that what was learned was useful in recognizing patterns induced by SM and TDE. Thus a possible endpoint for remediation would be the protein pattern at no effect levels of chemicals of interest.« less

Protein expression profiles of human lymph and plasma mapped by 2D-DIGE and 1D SDS–PAGE coupled with nanoLC–ESI–MS/MS bottom-up proteomics

PubMed Central

Clement, Cristina C.; Aphkhazava, David; Nieves, Edward; Callaway, Myrasol; Olszewski, Waldemar; Rotzschke, Olaf; Santambrogio, Laura

2013-01-01

In this study a proteomic approach was used to define the protein content of matched samples of afferent prenodal lymph and plasma derived from healthy volunteers. The analysis was performed using two analytical methodologies coupled with nanoliquid chromatography-tandem mass spectrometry: one-dimensional gel electrophoresis (1DEF nanoLC Orbitrap–ESI–MS/MS), and two-dimensional fluorescence difference-in-gel electrophoresis (2D-DIGE nanoLC–ESI–MS/MS). The 253 significantly identified proteins (p<0.05), obtained from the tandem mass spectrometry data, were further analyzed with pathway analysis (IPA) to define the functional signature of prenodal lymph and matched plasma. The 1DEF coupled with nanoLC–MS–MS revealed that the common proteome between the two biological fluids (144 out of 253 proteins) was dominated by complement activation and blood coagulation components, transporters and protease inhibitors. The enriched proteome of human lymph (72 proteins) consisted of products derived from the extracellular matrix, apoptosis and cellular catabolism. In contrast, the enriched proteome of human plasma (37 proteins) consisted of soluble molecules of the coagulation system and cell–cell signaling factors. The functional networks associated with both common and source-distinctive proteomes highlight the principal biological activity of these immunologically relevant body fluids. PMID:23202415

Evaluation of SPLAT with spinosad and methyl eugenol or cue-lure for "attract-and-kill" of oriental and melon fruit flies (Diptera: Tephritidae) in Hawaii.

PubMed

Vargas, Roger I; Stark, John D; Hertlein, Mark; Neto, Agenor Mafra; Coler, Reginald; Piñero, Jaime C

2008-06-01

Specialized Pheromone and Lure Application Technology (SPLAT) methyl eugenol (ME) and cue-lure (C-L) "attract-and-kill" sprayable formulations containing spinosad were compared with other formulations under Hawaiian weather conditions against oriental fruit fly, Bactrocera dorsalis (Hendel), and melon fly, Bactrocera cucurbitae (Coquillett) (Diptera: Tephritidae), respectively. Field tests were conducted with three different dispensers (Min-U-Gel, Acti-Gel, and SPLAT) and two different insecticides (naled and spinosad). SPLAT ME with spinosad was equal in performance to the standard Min-U-Gel ME with naled formulation up to 12 wk. SPLAT C-L with spinosad was equal in performance to the standard Min-U-Gel C-L with naled formulation during weeks 7 to12, but not during weeks 1-6. In subsequent comparative trials, SPLAT ME + spinosad compared favorably with the current standard of Min-U-Gel ME + naled for up to 6 wk, and it was superior from weeks 7 to 12 in two separate tests conducted in a papaya (Carica papaya L.) orchard and a guava (Psidium guajava L.) orchard, respectively. In outdoor paired weathering tests (fresh versus weathered), C-L dispensers (SPLAT + spinosad, SPLAT + naled, and Min-U-Gel + naled) were effective up to 70 d. Weathered ME dispensers with SPLAT + spinosad compared favorably with SPLAT + naled and Min-U-Gel + naled, and they were equal to fresh dispensers for 21-28 d, depending on location. Our current studies indicate that SPLAT ME and SPLAT C-L sprayable attract-and-kill dispensers containing spinosad are a promising substitute for current liquid organophosphate insecticide formulations used for areawide suppression of B. dorsalis and B. cucurbitae in Hawaii.

Comparative Proteomics of Rubber Latex Revealed Multiple Protein Species of REF/SRPP Family Respond Diversely to Ethylene Stimulation among Different Rubber Tree Clones.

PubMed

Tong, Zheng; Wang, Dan; Sun, Yong; Yang, Qian; Meng, Xueru; Wang, Limin; Feng, Weiqiang; Li, Ling; Wurtele, Eve Syrkin; Wang, Xuchu

2017-05-02

Rubber elongation factor (REF) and small rubber particle protein (SRPP) are two key factors for natural rubber biosynthesis. To further understand the roles of these proteins in rubber formation, six different genes for latex abundant REF or SRPP proteins, including REF 138,175,258 and SRPP 117,204,243 , were characterized from Hevea brasiliensis Reyan (RY) 7-33-97. Sequence analysis showed that REFs have a variable and long N-terminal, whereas SRPPs have a variable and long C-terminal beyond the REF domain, and REF 258 has a β subunit of ATPase in its N-terminal. Through two-dimensional electrophoresis (2-DE), each REF/SRPP protein was separated into multiple protein spots on 2-DE gels, indicating they have multiple protein species. The abundance of REF/SRPP proteins was compared between ethylene and control treatments or among rubber tree clones with different levels of latex productivity by analyzing 2-DE gels. The total abundance of each REF/SRPP protein decreased or changed a little upon ethylene stimulation, whereas the abundance of multiple protein species of the same REF/SRPP changed diversely. Among the three rubber tree clones, the abundance of the protein species also differed significantly. Especially, two protein species of REF 175 or REF 258 were ethylene-responsive only in the high latex productivity clone RY 8-79 instead of in RY 7-33-97 and PR 107. Some individual protein species were positively related to ethylene stimulation and latex productivity. These results suggested that the specific protein species could be more important than others for rubber production and post-translational modifications might play important roles in rubber biosynthesis.

Injectable Silica–Permanganate Gel as a Slow-Release MnO 4 - Source for Groundwater Remediation. Rheological Properties and Release Dynamics

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Shuo; Oostrom, Martinus; Truex, Michael J.

2016-01-12

Injectable slow-release permanganate gel (ISRPG), formed by mixing KMnO 4 solution with fumed silica powder, may have a potential application in remediating chlorinated solvent plumes in groundwater. A series of batch, column, and flow cell experiments has been completed to test the gel behavior under a variety of conditions. The experiments have provided information on ISRPG rheology, permanganate (MnO 4 - ) release dynamics and distribution, and trichloroethene (TCE) degradation by ISRPG-released oxidant. The gel possesses remarkable shear thinning characteristics, resulting in a relative low viscosity during mixing, and facilitating its subsurface injection and distribution. Batch tests revealed that MnOmore » 4 - was diffused out from ISRPG into water while the gel did not dissolve or disperse into water but maintained its initial shape. Column experiments showed that MnO 4 - release from ISRPG lasted considerably longer than the release from aqueous solution. TCE degradation by ISRPG-released MnO 4 - was much more effective than that when MnO 4 - was delivered using aqueous solution injection. In two-dimensional flow cell experiments, it was demonstrated that ISRPG slowly released a long-lasting low concentration MnO 4 - plume sufficient for remediation and sustainable in an aquifer for a long period of time.« less

Well-aligned Vertically Oriented ZnO Nanorod Arrays and their Application in Inverted Small Molecule Solar Cells.

PubMed

Lin, Ming-Yi; Wu, Shang-Hsuan; Hsiao, Li-Jen; Budiawan, Widhya; Chen, Shih-Lun; Tu, Wei-Chen; Lee, Chia-Yen; Chang, Yia-Chung; Chu, Chih-Wei

2018-04-25

This manuscript describes how to design and fabricate efficient inverted solar cells, which are based on a two-dimensional conjugated small molecule (SMPV1) and [6,6]-phenyl-C71-butyric acid methyl ester (PC71BM), by utilizing ZnO nanorods (NRs) grown on a high quality Al-doped ZnO (AZO) seed layer. The inverted SMPV1:PC71BM solar cells with ZnO NRs that grew on both a sputtered and sol-gel processed AZO seed layer are fabricated. Compared with the AZO thin film prepared by the sol-gel method, the sputtered AZO thin film exhibits better crystallization and lower surface roughness, according to X-ray diffraction (XRD) and atomic force microscope (AFM) measurements. The orientation of the ZnO NRs grown on a sputtered AZO seed layer shows better vertical alignment, which is beneficial for the deposition of the subsequent active layer, forming better surface morphologies. Generally, the surface morphology of the active layer mainly dominates the fill factor (FF) of the devices. Consequently, the well-aligned ZnO NRs can be used to improve the carrier collection of the active layer and to increase the FF of the solar cells. Moreover, as an anti-reflection structure, it can also be utilized to enhance the light harvesting of the absorption layer, with the power conversion efficiency (PCE) of solar cells reaching 6.01%, higher than the sol-gel based solar cells with an efficiency of 4.74%.

Proteomic Profile of Mabuya sp. (Squamata: Scincidae) Ovary and Placenta During Gestation.

PubMed

Hernández-Díaz, Nathaly; Torres, Rodrigo; Ramírez-Pinilla, Martha Patricia

2017-06-01

Reptiles are one of the most diverse groups of vertebrates, providing an integrated system for comparative studies on metabolic, animal physiology, and developmental biology. However, the molecular data available are limited and only recently have started to call attention in the "omics" sciences. Mabuya sp. is a viviparous placentrotrophic skink with particular reproductive features, including microlecithal eggs, early luteolysis, prolonged gestation, and development of a highly specialized placenta. This placenta is responsible for respiratory exchange and the transference of all nutrients necessary for embryonic development. Our aim was to identify differentially expressed proteins in the ovary and placenta of Mabuya sp. during early, mid, and late gestation; their possible metabolic pathways; and biological processes. We carried out a comparative proteomic analysis during gestation in both tissues by sodium dodecyl sulfate polyacrylamide gel electrophoresis, two-dimensional gel electrophoresis, and matrix-assisted laser desorption/ionization. Differential protein expression in both tissues (Student's t-test P < 0.05) was related to several processes such as cell structure, cell movement, and energy. Proteins found in ovary are mainly associated with follicular development and its regulation. In the placenta, particularly during mid and late gestation, protein expression is involved in nutrient metabolism, transport, protein synthesis, and embryonic development. This work provides new insights about the proteins expressed and their physiological mechanisms in Mabuya sp. placenta and ovary during gestation. © 2017 Wiley Periodicals, Inc.

Aldehyde dehydrogenase induction in arsenic-exposed rat bladder epithelium.

PubMed

Huang, Ya-Chun; Yu, Hsin-Su; Chai, Chee-Yin

2016-01-01

Arsenic is widely distributed in the environment. Many human cancers, including urothelial carcinoma (UC), show a dose-dependent relationship with arsenic exposure in the south-west coast of Taiwan (also known as the blackfoot disease (BFD) areas). However, the molecular mechanisms of arsenic-mediated UC carcinogenesis has not yet been defined. In vivo study, the rat bladder epithelium were exposed with arsenic for 48 h. The proteins were extracted from untreated and arsenic-treated rat bladder cells and utilized two-dimensional gel electrophoresis and mass spectrometry. Selected peptides were extracted from the gel and identified by quadrupole-time of flight (Q-TOF) Ultima-Micromass spectra. The significantly difference expression of proteins in arsenic-treated groups as compared with untreated groups was confirmed by immunohistochemistry (IHC) and western blotting. We found that thirteen proteins were down-regulated and nine proteins were up-regulated in arsenic-treated rat bladder cells when compared with untreated groups. The IHC and western blotting results confirmed that aldehyde dehydrogenase (ALDH) protein was up-regulated in arsenic-treated rat bladder epithelium. Expression of ALDH protein was significantly higher in UC patients from BFD areas than those from non-BFD areas using IHC (p=0.018). In conclusion, the ALDH protein expression could be used as molecular markers for arsenic-induced transformation. Copyright © 2015 Elsevier GmbH. All rights reserved.

Cloning and characterization of two novel DNases from Streptococcus pyogenes.

PubMed

Hasegawa, Tadao; Torii, Keizo; Hashikawa, Shinnosuke; Iinuma, Yoshitsugu; Ohta, Michio

2002-06-01

The proteins in the culture supernatant (exoproteins) from Streptococcus pyogenes serotype M1 were separated by two-dimensional gel electrophoresis, and their N-terminal amino acid sequences were determined. The amino acid sequences were compared to sequences in the S. pyogenes genome database. The coding sequence showed similarity to sequences of two genes, mf2-v ( mf2 variant) and mf3, which had sequence similarity to genes encoding mitogenic factor (MF); MF has DNase activity. The recombinant genes were expressed in Escherichia coli and the proteins were synthesized. Mf2-v and Mf3 had DNase activity. The activity of Mf2-v was localized to the C-terminal half of the protein. The mf3 gene was shown to be present in most clinically isolated strains of S. pyogenes tested, and the mf2gene was detected in 20% of the isolates. The products of the mf2 and mf3 genes in clinically isolated S. pyogenes strains were thus shown to be DNases.

Urine Collected From Diapers Can Be Used for 2-Dimensional Polyacrylamide Gel Electrophoresis (2D-PAGE) in Infants and Young Children

PubMed Central

Kennedy, Mary Jayne; Griffin, Angela; Su, Ruifeng; Merchant, Michael; Klein, Jon

2011-01-01

Urinary proteomic profiling has potential to identify candidate biomarkers of renal injury in infants provided an adequate urine sample can be obtained. Although diapers are used to obtain urine for clinical evaluation, their use for proteomic analysis has not been investigated. We therefore performed feasibility studies on the use of diaper-extracted urine for 2-dimensional polyacrylamide gel electrophoresis (2D-PAGE). Pediatric waste urine (2–20 mL) was applied to gel-containing, non-gel and cotton-gauze diapers and then mechanically expressed. Urine volume and total protein were measured pre- and post-extraction. Proteins were separated via 2D-PAGE following application of urine (20–40 mL) to each matrix. 2D-PAGE was also performed on clinical specimens collected using each diaper type. Differences in the adsorption and retention of urine volume and protein were noted between matrices. Non-gel and cotton-gauze diapers provided the best protein/volume recovery and the lowest interference with the Bradford assay. 2D-PAGE was also successfully completed using urine samples from both cotton fiber matrices. Conversely, samples from low-gel diapers demonstrated poor protein separation and reproducibility. Diapers containing cotton-fiber matrices appear adequate for 2D-PAGE. Qualitative and quantitative analyses of resolved proteins using replicate, high resolution gels will be required, however, before diaper-extracted urine can be applied in proteomic profiling. PMID:21137001

Process of forming a sol-gel/metal hydride composite

DOEpatents

Congdon, James W [Aiken, SC

2009-03-17

An external gelation process is described which produces granules of metal hydride particles contained within a sol-gel matrix. The resulting granules are dimensionally stable and are useful for applications such as hydrogen separation and hydrogen purification. An additional coating technique for strengthening the granules is also provided.

Two-Dimensional Differential In-Gel Electrophoresis Proteomic Approaches Reveal Urine Candidate Biomarkers in Pediatric Obstructive Sleep Apnea

PubMed Central

Gozal, David; Jortani, Saeed; Snow, Ayelet B.; Kheirandish-Gozal, Leila; Bhattacharjee, Rakesh; Kim, Jinkwan; Capdevila, Oscar Sans

2009-01-01

Rationale: Sleep studies are laborious, expensive, inaccessible, and inconvenient for diagnosing obstructive sleep apnea (OSA) in children. Objectives: To examine whether the urinary proteome uncovers specific clusters that are differentially expressed in the urine of children with OSA. Methods: Two-dimensional differential in-gel electrophoresis (2D-DIGE) and mass spectrometry proteomics followed by validation with western blot of ELISA. Measurements and Main Results: Morning urine proteins from 60 children with polysomnographically confirmed OSA and from matched children with primary snoring (n = 30) and control subjects (n = 30) were assessed. A total of 16 proteins that are differentially expressed in OSA were identified, and 7 were confirmed by either immunoblots or ELISA. Among the latter, receiver–operator curve analyses of urinary concentrations of uromodulin, urocortin-3, orosomucoid-1, and kallikrein assigned favorable predictive properties to these proteins. Furthermore, combinatorial approaches indicated that the presence of values beyond the calculated cutoff concentrations for three or more of the proteins yielded a sensitivity of 95% and a specificity of 100%. Conclusions: Proteomic approaches reveal that pediatric OSA is associated with specific and consistent alterations in urinary concentrations of specific protein clusters. Future studies aiming to validate this approach as a screening method of habitually snoring children appears warranted. PMID:19797158

Identification of DNA-binding proteins that interact with the 5'-flanking region of the human D-amino acid oxidase gene by pull-down assay coupled with two-dimensional gel electrophoresis and mass spectrometry.

PubMed

Tran, Diem Hong; Shishido, Yuji; Chung, Seong Pil; Trinh, Huong Thi Thanh; Yorita, Kazuko; Sakai, Takashi; Fukui, Kiyoshi

2015-12-10

D-Amino acid oxidase (DAO) is a flavoenzyme that metabolizes D-amino acids and is expected to be a promising therapeutic target of schizophrenia and glioblastoma. The study of DNA-binding proteins has yielded much information in the regulation of transcription and other biological processes. However, proteins interacting with DAO gene have not been elucidated. Our assessment of human DAO promoter activity using luciferase reporter system indicated the 5'-flanking region of this gene (-4289 bp from transcription initiation site) has a regulatory sequence for gene expression, which is regulated by multi-protein complexes interacting with this region. By using pull-down assay coupled with two-dimensional gel electrophoresis and mass spectrometry, we identified six proteins binding to the 5'-flanking region of the human DAO gene (zinc finger C2HC domain-containing protein 1A; histidine-tRNA ligase, cytoplasmic; molybdenum cofactor biosynthesis protein; 60S ribosomal protein L37; calponin-1; calmodulin binding protein and heterogeneous nuclear ribonucleoprotein A2/B1). These preliminary results will contribute to the advance in the understanding of the potential factors associated with the regulatory mechanism of DAO expression. Copyright © 2015 Elsevier B.V. All rights reserved.

Mobility of long-chain DNA in two-dimensional artificial gels

NASA Astrophysics Data System (ADS)

Turner, Stephen W. P.; Han, Jongyoon; Craighead, Harold G.

2000-03-01

In this study, a two-dimensional array of nanofabricated obstacles is used as an artificial gel to study the electrophoretic mobility dependence of DNA as a function of pore size, molecule length and electric field. Limitations in feature size have prevented previous studies from testing the crossover from the separating to the non-separating regime predicted by the biased reptation model of Lumpkin, Dejardin and Zimm[1] and the modified model of Duke, Semenov and Viovy.[2] That limitation is overcome in this work with the use of electron beam lithography to define features as small as 30 nm. Attainment of these feature sizes was made possible by the use of a sacrificial-layer-based technique for fluidics fabrication.[3] A novel band-launching strategy is used to provide band separation data for the first time in this system. Molecule lengths between 5 and 150 kilobases are studied for electric field strengths from 0.1 to 20 Volts per meter. [1] O. Lumpkin, P. Dejardin and B. Zimm, Biopolymers, Vol. 24, 1573-1593 (1985) [2] T. Duke, A. Semenov and J. Viovy, Phys. Rev. Lett. Vol. 69, No. 22, 3260-3263 (1992) [3] S. Turner, A. Perez, A. Lopez, and H. Craighead, J. Vac. Sci. Technol. B 16(6) 3835-3840 (1998)

Room-temperature synthesis of two-dimensional ultrathin gold nanowire parallel array with tunable spacing.

PubMed

Morita, Clara; Tanuma, Hiromitsu; Kawai, Chika; Ito, Yuki; Imura, Yoshiro; Kawai, Takeshi

2013-02-05

A series of long-chain amidoamine derivatives with different alkyl chain lengths (CnAA where n is 12, 14, 16, or 18) were synthesized and studied with regard to their ability to form organogels and to act as soft templates for the production of Au nanomaterials. These compounds were found to self-assemble into lamellar structures and exhibited gelation ability in some apolar solvents. The gelation concentration, gel-sol phase transition temperature, and lattice spacing of the lamellar structures in organic solvent all varied on the basis of the alkyl chain length of the particular CnAA compound employed. The potential for these molecules to function as templates was evaluated through the synthesis of Au nanowires (NWs) in their organogels. Ultrathin Au NWs were obtained from all CnAA/toluene gel systems, each within an optimal temperature range. Interestingly, in the case of C12AA and C14AA, it was possible to fabricate ultrathin Au NWs at room temperature. In addition, two-dimensional parallel arrays of ultrathin Au NWs were self-assembled onto TEM copper grids as a result of the drying of dispersion solutions of these NWs. The use of CnAA compounds with differing alkyl chain lengths enabled precise tuning of the distance between the Au NWs in these arrays.

Long-Acting Phospholipid Gel of Exenatide for Long-Term Therapy of Type II Diabetes.

PubMed

Hu, Mei; Zhang, Yu; Xiang, Nanxi; Zhong, Ying; Gong, Tao; Zhang, Zhi-Rong; Fu, Yao

2016-06-01

This study aimed to develop a sustained-release formulation of exenatide (EXT) for the long-term therapeutic efficacy in the treatment of type II diabetes. In this study, we present an injectable phospholipid gel by mixing biocompatible phospholipid S100, medium chain triglyceride (MCT) with 85% (w/w) ethanol. A systemic pre-formulation study has been carried out to improve the stability of EXT during formulation fabrication. With the optimized formulation, the pharmacokinetic profiles in rats were studied and two diabetic animal models were employed to evaluate the therapeutic effect of EXT phospholipid gel via a single subcutaneous injection versus repeated injections of normal saline and EXT solution. With optimized formulation, sustained release of exenatide in vivo for over three consecutive weeks was observed after one single subcutaneous injection. Moreover, the pharmacodynamic study in two diabetic models justified that the gel formulation displayed a comparable hypoglycemic effect and controlled blood glucose level compared with exenatide solution treated group. EXT-loaded phospholipid gel represents a promising controlled release system for long-term therapy of type II diabetes.

Influence of Diet on the Proteome of Drosophila Melanogaster as Assessed by Two-Dimensional Gel Electrophoresis and Capillary Liquid Chromatography–Mass Spectrometry: The Hamburger Effect Revisited

PubMed Central

Culwell, Thomas F.; Thulin, Craig D.; Merrell, Karen J.; Graves, Steven W.

2008-01-01

Proteomic biomarker discovery has been called into question. Diamandis hypothesized that seemingly trivial factors, such as eating a hamburger, may cause sufficient proteomic change as to confound proteomic differences. This has been termed the hamburger effect. Little is known about the variability of complex proteomes in response to the environment. Two methods—two-dimensional gel electrophoresis (2DGE) and capillary liquid chromatography–electrospray ionization time-of-flight mass spectrometry (LCMS)—were used to study the hamburger effect in two cross-sections of the soluble fruit fly proteome. 2DGE measured abundant proteins, whereas LCMS measured small proteins and peptides. Proteomic differences between males and females were first evaluated to assess the discriminatory capability of the methods. Likewise, wild-type and white-eyed flies were analyzed as a further demonstration that genetically based proteomic differences could be observed above the background analytical variation. Then dietary interventions were imposed. Ethanol was added to the diet of some populations without significant proteomic effect. However, after a 24-h fast, proteomic differences were found using LCMS but not 2DGE. Even so, only three of ~1000 molecular species were altered significantly, suggesting that the influence of even an extreme diet change produced only modest proteomic variability, and that much of the fruit fly proteome remains relatively constant in response to diet. These experiments suggest that proteomics can be a viable approach to biomarker discovery. PMID:19137114

Sensitivity calibration procedures in optical-CT scanning of BANG 3 polymer gel dosimeters

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Y.; Wuu, Cheng-Shie; Maryanski, Marek J.

2010-02-15

The dose response of the BANG 3 polymer gel dosimeter (MGS Research Inc., Madison, CT) was studied using the OCTOPUS laser CT scanner (MGS Research Inc., Madison, CT). Six 17 cm diameter and 12 cm high Barex cylinders, and 18 small glass vials were used to house the gel. The gel phantoms were irradiated with 6 and 10 MV photons, as well as 12 and 16 MeV electrons using a Varian Clinac 2100EX. Three calibration methods were used to obtain the dose response curves: (a) Optical density measurements on the 18 glass vials irradiated with graded doses from 0 tomore » 4 Gy using 6 or 10 MV large field irradiations; (b) optical-CT scanning of Barex cylinders irradiated with graded doses (0.5, 1, 1.5, and 2 Gy) from four adjacent 4x4 cm{sup 2} photon fields or 6x6 cm{sup 2} electron fields; and (c) percent depth dose (PDD) comparison of optical-CT scans with ion chamber measurements for 6x6 cm{sup 2}, 12 and 16 MeV electron fields. The dose response of the BANG 3 gel was found to be linear and energy independent within the uncertainties of the experimental methods (about 3%). The slopes of the linearly fitted dose response curves (dose sensitivities) from the four field irradiations (0.0752{+-}3%, 0.0756{+-}3%, 0.0767{+-}3%, and 0.0759{+-}3% cm{sup -1} Gy{sup -1}) and the PDD matching methods (0.0768{+-}3% and 0.0761{+-}3% cm{sup -1} Gy{sup -1}) agree within 2.2%, indicating a good reproducibility of the gel dose response within phantoms of the same geometry. The dose sensitivities from the glass vial approach are different from those of the cylindrical Barex phantoms by more than 30%, owing probably to the difference in temperature inside the two types of phantoms during gel formation and irradiation, and possible oxygen contamination of the glass vial walls. The dose response curve obtained from the PDD matching approach with 16 MeV electron field was used to calibrate the gel phantom irradiated with the 12 MeV, 6x6 cm{sup 2} electron field. Three-dimensional dose distributions from the gel measurement and the Eclipse planning system (Varian Corporation, Palo Alto, CA) were compared and evaluated using 3% dose difference and 2 mm distance-to-agreement criteria.« less

Analysis of branched DNA replication and recombination intermediates from prokaryotic cells by two-dimensional (2D) native-native agarose gel electrophoresis.

PubMed

Robinson, Nicholas P

2013-01-01

Branched DNA molecules are generated by the essential processes of replication and recombination. Owing to their distinctive extended shapes, these intermediates migrate differently from linear double-stranded DNA under certain electrophoretic conditions. However, these branched species exist in the cell at much low abundance than the bulk linear DNA. Consequently, branched molecules cannot be visualized by conventional electrophoresis and ethidium bromide staining. Two-dimensional native-native agarose electrophoresis has therefore been developed as a method to facilitate the separation and visualization of branched replication and recombination intermediates. A wide variety of studies have employed this technique to examine branched molecules in eukaryotic, archaeal, and bacterial cells, providing valuable insights into how DNA is duplicated and repaired in all three domains of life.

Double phosphorylation of the myosin regulatory light chain during rigor mortis of bovine Longissimus muscle.

PubMed

Muroya, Susumu; Ohnishi-Kameyama, Mayumi; Oe, Mika; Nakajima, Ikuyo; Shibata, Masahiro; Chikuni, Koichi

2007-05-16

To investigate changes in myosin light chains (MyLCs) during postmortem aging of the bovine longissimus muscle, we performed two-dimensional gel electrophoresis followed by identification with matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The results of fluorescent differential gel electrophoresis showed that two spots of the myosin regulatory light chain (MyLC2) at pI values of 4.6 and 4.7 shifted toward those at pI values of 4.5 and 4.6, respectively, by 24 h postmortem when rigor mortis was completed. Meanwhile, the MyLC1 and MyLC3 spots did not change during the 14 days postmortem. Phosphoprotein-specific staining of the gels demonstrated that the MyLC2 proteins at pI values of 4.5 and 4.6 were phosphorylated. Furthermore, possible N-terminal region peptides containing one and two phosphoserine residues were detected in each mass spectrum of the MyLC2 spots at pI values of 4.5 and 4.6, respectively. These results demonstrated that MyLC2 became doubly phosphorylated during rigor formation of the bovine longissimus, suggesting involvement of the MyLC2 phosphorylation in the progress of beef rigor mortis. Bovine; myosin regulatory light chain (RLC, MyLC2); phosphorylation; rigor mortis; skeletal muscle.

Tretinoin Photostability

PubMed Central

Harper, Julie; Pillai, Radhakrishnan; Moore, Robert

2012-01-01

Background: Various formulations of tretinoin (gel, liquid, cream) have been reported to be unstable on the skin under bright artificial light or sunlight. This photodegradation can potentially influence treatment regimens and possibly modify efficacy. The maximum light energy absorption of tretinoin is in the ultraviolet A region. Objective: To compare the photostability of a micronized formulation of tretinoin (0.05%) aqueos gel with tretinoin (0.025%) gel following exposure to ultraviolet A light for eight hours. Methods: Micronized tretinoin (0.05%) gel and tretinoin (0.025%) gel were then exposed to ultraviolet A light with an integrated intensity from 315 to 400nm of 22watt/m2. Samples of both products were prepared and analyzed for tretinoin and its degradation products using a high-performance liquid chromatography method. Additional duplicate samples were similarly prepared and analyzed after 2,4,6, and 8 hours. Results: There was a nine-percent degradation of micronized tretinoin in the 0.05% aqueous gel compared to 72-percent degradation of tretinoin in the 0.025% gel following eight-hour ultraviolet A light exposure. The small increase in tretinoin degradation products with micronized tretinoin (0.05%) aqueous gel remained below six percent of the labeled concentration compared to a marked increase in tretinoin degradation products with tretinoin 0.025% gel at two hours that increased to over 66-percent labeled concentration at eight hours. Conclusion: Micronized tretinoin (0.05%) aqueous gel showed less than 10-percent degradation when exposed to eight hours of ultraviolet A light, while tretinoin (0.025%) gel showed significant tretinoin degradation. PMID:22328956

Spiral pattern in a radial displacement in a Hele-Shaw cell

NASA Astrophysics Data System (ADS)

Ban, Mitsumasa; Nagatsu, Yuichiro; Hayashi, Atsushi; Kato, Yoshihiro; Tada, Yutaka

2008-11-01

When a reactive and miscible less-viscous liquid displaces a more-viscous liquid in a Hele-Shaw cell, reactive miscible viscous fingering takes place. We have experimentally shown that the pattern created by the displacement of a more-viscous fluid by a less-viscous one in a radial Hele-Shaw cell develops not radially but spirally when a more-viscous sodium polyacrylate solution is displaced by a less-viscous trivalent iron ion (Fe^3+) solution with a sufficiently high concentration of Fe^3+. Another experiment in order to investigate the mechanism of spiral pattern formation revealed that an instantaneous chemical reaction takes place between the two fluids and at high Fe^3+ concentrations it produces a film of the gel at the contact plane. The gel is formed by three-dimensional network structures between the polyacrylate solution and the trivalent iron ion (Fe^3+) solution. We have proposed a physical model that the gel's film is responsible for the form of the spiral pattern.

DIGE Analysis Software and Protein Identification Approaches.

PubMed

Hmmier, Abduladim; Dowling, Paul

2018-01-01

DIGE is a high-resolution two-dimensional gel electrophoresis method, with excellent dynamic range obtained by fluorescent tag labeling of protein samples. Scanned images of DIGE gels show thousands of protein spots, each spot representing a single or a group of protein isoforms. By using commercially available software, each protein spot is defined by an outline, which is digitized and correlated with the quantity of proteins present in each spot. Software packages include DeCyder, SameSpots, and Dymension 3. In addition, proteins of interest can be excised from post-stained gels and identified with conventional mass spectrometry techniques. High-throughput mass spectrometry is performed using sophisticated instrumentation including matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF), MALDI-TOF/TOF, and liquid chromatography tandem mass spectrometry (LC-MS/MS). Tandem MS (MALDI-TOF/TOF or LC-MS/MS), analyzes fragmented peptides, resulting in amino acid sequence information, especially useful when protein spots are low abundant or where a mixture of proteins is present.

Variables affecting resolution of lung phospholipids in one-dimensional thin-layer chromatography.

PubMed

Krahn, J

1987-01-01

Resolution of the confusion in the literature about the separation of lung phospholipids in thin-layer chromatographic systems has awaited a systematic study of the variables that potentially affect this separation. In this study I show that: incorporation of ammonium sulfate into silica gel "GHL" has a dramatic effect on separation of lung phospholipids; this effect is equally dramatic but different in activated and nonactivated gels; when it picks up moisture, ammonium sulfate-activated gel very rapidly loses its ability to resolve lecithin from phosphatidylinositol; in gel containing ammonium sulfate, small amounts of phosphatidylethanolamine are hydrolyzed to lyso-phosphatidylethanolamine.

Lesson learned and dispelled myths: three-dimensional imaging of the human vagina.

PubMed

Barnhart, Kurt T; Pretorius, E Scott; Malamud, Daniel

2004-05-01

Three-dimensional imaging of the human vagina demonstrates that the cross section can be a "W," rather than an "H," and that intravaginal gel can ascend into the endocervix and presumably into the endometrium.

Cell mechanics, structure, and function are regulated by the stiffness of the three-dimensional microenvironment.

PubMed

Chen, J; Irianto, J; Inamdar, S; Pravincumar, P; Lee, D A; Bader, D L; Knight, M M

2012-09-19

This study adopts a combined computational and experimental approach to determine the mechanical, structural, and metabolic properties of isolated chondrocytes cultured within three-dimensional hydrogels. A series of linear elastic and hyperelastic finite-element models demonstrated that chondrocytes cultured for 24 h in gels for which the relaxation modulus is <5 kPa exhibit a cellular Young's modulus of ∼5 kPa. This is notably greater than that reported for isolated chondrocytes in suspension. The increase in cell modulus occurs over a 24-h period and is associated with an increase in the organization of the cortical actin cytoskeleton, which is known to regulate cell mechanics. However, there was a reduction in chromatin condensation, suggesting that changes in the nucleus mechanics may not be involved. Comparison of cells in 1% and 3% agarose showed that cells in the stiffer gels rapidly develop a higher Young's modulus of ∼20 kPa, sixfold greater than that observed in the softer gels. This was associated with higher levels of actin organization and chromatin condensation, but only after 24 h in culture. Further studies revealed that cells in stiffer gels synthesize less extracellular matrix over a 28-day culture period. Hence, this study demonstrates that the properties of the three-dimensional microenvironment regulate the mechanical, structural, and metabolic properties of living cells. Copyright © 2012 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Separation and characterization of needle and xylem maritime pine proteins.

PubMed

Costa, P; Pionneau, C; Bauw, G; Dubos, C; Bahrmann, N; Kremer, A; Frigerio, J M; Plomion, C

1999-01-01

Two-dimensional gel electrophoresis (2-DE) and image analysis are currently used for proteome analysis in maritime pine (Pinus pinaster Ait.). This study presents a database of expressed proteins extracted from needles and xylem, two important tissues for growth and wood formation. Electrophoresis was carried out by isoelectric focusing (IEF) in the first dimension and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in the second. Silver staining made it possible to detect an average of 900 and 600 spots on 2-DE gels from needles and xylem, respectively. A total of 28 xylem and 35 needle proteins were characterized by internal peptide microsequencing. Out of these 63 proteins, 57 (90%) could be identified based on amino acid similarity with known proteins, of which 24 (42%) have already been described in conifers. Overall comparison of both tissues indicated that 29% and 36% of the spots were specific to xylem and needles, respectively, while the other spots were of identical molecular weight and isoelectric point. The homology of spot location in 2-DE patterns was further validated by sequence analysis of proteins present in both tissues. A proteomic database of maritime pine is accessible on the internet (http://www.pierroton.inra.fr/genetics/2D/).

High solid loading aqueous base metal/ceramic feedstock for injection molding

NASA Astrophysics Data System (ADS)

Behi, Mohammad

2001-07-01

Increasing volume fraction of metal powder in feedstock provided lower shrinkage. Reduction of the shrinkage results in better dimensional precision. The rheology of the feedstock material plays an important role to allowing larger volume fractions of the metal powder to be incorporated in the feedstock formulations. The viscosity of the feedstock mainly depends on the binder viscosity, powder volume fraction and characteristics of metal powder. Aqueous polysaccharide agar was used as a baseline binder system for this study. The effect of several gel-strengthening additives on 1.5wt% and 2wt% agar gel was evaluated. A new gel-strengthening additive was found to be the most effective among the others. The effect of other additives such as glucose, sucrose and fructose on viscosity of baseline binder and feedstock was investigated. Two new agar based binder compositions were developed. The use of these new binder formulations significantly improved the volume fraction of the metal powder, the stability of the feedstock, and reduced the final shrinkage of the molded articles. Two types of 17-4PH stainless steel metal powders, one gas atomized and, the other water atomized, were used for this research.

Gel nanostructure in alkali-activated binders based on slag and fly ash, and effects of accelerated carbonation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bernal, Susan A., E-mail: s.bernal@sheffield.ac.uk; Department of Materials Science and Engineering, The University of Sheffield, Sheffield S1 3JD; Provis, John L., E-mail: j.provis@sheffield.ac.uk

2013-11-15

Binders formed through alkali-activation of slags and fly ashes, including ‘fly ash geopolymers’, provide appealing properties as binders for low-emissions concrete production. However, the changes in pH and pore solution chemistry induced during accelerated carbonation testing provide unrealistically low predictions of in-service carbonation resistance. The aluminosilicate gel remaining in an alkali-activated slag system after accelerated carbonation is highly polymerised, consistent with a decalcification mechanism, while fly ash-based binders mainly carbonate through precipitation of alkali salts (bicarbonates at elevated CO{sub 2} concentrations, or carbonates under natural exposure) from the pore solution, with little change in the binder gel identifiable by nuclearmore » magnetic resonance spectroscopy. In activated fly ash/slag blends, two distinct gels (C–A–S–H and N–A–S–H) are formed; under accelerated carbonation, the N–A–S–H gel behaves comparably to fly ash-based systems, while the C–A–S–H gel is decalcified similarly to alkali-activated slag. This provides new scope for durability optimisation, and for developing appropriate testing methodologies. -- Highlights: •C-A-S-H gel in alkali-activated slag decalcifies during accelerated carbonation. •Alkali-activated fly ash gel changes much less under CO{sub 2} exposure. •Blended slag-fly ash binder contains two coexisting gel types. •These two gels respond differently to carbonation. •Understanding of carbonation mechanisms is essential in developing test methods.« less

Modeling the effects of pH and ionic strength on swelling of anionic polyelectrolyte gels

NASA Astrophysics Data System (ADS)

Drozdov, A. D.; deClaville Christiansen, J.

2015-07-01

A constitutive model is developed for the elastic response of an anionic polyelectrolyte gel under swelling in water with an arbitrary pH and an arbitrary molar fraction of dissolved monovalent salt. A gel is treated as a three-phase medium consisting of a solid phase (polymer network), solvent (water), and solute (mobile ions). Transport of solvent and solute is thought of as their diffusion through the polymer network accelerated by an electric field formed by mobile and fixed ions and accompanied by chemical reactions (dissociation of functional groups attached to polymer chains and formation of ion pairs between bound charges and mobile counter-ions). Constitutive equations are derived by means of the free energy imbalance inequality for an arbitrary three-dimensional deformation with finite strains. These relations are applied to analyze equilibrium swelling diagrams on poly(acrylic acid) gel, poly(methacrylic acid) gel, and three composite hydrogels under water uptake in a bath (i) with a fixed molar fraction of salt and varied pH, and (ii) with a fixed pH and varied molar fraction of salt. To validate the ability of the model to predict observations quantitatively, material constants are found by matching swelling curves under one type of experimental conditions and results of simulation are compared with experimental data in the other type of tests.

Dynamic swelling of tunable full-color block copolymer photonic gels via counterion exchange.

PubMed

Lim, Ho Sun; Lee, Jae-Hwang; Walish, Joseph J; Thomas, Edwin L

2012-10-23

One-dimensionally periodic block copolymer photonic lamellar gels with full-color tunability as a result of a direct exchange of counteranions were fabricated via a two-step procedure comprising the self-assembly of a hydrophobic block-hydrophilic polyelectrolyte block copolymer, polystyrene-b-poly(2-vinyl pyridine) (PS-b-P2VP), followed by sequential quaternization of the P2VP layers in 1-bromoethane solution. Depending on the hydration characteristics of each counteranion, the selective swelling of the block copolymer lamellar structures leads to large tunability of the photonic stop band from blue to red wavelengths. More extensive quaternization of the P2VP block allows the photonic lamellar gels to swell more and red shift to longer wavelength. Here, we investigate the dynamic swelling behavior in the photonic gel films through time-resolved in situ measurement of UV-vis transmission. We model the swelling behavior using the transfer matrix method based on the experimentally observed reflectivity data with substitution of appropriate counterions. These tunable structural color materials may be attractive for numerous applications such as high-contrast displays without using a backlight, color filters, and optical mirrors for flexible lasing.

A clinical evaluation comparing two H2O2 concentrations used with a light-assisted chairside tooth whitening system.

PubMed

Ward, Marilyn; Felix, Heather

2012-04-01

The purpose of this study was to assess the efficacy of two different BriteSmile hydrogen peroxide (H2O2) gels in a split-arch protocol for whitening teeth in a clinical setting when used in conjunction with a BriteSmile BS4000 lamp. Fifteen subjects were enrolled into a single-center clinical trial. The efficacy of the BriteSmile BS4000 lamp using both 15% H2O2 and 25% H2O2 gel formulations was tested. Study subjects were concurrently exposed to the whitening lamp with the 15% H2O2 gel placed on half of their anterior teeth and the 25% H2O2 gel on the other half for a total light and gel exposure of 60 minutes. The clinical data collected were shade score, gingival health, and dentinal hypersensitivity self-assessment. Changes in tooth shade were better for subjects exposed to the 25% gel and the dental whitening lamp (average 8.0 shade changes) compared to subjects exposed to the 15% gel and dental whitening lamp (average 7.6 shade changes) immediately after treatment. The same held true at the 7-day follow-up (25% gel average 7.4 shade changes versus 15% gel average 7.3 shade changes). However, these differences were not statistically significant. No reports of irritation of gingival soft tissues were documented. The relative changes in mean sensitivity scores were similar for both groups with no significant differences in mean sensitivity scores between the groups. Both concentrations of H2O2 gel and the whitening lamp combined gave study subjects an average of 8.0 (25% gel) and 7.6 (15% gel) shade changes immediately after treatment. The 7-day follow-up examination resulted in a regression of lightest to an average of 7.4 (25% gel) and 7.3 (15% gel). It was concluded that the use of the chairside whitening light and either 15% or 25% hydrogen peroxide gel is safe and effective for whitening teeth in 1 hour.

Effect of nano-sized, elemental selenium supplement on the proteome of chicken liver.

PubMed

Gulyas, G; Csosz, E; Prokisch, J; Javor, A; Mezes, M; Erdelyi, M; Balogh, K; Janaky, T; Szabo, Z; Simon, A; Czegledi, L

2017-06-01

The nano-sized (100-500 nm) selenium has higher bioavailability and relatively lower toxicity compared to other selenium forms. The objective of the present study was to compare liver proteome profiles of broiler chicken fed with control diet without Se supplementation and diet supplemented with nano-Se with 4.25 mg/kg DM. Differential proteome analyses were performed by two-dimensional gel electrophoresis (2D-PAGE) followed by tryptic digestion and protein identification by liquid chromatography-mass spectrometry (LC-MS). Seven hundred and eight spots were detected, and 18 protein spots showed significant difference in their intensity (p < 0.05) between the two groups. In response to nano-Se supplementation, the expression of 8 proteins was higher, and 5 proteins were lower in nano-Se supplemented group compared to control group. The functions of the differentially expressed proteins indicate that the high dose of selenium supplementation induced a dietary stress. Selenium supplementation may influence the metabolism of fatty acids and carbohydrates and antioxidant system, and increase the quantity of cytoskeletal actin and the expression of actin regulatory protein as well. Journal of Animal Physiology and Animal Nutrition © 2016 Blackwell Verlag GmbH.

On-line comprehensive two-dimensional normal-phase liquid chromatography × reversed-phase liquid chromatography for preparative isolation of Peucedanum praeruptorum.

PubMed

Wang, Xin-Yuan; Li, Jia-Fu; Jian, Ya-Mei; Wu, Zhen; Fang, Mei-Juan; Qiu, Ying-Kun

2015-03-27

A new on-line comprehensive preparative two-dimensional normal-phase liquid chromatography × reversed-phase liquid chromatography (2D NPLC × RPLC) system was developed for the separation of complicated natural products. It was based on the use of a silica gel packed medium-pressure column as the first dimension and an ODS preparative HPLC column as the second dimension. The two dimensions were connected with normal-phase (NP) and reversed-phase (RP) enrichment units, involving a newly developed airflow assisted adsorption (AAA) technique. The instrument operation and the performance of this NPLC × RPLC separation method were illustrated by gram-scale isolation of ethanol extract from the roots of Peucedanum praeruptorum. In total, 19 compounds with high purity were obtained via automated multi-step preparative separation in a short period of time using this system, and their structures were comprehensively characterized by ESI-MS, (1)H NMR, and (13)C NMR. Including two new compounds, five isomers in two groups with identical HPLC and TLC retention values were also obtained and identified by 1D NMR and 2D NMR. This is the first report of an NPLC × RPLC system successfully applied in an on-line preparative process. This system not only solved the interfacing problem of mobile-phase immiscibility caused by NP and RP separation, it also exhibited apparent advantages in separation efficiency and sample treatment capacity compared with conventional methods. Copyright © 2015 Elsevier B.V. All rights reserved.

Production of Composite Scaffold Containing Silk Fibroin, Chitosan, and Gelatin for 3D Cell Culture and Bone Tissue Regeneration.

PubMed

Li, Jianqing; Wang, Qiuke; Gu, Yebo; Zhu, Yu; Chen, Liang; Chen, Yunfeng

2017-11-08

BACKGROUND Bone tissue engineering, a powerful tool to treat bone defects, is highly dependent on use of scaffolds. Both silk fibroin (SF) and chitosan (Cs) are biocompatible and actively studied for reconstruction of tissue engineering. Gelatin (Gel) is also widely applied in the biomedical field due to its low antigenicity and physicochemical stability. MATERIAL AND METHODS In this study, 4 different types of scaffolds were constructed - SF, SF/Cs, SF/Gel, and SF/Cs/Gel - and we compared their physical and chemical properties as well as biological characterization of these scaffolds to determine the most suitable scaffold for use in bone regeneration. First, these scaffolds were produced via chemical cross-linking method and freeze-drying technique. Next, the characterization of internal structure was studied using scanning electron microscopy and the porosity was evaluated by liquid displacement method. Then, we compared physicochemical properties such as water absorption rate and degradation property. Finally, MC3T3-E1 cells were inoculated on the scaffolds to study the biocompatibility and osteogenesis of the three-dimensional (3D) scaffolds in vitro. RESULTS The composite scaffold formed by all 3 components was the best for use in bone regeneration. CONCLUSIONS We conclude that the best scaffold among the 4 studied for MC3T3-E1 cells is our SF/Cs/Gel scaffold, suggesting a new choice for bone regeneration that can be used to treat bone defects or fractures in clinical practice.

All-trans-retinoic acid inhibition of transforming growth factor-β-induced collagen gel contraction mediated by human Tenon fibroblasts: role of matrix metalloproteinases.

PubMed

Liu, Yang; Kimura, Kazuhiro; Orita, Tomoko; Teranishi, Shinichiro; Suzuki, Katsuyoshi; Sonoda, Koh-Hei

2015-04-01

Scarring and contraction of the conjunctiva are common complications of many ocular diseases. We investigated the effects of all-trans-retinoic acid (ATRA) on the contractility of human Tenon's capsule fibroblasts (HTFs) cultured in a three-dimensional collagen gel. HTFs were cultured in a three-dimensional gel of type I collagen and in the absence or presence of transforming growth factor (TGF)-β, ATRA, or an inhibitor of matrix metalloproteinases (MMPs). Collagen gel contraction was evaluated by measurement of gel diameter. The release of MMPs and tissue inhibitors of metalloproteinases (TIMPs) into culture supernatants was assessed by immunoblot analysis and gelatin zymography. The release of lactate dehydrogenase activity from HTFs was measured with a colorimetric assay kit. ATRA inhibited TGF-β-induced collagen gel contraction mediated by HTFs in a concentration- and time-dependent manner. TGF-β induced the release of MMP-1, MMP-2 and MMP-3 by HTFs, and ATRA inhibited these effects of TGF-β on MMP-1 and MMP-3 release. ATRA also stimulated TIMP-1 release from HTFs in the presence of TGF-β. Furthermore, TGF-β-induced collagen gel contraction was blocked by the MMP inhibitor GM6001. ATRA did not exhibit cytotoxicity for HTFs. ATRA inhibited TGF-β-induced collagen gel contraction mediated by HTFs, likely in part by attenuating the production of MMP-1 and MMP-3 and by stimulating the production of TIMP-1. ATRA may therefore prove to be of clinical value for inhibition of scar formation in the conjunctiva. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

A Study of Dip-Coatable, High-Capacitance Ion Gel Dielectrics for 3D EWOD Device Fabrication

PubMed Central

Clement, Carlos E.; Jiang, Dongyue; Thio, Si Kuan; Park, Sung-Yong

2017-01-01

We present a dip-coatable, high-capacitance ion gel dielectric for scalable fabrication of three-dimensional (3D) electrowetting-on-dielectric (EWOD) devices such as an n × n liquid prism array. Due to the formation of a nanometer-thick electric double layer (EDL) capacitor, an ion gel dielectric offers two to three orders higher specific capacitance (c ≈ 10 μF/cm2) than that of conventional dielectrics such as SiO2. However, the previous spin-coating method used for gel layer deposition poses several issues for 3D EWOD device fabrication, particularly when assembling multiple modules. Not only does the spin-coating process require multiple repetitions per module, but the ion gel layer also comes in risks of damage or contamination due to handling errors caused during assembly. In addition, it was observed that the chemical formulation previously used for the spin-coating method causes the surface defects on the dip-coated gel layers and thus leads to poor EWOD performance. In this paper, we alternatively propose a dip-coating method with modified gel solutions to obtain defect-free, functional ion gel layers without the issues arising from the spin-coating method for 3D device fabrication. A dip-coating approach offers a single-step coating solution with the benefits of simplicity, scalability, and high throughput for deposition of high-capacitance gel layers on non-planar EWOD devices. An ion gel solution was prepared by combining the [EMIM][TFSI] ionic liquid and the [P(VDF-HFP)] copolymer at various wt % ratios in acetone solvent. Experimental studies were conducted to fully understand the effects of chemical composition ratios in the gel solution and how varying thicknesses of ion gel and Teflon layers affects EWOD performance. The effectiveness and potentiality of dip-coatable gel layers for 3D EWOD devices have been demonstrated through fabricating 5 × 1 arrayed liquid prisms using a single-step dip-coating method. Each prism module has been individually controlled to achieve spatial beam steering without the need for bulky mechanical moving parts. PMID:28772400

A Study of Dip-Coatable, High-Capacitance Ion Gel Dielectrics for 3D EWOD Device Fabrication.

PubMed

Clement, Carlos E; Jiang, Dongyue; Thio, Si Kuan; Park, Sung-Yong

2017-01-05

We present a dip-coatable, high-capacitance ion gel dielectric for scalable fabrication of three-dimensional (3D) electrowetting-on-dielectric (EWOD) devices such as an n × n liquid prism array. Due to the formation of a nanometer-thick electric double layer (EDL) capacitor, an ion gel dielectric offers two to three orders higher specific capacitance ( c ≈ 10 μF/cm²) than that of conventional dielectrics such as SiO₂. However, the previous spin-coating method used for gel layer deposition poses several issues for 3D EWOD device fabrication, particularly when assembling multiple modules. Not only does the spin-coating process require multiple repetitions per module, but the ion gel layer also comes in risks of damage or contamination due to handling errors caused during assembly. In addition, it was observed that the chemical formulation previously used for the spin-coating method causes the surface defects on the dip-coated gel layers and thus leads to poor EWOD performance. In this paper, we alternatively propose a dip-coating method with modified gel solutions to obtain defect-free, functional ion gel layers without the issues arising from the spin-coating method for 3D device fabrication. A dip-coating approach offers a single-step coating solution with the benefits of simplicity, scalability, and high throughput for deposition of high-capacitance gel layers on non-planar EWOD devices. An ion gel solution was prepared by combining the [EMIM][TFSI] ionic liquid and the [P(VDF-HFP)] copolymer at various wt % ratios in acetone solvent. Experimental studies were conducted to fully understand the effects of chemical composition ratios in the gel solution and how varying thicknesses of ion gel and Teflon layers affects EWOD performance. The effectiveness and potentiality of dip-coatable gel layers for 3D EWOD devices have been demonstrated through fabricating 5 × 1 arrayed liquid prisms using a single-step dip-coating method. Each prism module has been individually controlled to achieve spatial beam steering without the need for bulky mechanical moving parts.

Estimation of the influence of radical effect in the proton beams using a combined approach with physical data and gel data

NASA Astrophysics Data System (ADS)

Haneda, K.

2016-04-01

The purpose of this study was to estimate an impact on radical effect in the proton beams using a combined approach with physical data and gel data. The study used two dosimeters: ionization chambers and polymer gel dosimeters. Polymer gel dosimeters have specific advantages when compared to other dosimeters. They can measure chemical reaction and they are at the same time a phantom that can map in three dimensions continuously and easily. First, a depth-dose curve for a 210 MeV proton beam measured using an ionization chamber and a gel dosimeter. Second, the spatial distribution of the physical dose was calculated by Monte Carlo code system PHITS: To verify of the accuracy of Monte Carlo calculation, and the calculation results were compared with experimental data of the ionization chamber. Last, to evaluate of the rate of the radical effect against the physical dose. The simulation results were compared with the measured depth-dose distribution and showed good agreement. The spatial distribution of a gel dose with threshold LET value of proton beam was calculated by the same simulation code. Then, the relative distribution of the radical effect was calculated from the physical dose and gel dose. The relative distribution of the radical effect was calculated at each depth as the quotient of relative dose obtained using physical and gel dose. The agreement between the relative distributions of the gel dosimeter and Radical effect was good at the proton beams.

Defect Detection on Carbon Fibre Reinforced Plastics (cfrp) with Laser Generated Lamb Waves

NASA Astrophysics Data System (ADS)

Focke, O.; Huke, P.; Hildebrandt, A.

2011-06-01

Standard ultrasound methods using a phased-array or a single transducer are commonly used for non-destructive evaluation (NDE) during manufacturing of carbon fiber reinforced plastics (CFRP) parts and certificated testing schemes were developed for individual parts and geometries. However, most testing methods need direct contact, matching gels and remain therefore time consuming. Laser-Ultrasonics is advantageous due to the contactless measurement technology and high accessibility even on complex parts. Despite the non-destructive testing with body waves, we show that the NDE can be expanded using two-dimensional surface (Lamb) waves for detection of delaminations close to the surface or small deteriorations caused by e.g. impacts. Lamb waves have been excited with a single transducer and with a short-pulse Laser with additionally producing A0-and S0-Lamb waves. The waves were detected with a shearography setup that allows for measuring two-dimensionally the displacement of a surface. Short integration times of the camera were realized using a pulsed ruby laser for illumination. As a consequence to the anisotropy the propagation in different directions exhibits individual characteristics like amplitude, damping and velocity. This has motivated to build up models for the propagation of Lamb waves and to compare them with experimental results.

Two-dimensional chromatographic analysis using three second-dimension columns for continuous comprehensive analysis of intact proteins.

PubMed

Zhu, Zaifang; Chen, Huang; Ren, Jiangtao; Lu, Juan J; Gu, Congying; Lynch, Kyle B; Wu, Si; Wang, Zhe; Cao, Chengxi; Liu, Shaorong

2018-03-01

We develop a new two-dimensional (2D) high performance liquid chromatography (HPLC) approach for intact protein analysis. Development of 2D HPLC has a bottleneck problem - limited second-dimension (second-D) separation speed. We solve this problem by incorporating multiple second-D columns to allow several second-D separations to be proceeded in parallel. To demonstrate the feasibility of using this approach for comprehensive protein analysis, we select ion-exchange chromatography as the first-dimension and reverse-phase chromatography as the second-D. We incorporate three second-D columns in an innovative way so that three reverse-phase separations can be performed simultaneously. We test this system for separating both standard proteins and E. coli lysates and achieve baseline resolutions for eleven standard proteins and obtain more than 500 peaks for E. coli lysates. This is an indication that the sample complexities are greatly reduced. We see less than 10 bands when each fraction of the second-D effluents are analyzed by sodium dodecyl sulfate - polyacrylamide gel electrophoresis (SDS-PAGE), compared to hundreds of SDS-PAGE bands as the original sample is analyzed. This approach could potentially be an excellent and general tool for protein analysis. Copyright © 2017 Elsevier B.V. All rights reserved.

Material Properties of Silicon Carbide Fibers with Continuously Applied Sol-Gel Alumina Coatings

DTIC Science & Technology

1990-12-01

71 Coating Characterization ...................... 73 iii Two-Dimensional Plane Strain Analysis .................. 78 VI ...Axial Load in the Coating of Fiber Serie T ...... .82 vi List of Figures (continued) Figure Page 39. Tangential Stress Due to Axial Load in the Coating...residual stress will be presented 17 Fiur Vi o a CaFber EfIVfef Sic=’/. Figure 1. Sectional View of a Coated Fiber first, since these stresses are of

Decreased expression of the stress protein HSP70 is an early event in murine erythroleukemic cell differentiation.

PubMed Central

Hensold, J O; Housman, D E

1988-01-01

Two-dimensional protein gels were used to systematically assess changes in gene expression in Friend erythroleukemia cells after exposure to inducers of differentiation. A rapid decrease in expression of the stress protein HSP70 was observed after exposure to inducers. The kinetics of this change suggest that it may be related to the cellular events that regulate the onset of differentiation. Images PMID:3164440

Measurement of consumer preference for treatments used to induce labour: a willingness-to-pay approach.

PubMed

Taylor, Susan J.; Armour, Carol L.

2000-09-01

AIM: The purpose of the study was to assess the acceptability to consumers of two methods of induction of labour using a willingness-to-pay (WTP) approach. The methods compared were amniotomy plus oxytocin and prostaglandin E2 vaginal gel, followed by oxytocin if necessary. METHODS: A description of each method was presented, in questionnaire format, to pregnant women attending a public hospital ante-natal clinic. Women were asked to choose one of the two treatments, then give a valuation in dollar terms for both their preferred treatment and the alternative. RESULTS: It was found that 73.7% of patients preferred gel. The mean maximum WTP for amniotomy plus oxytocin was Aus$133 while that for gel was Aus$178 (P=0.0001). Those who chose amniotomy plus oxytocin were WTP 90% more for their preferred treatment compared with the alternative (Aus$180 vs. Aus$95). Similarly, those who preferred gel were WTP 90% more for their preferred treatment compared with the alternative (Aus$222 vs. Aus$119). CONCLUSION: Consumers were able to assess drug information provided on the two therapies, make an informed choice and to value that choice. Information obtained in this way, combined with information on costs, could be used in policy decision-making.

Measurement of consumer preference for treatments used to induce labour: a willingness‐to‐pay approach

PubMed Central

Taylor, Susan J.; Armour, Carol L.

2001-01-01

Aim The purpose of the study was to assess the acceptability to consumers of two methods of induction of labour using a willingness‐to‐pay (WTP) approach. The methods compared were amniotomy plus oxytocin and prostaglandin E2 vaginal gel, followed by oxytocin if necessary. Methods A description of each method was presented, in questionnaire format, to pregnant women attending a public hospital ante‐natal clinic. Women were asked to choose one of the two treatments, then give a valuation in dollar terms for both their preferred treatment and the alternative. Results It was found that 73.7% of patients preferred gel. The mean maximum WTP for amniotomy plus oxytocin was Aus$133 while that for gel was Aus$178 (P=0.0001). Those who chose amniotomy plus oxytocin were WTP 90% more for their preferred treatment compared with the alternative (Aus$180 vs. Aus$95). Similarly, those who preferred gel were WTP 90% more for their preferred treatment compared with the alternative (Aus$222 vs. Aus$119). Conclusion Consumers were able to assess drug information provided on the two therapies, make an informed choice and to value that choice. Information obtained in this way, combined with information on costs, could be used in policy decision‐making. PMID:11281930

A bead-spring chain as a one-dimensional polyelectrolyte gel.

PubMed

Manning, Gerald S

2018-05-23

The physical principles underlying expansion of a single-chain polyelectrolyte coil caused by Coulomb repulsions among its ionized groups, and the expansion of a cross-linked polyelectrolyte gel, are probably the same. In this paper, we analyze a "one-dimensional" version of a gel, namely, a linear chain of charged beads connected by Hooke's law springs. In the Debye-Hückel range of relatively weak Coulomb strength, where counterion condensation does not occur, the springs are realistically stretched on a nanolength scale by the repulsive interactions among the beads, if we use a spring constant normalized by the inverse square of the solvent Bjerrum length. The persistence length and radius of gyration counter-intuitively decrease when Coulomb strength is increased, if analyzed in the framework of an OSF-type theory; however, a buckling theory generates the increase that is consistent with bead-spring simulations.

TCA precipitation and ethanol/HCl single-step purification evaluation: One-dimensional gel electrophoresis, bradford assays, spectrofluorometry and Raman spectroscopy data on HSA, Rnase, lysozyme - Mascots and Skyline data.

PubMed

Eddhif, Balkis; Guignard, Nadia; Batonneau, Yann; Clarhaut, Jonathan; Papot, Sébastien; Geffroy-Rodier, Claude; Poinot, Pauline

2018-04-01

The data presented here are related to the research paper entitled "Study of a Novel Agent for TCA Precipitated Proteins Washing - Comprehensive Insights into the Role of Ethanol/HCl on Molten Globule State by Multi-Spectroscopic Analyses" (Eddhif et al., submitted for publication) [1]. The suitability of ethanol/HCl for the washing of TCA-precipitated proteins was first investigated on standard solution of HSA, cellulase, ribonuclease and lysozyme. Recoveries were assessed by one-dimensional gel electrophoresis, Bradford assays and UPLC-HRMS. The mechanistic that triggers protein conformational changes at each purification stage was then investigated by Raman spectroscopy and spectrofluorometry. Finally, the efficiency of the method was evaluated on three different complex samples (mouse liver, river biofilm, loamy soil surface). Proteins profiling was assessed by gel electrophoresis and by UPLC-HRMS.

Macroporous Hydrogel Scaffolds for Three-Dimensional Cell Culture and Tissue Engineering.

PubMed

Fan, Changjiang; Wang, Dong-An

2017-10-01

Hydrogels have been promising candidate scaffolds for cell delivery and tissue engineering due to their tissue-like physical properties and capability for homogeneous cell loading. However, the encapsulated cells are generally entrapped and constrained in the submicron- or nanosized gel networks, seriously limiting cell growth and tissue formation. Meanwhile, the spatially confined settlement inhibits attachment and spreading of anchorage-dependent cells, leading to their apoptosis. In recent years, macroporous hydrogels have attracted increasing attention in use as cell delivery vehicles and tissue engineering scaffolds. The introduction of macropores within gel scaffolds not only improves their permeability for better nutrient transport but also creates space/interface for cell adhesion, proliferation, and extracellular matrix deposition. Herein, we will first review the development of macroporous gel scaffolds and outline the impact of macropores on cell behaviors. In the first part, the advantages and challenges of hydrogels as three-dimensional (3D) cell culture scaffolds will be described. In the second part, the fabrication of various macroporous hydrogels will be presented. Third, the enhancement of cell activities within macroporous gel scaffolds will be discussed. Finally, several crucial factors that are envisaged to propel the improvement of macroporous gel scaffolds are proposed for 3D cell culture and tissue engineering.

Comparative Proteomic Analysis of Three Gelatinous Chinese Medicines and Their Authentications by Tryptic-digested Peptides Profiling using Matrix-assisted Laser Desorption/Ionization-time of Flight/Time of Flight Mass Spectrometry.

PubMed

Yang, Huan; Zheng, Jie; Wang, Hai-Yan; Li, Nan; Yang, Ya-Ya; Shen, Yu-Ping

2017-01-01

Gelatinous Chinese medicines (GCMs) including Asini Corii Colla, Testudinis Carapacis ET Plastri Colla, and Cervi Cornus Colla, were made from reptile shell or mammalian skin or deer horn, and consumed as a popular tonic, as well as hemopoietic and hemostatic agents. Misuse of them would not exert their functions, and fake or adulterate products have caused drug market disorder and affected food and drug safety. GCMs are rich in denatured proteins, but insufficient in available DNA fragments, hence commonly used cytochrome c oxidase I barcoding was not successful for their authentication. In this study, we performed comparative proteomic analysis of them and their animal origins to identify the composition of intrinsic proteins for the first time. A reliable and convenient approach was proposed for their authentication, by the incorporation of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, two-dimensional electrophoresis, and matrix-assisted laser desorption/ionization-time of flight/time of flight mass spectrometry (MALDI-TOF/TOF-MS). A total of 26 proteins were identified from medicinal parts of original animals, and GCMs proteins presented in a dispersive manner in electrophoresis analyses due to complicated changes in the structure of original proteins caused by long-term decoction and the addition of ingredients during their manufacturing. In addition, by comparison of MALDI-TOF/TOF-MS profiling, 19 signature peptide fragments originated from the protein of GCM products were selected according to criteria. These could assist in the discrimination and identification of adulterates of GCMs and other ACMs for their form of raw medicinal material, the pulverized, and even the complex. Comparative proteomic analysis of three gelatinous Chinese medicines was conducted, and their authentications were based on tryptic-digested peptides profiling using matrix-assisted laser desorption/ionization-time of flight/time of flight mass spectrometry. Abbreviations used: GCMs: Gelatinous Chinese medicines, COI: Cytochrome c oxidase I, SDS-PAGE: Sodium dodecyl sulfate polyacrylamide gel electrophoresis, 2-DE: Two-dimensional electrophoresis, MALDI-TOF/TOF-MS: Matrix-assisted laser desorption/ionization-time of flight/time of flight mass spectrometry, LC: Liquid chromatography, ChP: Chinese Pharmacopoeia, HPLC: High performance liquid chromatography, LC-ESI + -MS: Liquid chromatography-electro spray ionization-mass spectrometry, IEF: isoelectric focusing, HCCA: α-Cyano-4-hydroxycinnamic acid.

Dynamic analysis of a needle insertion for soft materials: Arbitrary Lagrangian-Eulerian-based three-dimensional finite element analysis.

PubMed

Yamaguchi, Satoshi; Tsutsui, Kihei; Satake, Koji; Morikawa, Shigehiro; Shirai, Yoshiaki; Tanaka, Hiromi T

2014-10-01

Our goal was to develop a three-dimensional finite element model that enables dynamic analysis of needle insertion for soft materials. To demonstrate large deformation and fracture, we used the arbitrary Lagrangian-Eulerian (ALE) method for fluid analysis. We performed ALE-based finite element analysis for 3% agar gel and three types of copper needle with bevel tips. To evaluate simulation results, we compared the needle deflection and insertion force with corresponding experimental results acquired with a uniaxial manipulator. We studied the shear stress distribution of agar gel on various time scales. For 30°, 45°, and 60°, differences in deflections of each needle between both sets of results were 2.424, 2.981, and 3.737mm, respectively. For the insertion force, there was no significant difference for mismatching area error (p<0.05) between simulation and experimental results. Our results have the potential to be a stepping stone to develop pre-operative surgical planning to estimate an optimal needle insertion path for MR image-guided microwave coagulation therapy and for analyzing large deformation and fracture in biological tissues. Copyright © 2014 Elsevier Ltd. All rights reserved.

Why do different criteria for 'cure' yield different conclusions in comparing two treatments for bacterial vaginosis?

PubMed

Thomas, Katherine K; Sanchez, Sixto; Garcia, Patricia J; Holmes, King K

2005-09-01

The objective of this study was to determine why different criteria for response to treatment of bacterial vaginosis (BV) led to markedly different conclusions about treatment efficacy in a randomized trial comparing metronidazole gel versus metronidazole/nystatin ovules. We compared the impact of two treatment regimens on individual components of Amsel and Nugent criteria at follow-up visits 14, 42, and 104 days after initiating treatment. Compared with gel, ovules more effectively eliminated amines, clue cells, and Gardnerella, Prevotella, or Mobiluncus morphotypes from vaginal fluid, thus achieving cure based on "usual" criteria (absence of BV by Amsel or Nugent criteria), but did not more effectively restore Lactobacillus morphotypes or lower vaginal pH, thus not meeting Federal Drug Administration (FDA) criteria for cure. Because early vaginal recolonization by lactobacilli was poor after both gel and ovules, FDA draft criteria for cure missed marked differences in treatment efficacies against Gardnerella, clue cells, and amines. Cure defined more "usually" may give more useful information.

A placebo controlled clinical trial investigating the efficacy of a homeopathic after-bite gel in reducing mosquito bite induced erythema.

PubMed

Hill, N; Stam, C; Tuinder, S; van Haselen, R A

1995-01-01

A randomised, placebo controlled clinical trial was conducted to examine the efficacy of a homeopathic after-bite gel in the symptomatic relief of mosquito bites. Sixty eight healthy volunteers were bitten under laboratory conditions by Aedes aegypti mosquitoes at three spots, on the ventral aspect of the forearm. One bite was treated with the homeopathic after-bite gel, another bite with a placebo gel which was identical in appearance and smell to the homeopathic after-bite gel, and the third bite remained untreated. Immediately after the bites and 1, 3, 6, 26 and 31 hours post-bite, the length and width of the erythema were measured with a calliper, and photographs were taken of the bite sites from which the size of the erythema was subsequently determined. This was followed by assessment of the extent of itching with a verbal analogue scale, and finally treatment took place. For each spot the total erythema was calculated as the area under the plotted curve of the erythema at different time points (mm2*h) and the total sum of the itch scores was determined. For the bites treated with the homeopathic after-bite gel the median total erythema was 10.500 mm2*h. For the spots treated with the placebo gel and the untreated spots the median total erythema was 12.900 mm2*h and 13.300 mm2*h, respectively. The difference between the spots treated with the homeopathic after-bite gel and the untreated spots came close to significance (two-tailed P = 0.06), which was not the case for the difference between the spots treated with the homeopathic after-bite gel and the spots treated with placebo gel (P = 0.13). After pooling the data of a very similar previous pilot study and the present study (ntotal = 83), the homeopathic after-bite gel was significantly superior to no treatment (two-tailed P = 0.003) as well as to placebo gel (two-tailed P = 0.03). Comparing itching after the three treatments, no significant differences could be demonstrated. The extent of itching was positively correlated with the area of the erythema (r = 0.63). Treatment of mosquito bites with the homeopathic after-bite gel will reduce the erythema compared to no treatment. Comparison with the placebo gel suggests it is the plant extracts which are the active components of this gel.

Integration of gel-based proteome data with pProRep.

PubMed

Laukens, Kris; Matthiesen, Rune; Lemière, Filip; Esmans, Eddy; Onckelen, Harry Van; Jensen, Ole Nørregaard; Witters, Erwin

2006-11-15

pProRep is a web application integrating electrophoretic and mass spectral data from proteome analyses into a relational database. The graphical web-interface allows users to upload, analyse and share experimental proteome data. It offers researchers the possibility to query all previously analysed datasets and can visualize selected features, such as the presence of a certain set of ions in a peptide mass spectrum, on the level of the two-dimensional gel. The pProRep package and instructions for its use can be downloaded from http://www.ptools.ua.ac.be/pProRep. The application requires a web server that runs PHP 5 (http://www.php.net) and MySQL. Some (non-essential) extensions need additional freely available libraries: details are described in the installation instructions.

[Polymer Gel Dosimeter].

PubMed

Hayashi, Shin-Ichiro

2017-01-01

With rapid advances being made in radiotherapy treatment, three-dimensional (3D) dose measurement techniques of great precision are required more than ever before. It is expected that 3D polymer gel dosimeters will satisfy clinical needs for an effective detector that can measure the complex 3D dose distributions. Polymer gel dosimeters are devices that utilize the radiation-induced polymerization reactions of vinyl monomers in a gel to store information about radiation dose. The 3D absorbed dose distribution can be deduced from the resulting polymer distribution using several imaging modalities, such as MRI, X-ray and optical CTs. In this article, the fundamental characteristics of polymer gel dosimeter are reviewed and some challenging keys are also suggested for the widely spread in clinical use.

Phosphorylation of hepatocyte growth factor receptor and epidermal growth factor receptor of human hepatocytes can be maintained in a (3D) collagen sandwich culture system.

PubMed

Engl, Tobias; Boost, Kim A; Leckel, Kerstin; Beecken, Wolf-Dietrich; Jonas, Dietger; Oppermann, Elsie; Auth, Marcus K H; Schaudt, André; Bechstein, Wolf-Otto; Blaheta, Roman A

2004-08-01

In vitro culture models that employ human liver cells could be potent tools for predictive studies on drug toxicity and metabolism in the pharmaceutical industry. However, an adequate receptor responsiveness is necessary to allow intracellular signalling and metabolic activity. We tested the ability of three-dimensionally arranged human hepatocytes to respond to the growth factors hepatocyte growth factor (HGF) or epidermal growth factor (EGF). Isolated adult human hepatocytes were cultivated within a three-dimensional collagen gel (sandwich) or on a two-dimensional collagen matrix. Cells were treated with HGF or EGF and expression and phosphorylative activity of HGF receptors (HGFr, c-met) or EGF receptors (EGFr) were measured by flow cytometry and Western blot. Increasing HGFr and EGFr levels were detected in hepatocytes growing two-dimensionally. However, both receptors were not activated in presence of growth factors. In contrast, when hepatocytes were plated within a three-dimensional matrix, HGFr and EGFr levels remained constantly low. However, both receptors became strongly phosphorylated by soluble HGF or EGF. We conclude that cultivation of human hepatocytes in a three-dimensionally arranged in vitro system allows the maintenance of specific functional activities. The necessity of cell dimensionality for HGFr and EGFr function should be considered when an adequate in vitro system has to be introduced for drug testing.

Proteome analysis to assess physiological changes in Escherichia coli grown under glucose-limited fed-batch conditions.

PubMed

Raman, Babu; Nandakumar, M P; Muthuvijayan, Vignesh; Marten, Mark R

2005-11-05

Proteome analysis was used to compare global protein expression changes in Escherichia coli fermentation between exponential and glucose-limited fed-batch phase. Two-dimensional gel electrophoresis and MALDI-TOF mass spectrometry were used to separate and identify 49 proteins showing >2-fold difference in expression. Proteins upregulated during exponential phase include ribonucleotide biosynthesis enzymes and ribosomal recycling factor. Proteins upregulated during fed-batch phase include those involved in high-affinity glucose uptake, transport and degradation of alternate carbon sources and TCA cycle, suggesting an enhanced role of the cycle under glucose- and energy-limited conditions. We report the upregulation of several putative proteins (ytfQ, ygiS, ynaF, yggX, yfeX), not identified in any previous study under carbon-limited conditions. Copyright (c) 2005 Wiley Periodicals, Inc.

Characterization of the Tumor Secretome from Tumor Interstitial Fluid (TIF).

PubMed

Gromov, Pavel; Gromova, Irina

2016-01-01

Tumor interstitial fluid (TIF) surrounds and perfuses bodily tumorigenic tissues and cells, and can accumulate by-products of tumors and stromal cells in a relatively local space. Interstitial fluid offers several important advantages for biomarker and therapeutic target discovery, especially for cancer. Here, we describe the most currently accepted method for recovering TIF from tumor and nonmalignant tissues that was initially performed using breast cancer tissue. TIF recovery is achieved by passive extraction of fluid from small, surgically dissected tissue specimens in phosphate-buffered saline. We also present protocols for hematoxylin and eosin (H&E) staining of snap-frozen and formalin-fixed, paraffin-embedded (FFPE) tumor sections and for proteomic profiling of TIF and matched tumor samples by high-resolution two-dimensional gel electrophoresis (2D-PAGE) to enable comparative analysis of tumor secretome and paired tumor tissue.

[Plasma proteomic analysis in children with infectious mononucleosis].

PubMed

Ran, Zhi-Ling; Xiao, Bin; Liu, Hong-Rui; Liu, You-Ping; Sheng, Qiao-Ni

2015-03-01

To explore the abnormal expression of plasma proteins by analysis of proteomic expression profile in children with infectious mononucleosis (IM). Two dimensional gel electrophoresis (2-DE) followed by the mass spectrometry was used to examine important protein spots with different expression levels between children with IM and normal controls. Seven differential proteins were obtained: hemopexin, vitamin D binding protein, fetuin A, C-reactive protein, apolipoprotein A, haptoglobin and transthyretin. Compared with the control group, haptoglobin showed a higher expression level in children with IM, and the expression levels of the other proteins were obviously down-regulated. The expression changes of differential proteins identified in this study are all related with the liver acute injury, suggesting that children with IM are associated with acute liver injury. Further studies on the characteristics of above proteins will contribute to the diagnosis and treatment of pediatric IM.

Nose-to-brain transport of melatonin from polymer gel suspensions: a microdialysis study in rats.

PubMed

Jayachandra Babu, R; Dayal, Pankaj Patrick; Pawar, Kasturi; Singh, Mandip

2011-11-01

Exogenous melatonin (MT) has significant neuroprotective roles in Alzheimer's and Parkinson's diseases. This study investigates the delivery MT to brain via nasal route as a polymeric gel suspension using central brain microdialysis in anesthetized rats. Micronized MT suspensions using polymers [carbopol, carboxymethyl cellulose (CMC)] and polyethylene glycol 400 (PEG400) were prepared and characterized for nasal administration. In vitro permeation of the formulations was measured across a three-dimensional tissue culture model EpiAirway(™). The central brain delivery into olfactory bulb of nasally administered MT gel suspensions was studied using brain microdialysis in male Wistar rats. The MT content of microdialysis samples was analyzed by high performance liquid chromatography (HPLC) using electrochemical detection. The nose-to-brain delivery of MT formulations was compared with intravenously administered MT solution. MT suspensions in carbopol and CMC vehicles have shown significantly higher permeability across Epiairway(™) as compared to control, PEG400 (P < 0.05). The brain (olfactory bulb) levels of MT after intranasal administration were 9.22, 6.77 and 4.04-fold higher for carbopol, CMC and PEG400, respectively, than that of intravenous MT in rats. In conclusion, microdialysis studies demonstrated increased brain levels of MT via nasal administration in rats.

Nose-to-brain transport of melatonin from polymer gel suspensions: a microdialysis study in rats

PubMed Central

Babu, R. Jayachandra; Dayal, Pankaj Patrick; Pawar, Kasturi; Singh, Mandip

2012-01-01

Purpose Exogenous melatonin (MT) has significant neuroprotective roles in Alzheimer’s and Parkinson’s diseases. This study investigates the delivery MT to brain via nasal route as a polymeric gel suspension using central brain microdialysis in anesthetized rats. Methods Micronized MT suspensions using polymers [carbopol, carboxymethyl cellulose (CMC)] and polyethylene glycol 400 (PEG400) were prepared and characterized for nasal administration. In vitro permeation of the formulations was measured across a three-dimensional tissue culture model EpiAirway™. The central brain delivery into olfactory bulb of nasally administered MT gel suspensions was studied using brain microdialysis in male Wistar rats. The MT content of microdialysis samples was analyzed by high performance liquid chromatography (HPLC) using electrochemical detection. The nose-to-brain delivery of MT formulations was compared with intravenously administered MT solution. Results MT suspensions in carbopol and CMC vehicles have shown significantly higher permeability across Epiairway™ as compared to control, PEG400 (P < 0.05). The brain (olfactory bulb) levels of MT after intranasal administration were 9.22, 6.77 and 4.04-fold higher for carbopol, CMC and PEG400, respectively, than that of intravenous MT in rats. In conclusion, microdialysis studies demonstrated increased brain levels of MT via nasal administration in rats. PMID:21428693

Sol-Gel Manufactured Energetic Materials

DOEpatents

Simpson, Randall L.; Lee, Ronald S.; Tillotson, Thomas M.; Hrubesh, Lawrence W.; Swansiger, Rosalind W.; Fox, Glenn A.

2005-05-17

Sol-gel chemistry is used for the preparation of energetic materials (explosives, propellants and pyrotechnics) with improved homogeneity, and/or which can be cast to near-net shape, and/or made into precision molding powders. The sol-gel method is a synthetic chemical process where reactive monomers are mixed into a solution, polymerization occurs leading to a highly cross-linked three dimensional solid network resulting in a gel. The energetic materials can be incorporated during the formation of the solution or during the gel stage of the process. The composition, pore, and primary particle sizes, gel time, surface areas, and density may be tailored and controlled by the solution chemistry. The gel is then dried using supercritical extraction to produce a highly porous low density aerogel or by controlled slow evaporation to produce a xerogel. Applying stress during the extraction phase can result in high density materials. Thus, the sol-gel method can be used for precision detonator explosive manufacturing as well as producing precision explosives, propellants, and pyrotechnics, along with high power composite energetic materials.

Sol-gel manufactured energetic materials

DOEpatents

Simpson, Randall L.; Lee, Ronald S.; Tillotson, Thomas M.; Hrubesh, Lawrence W.; Swansiger, Rosalind W.; Fox, Glenn A.

2003-12-23

Sol-gel chemistry is used for the preparation of energetic materials (explosives, propellants and pyrotechnics) with improved homogeneity, and/or which can be cast to near-net shape, and/or made into precision molding powders. The sol-gel method is a synthetic chemical process where reactive monomers are mixed into a solution, polymerization occurs leading to a highly cross-linked three dimensional solid network resulting in a gel. The energetic materials can be incorporated during the formation of the solution or during the gel stage of the process. The composition, pore, and primary particle sizes, gel time, surface areas, and density may be tailored and controlled by the solution chemistry. The gel is then dried using supercritical extraction to produce a highly porous low density aerogel or by controlled slow evaporation to produce a xerogel. Applying stress during the extraction phase can result in high density materials. Thus, the sol-gel method can be used for precision detonator explosive manufacturing as well as producing precision explosives, propellants, and pyrotechnics, along with high power composite energetic materials.

A Phase 1 Trial to Assess the Safety, Acceptability, Pharmacokinetics, and Pharmacodynamics of a Novel Dapivirine Vaginal Film.

PubMed

Bunge, Katherine E; Dezzutti, Charlene S; Rohan, Lisa C; Hendrix, Craig W; Marzinke, Mark A; Richardson-Harman, Nicola; Moncla, Bernard J; Devlin, Brid; Meyn, Leslie A; Spiegel, Hans M L; Hillier, Sharon L

2016-04-15

Films may deliver antiretroviral drugs efficiently to mucosal tissues. In this first in-human trial of a vaginal film for delivering the nonnucleoside reverse transcriptase inhibitor dapivirine, safety, pharmacokinetics, and pharmacodynamics of film and gel formulations were compared with placebo. Sixty-one healthy HIV-negative women were randomized to daily dapivirine (0.05%) or placebo gel, or dapivirine (1.25 mg) or placebo film for seven days. The proportion of participants experiencing grade 2 and higher adverse events related to study product were compared. Plasma dapivirine concentrations were quantified. Paired cervical and vaginal tissue biopsies obtained ∼2 hours after the last dose were measured for tissue drug concentration and exposed to HIV in an ex vivo challenge assay. Two grade 2 related adverse events occurred in the placebo film group. Women randomized to gel and film products had 4 log10 higher of dapivirine in cervical and vaginal tissues than plasma. Although gel and film users had comparable plasma dapivirine concentrations, tissue concentrations of dapivirine were 3-5 times higher in the gel users when compared with film users. HIV replication in the ex vivo challenge assay was significantly reduced in vaginal tissues from women randomized to dapivirine film or gel; furthermore, tissue drug concentrations were highly correlated with HIV protection. Women rated the film more comfortable with less leakage but found it more difficult to insert than gel. Both film and gel delivered dapivirine at concentrations sufficient to block HIV ex vivo. This proof-of-concept study suggests film formulations for microbicides merit further investigation.

Effect of acidity of in-office bleaching gels on tooth sensitivity and whitening: a two-center double-blind randomized clinical trial.

PubMed

Loguercio, A D; Servat, F; Stanislawczuk, R; Mena-Serrano, A; Rezende, M; Prieto, M V; Cereño, V; Rojas, M F; Ortega, K; Fernandez, E; Reis, A

2017-12-01

The study aimed to compare the tooth sensitivity (TS) and bleaching efficacy of two hydrogen peroxide gels with different pHs (acid pH [Pola Office, SDI] and the neutral pH [Pola Office+, SDI]) used for in-office bleaching. Fifty-four patients from Brazil and Chile, with right superior incisor darker than A2, were selected for this double-blind, split-mouth randomized trial. Teeth were bleached in two sessions, with 1-week interval. Each session had three applications of 8 min each, according to the manufacturer's instructions. The color changes were evaluated by subjective (Vita Classical and Vita Bleachedguide) and objective (Easy shade spectrophotometer) methods. Participants recorded TS with 0-10 visual analog scale. Color change in shade guide units (SGU) and ΔE was analyzed by Student's t test (α = 0.05). The absolute risk and intensity of TS were evaluated by McNemar's test and Wilcoxon-paired test, respectively (α = 0.05). All groups achieved the same level of whitening after 30 days of clinical evaluation. The use of a neutral in-office bleaching gel significantly decreases the absolute risk of TS (28%, 95% CI 18-41) and intensity of TS when compared to the acid bleaching gel (absolute risk of 50%, 95% CI 37-63). The use of a neutral in-office bleaching agent gel produced the same whitening degree than an acid bleaching gel but with reduced risk and intensity of tooth sensitivity. Clinicians should opt to use in-office bleaching with a neutral gel than an acid product because the former causes a significant lower risk and intensity of tooth sensitivity.

Modeling the Dynamics of Gel Electrophorresis in the High School Classroom

ERIC Educational Resources Information Center

Saucedo, Skyler R.

2013-01-01

Gel electrophoresis, used by geneticists and forensic experts alike, is an immensely popular technique that utilizes an electric field to separate molecules and proteins by size and charge. At the microscopic level, a dye or complex protein like DNA is passed through agarose, a gelatinous three-dimensional matrix of pores and nano-sized tunnels.…

From a 2DE-gel spot to protein function: lesson learned from HS1 in chronic lymphocytic leukemia.

PubMed

Apollonio, Benedetta; Bertilaccio, Maria Teresa Sabrina; Restuccia, Umberto; Ranghetti, Pamela; Barbaglio, Federica; Ghia, Paolo; Caligaris-Cappio, Federico; Scielzo, Cristina

2014-10-19

The identification of molecules involved in tumor initiation and progression is fundamental for understanding disease's biology and, as a consequence, for the clinical management of patients. In the present work we will describe an optimized proteomic approach for the identification of molecules involved in the progression of Chronic Lymphocytic Leukemia (CLL). In detail, leukemic cell lysates are resolved by 2-dimensional Electrophoresis (2DE) and visualized as "spots" on the 2DE gels. Comparative analysis of proteomic maps allows the identification of differentially expressed proteins (in terms of abundance and post-translational modifications) that are picked, isolated and identified by Mass Spectrometry (MS). The biological function of the identified candidates can be tested by different assays (i.e. migration, adhesion and F-actin polymerization), that we have optimized for primary leukemic cells.

Treatment with captopril abrogates the altered expression of alpha1 macroglobulin and alpha1 antiproteinase in sera of spontaneously hypertensive rats

PubMed Central

2012-01-01

Background Proteins that are associated with hypertension may be identified by comparing the 2-dimensional gel electrophoresis (2-DE) profiles of the sera of spontaneously hypertensive rats (SHR) with those generated from normotensive Spraque-Dawley rats (SDR). Results Five proteins of high abundance were found to be significantly altered when the 2-DE serum profiles of the SHR were compared to those that were similarly generated from the SDR. Analysis by mass spectrometry and database search identified the proteins as retinol binding protein 4, complement C3, albumin (19.9 kDa fragment), alpha1 macroglobulin and alpha1 antiproteinase, which are all known to be associated with hypertension. The altered expression of the two latter proteins was found to be abrogated when similar analysis was performed on sera of the SHR that were treated with captopril. Conclusion Our data suggests that serum alpha1 macroglobulin and alpha1 antiproteinase are potentially useful complementary biomolecular indicators for monitoring of hypertension. PMID:22416803

Assessing Amide Proton Transfer (APT) MRI Contrast Origins in 9 L Gliosarcoma in the Rat Brain Using Proteomic Analysis.

PubMed

Yan, Kun; Fu, Zongming; Yang, Chen; Zhang, Kai; Jiang, Shanshan; Lee, Dong-Hoon; Heo, Hye-Young; Zhang, Yi; Cole, Robert N; Van Eyk, Jennifer E; Zhou, Jinyuan

2015-08-01

To investigate the biochemical origin of the amide photon transfer (APT)-weighted hyperintensity in brain tumors. Seven 9 L gliosarcoma-bearing rats were imaged at 4.7 T. Tumor and normal brain tissue samples of equal volumes were prepared with a coronal rat brain matrix and a tissue biopsy punch. The total tissue protein and the cytosolic subproteome were extracted from both samples. Protein samples were analyzed using two-dimensional gel electrophoresis, and the proteins with significant abundance changes were identified by mass spectrometry. There was a significant increase in the cytosolic protein concentration in the tumor, compared to normal brain regions, but the total protein concentrations were comparable. The protein profiles of the tumor and normal brain tissue differed significantly. Six cytosolic proteins, four endoplasmic reticulum proteins, and five secreted proteins were considerably upregulated in the tumor. Our experiments confirmed an increase in the cytosolic protein concentration in tumors and identified several key proteins that may cause APT-weighted hyperintensity.

Analysis of electrophoresis performance

NASA Technical Reports Server (NTRS)

Roberts, G. O.

1984-01-01

The SAMPLE computer code models electrophoresis separation in a wide range of conditions. Results are included for steady three dimensional continuous flow electrophoresis (CFE), time dependent gel and acetate film experiments in one or two dimensions and isoelectric focusing in one dimension. The code evolves N two dimensional radical concentration distributions in time, or distance down a CFE chamber. For each time or distance increment, there are six stages, successively obtaining the pH distribution, the corresponding degrees of ionization for each radical, the conductivity, the electric field and current distribution, and the flux components in each direction for each separate radical. The final stage is to update the radical concentrations. The model formulation for ion motion in an electric field ignores activity effects, and is valid only for low concentrations; for larger concentrations the conductivity is, therefore, also invalid.

Phase diagram of a reentrant gel of patchy particles

NASA Astrophysics Data System (ADS)

Roldán-Vargas, Sándalo; Smallenburg, Frank; Kob, Walter; Sciortino, Francesco

2013-12-01

We study the phase diagram of a binary mixture of patchy particles which has been designed to form a reversible gel. For this we perform Monte Carlo and molecular dynamics simulations to investigate the thermodynamics of such a system and compare our numerical results with predictions based on the analytical parameter-free Wertheim theory. We explore a wide range of the temperature-density-composition space that defines the three-dimensional phase diagram of the system. As a result, we delimit the region of thermodynamic stability of the fluid. We find that for a large region of the phase diagram the Wertheim theory is able to give a quantitative description of the system. For higher densities, our simulations show that the system is crystallizing into a BCC structure. Finally, we study the relaxation dynamics of the system by means of the density and temperature dependences of the diffusion coefficient. We show that there exists a density range where the system passes reversibly from a gel to a fluid upon both heating and cooling, encountering neither demixing nor phase separation.

Cell-Responsive Hydrogel for Encapsulation of Vascular Cells

PubMed Central

Kraehenbuehl, Thomas P.; Ferreira, Lino S.; Zammaretti, Prisca; Hubbell, Jeffrey A.; Langer, Robert

2014-01-01

The in vitro potential of a synthetic matrix metalloproteinase (MMP)-responsive polyethylene glycol) (PEG)-based hydrogel as a bioactive co-encapsulation system for vascular cells and a small bioactive peptide, thymosin β4 (Tp4), was examined. We show that the physical incorporation of Tβ4 in this bioactive matrix creates a three-dimensional (3D) environment conducive for human umbilical vein endothelial cell (HUVEC) adhesion, survival, migration and organization. Gels with entrapped Tβ4 increased the survival of HUVEC compared to gels without Tp4, and significantly up-regulated the endothelial genes vascular endothelial-cadherin and angiopoietin-2, whereas von Willebrand factor was significantly down-regulated. Incorporation of Tβ4 significantly increased MMP-2 and MMP-9 secretion of encapsulated HUVEC. The gel acts as a controlled Tβ4-release system, as MMP-2 and MMP-9 enzymes trigger the release. In addition, Tβ4 facilitated HUVEC attachment and induced vascular-like network formation upon the PEG-hydrogels. These MMP-responsive PEG-hydrogels may thus serve as controlled co-encapsulation system of vascular cells and bioactive factors for in situ regeneration of ischemic tissues. PMID:19500842

Lamellar biogels comprising fluid membranes with a newly synthesized class of polyethylene glycol-surfactants

NASA Astrophysics Data System (ADS)

Warriner, Heidi E.; Davidson, Patrick; Slack, Nelle L.; Schellhorn, Matthias; Eiselt, Petra; Idziak, Stefan H. J.; Schmidt, Hans-Werner; Safinya, Cyrus R.

1997-09-01

A series of four polymer-surfactant macromolecules, each consisting of a double-chain hydrophobic moiety attached onto a monofunctional polyethylene glycol (PEG) polymer chain, were synthesized in order to study their effect upon the fluid lamellar liquid crystalline (Lα) phase of the dimyristoylphosphatidylcholine/pentanol/water system. The main finding of this study is that the addition of these compounds induces a new lamellar gel, called Lα,g. We have determined the phase diagrams as a function of PEG-surfactant concentration, cPEG, and weight fraction water, ΦW. All phase diagrams are qualitatively similar and show the existence of the gel. Unlike more common polymer physical gels, this gel can be induced either by increasing cPEG or by adding water at constant cPEG. In particular, less polymer is required for gelation as water concentration increases. Moreover, the gel phase is attained at concentrations of PEG-surfactant far below that required for classical polymer gels and is stable at temperatures comparable to the lower critical solution temperature of free PEG-water mixtures. Small angle x-ray experiments demonstrate the lamellar structure of the gel phase, while wide angle x-ray scattering experiments prove that the structure is Lα, not Lβ' (a common chain-ordered phase which is also a gel). The rheological behavior of the Lα,g phase demonstrates the existence of three dimensional elastic properties. Polarized light microscopy of Lα,g samples reveals that the Lα,g is induced by a proliferation of defect structures, including whispy lines, spherulitic defects, and a nematiclike Schlieren texture. We propose a model of topological defects created by the aggregation of PEG-surfactant into highly curved regions within the membranes. This model accounts for both the inverse relationship between ΦW and cPEG observed along the gel transition line and the scaling dependence of the interlayer spacing at the gel transition with the PEG molecular weight. These Lα hydrogels could serve as the matrix for membrane-anchored peptides, proteins or other drug molecules, creating a "bioactive gel" with mechanical stability deriving from the polymer-lipid minority component.

Is instillation of anesthetic gel necessary in flexible cystoscopic examination? A prospective randomized study.

PubMed

Kobayashi, Takashi; Nishizawa, Koji; Ogura, Keiji

2003-01-01

To determine whether urethral injection of anesthetic and lubricating agent before outpatient flexible cystoscopic examination is worthwhile regarding patient tolerance of pain. A randomized prospective study was conducted. A total of 133 consecutive men scheduled to undergo flexible cystoscopy were randomized to receive 11 mL of 0.2% oxybuprocaine hydrochloride gel (group 1), 11 mL of plain lubricating gel (group 2), or no gel injection (group 3). In every group, 2% lidocaine gel was applied to the fiberscope. Patients recorded the level of pain during gel instillation, scope insertion, and intravesical observation separately on a 100-mm visual analog self-assessment scale. Pain scores for gel instillation were approximately two thirds those for scope insertion and intravesical observation in groups 1 and 2. No significant difference was noted in the pain score of each group during either scope insertion or intravesical observation. Pain during intraurethral gel instillation is significant. Anesthetic gel instillation has no advantage compared with no-gel injection in men when lubricating gel is applied to a flexible fiberscope.

SU-G-BRB-17: Dosimetric Evaluation of the Respiratory Interplay Effect During VMAT Delivery Using IPAGAT Polymer Gel Dosimeter

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ono, K; Fujimoto, S; Akagi, Y

Purpose: To evaluate the dosimetric impact of the interplay effect between multileaf collimator (MLC) movement and tumor respiratory motion during delivery of volumetric modulate arc therapy (VMAT) by using customized polymer gel dosimeter. Methods: Polyacrylamide-based gel dosimeter contained magnesium chloride as a sensitizer (iPAGAT) was used in this study. An excellent gas barrier PAN (BAREX) techno bottle (φ8 cm, 650 mL) filled with iPAGAT was set to the QUASAR™ respiratory motion phantom, and was moved with motion amplitudes of 1 and 2 cm with a 4 second period during VMAT delivery by the Novalis Tx linear accelerator (Varian/BrainLAB). Two sphericalmore » tumors with a 2 cm diameter (GTV1 and GTV2) were defined, and ITV1 (GTV1+1 cm) and ITV2 (GTV2+2 cm) with expansion in the superior-inferior (S-I) direction were also defined with simulated respiratory motion. PTV margin was 2 mm around the ITV considering the setup uncertainty. Two single arc VMAT plans with 30 Gy at 3 Gy per fraction (GTV: D98>100%, PTV: D95=100%) were generated by the Varian Eclipse treatment planning system. Three-dimensional dose distribution in iPAGAT was read out by the Signa 1.5T MRI system (GE), and was evaluated by dose-volume histogram (DVH) using in-house developed software. Results: According to DVH analysis by iPAGAT, D98 of GTV1 and GTV2 were more than 100% of the prescribed dose. In contrast, D95 of PTV1 and PTV2 were about 85% and 65%, respectively. Furthermore, low-to-intermediate dose was widespread with motion amplitude of 2 cm. Conclusion: DVH analysis using iPAGAT polymer gel dosimeter was performed in this study. As a result, interplay effect was negligible, since dose coverage of GTV was sufficient during VMAT delivery with simulated respiratory motion. However, the dose reduction of PTV and the spread of low-to-intermediate dose compared to the planned dose require scrupulous attention for large tumor respiratory motion.« less

Cacao Phylloplane: The First Battlefield against Moniliophthora perniciosa, Which Causes Witches' Broom Disease.

PubMed

Almeida, D S M; Gramacho, K P; Cardoso, T H S; Micheli, F; Alvim, F C; Pirovani, C P

2017-07-01

The phylloplane is the first contact surface between Theobroma cacao and the fungus Moniliophthora perniciosa, which causes witches' broom disease (WBD). We evaluated the index of short glandular trichomes (SGT) in the cacao phylloplane and the effect of irrigation on the disease index of cacao genotypes with or without resistance to WBD, and identified proteins present in the phylloplane. The resistant genotype CCN51 and susceptible Catongo presented a mean index of 1,600 and 700 SGT cm -2 , respectively. The disease index in plants under drip irrigation was reduced by approximately 30% compared with plants under sprinkler irrigation prior to inoculation. Leaf water wash (LWW) of the cacao inhibited the germination of spores by up to 98%. Proteins from the LWW of CCN51 were analyzed by two-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis followed by tandem mass spectrometry. The gel showed 71 spots and identified a total of 42 proteins (28 from the plant and 14 from bacteria). Proteins related to defense and synthesis of defense metabolites and involved in nucleic acid metabolism were identified. The results support the hypothesis that the proteins and water-soluble compounds secreted to the cacao phylloplane participate in the defense against pathogens. They also suggest that SGT can contribute to the resistance of cacao.

Laser Direct Writing Process for Making Electrodes and High-k Sol-Gel ZrO2 for Boosting Performances of MoS2 Transistors.

PubMed

Kwon, Hyuk-Jun; Jang, Jaewon; Grigoropoulos, Costas P

2016-04-13

A series of two-dimensional (2D) transition metal dichalcogenides (TMDCs), including molybdenum disulfide (MoS2), can be attractive materials for photonic and electronic applications due to their exceptional properties. Among these unique properties, high mobility of 2D TMDCs enables realization of high-performance nanoelectronics based on a thin film transistor (TFT) platform. In this contribution, we report highly enhanced field effect mobility (μ(eff) = 50.1 cm(2)/(V s), ∼2.5 times) of MoS2 TFTs through the sol-gel processed high-k ZrO2 (∼22.0) insulator, compared to those of typical MoS2/SiO2/Si structures (μ(eff) = 19.4 cm(2)/(V s)) because a high-k dielectric layer can suppress Coulomb electron scattering and reduce interface trap concentration. Additionally, in order to avoid costly conventional mask based photolithography and define the patterns, we employ a simple laser direct writing (LDW) process. This process allows precise and flexible control with reasonable resolution (up to ∼10 nm), depending on the system, and enables fabrication of arbitrarily patterned devices. Taking advantage of continuing developments in laser technology offers a substantial cost decrease, and LDW may emerge as a promising technology.

Calcium alginate gel as encapsulation matrix for coimmobilized enzyme systems.

PubMed

Blandino, A; Macías, M; Cantero, D

2003-07-01

Encapsulation within calcium alginate gel capsules was used to produce a coimmobilized enzyme system. Glucose oxidase (GOD) and catalase (CAT) were chosen as model enzymes. The same values of Vmax and Km app for the GOD encapsulated system and for the GOD-CAT coencapsulated system were calculated. When gel beads and capsules were compared, the same catalyst deactivation sequence for the two enzymes was observed. However, when capsules were employed as immobilization support, GOD efficiencies were higher than for the gel beads. These results were explained in terms of the structure of the capsules.

CHIP mediates down-regulation of nucleobindin-1 in preosteoblast cell line models.

PubMed

Xue, Fuying; Wu, Yanping; Zhao, Xinghui; Zhao, Taoran; Meng, Ying; Zhao, Zhanzhong; Guo, Junwei; Chen, Wei

2016-08-01

Nucleobindin-1 (NUCB1), also known as Calnuc, is a highly conserved, multifunctional protein widely expressed in tissues and cells. It contains two EF-hand motifs which have been shown to play a crucial role in binding Ca(2+) ions. In this study, we applied comparative two-dimensional gel electrophoresis to characterize differentially expressed proteins in HA-CHIP over-expressed and endogenous CHIP depleted MC3T3-E1 stable cell lines, identifying NUCB1 as a novel CHIP/Stub1 targeted protein. NUCB1 interacts with and is down-regulated by CHIP by both proteasomal dependent and independent pathways, suggesting that CHIP-mediated down-regulation of nucleobindin-1 might play a role in osteoblast differentiation. The chaperone protein Hsp70 was found to be important for CHIP and NUCB1 interaction as well as CHIP-mediated NUCB1 down-regulation. Our findings provide new insights into understanding the stability regulation of NUCB1. Copyright © 2016 Elsevier Inc. All rights reserved.

Electronic method for autofluorography of macromolecules on two-D matrices

DOEpatents

Davidson, Jackson B.; Case, Arthur L.

1983-01-01

A method for detecting, localizing, and quantifying macromolecules contained in a two-dimensional matrix is provided which employs a television-based position sensitive detection system. A molecule-containing matrix may be produced by conventional means to produce spots of light at the molecule locations which are detected by the television system. The matrix, such as a gel matrix, is exposed to an electronic camera system including an image-intensifier and secondary electron conduction camera capable of light integrating times of many minutes. A light image stored in the form of a charge image on the camera tube target is scanned by conventional television techniques, digitized, and stored in a digital memory. Intensity of any point on the image may be determined from the number at the memory address of the point. The entire image may be displayed on a television monitor for inspection and photographing or individual spots may be analyzed through selected readout of the memory locations. Compared to conventional film exposure methods, the exposure time may be reduced 100-1000 times.

Differential protein expression in ovaries of uninfected and Babesia-infected southern cattle ticks, Rhipicephalus (Boophilus) microplus.

PubMed

Rachinsky, Anna; Guerrero, Felix D; Scoles, Glen A

2007-12-01

We used gel electrophoresis and mass spectrometry to investigate differences in protein expression in ovarian tissues from Babesia bovis-infected and uninfected southern cattle tick, Rhipicephalus (Boophilus) microplus. Soluble and membrane proteins were extracted from ovaries of adult female ticks, and analyzed by isoelectric focusing (IEF) and one-dimensional or two-dimensional (2-D) gel electrophoresis. Protein patterns were analyzed for differences in expression between infected and uninfected ticks. 2-D separation of proteins revealed a number of proteins that appeared to be up- or down-regulated in response to infection with Babesia, in particular membrane/membrane-associated proteins and proteins in a low molecular mass range between 6 and 36kDa. A selection of differentially expressed proteins was subjected to analysis by capillary-HPLC-electrospray tandem mass spectrometry (HPLC-ESI-MS/MS). Among the ovarian proteins that were up-regulated in infected ticks were calreticulin, two myosin subunits, an endoplasmic reticulum protein, a peptidyl-prolyl cis-trans isomerase (PPIase), a cytochrome c oxidase subunit, a glutamine synthetase, and a family of Kunitz-type serine protease inhibitors. Among the down-regulated ovarian proteins were another PPIase, a hemoglobin subunit, and a lysozyme. This study is part of an ongoing effort to establish a proteome database that can be utilized to investigate specific proteins involved in successful pathogen transmission.

Growth hormone-releasing hormone stimulates and somatostatin inhibits the release of a novel protein by cultured rat pituitary cells.

PubMed

Tachibana, K; Marquardt, H; Yokoya, S; Friesen, H G

1988-10-01

We have reported that the secretion of at least 17 distinct peptides [including rat (rGH)] GH by cultured rat pituitary cells was stimulated by GH-releasing hormone and inhibited by somatostatin, when analyzed by two-dimensional polyacrylamide gel electrophoresis. Three of these peptides (no. 23, 24, and 25) were not rGH immunoreactive. In order to determine whether these three peptides are fragments, degradation products or posttranscriptionally modified forms of rGH, rGH and peptide no. 23 were characterized structurally. From partial peptide maps of rGH and peptide no. 23 by V8 protease or chymotrypsin, it appeared that these peptides were not related to each other. By N-terminal microsequencing of two-dimensional polyacrylamide gel electrophoresis purified peptide, we have obtained the sequence of 24 N-terminal amino acid residues of peptide no. 23. This sequence has no significant homology with rGH or any other reported protein sequence. Antiserum was generated against a synthetic oligopeptide corresponding to amino acid residues 3-24 of peptide no. 23. The antiserum cross-reacted with peptides no. 23, 24, and 25 upon Western blot analysis. These results indicate that peptide no. 23 has a novel structure unrelated to other pituitary hormones. Since its secretion is influenced by GH-releasing hormone and somatostatin, peptide no. 23 may represent a previously unrecognized structurally unique growth factor.

Isolation and Purification of Water Soluble Proteins from Ginger Root (Zingiber officinale) by Two Dimensional Liquid Chromatography

PubMed Central

Sandovall, A.O.; Andrews, K.; Wahab, A.; Choudhary, M.I.; Ahmed, A.

2014-01-01

The RI-INBRE Centralized Core Facility was established in 2003 and participates annually in Undergraduate Summer Research Program. It provides students hands on research experience in key technologies in biomedical sciences. We present here the isolation and purification of water soluble proteins from ginger, a rhizome of the plant, Zingiber officinale. It is an important ingredient of species used in traditional South Asian cuisines. In Indian, Pakistani and Chinese folk medicine, ginger is used for gastro-intestinal disorders, nausea, vomiting, inflammatory diseases, muscle and joint pain. Limited studies have been reported on the bioactive proteins from ginger extract. The water soluble proteins were extracted from ginger root and successfully purified to homogeneity by using two-dimensional liquid chromatography (FPLC/RP-HPLC) approach. The ginger root was washed with distilled water; skin removed and then emulsified using an electric blender. Sample was stirred for four days at 4°C with and without protease inhibitor. Purification of a 42kDa protein was achieved by employing gel filtration, ion-exchange and reversed phase HPLC. The homogeneity of the protein was confirmed by SDS-PAGE gel electrophoresis and MALDI-TOF mass spectrometry. Future work will be conducted on the protein characterization using mass spectrometry and Edman protein sequencing. Supported by grant 5P20GM103430 from the National Institute of General Medical Sciences, NIH, USA.

Analysis of the Aspergillus fumigatus proteome reveals metabolic changes and the activation of the pseurotin A biosynthesis gene cluster in response to hypoxia.

PubMed

Vödisch, Martin; Scherlach, Kirstin; Winkler, Robert; Hertweck, Christian; Braun, Hans-Peter; Roth, Martin; Haas, Hubertus; Werner, Ernst R; Brakhage, Axel A; Kniemeyer, Olaf

2011-05-06

The mold Aspergillus fumigatus is the most important airborne fungal pathogen. Adaptation to hypoxia represents an important virulence attribute for A. fumigatus. Therefore, we aimed at obtaining a comprehensive overview about this process on the proteome level. To ensure highly reproducible growth conditions, an oxygen-controlled, glucose-limited chemostat cultivation was established. Two-dimensional gel electrophoresis analysis of mycelial and mitochondrial proteins as well as two-dimensional Blue Native/SDS-gel separation of mitochondrial membrane proteins led to the identification of 117 proteins with an altered abundance under hypoxic in comparison to normoxic conditions. Hypoxia induced an increased activity of glycolysis, the TCA-cycle, respiration, and amino acid metabolism. Consistently, the cellular contents in heme, iron, copper, and zinc increased. Furthermore, hypoxia induced biosynthesis of the secondary metabolite pseurotin A as demonstrated at proteomic, transcriptional, and metabolite levels. The observed and so far not reported stimulation of the biosynthesis of a secondary metabolite by oxygen depletion may also affect the survival of A. fumigatus in hypoxic niches of the human host. Among the proteins so far not implicated in hypoxia adaptation, an NO-detoxifying flavohemoprotein was one of the most highly up-regulated proteins which indicates a link between hypoxia and the generation of nitrosative stress in A. fumigatus.

Pre-labeling of diverse protein samples with a fixed amount of Cy5 for sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis.

PubMed

Bjerneld, Erik J; Johansson, Johan D; Laurin, Ylva; Hagner-McWhirter, Åsa; Rönn, Ola; Karlsson, Robert

2015-09-01

A pre-labeling protocol based on Cy5 N-hydroxysuccinimide (NHS) ester labeling of proteins has been developed for one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis. We show that a fixed amount of sulfonated Cy5 can be used in the labeling reaction to label proteins over a broad concentration range-more than three orders of magnitude. The optimal amount of Cy5 was found to be 50 to 250pmol in 20μl using a Tris-HCl labeling buffer at pH 8.7. Labeling protein samples with a fixed amount of dye in this range balances the requirements of sub-nanogram detection sensitivity and low dye-to-protein (D/P) ratios for SDS-PAGE. Simulations of the labeling reaction reproduced experimental observations of both labeling kinetics and D/P ratios. Two-dimensional electrophoresis was used to examine the labeling of proteins in a cell lysate using both sulfonated and non-sulfonated Cy5. For both types of Cy5, we observed efficient labeling across a broad range of molecular weights and isoelectric points. Copyright © 2015 Elsevier Inc. All rights reserved.

Fabrication of 3D Reconstituted Organoid Arrays by DNA-programmed Assembly of Cells (DPAC)

PubMed Central

Todhunter, Michael E; Weber, Robert J; Farlow, Justin; Jee, Noel Y; Cerchiari, Alec E; Gartner, Zev J

2016-01-01

Tissues are the organizational units of function in metazoan organisms. Tissues comprise an assortment of cellular building blocks, soluble factors, and extracellular matrix (ECM) that are composed into specific three dimensional (3D) structures. The capacity to reconstitute tissues in vitro with the structural complexity observed in vivo is key to understanding processes such as morphogenesis, homeostasis, and disease. In this unit, we describe DNA-programmed Assembly of Cells (DPAC), a method to fabricate viable, functional arrays of organoid-like tissues within 3D ECM gels. In DPAC, dissociated cells are chemically functionalized with degradable oligonucleotide “velcro,” allowing rapid, specific, and reversible cell adhesion to a two-dimensional (2D) template patterned with complementary DNA. An iterative assembly process builds up organoids, layer-by-layer, from this initial 2D template and into the third dimension. Cleavage of the DNA releases the completed array of tissues that are captured and fully embedded in ECM gels for culture and observation. DPAC controls the size, shape, composition, and spatial heterogeneity of organoids, and permits positioning constituent cells with single-cell resolution even within cultures several centimeters long. PMID:27622567

Comparison between two forms of vaginally administered progesterone for luteal phase support in assisted reproduction cycles.

PubMed

Geber, Selmo; Moreira, Ana Carolina Ferreira; de Paula, Sálua Oliveira Calil; Sampaio, Marcos

2007-02-01

The use of progesterone for luteal phase support has been demonstrated to be beneficial in assisted reproduction cycles using gonadotrophin-releasing hormone analogues (GnRHa). Two micronized progesterone preparations are available for vaginal administration: capsules and gel. The objective of this study was to compare the efficacy of these two forms for luteal phase support in assisted reproduction cycles. A total of 244 couples undergoing IVF/intracytoplasmic sperm injection cycles were included in the study and were randomly allocated (sealed envelopes) into two groups: group 1 (122) received vaginal capsules of 200 mg of micronized progesterone (Utrogestan), 3 times daily, and group 2 (122) received micronized progesterone in gel (Crinone 8%), once daily. Both groups received progesterone for 13 days beginning day 1 after oocyte retrieval, continuing until the pregnancy test was performed and until 12 weeks of pregnancy. Groups were compared by clinical data and assisted reproduction results and had similar ages and causes of infertility. Although the pregnancy rate was higher for those receiving progesterone gel than capsules (44.26 and 36.06% respectively), this difference was not statistically significant. The study showed that vaginal progesterone gel and capsules used for luteal phase support in assisted reproduction cycles with long protocol GnRHa result in similar pregnancy rates.

In situ ultrasound imaging of silk hydrogel degradation and neovascularization.

PubMed

Leng, Xiaoping; Liu, Bin; Su, Bo; Liang, Min; Shi, Liangchen; Li, Shouqiang; Qu, Shaohui; Fu, Xin; Liu, Yue; Yao, Meng; Kaplan, David L; Wang, Yansong; Wang, Xiaoqin

2017-03-01

Ultrasound (US) is a useful technique to monitor morphological and functional changes of biomaterial implants without sacrificing the animal. Contrast-enhanced ultrasound (CEUS) along with two-dimensional (2D) US were used to characterize the biodegradation and neovascularization of silk protein (8 wt%) hydrogel implants in rats. Cylinder-shaped silk hydrogel plugs were implanted into the space between the hind limb thigh muscles in Wistar rats (n = 6). The increase of echogenicity in 2D US revealed tissue-ingrowth-accompanied gel degradation over 18 weeks. The shape and size of the implanted gels remained qualitatively unchanged until week 15, as confirmed by Bland and Altman analysis and visualization of retrieved samples. Using CEUS, neovascularization was monitored by the presence of microbubbles in the gel area, and the dynamic vascularization process was indicated by the contrast enhancement values, which showed a relatively low level (< 5 dB) during weeks 1-8 and significantly increased levels (around 20 dB at week 15 and > 35 dB at week 18), suggesting that major vascularization had occurred in the gel implants by this time point. Histological and scanning electron microscopic analysis of explants revealed time-dependent increases in the pore size of the gel matrix, the presence of endothelial and red blood cells and the number of blood vessels in the gel implants, indicating that degradation and vascularization did occur in silk gel implants during the time period. The present study demonstrates the use of US imaging for monitoring of in vivo degradation and vascularization of silk implants in a non-destructive way. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

Tubulin-isotype analysis of two grass species-resistant to dinitroaniline herbicides.

PubMed

Waldin, T R; Ellis, J R; Hussey, P J

1992-09-01

Trifluralin-resistant biotypes of Eleusine indica (L.) Gaertn. (goosegrass) and Setaria viridis (L.) Beauv. (green foxtail) exhibit cross-resistance to other dinitroaniline herbicides. Since microtubules are considered the primary target site for dinitroaniline herbicides we investigated whether the differential sensitivity of resistant and susceptible biotypes of these species results from modified tubulin polypeptides. One-dimensional and two-dimensional polyacrylamide gel electrophoresis combined with immunoblotting using well-characterised anti-tubulin monoclonal antibodies were used to display the family of tubulin isotypes in each species. Seedlings of E. indica exhibited four β-tubulin isotypes and one α-tubulin isotype, whereas those of S. viridis exhibited two β-tubulin and two α-tubulin isotypes. Comparison of the susceptible and resistant biotypes within each species revealed no differences in electrophoretic properties of the multiple tubulin isotypes. These results provide no evidence that resistance to dinitroaniline herbicides is associated with a modified tubulin polypeptide in these biotypes of E. indica or S. viridis.

Preliminary evaluation of a thermosensitive chitosan hydrogel for Echinococcus granulosus vaccine delivery.

PubMed

Umair, Saleh; Pernthaner, Anton; Deng, Qing; Gibson, Blake; Hook, Sarah; Heath, David

2017-03-15

The EG95 vaccine is effective in protecting grazing animals from infection with Echinococcus granulosus. Six male lambs were used in the study, two were each vaccinated subcutaneously with 50μg EG95/1mg Quil-A, two animals were each vaccinated with 50μg EG95/1mg Quil-A in 1% chitosan thermolabile gel subcutaneously, and two animals served as non-vaccinated controls. Two vaccinations were given at a 7 week interval. Two vaccinations induced a significantly higher antibody titre in the chitosan group compared with the Quil-A only group. The chitosan vaccine group also had a significantly higher antibody titre compared with a positive control sera from vaccinated and challenged sheep. Incorporating the EG95/Quil-A vaccine in a thermo-responsive chitosan sol-gel stimulated, after the second injection, a high level of antibody absorbance which remained high for at least one year. This response was significantly greater than the response to vaccine without the gel. Copyright © 2017 Elsevier B.V. All rights reserved.

Hybrid Hydroxyapatite Nanoparticle Colloidal Gels are Injectable Fillers for Bone Tissue Engineering

PubMed Central

Gu, Zhen; Jamal, Syed; Detamore, Michael S.

2013-01-01

Injectable bone fillers have emerged as an alternative to the invasive surgery often required to treat bone defects. Current bone fillers may benefit from improvements in dynamic properties such as shear thinning during injection and recovery of material stiffness after placement. Negatively charged inorganic hydroxyapatite (HAp) nanoparticles (NPs) were assembled with positively charged organic poly(d,l-lactic-co-glycolic acid) (PLGA) NPs to create a cohesive colloidal gel. This material is held together by electrostatic forces that may be disrupted by shear to facilitate extrusion, molding, or injection. Scanning electron micrographs of the dried colloidal gels showed a well-organized, three-dimensional porous structure. Rheology tests revealed that certain colloidal gels could recover after being sheared. Human umbilical cord mesenchymal stem cells were also highly viable when seeded on the colloidal gels. HAp/PLGA NP colloidal gels offer an attractive scheme for injectable filling and regeneration of bone tissue. PMID:23815275

Preliminary investigation of PAGAT polymer gel radionuclide dosimetry of Tc-99m

NASA Astrophysics Data System (ADS)

Braun, Kelly; Bailey, Dale; Hill, Brendan; Baldock, Clive

2009-05-01

PAGAT polymer gel was investigated as a suitable dosimeter materials for measuring absorbed dose from the unsealed source radionuclide Tc-99m. Differing amounts of Tc-99m over the range of 25-5000 MBq were introduced into a normoxic polymer gel mixture (PAGAT) in sealed nitrogen-filled P6 glass vials. After irradiation the gels were evaluated using MRI more than 48 hours after preparation to allow for radioactive decay. The dose delivered to the vial was also calculated empirically. R2 versus total activity curves were obtained over a number of experiments and these were used to evaluate the relationship between the amount of gel polymerization and the dose deposited by the radionuclide. A linear response up to 1000 MBq (corresponding to 20Gy) was displayed and was still behaving monotonically at 5000 MBq. Polymer gels offer the potential to measure radiation dose three-dimensionally using MRI.

A new approach combining different MRI methods to provide detailed view on swelling dynamics of xanthan tablets influencing drug release at different pH and ionic strength.

PubMed

Mikac, Ursa; Sepe, Ana; Kristl, Julijana; Baumgartner, Sasa

2010-08-03

The key element in drug release from hydrophilic matrix tablets is the gel layer that regulates the penetration of water and controls drug dissolution and diffusion. We have selected magnetic resonance imaging (MRI) as the method of choice for visualizing the dynamic processes occurring during the swelling of xanthan tablets in a variety of media. The aims were (i) to develop a new method using MRI for accurate determination of penetration, swelling and erosion fronts, (ii) to investigate the effects of pH and ionic strength on swelling, and (iii) to study the influence of structural changes in xanthan gel on drug release. Two dimensional (2D) MRI and one dimensional single point imaging (SPI) of swollen xanthan tablets were recorded, together with T(2) mapping. The border between dry and hydrated glassy xanthan-the penetration front-was determined from 1D SPI signal intensity profiles. The erosion front was obtained from signal intensity profiles of 2D MR images. The swelling front, where xanthan is transformed from a glassy to a rubbery state (gel formation), was determined from T(2) profiles. Further, the new combination of MRI methods for swelling front determination enables to explain the appearance of the unusual "bright front" observed on 2D MR images in tablets swollen in HCl pH 1.2 media, which represents the position of swelling front. All six media studied, differing in pH and ionic strength, penetrate through the whole tablet in 4h+/-0.3h, but formation of the gel layer is significantly delayed. Unexpectedly, the position of the swelling front was the same, independently of the different xanthan gel structures formed under different conditions of pH and ionic strength. The position of the erosion front, on the other hand, is strongly dependent on pH and ionic strength, as reflected in different thicknesses of the gel layers. The latter are seen to be the consequence of the different hydrodynamic radii of the xanthan molecules, which affect the drug release kinetics. The slowest release of pentoxifylline was observed in water where the thickest gel was formed, whereas the fastest release was observed in HCl pH 1.2, in which the gel layer was thinnest. Moreover, experiments simulating physiological conditions showed that changes of pH and ionic strength influence the xanthan gel structure relatively quickly, and consequently the drug release kinetics. It is therefore concluded that drug release is greatly influenced by changes in the xanthan molecular conformation, as reflected in changed thickness of the gel layer. A new method utilizing combination of SPI, multi-echo MRI and T(2) mapping eliminates the limitations of standard methods used in previous studies for determining moving fronts and improves current understanding of the dynamic processes involved in polymer swelling. Copyright (c) 2010 Elsevier B.V. All rights reserved.

Integration of gel-based and gel-free proteomic data for functional analysis of proteins through Soybean Proteome Database.

PubMed

Komatsu, Setsuko; Wang, Xin; Yin, Xiaojian; Nanjo, Yohei; Ohyanagi, Hajime; Sakata, Katsumi

2017-06-23

The Soybean Proteome Database (SPD) stores data on soybean proteins obtained with gel-based and gel-free proteomic techniques. The database was constructed to provide information on proteins for functional analyses. The majority of the data is focused on soybean (Glycine max 'Enrei'). The growth and yield of soybean are strongly affected by environmental stresses such as flooding. The database was originally constructed using data on soybean proteins separated by two-dimensional polyacrylamide gel electrophoresis, which is a gel-based proteomic technique. Since 2015, the database has been expanded to incorporate data obtained by label-free mass spectrometry-based quantitative proteomics, which is a gel-free proteomic technique. Here, the portions of the database consisting of gel-free proteomic data are described. The gel-free proteomic database contains 39,212 proteins identified in 63 sample sets, such as temporal and organ-specific samples of soybean plants grown under flooding stress or non-stressed conditions. In addition, data on organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored. Furthermore, the database integrates multiple omics data such as genomics, transcriptomics, metabolomics, and proteomics. The SPD database is accessible at http://proteome.dc.affrc.go.jp/Soybean/. The Soybean Proteome Database stores data obtained from both gel-based and gel-free proteomic techniques. The gel-free proteomic database comprises 39,212 proteins identified in 63 sample sets, such as different organs of soybean plants grown under flooding stress or non-stressed conditions in a time-dependent manner. In addition, organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored in the gel-free proteomics database. A total of 44,704 proteins, including 5490 proteins identified using a gel-based proteomic technique, are stored in the SPD. It accounts for approximately 80% of all predicted proteins from genome sequences, though there are over lapped proteins. Based on the demonstrated application of data stored in the database for functional analyses, it is suggested that these data will be useful for analyses of biological mechanisms in soybean. Furthermore, coupled with recent advances in information and communication technology, the usefulness of this database would increase in the analyses of biological mechanisms. Copyright © 2017 Elsevier B.V. All rights reserved.

Directing Induced Pluripotent Stem Cell Derived Neural Stem Cell Fate with a Three-Dimensional Biomimetic Hydrogel for Spinal Cord Injury Repair.

PubMed

Fan, Lei; Liu, Can; Chen, Xiuxing; Zou, Yan; Zhou, Zhengnan; Lin, Chenkai; Tan, Guoxin; Zhou, Lei; Ning, Chenyun; Wang, Qiyou

2018-05-30

Current treatment approaches for spinal cord injuries (SCIs) are mainly based on cellular transplantation. Induced pluripotent stem cells (iPSCs) without supply constraints and ethical concerns have emerged as a viable treatment option for repairing neurological disorders. However, the primarily limitations in the neuroregeneration field are uncontrolled cell differentiation, and low cell viability caused by the ischemic environment. The mechanical property of three-dimensional (3D) hydrogel can be easily controlled and shared similar characteristics with nerve tissue, thus promoting cell survival and controlled cell differentiation. We propose the combination of a 3D gelatin methacrylate (GelMA) hydrogel with iPSC-derived NSCs (iNSCs) to promote regeneration after SCI. In vitro, the iNSCs photoencapsulated in the 3D GelMA hydrogel survived and differentiated well, especially in lower-moduli hydrogels. More robust neurite outgrowth and more neuronal differentiation were detected in the soft hydrogel group. To further evaluate the in vivo neuronal regeneration effect of the GelMA hydrogels, a mouse spinal cord transection model was generated. We found that GelMA/iNSC implants significantly promoted functional recovery. Further histological analysis showed that the cavity areas were significantly reduced, and less collagen was deposited in the GelMA/iNSC group. Furthermore, the GelMA and iNSC combined transplantation decreased inflammation by reducing activated macrophages/microglia (CD68-positive cells). Additionally, GelMA/iNSC implantation showed striking therapeutic effects of inhibiting GFAP-positive cells and glial scar formation while simultaneously promoting axonal regeneration. Undoubtedly, use of this 3D hydrogel stem cell-loaded system is a promising therapeutic strategy for SCI repair.

The Application of a Three-Step Proteome Analysis for Identification of New Biomarkers of Pancreatic Cancer

PubMed Central

Abulaizi, Mayinuer; Tomonaga, Takeshi; Satoh, Mamoru; Sogawa, Kazuyuki; Matsushita, Kazuyuki; Kodera, Yoshio; Obul, Jurat; Takano, Shigetsugu; Yoshitomi, Hideyuki; Miyazaki, Masaru; Nomura, Fumio

2011-01-01

We searched for novel tumor markers of pancreatic cancer by three-step serum proteome analysis. Twelve serum abundant proteins were depleted using immunoaffinity columns followed by fractionation by reverse-phase high-performance liquid chromatography. Proteins in each fraction were separated by two-dimensional gel electrophoresis. Then the gel was stained by Coomassie Brilliant Blue. Protein spots in which the expression levels were significantly different between cancer and normal control were identified by LC-MS/MS. One hundred and two spots were upregulated, and 84 spots were downregulated in serum samples obtained from patients with pancreatic cancers, and 58 proteins were identified by mass spectrometry. These candidate proteins were validated using western blot analysis and enzyme-linked immunosorbent assay (ELISA). As a result of these validation process, we could confirm that the serum levels of apolipoprotein A-IV, vitamin D-binding protein, plasma retinol-binding protein 4, and tetranectin were significantly decreased in patients with pancreatic cancer. PMID:22091389

Accumulation of New Polypeptides in Ri T-DNA-Transformed Roots of Tomato (Lycopersicon esculentum) during the Development of Vesicular-Arbuscular Mycorrhizae.

PubMed

Simoneau, P; Louisy-Louis, N; Plenchette, C; Strullu, D G

1994-06-01

Root-inducing transferred-DNA (Ri T-DNA)-transformed roots of tomato (Lycopersicon esculentum) were in vitro inoculated with surface-sterilized vesicular-arbuscular mycorrhizal leek root pieces. About 1 week after inoculation, the infection of the transformed root culture by the fungal endophyte was confirmed by photonic microscopy. Total proteins were extracted from the mycorrhizal roots and analyzed by two-dimensional polyacrylamide gel electrophoresis. Control gels were run with proteins extracted from noninoculated roots mixed with purified intraradical vesicles and extraradical hyphae. Comparison of the resulting patterns revealed the presence of two polypeptides with estimated apparent masses of 24 and 39 kDa that were detected only in infected roots. Polypeptides with similar migration parameters were not detected in roots challenged with spore extracts, suggesting that the accumulation of the polypeptides was directly linked to root colonization by the fungus rather than to induction by fungus-derived elicitors.

Accumulation of New Polypeptides in Ri T-DNA-Transformed Roots of Tomato (Lycopersicon esculentum) during the Development of Vesicular-Arbuscular Mycorrhizae

PubMed Central

Simoneau, Philippe; Louisy-Louis, Nathalie; Plenchette, Christian; Strullu, Désiré Georges

1994-01-01

Root-inducing transferred-DNA (Ri T-DNA)-transformed roots of tomato (Lycopersicon esculentum) were in vitro inoculated with surface-sterilized vesicular-arbuscular mycorrhizal leek root pieces. About 1 week after inoculation, the infection of the transformed root culture by the fungal endophyte was confirmed by photonic microscopy. Total proteins were extracted from the mycorrhizal roots and analyzed by two-dimensional polyacrylamide gel electrophoresis. Control gels were run with proteins extracted from noninoculated roots mixed with purified intraradical vesicles and extraradical hyphae. Comparison of the resulting patterns revealed the presence of two polypeptides with estimated apparent masses of 24 and 39 kDa that were detected only in infected roots. Polypeptides with similar migration parameters were not detected in roots challenged with spore extracts, suggesting that the accumulation of the polypeptides was directly linked to root colonization by the fungus rather than to induction by fungus-derived elicitors. Images PMID:16349273

A Phase 1 trial to assess the safety, acceptability, pharmacokinetics and pharmacodynamics of a novel dapivirine vaginal film

PubMed Central

Bunge, Authors: Katherine E.; Dezzutti, Charlene S.; Rohan, Lisa C.; Hendrix, Craig W.; Marzinke, Mark A.; Richardson-Harman, Nicola; Moncla, Bernard J.; Devlin, Brid; Meyn, Leslie A.; M.L.Spiegel, Hans; Hillier, Sharon L.

2016-01-01

Background Films may deliver antiretroviral drugs efficiently to mucosal tissues. In this first in-human trial of a vaginal film for delivering the non-nucleoside reverse transcriptase inhibitor dapivirine, safety, pharmacokinetics, and pharmacodynamics of film and gel formulations were compared to placebo. Methods 61 healthy HIV negative women were randomized to daily dapivirine (0.05%) or placebo gel, or dapivirine (1.25mg) or placebo film for seven days. The proportion of participants experiencing Grade 2 and higher adverse events related to study product were compared. Plasma dapivirine concentrations were quantified. Paired cervical and vaginal tissue biopsies obtained ∼2 hours following the last dose were measured for tissue drug concentration and exposed to HIV in an ex vivo challenge assay. Results Two Grade 2 related AEs occurred in the placebo film group. Women randomized to gel and film products had 4 log10 higher of dapivirine in cervical and vaginal tissues than plasma. While gel and film users had comparable plasma dapivirine concentrations, tissue concentrations of dapivirine were 3 to 5 times higher in the gel users when compared to film users. HIV replication in the ex vivo challenge assay was significantly reduced in vaginal tissues from women randomized to dapivirine film or gel; furthermore, tissue drug concentrations were highly correlated with HIV protection. Women rated the film more comfortable with less leakage, but found it more difficult to insert than gel. Discussion Both film and gel delivered dapivirine at concentrations sufficient to block HIV ex vivo. This proof of concept study suggests film formulations for microbicides merit further investigation. PMID:26565716

Process for preparing energetic materials

DOEpatents

Simpson, Randall L [Livermore, CA; Lee, Ronald S [Livermore, CA; Tillotson, Thomas M [Tracy, CA; Hrubesh, Lawrence W [Pleasanton, CA; Swansiger, Rosalind W [Livermore, CA; Fox, Glenn A [Livermore, CA

2011-12-13

Sol-gel chemistry is used for the preparation of energetic materials (explosives, propellants and pyrotechnics) with improved homogeneity, and/or which can be cast to near-net shape, and/or made into precision molding powders. The sol-gel method is a synthetic chemical process where reactive monomers are mixed into a solution, polymerization occurs leading to a highly cross-linked three dimensional solid network resulting in a gel. The energetic materials can be incorporated during the formation of the solution or during the gel stage of the process. The composition, pore, and primary particle sizes, gel time, surface areas, and density may be tailored and controlled by the solution chemistry. The gel is then dried using supercritical extraction to produce a highly porous low density aerogel or by controlled slow evaporation to produce a xerogel. Applying stress during the extraction phase can result in high density materials. Thus, the sol-gel method can be used for precision detonator explosive manufacturing as well as producing precision explosives, propellants, and pyrotechnics, along with high power composite energetic materials.

Composition and method for self-assembly and mineralization of peptide-amphiphiles

DOEpatents

Stupp, Samuel I [Chicago, IL; Beniash, Elia [Newton, MA; Hartgerink, Jeffrey D [Pearland, TX

2012-02-28

The present invention is directed to a composition useful for making homogeneously mineralized self assembled peptide-amphiphile nanofibers and nanofiber gels. The composition is generally a solution comprised of a positively or negatively charged peptide-amphiphile and a like signed ion from the mineral. Mixing this solution with a second solution containing a dissolved counter-ion of the mineral and/or a second oppositely charged peptide amphiphile, results in the rapid self assembly of the peptide-amphiphiles into a nanofiber gel and templated mineralization of the ions. Templated mineralization of the initially dissolved mineral cations and anions in the mixture occurs with preferential orientation of the mineral crystals along the fiber surfaces within the nanofiber gel. One advantage of the present invention is that it results in homogenous growth of the mineral throughout the nanofiber gel. Another advantage of the present invention is that the nanofiber gel formation and mineralization reactions occur in a single mixing step and under substantially neutral or physiological pH conditions. These homogeneous nanostructured composite materials are useful for medical applications especially the regeneration of damaged bone in mammals. This invention is directed to the synthesis of peptide-amphiphiles with more than one amphiphilic moment and to supramolecular compositions comprised of such multi-dimensional peptide-amphiphiles. Supramolecular compositions can be formed by self assembly of multi-dimensional peptide-amphiphiles by mixing them with a solution comprising a monovalent cation.

Composition and method for self-assembly and mineralization of peptide amphiphiles

DOEpatents

Stupp, Samuel I [Chicago, IL; Beniash, Elia [Newton, MA; Hartgerink, Jeffrey D [Houston, TX

2009-06-30

The present invention is directed to a composition useful for making homogeneously mineralized self assembled peptide-amphiphile nanofibers and nanofiber gels. The composition is generally a solution comprised of a positively or negatively charged peptide-amphiphile and a like signed ion from the mineral. Mixing this solution with a second solution containing a dissolved counter-ion of the mineral and/or a second oppositely charged peptide amphiphile, results in the rapid self assembly of the peptide-amphiphiles into a nanofiber gel and templated mineralization of the ions. Templated mineralization of the initially dissolved mineral cations and anions in the mixture occurs with preferential orientation of the mineral crystals along the fiber surfaces within the nanofiber gel. One advantage of the present invention is that it results in homogenous growth of the mineral throughout the nanofiber gel. Another advantage of the present invention is that the nanofiber gel formation and mineralization reactions occur in a single mixing step and under substantially neutral or physiological pH conditions. These homogeneous nanostructured composite materials are useful for medical applications especially the regeneration of damaged bone in mammals. This invention is directed to the synthesis of peptide-amphiphiles with more than one amphiphilic moment and to supramolecular compositions comprised of such multi-dimensional peptide-amphiphiles. Supramolecular compositions can be formed by self assembly of multi-dimensional peptide-amphiphiles by mixing them with a solution comprising a monovalent cation.

Comparative Evaluation of Pain Scores during Periodontal Probing with or without Anesthetic Gels.

PubMed

Mishra, Ashank; Priyanka, Mandapathi; Pradeep, Koppolu; Reddy Pathakota, Krishnajaneya

2016-01-01

Context. The initial periodontal examination which includes full-mouth periodontal probing is one of the discomforting procedures for a patient. Aim. To evaluate the efficacy of two local anesthetic gels in the reduction of pain during periodontal probing using Florida probe in CGP patients in comparison with manual probing. Materials and Methods. Ninety systemically healthy patients with moderate to severe CGP patients were recruited. In each patient, the quadrants were randomly assigned to manual probing with UNC-15 probe, probing with Florida probe, and Florida probing with lidocaine 10% gel and with benzocaine 20% gel. In the quadrants undergoing probing with anesthetic gels, the sites were isolated and the gel was injected using syringe and a blunt-end cannula. Pain was measured using 10 mm horizontal VAS. Statistical Analysis. The analysis was carried out using SPSS version 18. The comparison of mean VAS scores was done using repeated measures ANOVA with post hoc Bonferroni test. Results. Mean VAS for manual probing was significantly more than Florida probing. Further, the mean VAS score for Florida probing was higher than the two gels. Conclusion. It is suggested that the gels might be useful in reducing pain experienced during full-mouth periodontal probing in patients with CGP.

Detection of seminal fluid proteins in the bed bug, Cimex lectularius, using two-dimensional gel electrophoresis and mass spectrometry

PubMed Central

REINHARDT, K.; WONG, C. H.; GEORGIOU, A. S.

2008-01-01

SUMMARY The global increase of the human parasite, the common bed bug Cimex lectularius, calls for specific pest control target sites. The bed bug is also a model species for sexual conflict theory which suggests seminal fluids may be highly diverse. The species has a highly unusual sperm biology and seminal proteins may have unique functions. 1-D PAGE gels showed 40 to 50% band sharing between C. lectularius and another cimicid species, Afrocimex constrictus. However, adult, sexually rested C. lectularius males were found to store 5 to 7μg of seminal protein and with only 60μg of protein we obtained informative 2-D PAGE gels. These showed 79% shared protein spots between two laboratory populations, and more than half of the shared protein spots were detected in the mated female. Further analysis using liquid chromatography electrospray ionisation tandem mass spectrometry revealed that 26.5% of the proteins had matches among arthropods in data bases and 14.5% matched Drosophila proteins. These included ubiquitous proteins but also those more closely associated with reproduction such as moj 29, ubiquitin, the stress-related elongation factor EF-1alpha, a protein disulfide isomerase and an antioxidant, Peroxiredoxin 6. PMID:19091156

29 Si NMR and SAXS investigation of the hybrid organic–inorganic glasses obtained by consolidation of the melting gels

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jitianu, Andrei; Cadars, Sylvian; Zhang, Fan

This study is focused on structural characterization of hybrid glasses obtained by consolidation of melting gels. The melting gels were prepared in molar ratios of methyltriethoxysilane (MTES) and dimethyldiethoxysilane (DMDES) of 75%MTES-25%DMDES and 65%MTES-35%DMDES. Following consolidation, the hybrid glasses were characterized using Raman, 29Si and 13C Nuclear Magnetic Resonance (NMR) spectroscopies, synchrotron Small Angle X-Ray Scattering (SAXS) and scanning electron microscopy (SEM). Raman spectroscopy revealed the presence of Si–C bonds in the hybrid glasses and 8-membered ring structures in the Si–O–Si network. Qualitative NMR spectroscopy identified the main molecular species, while quantitative NMR data showed that the ratio of trimersmore » (T) to dimers (D) varied between 4.6 and 3.8. Two-dimensional 29Si NMR data were used to identify two distinct types of T3 environments. SAXS data showed that the glasses are homogeneous across the nm to micrometer length scales. The scattering cross section was one thousand times lower than what is expected when phase separation occurs. The SEM images show a uniform surface without defects, in agreement with the SAXS results, which further supports that the hybrid glasses are nonporous.« less

29Si NMR and SAXS investigation of the hybrid organic-inorganic glasses obtained by consolidation of the melting gels

PubMed Central

Jitianu, Andrei; Cadars, Sylvian; Zhang, Fan; Rodriguez, Gabriela; Picard, Quentin; Aparicio, Mario; Mosa, Jadra; Klein, Lisa C.

2017-01-01

This study is focused on structural characterization of hybrid glasses obtained by consolidation of melting gels. The melting gels were prepared in molar ratios of methyltriethoxysilane (MTES) and dimethyldiethoxysilane (DMDES) of 75%MTES-25%DMDES and 65%MTES-35%DMDES. Following consolidation, the hybrid glasses were characterized using Raman, 29Si and 13C Nuclear Magnetic Resonance (NMR) spectroscopies, synchrotron Small Angle X-Ray Scattering (SAXS) and scanning electron microscopy (SEM). Raman spectroscopy revealed the presence of Si-C bonds in the hybrid glasses and 8-membered ring structures in the Si-O-Si network. Qualitative NMR spectroscopy identified the main molecular species, while quantitative NMR data showed that the ratio of trimers (T) to dimers (D) varied between 4.6 and 3.8. Two-dimensional 29Si NMR data were used to identify two distinct types of T3 environments. SAXS data showed that the glasses are homogeneous across the nm to micrometer length scales. The scattering cross section was one thousand times lower than what is expected when phase separation occurs. The SEM images show a uniform surface without defects, in agreement with the SAXS results, which further supports that the hybrid glasses are nonporous. PMID:28262904

Evaluation of different methods of protein extraction and identification of differentially expressed proteins upon ethylene-induced early-ripening in banana peels.

PubMed

Zhang, Li-Li; Feng, Ren-Jun; Zhang, Yin-Dong

2012-08-15

Banana peels (Musa spp.) are a good example of a plant tissue where protein extraction is challenging due to the abundance of interfering metabolites. Sample preparation is a critical step in proteomic research and is critical for good results. We sought to evaluate three methods of protein extraction: trichloroacetic acid (TCA)-acetone precipitation, phenol extraction, and TCA precipitation. We found that a modified phenol extraction protocol was the most optimal method. SDS-PAGE and two-dimensional gel electrophoresis (2-DE) demonstrated good protein separation and distinct spots of high quality protein. Approximately 300 and 550 protein spots were detected on 2-DE gels at pH values of 3-10 and 4-7, respectively. Several spots were excised from the 2-DE gels and identified by mass spectrometry. The protein spots identified were found to be involved in glycolysis, the tricarboxylic acid cycle, and the biosynthesis of ethylene. Several of the identified proteins may play important roles in banana ripening. Copyright © 2012 Society of Chemical Industry.

Improved techniques for fluid diversion in oil recovery. Final report

DOE Office of Scientific and Technical Information (OSTI.GOV)

Seright, R.

This three-year project had two technical objectives. The first objective was to compare the effectiveness of gels in fluid diversion (water shutoff) with those of other types of processes. Several different types of fluid-diversion processes were compared, including those using gels, foams, emulsions, particulates, and microorganisms. The ultimate goals of these comparisons were to (1) establish which of these processes are most effective in a given application and (2) determine whether aspects of one process can be combined with those of other processes to improve performance. Analyses and experiments were performed to verify which materials are the most effective inmore » entering and blocking high-permeability zones. The second objective of the project was to identify the mechanisms by which materials (particularly gels) selectively reduce permeability to water more than to oil. A capacity to reduce water permeability much more than oil or gas permeability is critical to the success of gel treatments in production wells if zones cannot be isolated during gel placement. Topics covered in this report include (1) determination of gel properties in fractures, (2) investigation of schemes to optimize gel placement in fractured systems, (3) an investigation of why some polymers and gels can reduce water permeability more than oil permeability, (4) consideration of whether microorganisms and particulates can exhibit placement properties that are superior to those of gels, and (5) examination of when foams may show placement properties that are superior to those of gels.« less

Modulating the forces between self-assembling molecules to control the shape of vesicles and the mechanics and alignment of nanofiber networks

NASA Astrophysics Data System (ADS)

Greenfield, Megan Ann

One of the great challenges in supramolecular chemistry is the design of molecules that can self-assemble into functional aggregates with well-defined three-dimensional structures and bulk material properties. Since the self-assembly of nanostructures is greatly influenced by both the nature of the self-assembling components and the environmental conditions in which the components assemble, this work explores how changes in the molecular design and the environment affect the properties of self-assembled structures. We first explore how to control the mechanical properties of self-assembled fibrillar networks by changing environmental conditions. We report here on how changing pH, screening ions, and solution temperature affect the gelation, stiffness, and response to deformation of peptide amphiphile gels. Although the morphology of PA gels formed by charge neutralization and salt-mediated charge screening are similar by electron microscopy, rheological measurements indicate that the calcium-mediated ionic bridges in CaCl2-PA gels form stronger intra- and inter-fiber crosslinks than the hydrogen bonds formed by the protonated carboxylic acid residues in HCl-PA gels. In contrast, the structure of PA gels changes drastically when the PA solution is annealed prior to gel formation. Annealed PA solutions are birefringent and can form viscoelastic strings of aligned nanofibers when manually dragged across a thin film of CaCl2. These aligned arrays of PA nanofibers hold great promise in controlling the orientation of cells in three-dimensions. Separately, we applied the principles of molecular design to create buckled membrane nanostructures that mimic the shape of viruses. When oppositely charged amphiphilic molecules are mixed they can form vesicles with a periodic two-dimensional ionic lattice that opposes the membrane's natural curvature and can result in vesicle buckling. Our results demonstrate that a large +3 to -1 charge imbalance between the cationic and anionic head groups of amphiphiles enables their co-assembly into small buckled vesicles. In contrast to previous reports, the structures described here form without the rigorous exclusion of salt and are tolerant to physiological salt concentrations. Our work opens a new path for exploring how ionic laterally correlated domains can influence the morphology of self-assembled nanostructures.

Surface-attached orthogonal gradient hydrogels

NASA Astrophysics Data System (ADS)

Chinnayan Kannan, Pandiyarajan; Genzer, Jan

Gradient materials play a significant role in the creation of artificial implants due to their potential to reduce stress concentration when two or more structures with different mechanical properties are joined together, e . g . , tendon, a fibrous protein that connects the soft and hard muscle tissues in our body. We employ free radical polymerization to synthesize random copolymers containing 90% of N-isopropyl acrylamide (NIPAAm), 5% photo-active methacrylyloxybenzophenone (MABP) and 5% thermally-active styrenesulfonylazide (SSAz) crosslinkers. The presence of MABP and SSAz facilitates adjusting gel density on a flat support in two orthogonal directions by spatially and independently controlling UV dosage and temperature. The swelling behavior (α) of the gels in water and methanol is examined using a spectroscopic ellipsometry and the degree of swelling depends on the extent of crosslinking that ranges from α = 1-1.2 (highly crosslinked gels) to α = 4-5 (loosely crosslinked gels). We compare the network properties surface-attached gels and un-attached identical counterparts and confirm that the linear swelling ratio of surface-attached networks is higher than that of the corresponding un-attached gels.

Isolation and characterization of nonhistone chromosomal protein C-14 which stimulates RNA synthesis.

PubMed

James, G T; Yeoman, L C; Matsui, S i; Goldberg, A H; Busch, H

1977-05-31

The nonhistone chromatin protein, C-14, was extracted from chromatin of Novikoff hepatoma ascites cells and isolated in high purity as shown by its migration as a single dense spot on two-dimensional polyacrylamide gels. Its mobility on sodium dodecyl sulfate gels is consistent with a molecular weight of approximately 70 000. The amino acid composition shows that protein C-14 has an acidic:basic amino acid ratio of 1.8. Its amino terminal amino acid is lysine. Protein C-14 stimulated the incorporation of [3H]UMP into RNA by approximately 30% when added to naked DNA and homologous RNA polymerase I. A 30% stimulation of [3H]UMP incorporation into RNA was also found when protein C-14 was added to an E. coli RNA polymerase system containing either E. coli or Novikoff hepatoma DNA.

Three-dimensional printing fiber reinforced hydrogel composites.

PubMed

Bakarich, Shannon E; Gorkin, Robert; in het Panhuis, Marc; Spinks, Geoffrey M

2014-09-24

An additive manufacturing process that combines digital modeling and 3D printing was used to prepare fiber reinforced hydrogels in a single-step process. The composite materials were fabricated by selectively pattering a combination of alginate/acrylamide gel precursor solution and an epoxy based UV-curable adhesive (Emax 904 Gel-SC) with an extrusion printer. UV irradiation was used to cure the two inks into a single composite material. Spatial control of fiber distribution within the digital models allowed for the fabrication of a series of materials with a spectrum of swelling behavior and mechanical properties with physical characteristics ranging from soft and wet to hard and dry. A comparison with the "rule of mixtures" was used to show that the swollen composite materials adhere to standard composite theory. A prototype meniscus cartilage was prepared to illustrate the potential application in bioengineering.

Blue native polyacrylamide gel electrophoresis and the monitoring of malate- and oxaloacetate-producing enzymes.

PubMed

Singh, R; Chénier, D; Bériault, R; Mailloux, R; Hamel, R D; Appanna, V D

2005-09-30

We demonstrate a facile blue native polyacrylamide gel electrophoresis (BN-PAGE) technique to detect two malate-generating enzymes, namely fumarase (FUM), malate synthase (MS) and four oxaloacetate-forming enzymes, namely pyruvate carboxylase (PC), phosphoenolpyruvate carboxykinase (PEPCK), citrate lyase (CL) and aspartate aminotransferase (AST). Malate dehydrogenase (MDH) was utilized as a coupling enzyme to detect either malate or oxaloacetate in the presence of their respective substrates and cofactors. The latter four oxaloacetate-forming enzymes were identified by 2,6-dichloroindophenol (DCIP) and p-iodonitrotetrazolium (INT) while the former two malate-producing enzymes were visualized by INT and phenazine methosulfate (PMS) in the reaction mixtures, respectively. The band formed at the site of enzymatic activity was easily quantified, while Coomassie staining provided information on the protein concentration. Hence, the expression and the activity of these enzymes can be readily evaluated. A two-dimensional (2D) BN-PAGE or SDS-PAGE enabled the rapid purification of the enzyme of interest. This technique also provides a quick and inexpensive means of quantifying these enzymatic activities in normal and stressed biological systems.

Mechanical Behavior of Collagen-Fibrin Co-Gels Reflects Transition From Series to Parallel Interactions With Increasing Collagen Content

PubMed Central

Lai, Victor K.; Lake, Spencer P.; Frey, Christina R.; Tranquillo, Robert T.; Barocas, Victor H.

2012-01-01

Fibrin and collagen, biopolymers occurring naturally in the body, are biomaterials commonly-used as scaffolds for tissue engineering. How collagen and fibrin interact to confer macroscopic mechanical properties in collagen-fibrin composite systems remains poorly understood. In this study, we formulated collagen-fibrin co-gels at different collagen-tofibrin ratios to observe changes in the overall mechanical behavior and microstructure. A modeling framework of a two-network system was developed by modifying our micro-scale model, considering two forms of interaction between the networks: (a) two interpenetrating but noninteracting networks (“parallel”), and (b) a single network consisting of randomly alternating collagen and fibrin fibrils (“series”). Mechanical testing of our gels show that collagen-fibrin co-gels exhibit intermediate properties (UTS, strain at failure, tangent modulus) compared to those of pure collagen and fibrin. The comparison with model predictions show that the parallel and series model cases provide upper and lower bounds, respectively, for the experimental data, suggesting that a combination of such interactions exists between the collagen and fibrin in co-gels. A transition from the series model to the parallel model occurs with increasing collagen content, with the series model best describing predominantly fibrin co-gels, and the parallel model best describing predominantly collagen co-gels. PMID:22482659

Global Analysis of Viral Infection in an Archaeal Model System

PubMed Central

Maaty, Walid S.; Steffens, Joseph D.; Heinemann, Joshua; Ortmann, Alice C.; Reeves, Benjamin D.; Biswas, Swapan K.; Dratz, Edward A.; Grieco, Paul A.; Young, Mark J.; Bothner, Brian

2012-01-01

The origin and evolutionary relationship of viruses is poorly understood. This makes archaeal virus-host systems of particular interest because the hosts generally root near the base of phylogenetic trees, while some of the viruses have clear structural similarities to those that infect prokaryotic and eukaryotic cells. Despite the advantageous position for use in evolutionary studies, little is known about archaeal viruses or how they interact with their hosts, compared to viruses of bacteria and eukaryotes. In addition, many archaeal viruses have been isolated from extreme environments and present a unique opportunity for elucidating factors that are important for existence at the extremes. In this article we focus on virus-host interactions using a proteomics approach to study Sulfolobus Turreted Icosahedral Virus (STIV) infection of Sulfolobus solfataricus P2. Using cultures grown from the ATCC cell stock, a single cycle of STIV infection was sampled six times over a 72 h period. More than 700 proteins were identified throughout the course of the experiments. Seventy one host proteins were found to change their concentration by nearly twofold (p < 0.05) with 40 becoming more abundant and 31 less abundant. The modulated proteins represent 30 different cell pathways and 14 clusters of orthologous groups. 2D gel analysis showed that changes in post-translational modifications were a common feature of the affected proteins. The results from these studies showed that the prokaryotic antiviral adaptive immune system CRISPR-associated proteins (CAS proteins) were regulated in response to the virus infection. It was found that regulated proteins come from mRNAs with a shorter than average half-life. In addition, activity-based protein profiling (ABPP) profiling on 2D-gels showed caspase, hydrolase, and tyrosine phosphatase enzyme activity labeling at the protein isoform level. Together, this data provides a more detailed global view of archaeal cellular responses to viral infection, demonstrates the power of quantitative two-dimensional differential gel electrophoresis and ABPP using 2D gel compatible fluorescent dyes. PMID:23233852

Comparison of gel contraction mediated by airway smooth muscle cells from patients with and without asthma.

PubMed

Matsumoto, Hisako; Moir, Lyn M; Oliver, Brian G G; Burgess, Janette K; Roth, Michael; Black, Judith L; McParland, Brent E

2007-10-01

Exaggerated bronchial constriction is the most significant and life threatening response of patients with asthma to inhaled stimuli. However, few studies have investigated the contractility of airway smooth muscle (ASM) from these patients. The purpose of this study was to establish a method to measure contraction of ASM cells by embedding them into a collagen gel, and to compare the contraction between subjects with and without asthma. Gel contraction to histamine was examined in floating gels containing cultured ASM cells from subjects with and without asthma following overnight incubation while unattached (method 1) or attached (method 2) to casting plates. Smooth muscle myosin light chain kinase protein levels were also examined. Collagen gels containing ASM cells reduced in size when stimulated with histamine in a concentration-dependent manner and reached a maximum at a mean (SE) of 15.7 (1.2) min. This gel contraction was decreased by inhibitors for phospholipase C (U73122), myosin light chain kinase (ML-7) and Rho kinase (Y27632). When comparing the two patient groups, the maximal decreased area of gels containing ASM cells from patients with asthma was 19 (2)% (n = 8) using method 1 and 22 (3)% (n = 6) using method 2, both of which were greater than that of cells from patients without asthma: 13 (2)% (n = 9, p = 0.05) and 10 (4)% (n = 5, p = 0.024), respectively. Smooth muscle myosin light chain kinase levels were not different between the two groups. The increased contraction of asthmatic ASM cells may be responsible for exaggerated bronchial constriction in asthma.

Titanium-Enriched Hydroxyapatite–Gelatin Scaffolds with Osteogenically Differentiated Progenitor Cell Aggregates for Calvaria Bone Regeneration

PubMed Central

Padilla, Ricardo; Urkasemsin, Ganokon; Yoon, Kun; Goeckner, Kelly; Hu, Wei-Shou

2013-01-01

Adequate bony support is the key to re-establish both function and esthetics in the craniofacial region. Autologous bone grafting has been the gold standard for regeneration of problematic large bone defects. However, poor graft availability and donor-site complications have led to alternative bone tissue-engineering approaches combining osteoinductive biomaterials and three-dimensional cell aggregates in scaffolds or constructs. The goal of the present study was to generate novel cell aggregate-loaded macroporous scaffolds combining the osteoinductive properties of titanium dioxide (TiO2) with hydroxyapatite–gelatin nanocomposites (HAP-GEL) for regeneration of craniofacial defects. Here we investigated the in vivo applicability of macroporous (TiO2)-enriched HAP-GEL scaffolds with undifferentiated and osteogenically differentiated multipotent adult progenitor cell (MAPC and OD-MAPC, respectively) aggregates for calvaria bone regeneration. The silane-coated HAP-GEL with and without TiO2 additives were polymerized and molded to produce macroporous scaffolds. Aggregates of the rat MAPC were precultured, loaded into each scaffold, and implanted to rat calvaria critical-size defects to study bone regeneration. Bone autografts were used as positive controls and a poly(lactic-co-glycolic acid) (PLGA) scaffold for comparison purposes. Preimplanted scaffolds and calvaria bone from pig were tested for ultimate compressive strength with an Instron 4411® and for porosity with microcomputerized tomography (μCT). Osteointegration and newly formed bone (NFB) were assessed by μCT and nondecalcified histology, and quantified by calcium fluorescence labeling. Results showed that the macroporous TiO2-HAP-GEL scaffold had a comparable strength relative to the natural calvaria bone (13.8±4.5 MPa and 24.5±8.3 MPa, respectively). Porosity was 1.52±0.8 mm and 0.64±0.4 mm for TiO2-HAP-GEL and calvaria bone, respectively. At 8 and 12 weeks postimplantation into rat calvaria defects, greater osteointegration and NFB were significantly present in the TiO2-enriched HAP-GEL constructs with OD-MAPCs, compared to the undifferentiated MAPC-loaded constructs, cell-free HAP-GEL with and without titanium, and PLGA scaffolds. The tissue-engineered TiO2-enriched HAP-GEL constructs with OD-MAPC aggregates present a potential useful therapeutic approach for calvaria bone regeneration. PMID:23495972

Effectiveness of 1% versus 0.2% chlorhexidine gels in reducing alveolar osteitis from mandibular third molar surgery: A randomized, double-blind clinical trial

PubMed Central

Bravo-Pérez, Manuel; Sánchez-López, José D.; Muñoz-Soto, Esther; Romero-Olid, María N.; Baca-García, Pilar

2013-01-01

Purpose: Alveolar osteitis (AO) is the most common postoperative complication of dental extractions. The purpose of this study was to compare the effectiveness of 1% versus 0.2% chlorhexidine (CHX) gel in reducing postoperative AO after surgical extraction of mandibular third molars, and assess the impact of treatment on the Oral HealthRelated Quality of Life (OHRQoL). Material and Methods: This clinical study was a randomized, double-blind clinical trial. Eighty eight patients underwent surgical extraction of one retained mandibular third molar with the intra-alveolar application of 0.2% CHX gel. Afterwards, they were assigned to one of two groups: 1% CHX gel (n=42) or 0.2% CHX gel (n=46). The patients applied the gel twice a day to the wound for one week. All patients were evaluated for AO. Results: In the 0.2% CHX gel group, 13% of AO incidence was found, while in the 1% CHX gel group, AO incidence was 7%, a difference that was not statistically significant. Variables such as sensation of pain and inflammation at baseline and during one week, as well as OHRQoL of the patients at 24 hours and 7 days post-extraction, gave no statistically significant differences. Conclusions: There are no significant differences in AO after surgical extraction of mandibular third molars, when comparing applying 1% CHX gel twice a day for 7 days with 0.2% CHX gel. Key words:Alveolar osteitis, chlorhexidine gel, third molar. PMID:23722126

Changes in the myosin secondary structure and shrimp surimi gel strength induced by dense phase carbon dioxide.

PubMed

Guo, Minghui; Liu, Shucheng; Ismail, Marliya; Farid, Mohammed M; Ji, Hongwu; Mao, Weijie; Gao, Jing; Li, Chengyong

2017-07-15

Dense phase carbon dioxide (DPCD) could induce protein conformation changes. Myosin and shrimp surimi from Litopenaeus vannamei were treated with DPCD at 5-25MPa and 40-60°C for 20min. Myosin secondary structure was investigated by circular dichroism and shrimp surimi gel strength was determined using textural analysis to develop correlations between them. DPCD had a greater effect on secondary structure and gel strength than heating. With increasing pressure and temperature, the α-helix content of DPCD-treated myosin decreased, while the β-sheet, β-turn and random coil contents increased, and the shrimp surimi gel strength increased. The α-helix content was negatively correlated with gel strength, while the β-sheet, β-turn and random coil contents were positively correlated with gel strength. Therefore, when DPCD induced myosin to form a gel, the α-helix of myosin was unfolded and gradually converted to a β-sheet. Such transformations led to protein-protein interactions and cross-linking, which formed a three-dimensional network to enhance the gel strength. Copyright © 2017 Elsevier Ltd. All rights reserved.

Biphasic response of cell invasion to matrix stiffness in 3-dimensional biopolymer networks

PubMed Central

Lang, Nadine R.; Skodzek, Kai; Hurst, Sebastian; Mainka, Astrid; Steinwachs, Julian; Schneider, Julia; Aifantis, Katerina E.; Fabry, Ben

2015-01-01

When cells come in contact with an adhesive matrix, they begin to spread and migrate with a speed that depends on the stiffness of the extracellular matrix. On a flat surface, migration speed decreases with matrix stiffness mainly due to an increased stability of focal adhesions. In a 3-dimensional (3D) environment, cell migration is thought to be additionally impaired by the steric hindrance imposed by the surrounding matrix. For porous 3D biopolymer networks such as collagen gels, however, the effect of matrix stiffness on cell migration is difficult to separate from effects of matrix pore size and adhesive ligand density, and is therefore unknown. Here we used glutaraldehyde as a crosslinker to increase the stiffness of self-assembled collagen biopolymer networks independently of collagen concentration or pore size. Breast carcinoma cells were seeded onto the surface of 3D collagen gels, and the invasion depth was measured after 3 days of culture. Cell invasion in gels with pore sizes larger than 5 μm increased with higher gel stiffness, whereas invasion in gels with smaller pores decreased with higher gel stiffness. These data show that 3D cell invasion is enhanced by higher matrix stiffness, opposite to cell behavior in 2D, as long as the pore size does not fall below a critical value where it causes excessive steric hindrance. These findings may be important for optimizing the recellularization of soft tissue implants or for the design of 3D invasion models in cancer research. PMID:25462839

[The effect of locally applied Grisaldone or choline salicylate gel on bone healing after tooth extraction in animal experiments].

PubMed

Cendelin, E; Fröhlich, M

1977-01-01

Comparative histological and experimental animal studies of the effects upon bone wound healing of topically applied choline salicylate gel and Grisaldon showed that Grisaldon tends to hinder the course of reparation of bone, whereas choline salicylate gel has no appreciable influence upon the time course of wound healing. This essential difference is considered to be due predominantly to the exactly opposite behavior shown by the two pharmaceutical preparations in regard to their solubility in water. The difficultly watersoluble Grisaldon tends to exert a longer-drawn-out irritant effect upon the tissue and can be detected in alveoli even after twenty-eight days from administration thereof. By contrast, choline salicylate gel, which is known to be readily soluble in water, will be eliminated already after two days from administration thereof.

Blood Sampling and Preparation Procedures for Proteomic Biomarker Studies of Psychiatric Disorders.

PubMed

Guest, Paul C; Rahmoune, Hassan

2017-01-01

A major challenge in proteomic biomarker discovery and validation for psychiatric diseases is the inherent biological complexity underlying these conditions. There are also many technical issues which hinder this process such as the lack of standardization in sampling, processing and storage of bio-samples in preclinical and clinical settings. This chapter describes a reproducible procedure for sampling blood serum and plasma that is specifically designed for maximizing data quality output in two-dimensional gel electrophoresis, multiplex immunoassay and mass spectrometry profiling studies.

Biochemical Markers of Brain Injury: An Integrated Proteomics-Based Approach

DTIC Science & Technology

2006-02-01

Anthony J Williams, X-C May Lu, Renwu Chen, Zhilin Liao, Rebeca Connors, Kevin K Wang, Ron L Hayes, Frank C Tortella, Jitendra R Dave. High throughput... YANG , A., et al. (2002). Evalu- ation of two-dimensional differential gel electrophoresis for proteomic expression analysis of a model breast cancer cell...apoptosis. J. Biol. Chem. 279, 1030–1039. Kuida K., Zheng T. S., Na S., Kuan C., Yang D., Karasuyama H., Rakic P. and Flavell R. A. (1996) Decreased apoptosis

Purification of barley dimeric α-amylase inhibitor-1 (BDAI-1) and avenin-like protein-a (ALP) from beer and their impact on beer foam stability.

PubMed

Iimure, Takashi; Kihara, Makoto; Sato, Kazuhiro; Ogushi, Kensuke

2015-04-01

Foam stability is a key factor of beer quality for consumers and brewers. Recent beer proteome analyses have suggested that barley dimeric α-amylase inhibitor-1 (BDAI-1) and avenin-like protein-a (ALP) derived from barley are important for beer foam stability. In this study, BDAI-1 and ALP were purified from a Japanese commercial beer sample using salt precipitation and column chromatography. The purification level was verified using two-dimensional gel electrophoresis, mass spectrometry, and database searches. Purified BDAI-1 and ALP were added to a beer sample to compare the foam stability to that of a control beer sample. As a result, beer foam stability was significantly improved by BDAI-1 but not by ALP, thereby suggesting that BDAI-1 affects beer foam stability whereas ALP does not. Copyright © 2014 Elsevier Ltd. All rights reserved.

Proteomic analysis of Trichoderma atroviride mycelia stressed by organophosphate pesticide dichlorvos.

PubMed

Tang, Jun; Liu, Lixing; Huang, Xiuli; Li, Yingying; Chen, Yunpeng; Chen, Jie

2010-02-01

The proteomic approach is a powerful tool to study microbial response to environmental stress. To evaluate the responses of Trichoderma spp. to the organophosphate pesticide dichlorvos, mycelia of Trichoderma atroviride T23 were exposed to dichlorvos at concentrations of 0, 100, 300, 500, and 1000 microg/mL, respectively. Changes in protein expression were investigated using two-dimensional sodium dodecyl sulfate - polyacrylamide gel electrophoresis. Sixteen protein spots were differentially expressed. They were identified by MALDI-TOF/TOF MS and were found to be linked to energy metabolism, transport, signal transduction, and stress tolerance. Among stress-related proteins, glutathione peroxidase-like protein (GPX), 1,4-benzoquinone reductase, and HEX1 were upregulated by and cyclophilin A induced by 1000 microg/mL dichlorvos when compared with the control. These proteins were considered to be associated with fungal adaptation to adverse conditions. The results will help us to understand molecular mechanisms through which Trichoderma responds to organophosphate pesticides.

Effect of lipopolysaccharide on protein accumulation by murine peritoneal macrophages: the correlation to activation for macrophage tumoricidal function

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tannenbaum, C.S.

1987-01-01

The protein synthetic patterns of tumoricidal murine peritoneal macrophage populations have been compared to those of non-tumoricidal populations utilizing two dimensional polyacrylamide gel electrophoresis (2D PAGE) of (/sup 35/S)-methionine-labeled proteins. While the protein synthetic patterns exhibited by resident, inflammatory and activated macrophages had numerous common features which distinguished them from the other normal non-macrophage cell types examined, unique proteins also distinguished each macrophage population from the others. Peritoneal macrophages elicited by treatment with heat killed Propionibacterium acnes, the live, attenuated Mycobacterium bovis strain BCG, Listeria monocytogenes and the protozoan flagellate Trypanosoma rhodesiense, all exhibited tumoricidal activity in 16h or 72hmore » functional assays, and shared a common protein synthetic profile which differentiated them from the synthetic patterns characteristic of the non-tumoricidal resident and inflammatory macrophages.« less

Physiological and proteomic analysis of plant growth enhancement by the rhizobacteria Bacillus sp. JS.

PubMed

Kim, Ji Seong; Lee, Jeong Eun; Nie, Hualin; Lee, Yong Jae; Kim, Sun Tae; Kim, Sun-Hyung

2018-02-01

In this study, the effects of the plant growth-promoting rhizobacterium (PGPR), Bacillus sp. JS on the growth of tobacco (Nicotiana tabacum 'Xanthi') and lettuce (Lactuca sativa 'Crispa'), were evaluated by comparing various growth parameters between plants treated with the bacterium and those exposed to water or nutrient broth as control. In both tobacco and lettuce, fresh weight and length of shoots were increased upon exposure to Bacillus sp. JS. To explain the overall de novo expression of plant proteins by bacterial volatiles, two-dimensional gel electrophoresis was performed on samples from PGPR-treated tobacco plants. Our results showed that chlorophyll a/b binding proteins were significantly up-regulated, and total chlorophyll content was also increased. Our findings indicate the potential benefits of using Bacillus sp. JS as a growth-promoting factor in agricultural practice, and highlight the need for further research to explore these benefits.

Ethanolic extraction, purification, and partial characterization of a fluorescent toxin from the coral reef crab, Lophozozymus pictor.

PubMed

Lau, C O; Tan, C H; Khoo, H E; Li, Q T; Yuen, R

1995-01-01

A purification procedure for Lophozozymus pictor toxin (LPTX) following ethanolic extraction of whole crab homogenate is described. The ethanol-extracted toxin (LPTX-E) had higher yield and specific activity than the hot aqueous-extracted one (LPTX-H). It was found that LPTX-E was fluorescent and cochromatographed with LPTX-H on two-dimensional thin-layer chromatography. Although LPTX-E, LPTX-H, and palytoxin (P. caribaeorum, PTX) had similar migration and retention times when analysed on high performance capillary electrophoresis and gel permeation-high performance liquid chromatography respectively, LPTX-E and LPTX-H were both fluorescent in contrast to PTX. In addition, LPTX-E had a different retention time compared with PTX when chromatographed on reversed phase high performance liquid chromatography in the solvent system 80% acetonitrile and 0.02 M Tris-HCl, pH 7.2, at a 4:1 ratio, respectively, indicating some differences in their chemical structures.

The evaluation of doxycycline controlled release gel versus doxycycline controlled release implant in the management of periodontitis

PubMed Central

Chadha, Vandana Srikrishna; Bhat, Khandige Mahalinga

2012-01-01

Background: Investigators have sought different methods to deliver antimicrobials to periodontal pockets. This study was designed to assess the efficacy of locally made doxycycline gel versus locally made doxycycline implant as biodegradable controlled local delivery systems, by evaluating the pharmacological drug release and improvement in gingival status, gain in attachment, and reduction in pocket depth. Materials and Methods: Thirty patients with localized periodontal pockets ≥5 mm were randomly divided into three groups. The first group received the doxycycline gel, the second the doxycycline implant, and the third received only scaling and root planing (the control group). The patients in the first two groups were selected for the drug release. Clinical parameters such as gingival index, plaque index, probing depth, and attachment levels were recorded at baseline and the 90th day. Gingival crevicular fluid (GCF) and saliva samples were collected 1 hour following gel and implant placement and then on the 10th, 30th, and 60th days. Results: There was a statistically significant difference in the release of doxycycline from the gel when compared with the implant in the GCF and saliva on the 10th and 30th days. All the three groups showed improvement in clinical parameters. The improvements in both gel and implant groups were greater when compared with the control group with no statistically significant difference between the implant and gel systems. Conclusion: The use of local delivery of doxycycline through gel and Implant media further enhances the positive changes obtained following scaling and root planing. The release of doxycycline from the implant and the gel was comparable. PMID:23055585

Two-dimensional electrophoretic analysis of human leukocyte proteins from patients with rheumatoid arthritis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Willard, K.E.; Thorsrud, A.K.; Munthe, E.

Human leukocyte proteins from more than 150 patients with rheumatoid arthritis, together with age- and sex-matched controls, were analyzed by use of the ISO-DALT technique of two-dimensional polyacrylamide gel electrophoresis. Patients with ankylosing spondylitis, polymyalgia rheumatica, psoriatic arthritis, calcium tendinitis, post-infectious arthritis, and asymmetrical seronegative arthritis were also included as positive controls. Synthesis of several proteins, referred to by number as members of the Rheuma set, is shown to increase in the leukocyte preparations from patients with classical rheumatoid arthritis. Several of these proteins are specific to monocytes or granulocytes; others are of unknown cellular origin, but appear to bemore » unique to rheumatoid arthritis. The Rheuma proteins appear to be indicators of disease activity, because their increased synthesis can be correlated with sedimentation rate and other clinical indices of rheumatoid disease activity.« less

Moesin Is a Biomarker for the Assessment of Genotoxic Carcinogens in Mouse Lymphoma

PubMed Central

Lee, Yoen Jung; Choi, In-Kwon; Sheen, Yhun Yhong; Park, Sue Nie; Kwon, Ho Jeong

2012-01-01

1,2-Dibromoethane and glycidol are well known genotoxic carcinogens, which have been widely used in industry. To identify a specific biomarker for these carcinogens in cells, the cellular proteome of L5178Y mouse lymphoma cells treated with these compounds was analyzed by 2-dimensional gel electrophoresis (2-DE) and MALDI-TOF mass spectrometry (MS). Of 50 protein spots showing a greater than 1.5-fold increase or decrease in intensity compared to control cells on a 2-D gel, we focused on the candidate biomarker moesin. Western analysis using monoclonal rabbit anti-moesin confirmed the identity of the protein and its increased level of expression upon exposure to the carcinogenic compounds. Moesin expression also increased in cells treated with six additional genotoxic carcinogens, verifying that moesin could serve as a biomarker to monitor phenotypic change upon exposure to genotoxic carcinogens in L5178Y mouse lymphoma cells. PMID:22358511

Magnetically levitated mesenchymal stem cell spheroids cultured with a collagen gel maintain phenotype and quiescence

PubMed Central

Lewis, Natasha S; Lewis, Emily EL; Mullin, Margaret; Wheadon, Helen; Dalby, Matthew J; Berry, Catherine C

2017-01-01

Multicellular spheroids are an established system for three-dimensional cell culture. Spheroids are typically generated using hanging drop or non-adherent culture; however, an emerging technique is to use magnetic levitation. Herein, mesenchymal stem cell spheroids were generated using magnetic nanoparticles and subsequently cultured within a type I collagen gel, with a view towards developing a bone marrow niche environment. Cells were loaded with magnetic nanoparticles, and suspended beneath an external magnet, inducing self-assembly of multicellular spheroids. Cells in spheroids were viable and compared to corresponding monolayer controls, maintained stem cell phenotype and were quiescent. Interestingly, core spheroid necrosis was not observed, even with increasing spheroid size, in contrast to other commonly used spheroid systems. This mesenchymal stem cell spheroid culture presents a potential platform for modelling in vitro bone marrow stem cell niches, elucidating interactions between cells, as well as a useful model for drug delivery studies. PMID:28616152

Moesin is a biomarker for the assessment of genotoxic carcinogens in mouse lymphoma.

PubMed

Lee, Yoen Jung; Choi, In-Kwon; Sheen, Yhun Yhong; Park, Sue Nie; Kwon, Ho Jeong

2012-02-01

1,2-Dibromoethane and glycidol are well known genotoxic carcinogens, which have been widely used in industry. To identify a specific biomarker for these carcinogens in cells, the cellular proteome of L5178Y mouse lymphoma cells treated with these compounds was analyzed by 2-dimensional gel electrophoresis (2-DE) and MALDI-TOF mass spectrometry (MS). Of 50 protein spots showing a greater than 1.5-fold increase or decrease in intensity compared to control cells on a 2-D gel, we focused on the candidate biomarker moesin. Western analysis using monoclonal rabbit anti-moesin confirmed the identity of the protein and its increased level of expression upon exposure to the carcinogenic compounds. Moesin expression also increased in cells treated with six additional genotoxic carcinogens, verifying that moesin could serve as a biomarker to monitor phenotypic change upon exposure to genotoxic carcinogens in L5178Y mouse lymphoma cells.

Data on DNA gel sample load, gel electrophoresis, PCR and cost analysis.

PubMed

Kuhn, Ramona; Böllmann, Jörg; Krahl, Kathrin; Bryant, Isaac Mbir; Martienssen, Marion

2018-02-01

The data presented in this article provide supporting information to the related research article "Comparison of ten different DNA extraction procedures with respect to their suitability for environmental samples" (Kuhn et al., 2017) [1]. In that article, we compared the suitability of ten selected DNA extraction methods based on DNA quality, purity, quantity and applicability to universal PCR. Here we provide the data on the specific DNA gel sample load, all unreported gel images of crude DNA and PCR results, and the complete cost analysis for all tested extraction procedures and in addition two commercial DNA extraction kits for soil and water.

A New Combination of Testosterone and Nestorone Transdermal Gels for Male Hormonal Contraception

PubMed Central

Ilani, Niloufar; Roth, Mara Y.; Amory, John K.; Swerdloff, Ronald S.; Dart, Clint; Page, Stephanie T.; Bremner, William J.; Sitruk-Ware, Regine; Kumar, Narender; Blithe, Diana L.

2012-01-01

Context: Combinations of testosterone (T) and nestorone (NES; a nonandrogenic progestin) transdermal gels may suppress spermatogenesis and prove appealing to men for contraception. Objective: The objective of the study was to determine the effectiveness of T gel alone or combined with NES gel in suppressing spermatogenesis. Design and Setting: This was a randomized, double-blind, comparator clinical trial conducted at two academic medical centers. Participants: Ninety-nine healthy male volunteers participated in the study. Interventions: Volunteers were randomized to one of three treatment groups applying daily transdermal gels (group 1: T gel 10 g + NES 0 mg/placebo gel; group 2: T gel 10 g + NES gel 8 mg; group 3: T gel 10 g + NES gel 12 mg). Main Outcome Variable: The main outcome variable of the study was the percentage of men whose sperm concentration was suppressed to 1 million/ml or less by 20–24 wk of treatment. Results: Efficacy data analyses were performed on 56 subjects who adhered to the protocol and completed at least 20 wk of treatment. The percentage of men whose sperm concentration was 1 million/ml or less was significantly higher for T + NES 8 mg (89%, P < 0.0001) and T + NES 12 mg (88%, P = 0.0002) compared with T + NES 0 mg group (23%). The median serum total and free T concentrations in all groups were maintained within the adult male range throughout the treatment period. Adverse effects were minimal in all groups. Conclusion: A combination of daily NES + T gels suppressed sperm concentration to 1 million/ml or less in 88.5% of men, with minimal adverse effects, and may be further studied as a male transdermal hormonal contraceptive. PMID:22791756

A new sol-gel synthesis of 45S5 bioactive glass using an organic acid as catalyst.

PubMed

Faure, J; Drevet, R; Lemelle, A; Ben Jaber, N; Tara, A; El Btaouri, H; Benhayoune, H

2015-02-01

In this paper a new sol-gel approach was explored for the synthesis of the 45S5 bioactive glass. We demonstrate that citric acid can be used instead of the usual nitric acid to catalyze the sol-gel reactions. The substitution of nitric acid by citric acid allows to reduce strongly the concentration of the acid solution necessary to catalyze the hydrolysis of silicon and phosphorus alkoxides. Two sol-gel powders with chemical compositions very close to that of the 45S5 were obtained by using either a 2M nitric acid solution or either a 5mM citric acid solution. These powders were characterized and compared to the commercial Bioglass®. The surface properties of the two bioglass powders were assessed by scanning electron microscopy (SEM) and by Brunauer-Emmett-Teller method (BET). The Fourier transformed infrared spectroscopy (FTIR) and the X-ray diffraction (XRD) revealed a partial crystallization associated to the formation of crystalline phases on the two sol-gel powders. The in vitro bioactivity was then studied at the key times during the first hours of immersion into acellular Simulated Body Fluid (SBF). After 4h immersion into SBF we clearly demonstrate that the bioactivity level of the two sol-gel powders is similar and much higher than that of the commercial Bioglass®. This bioactivity improvement is associated to the increase of the porosity and the specific surface area of the powders synthesized by the sol-gel process. Moreover, the nitric acid is efficiently substituted by the citric acid to catalyze the sol-gel reactions without alteration of the bioactivity of the 45S5 bioactive glass. Copyright © 2014 Elsevier B.V. All rights reserved.

Mesenchymal stem cells and myoblast differentiation under HGF and IGF-1 stimulation for 3D skeletal muscle tissue engineering.

PubMed

Witt, R; Weigand, A; Boos, A M; Cai, A; Dippold, D; Boccaccini, A R; Schubert, D W; Hardt, M; Lange, C; Arkudas, A; Horch, R E; Beier, J P

2017-02-28

Volumetric muscle loss caused by trauma or after tumour surgery exceeds the natural regeneration capacity of skeletal muscle. Hence, the future goal of tissue engineering (TE) is the replacement and repair of lost muscle tissue by newly generating skeletal muscle combining different cell sources, such as myoblasts and mesenchymal stem cells (MSCs), within a three-dimensional matrix. Latest research showed that seeding skeletal muscle cells on aligned constructs enhance the formation of myotubes as well as cell alignment and may provide a further step towards the clinical application of engineered skeletal muscle. In this study the myogenic differentiation potential of MSCs upon co-cultivation with myoblasts and under stimulation with hepatocyte growth factor (HGF) and insulin-like growth factor-1 (IGF-1) was evaluated. We further analysed the behaviour of MSC-myoblast co-cultures in different 3D matrices. Primary rat myoblasts and rat MSCs were mono- and co-cultivated for 2, 7 or 14 days. The effect of different concentrations of HGF and IGF-1 alone, as well as in combination, on myogenic differentiation was analysed using microscopy, multicolour flow cytometry and real-time PCR. Furthermore, the influence of different three-dimensional culture models, such as fibrin, fibrin-collagen-I gels and parallel aligned electrospun poly-ε-caprolacton collagen-I nanofibers, on myogenic differentiation was analysed. MSCs could be successfully differentiated into the myogenic lineage both in mono- and in co-cultures independent of HGF and IGF-1 stimulation by expressing desmin, myocyte enhancer factor 2, myosin heavy chain 2 and alpha-sarcomeric actinin. An increased expression of different myogenic key markers could be observed under HGF and IGF-1 stimulation. Even though, stimulation with HGF/IGF-1 does not seem essential for sufficient myogenic differentiation. Three-dimensional cultivation in fibrin-collagen-I gels induced higher levels of myogenic differentiation compared with two-dimensional experiments. Cultivation on poly-ε-caprolacton-collagen-I nanofibers induced parallel alignment of cells and positive expression of desmin. In this study, we were able to myogenically differentiate MSC upon mono- and co-cultivation with myoblasts. The addition of HGF/IGF-1 might not be essential for achieving successful myogenic differentiation. Furthermore, with the development of a biocompatible nanofiber scaffold we established the basis for further experiments aiming at the generation of functional muscle tissue.

Growth of Normal Mouse Vaginal Epithelial Cells in and on Collagen Gels

NASA Astrophysics Data System (ADS)

Iguchi, Taisen; Uchima, Francis-Dean A.; Ostrander, Patricia L.; Bern, Howard A.

1983-06-01

Sustained growth in primary culture of vaginal epithelial cells from ovariectomized adult BALB/cCrg1 mice embedded within or seeded on collagen gel matrix was achieved in a serum-free medium composed of Ham's F-12 medium/Dulbecco's modified Eagle's medium, 1:1 (vol/vol), supplemented with insulin, bovine serum albumin fraction V, epidermal growth factor, cholera toxin, and transferrin. Three-dimensional growth of vaginal epithelial cells occurred inside the collagen gel matrix. Cell numbers increased 4- to 8-fold in collagen gel and about 4-fold on collagen gel after 9-10 days in culture. The effect of 17β -estradiol (0.00018-180 nM in gel or 0.018-180 nM on gel) and diethylstilbestrol (DES; 0.0186-186 nM in gel) on the growth of vaginal epithelial cells was examined. The addition of estrogen did not enhance the growth of vaginal epithelial cells during this time period either in the complete medium or in a suboptimal medium. Cultures on floating collagen gels in the serum-free medium are composed of 1-3 cell layers with superficial cornification. Estrogen does not appear to be a direct mitogen for vaginal epithelial cells, at least in this system.

Tretinoin Photostability

PubMed Central

Rosso, James Del; Harper, Julie; Pillai, Radhakrishnan; Moore, Robert

2013-01-01

Background: Various formulations of tretinoin have been reported to be unstable after exposure to artificial light or sunlight. The observation that tretinoin is photolabile in the presence of light led to the recommendation that tretinoin be applied in the evening in order to avoid photodegradation, which could potentially reduce efficacy. More recently, the development of innovative vehicle formulations has led, in some cases, to a marked decrease in the photodegradation of tretinoin. Objective: To compare the photostability of a micronized aqueous-based formulation of tretinoin gel 0.05% with tretinoin gel 0.025% following exposure to fluorescent and simulated solar light conditions in vitro. Methods: Micronized tretinoin gel 0.05% and tretinoin gel 0.025% were exposed to fluorescent light over eight hours or simulated solar light up to 600mJ/cm2 (equivalent to 30 minimal erythemal dose). Product samples were prepared and analyzed for tretinoin concentration using high-performance liquid chromatography. Additional duplicate samples were similarly prepared and analyzed after 2, 4, 6, and 8 hours. Results: There was an 11-percent degradation of tretinoin 0.05% formulated as the micronized gel compared to an 86-percent degradation of tretinoin 0.025% formulated as the conventional gel following eight hours of exposure to fluorescent light in vitro. The degradation of tretinoin 0.025% in the conventional gel was greater than 83 percent within two hours. In the second light exposure study, in vitro exposure to simulated solar light provided a gradual dose-response effect with tretinoin 0.05% formulated as the micronized tretinoin gel. The photodegradation of tretinoin 0.025% in the conventional gel was more immediate and of substantial magnitude (>85%) after exposures at all minimal erythemal dose levels. Conclusion: Tretinoin 0.05% formulated as a micronized gel 0.05% showed minimal degradation when exposed to fluorescent light over eight hours. This same formulation exhibited a clear dose-response degradation pattern when exposed to simulated solar light. In contrast, tretinoin 0.025% formulated in a conventional gel exhibited marked photodegradation within the first two hours when exposed to both light conditions in vitro. This information adds to the body of evidence that supports the observation that certain vehicle formulations may reduce the potential for photodegradation of tretinoin. PMID:23441237

Proteomic analysis identifies differentially expressed proteins after red propolis treatment in Hep-2 cells.

PubMed

Frozza, Caroline Olivieri da Silva; Ribeiro, Tanara da Silva; Gambato, Gabriela; Menti, Caroline; Moura, Sidnei; Pinto, Paulo Marcos; Staats, Charley Christian; Padilha, Francine Ferreira; Begnini, Karine Rech; de Leon, Priscila Marques Moura; Borsuk, Sibele; Savegnago, Lucielli; Dellagostin, Odir; Collares, Tiago; Seixas, Fabiana Kömmling; Henriques, João Antonio Pêgas; Roesch-Ely, Mariana

2014-01-01

Here we investigated alterations in the protein profile of Hep-2 treated with red propolis using two-dimensional electrophoresis associated to mass spectrometry and apoptotic rates of cells treated with and without red propolis extracts through TUNEL and Annexin-V assays. A total of 325 spots were manually excised from the two-dimensional gel electrophoresis and 177 proteins were identified using LC-MS-MS. Among all proteins identified that presented differential expression, most were down-regulated in presence of red propolis extract at a concentration of 120 μg/mL (IC50): GRP78, PRDX2, LDHB, VIM and TUBA1A. Only two up-regulated proteins were identified in this study in the non-cytotoxic (6 μg/mL) red propolis treated group: RPLP0 and RAD23B. TUNEL staining assay showed a markedly increase in the mid- to late-stage apoptosis of Hep-2 cells induced by red propolis at concentrations of 60 and 120 μg/mL when compared with non-treated cells. The increase of late apoptosis was confirmed by in situ Annexin-V analysis in which red propolis extract induced late apoptosis in a dose-dependent manner. The differences in tumor cell protein profiles warrant further investigations including isolation of major bioactive compounds of red propolis in different cell lines using proteomics and molecular tests to validate the protein expression here observed. Copyright © 2013 Elsevier Ltd. All rights reserved.

Differential expression profiling of the hepatic proteome in a rat model of dioxin resistance: correlation with genomic and transcriptomic analyses.

PubMed

Pastorelli, Roberta; Carpi, Donatella; Campagna, Roberta; Airoldi, Luisa; Pohjanvirta, Raimo; Viluksela, Matti; Hakansson, Helen; Boutros, Paul C; Moffat, Ivy D; Okey, Allan B; Fanelli, Roberto

2006-05-01

One characteristic feature of acute 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) toxicity is dramatic interspecies and interstrain variability in sensitivity. This complicates dioxin risk assessment for humans. However, this variability also provides a means of characterizing mechanisms of dioxin toxicity. Long-Evans (Turku/AB) rats are orders of magnitude more susceptible to TCDD lethality than Han/Wistar (Kuopio) rats, and this difference constitutes a very useful model for identifying mechanisms of dioxin toxicity. We adopted a proteomic approach to identify the differential effects of TCDD exposure on liver protein expression in Han/Wistar rats as compared with Long-Evans rats. This allows determination of which, if any, protein markers are indicative of differences in dioxin susceptibility and/or responsible for conferring resistance. Differential protein expression in total liver protein was assessed using two-dimensional gel electrophoresis, computerized gel image analysis, in-gel digestion, and mass spectrometry. We observed significant changes in the abundance of several proteins, which fall into three general classes: (i) TCDD-independent and exclusively strain-specific (e.g. isoforms of the protein-disulfide isomerase A3, regucalcin, and agmatine ureohydrolase); (ii) strain-independent and only dependent on TCDD exposure (e.g. aldehyde dehydrogenase 3A1 and rat selenium-binding protein 2); (iii) dependent on both TCDD exposure and strain (e.g. oxidative stress-related proteins, apoptosis-inducing factor, and MAWD-binding protein). By integrating transcriptomic (microarray) data and genomic data (computational search of regulatory elements), we found that protein expression levels were mainly controlled at the level of transcription. These results reveal, for the first time, a subset of hepatic proteins that are differentially regulated in response to TCDD in a strain-specific manner. Some of these differential responses may play a role in establishing the major differences in TCDD response between these two strains of rats. As such, our work is expected to lead to new insights into the mechanism of TCDD toxicity and resistance.

Comparative proteomic analysis of two pathogenic Tritrichomonas foetus genotypes: there is more to the proteome than meets the eye.

PubMed

Stroud, Leah J; Šlapeta, Jan; Padula, Matthew P; Druery, Dylan; Tsiotsioras, George; Coorssen, Jens R; Stack, Colin M

2017-03-01

Certain clinical isolates of Tritrichomonas foetus infect the urogenital tract of cattle while others infect the gastrointestinal tract of cats. Previous studies have identified subtle genetic differences between these isolates with the term "genotype" adopted to reflect host origin. The aim of this work was to seek evidence of host-specific adaptation and to clarify the relationship between T. foetus genotypes. To do this we characterised the proteomes of both genotypes using two-dimensional gel electrophoresis (2DE) coupled with LC-MS/MS. Our comparative analysis of the data revealed that both genotypes exhibited largely similar proteoform profiles; however differentiation was possible with 24 spots identified as having a four-fold or greater change. Deeper analysis using 2DE zymography and protease-specific fluorogenic substrates revealed marked differences in cysteine protease (CP) expression profiles between the two genotypes. These variances in CP activities could also account for the pathogenic and histopathological differences previously observed between T. foetus genotypes in cross-infection studies. Our findings highlight the importance of CPs as major determinants of parasite virulence and provide a foundation for future host-parasite interaction studies, with direct implications for the development of vaccines or drugs targeting T. foetus. Copyright © 2017 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.

Comparison of theoretical and flight-measured local flow aerodynamics for a low-aspect-ratio fin

NASA Technical Reports Server (NTRS)

Johnson, J. B.; Sandlin, D. R.

1984-01-01

Flight test and theoretical aerodynamic data were obtained for a flight test fixture mounted on the underside of an F-104G aircraft. The theoretical data were generated using two codes, a two dimensional transonic code called Code H, and a three dimensional subsonic and supersonic code call wing-body. Pressure distributions generated by the codes for the flight test fixture as well as boundary layer displacement thickness generated by the two dimensional code were compared to the flight test data. The two dimensional code pressure distributions compared well except at the minimum pressure point and trailing edge. Shock locations compared well except at high transonic speeds. The three dimensional code pressure distributions compared well except at the trailing edge of the flight test fixture. The two dimensional code does not predict displacement thickness of the flight test fixture well.

Structural changes of malt proteins during boiling.

PubMed

Jin, Bei; Li, Lin; Liu, Guo-Qin; Li, Bing; Zhu, Yu-Kui; Liao, Liao-Ning

2009-03-09

Changes in the physicochemical properties and structure of proteins derived from two malt varieties (Baudin and Guangmai) during wort boiling were investigated by differential scanning calorimetry, SDS-PAGE, two-dimensional electrophoresis, gel filtration chromatography and circular dichroism spectroscopy. The results showed that both protein content and amino acid composition changed only slightly during boiling, and that boiling might cause a gradual unfolding of protein structures, as indicated by the decrease in surface hydrophobicity and free sulfhydryl content and enthalpy value, as well as reduced alpha-helix contents and markedly increased random coil contents. It was also found that major component of both worts was a boiling-resistant protein with a molecular mass of 40 kDa, and that according to the two-dimensional electrophoresis and SE-HPLC analyses, a small amount of soluble aggregates might be formed via hydrophobic interactions. It was thus concluded that changes of protein structure caused by boiling that might influence beer quality are largely independent of malt variety.

External and internal gelation of pectin solutions: microscopic dynamics versus macroscopic rheology

NASA Astrophysics Data System (ADS)

Secchi, E.; Munarin, F.; Alaimo, M. D.; Bosisio, S.; Buzzaccaro, S.; Ciccarella, G.; Vergaro, V.; Petrini, P.; Piazza, R.

2014-11-01

Pectin is a natural biopolymer that forms, in the presence of divalent cations, ionic-bound gels typifying a large class of biological gels stabilized by non-covalent cross-links. We investigate and compare the kinetics of formation and aging of pectin gels obtained either through external gelation via perfusion of free Ca2+ ions, or by internal gelation due to the supply of the same ions from the dissolution of CaCO3 nanoparticles. The microscopic dynamics obtained with photon correlation imaging, a novel optical technique that allows obtaining the microscopic dynamics of the sample while retaining the spatial resolution of imaging techniques, is contrasted with macroscopic rheological measurements at constant strain. Pectin gelation is found to display peculiar two-stage kinetics, highlighted by non-monotonic growth in time of both microscopic correlations and gel mechanical strength. These results are compared to those found for alginate, another biopolymer extensively used in food formulation.

Reducing pain from palatal needle stick by topical anesthetics: a comparative study between two lidocaine/prilocaine substances.

PubMed

Al-Melh, Manal Abu; Andersson, Lars

2008-01-01

The aims of this study were to compare the topical anesthetic effect of two different preparations of lidocaine/prilocaine, cream versus thermosetting gel, on the reduction of pain from needle stick in the palate. The study also compared the subjects' preferences with regard to the substances. Two preparations of 2.5% lidocaine/2.5% prilocaine topical anesthetic agents were applied on the palatal mucosa in the canine region bilaterally and blindly to forty subjects. In the same visit, the cream (EMLA) was applied on one side and the thermosetting gel (Oraqix) on the other side, simultaneously. After that, a needle stick was given on each side using a 27-gauge needle inserted to bone contact through the palatal mucosa every second minute during a ten-minute period. Immediately after the needle stick, the subjects started recording their findings using a verbal scale, a visual analogue scale, and a questionnaire form. Significant differences were set at p < 0.05. Both substances reduced or eliminated pain from needle stick in the palate. Significant differences in pain reduction were observed between the two substances. EMLA reduced pain significantly better than Oraqix. With regard to comfort, taste, anesthetic effect, personal preference, and recommendation, the majority of the subjects favored the EMLA cream to the thermosetting gel. No adverse effects were reported by the subjects. Although both topical anesthetic agents are similar in content, the cream seemed to be preferred by the patients and reduced pain from needle stick in the palate significantly better than the thermosetting gel.

Versatility of azelaic acid 15% gel in treatment of inflammatory acne vulgaris.

PubMed

Thiboutot, Diane

2008-01-01

Azelaic acid (AzA) 15% gel is approved for the treatment of rosacea in the US, but also has approval for the treatment of acne vulgaris in many European countries where it has demonstrated success. Two randomized, multicenter, controlled clinical trials compared the effects of AzA 15% gel with those of topical benzoyl peroxide 5% or topical clindamycin 1%, all using a twice-daily dosing regimen. The primary endpoint in the intent-to-treat analysis was a reduction in inflammatory papules and pustules. AzA 15% gel resulted in a 70% to 71% median reduction of facial papules and pustules compared with a 77% reduction with benzoyl peroxide 5% gel and a 63% reduction with clindamycin. AzA 15% gel was well-tolerated. In addition, a 1-year European observational study conducted by dermatologists in private practice evaluated the safety and efficacy of AzA 15% gel used as monotherapy or in combination with other agents in more than 1200 patients with acne. Most physicians (81.9%) described an improvement in patients' symptoms after an average of 34.6 days, and 93.9% of physicians reported patient improvement after an average of 73.1 days. Both physicians and patients assessed AzA 15% gel to be effective with 74% of patients being "very satisfied" at the end of therapy. AzA 15% gel was considered "well-tolerated" or "very well-tolerated" by 95.7% of patients. The majority of patients were more satisfied with AzA than with previous therapies. AzA 15% gel represents a new therapeutic option for the treatment of acne vulgaris.

Role of Somatic Testicular Cells during Mouse Spermatogenesis in Three-Dimensional Collagen Gel Culture System

PubMed Central

Khajavi, Noushafarin; Akbari, Mohammad; Abolhassani, Farid; Dehpour, Ahmad Reza; Koruji, Morteza; Habibi Roudkenar, Mehryar

2014-01-01

Objective Spermatogonial stem cells (SSCs) are the only cell type that can restore fertility to an infertile recipient following transplantation. Much effort has been made to develop a protocol for differentiating isolated SSCs in vitro. Recently, three-dimensional (3D) culture system has been introduced as an appropriate microenvironment for clonal expansion and differentiation of SSCs. This system provides structural support and multiple options for several manipulation such as addition of different cells. Somatic cells have a critical role in stimulating spermatogenesis. They provide complex cell to cell interaction, transport proteins and produce enzymes and regulatory factors. This study aimed to optimize the culture condition by adding somatic testicular cells to the collagen gel culture system in order to induce spermatogenesis progression. Materials and Methods In this experimental study, the disassociation of SSCs was performed by using a two-step enzymatic digestion of type I collagenase, hyaluronidase and DNase. Somatic testicular cells including Sertoli cells and peritubular cells were obtained after the second digestion. SSCs were isolated by Magnetic Activated Cell Sorting (MACS) using GDNF family receptor alpha-1 (Gfrα-1) antibody. Two experimental designs were investigated. 1. Gfrα-1 positive SSCs were cultured in a collagen solution. 2. Somatic testicular cells were added to the Gfrα-1 positive SSCs in a collagen solution. Spermatogenesis progression was determined after three weeks by staining of synaptonemal complex protein 3 (SCP3)-positive cells. Semi-quantitative Reverse Transcription PCR was undertaken for SCP3 as a meiotic marker and, Crem and Thyroid transcription factor-1 (TTF1) as post meiotic markers. For statistical analysis student t test was performed. Results Testicular supporter cells increased the expression of meiotic and post meiotic markers and had a positive effect on extensive colony formation. Conclusion Collagen gel culture system supported by somatic testicular cells provides a microenvironment that mimics seminiferous epithelium and induces spermatogenesis in vitro. PMID:24518977

Comparison of topical anesthetics (EMLA/Oraqix vs. benzocaine) on pain experienced during palatal needle injection.

PubMed

Al-Melh, Manal Abu; Andersson, Lars

2007-05-01

The aim of this study was to compare the topical anesthetic effect of 20% benzocaine gel with 2.5% lidocaine/2.5% prilocaine (L/P) cream and gel on the pain experienced during palatal anesthetic infiltration. Two groups were studied, each containing 20 subjects. Two types of L/P mixtures were tested, an anesthetic cream (EMLA) and a thermosetting gel (Oraqix), and benzocaine was used as control. The topical agents were applied on the palatal mucosa at the canine region. A needle prick was given on each side every 2 minutes during a period of 10 minutes. The subjects recorded their findings using verbal and VAS scales. Pain scores were significantly less (P < .05) with EMLA and Oraqix than with benzocaine. Topical application of EMLA and Oraqix before palatal anesthetic infiltration is associated with less pain than with benzocaine gel.

A simplified model for equilibrium and transient swelling of thermo-responsive gels.

PubMed

Drozdov, A D; deClaville Christiansen, J

2017-11-01

A simplified model is developed for the elastic response of thermo-responsive gels subjected to swelling under an arbitrary deformation with finite strains. The constitutive equations involve five adjustable parameters that are determined by fitting observations in equilibrium water uptake tests and T-jump transient tests on thin gel disks. Two scenarios for water release under heating are revealed by means of numerical simulation. When the final temperature in a T-jump test is below the volume-phase transition temperature, deswelling is characterized by smooth distribution of water molecules and small tensile stresses. When the final temperature exceeds the critical temperature, a gel disk is split into three regions (central part with a high concentration of water molecules and two domains near the boundaries with low water content) separated by sharp interfaces, whose propagation is accompanied by development of large (comparable with the elastic modulus) tensile stresses. Copyright © 2017 Elsevier Ltd. All rights reserved.

Automation in clinical microbiology: a new approach to identifying micro-organisms by automated pattern matching of proteins labelled with 35S-methionine.

PubMed Central

Tabaqchali, S; Silman, R; Holland, D

1987-01-01

A new rapid automated method for the identification and classification of microorganisms is described. It is based on the incorporation of 35S-methionine into cellular proteins and subsequent separation of the radiolabelled proteins by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The protein patterns produced were species specific and reproducible, permitting discrimination between the species. A large number of Gram negative and Gram positive aerobic and anaerobic organisms were successfully tested. Furthermore, there were sufficient differences within species between the protein profiles to permit subdivision of the species. New typing schemes for Clostridium difficile, coagulase negative staphylococci, and Staphylococcus aureus, including the methicillin resistant strains, could thus be introduced; this has provided the basis for useful epidemiological studies. To standardise and automate the procedure an automated electrophoresis system and a two dimensional scanner were developed to scan the dried gels directly. The scanner is operated by a computer which also stores and analyses the scan data. Specific histograms are produced for each bacterial species. Pattern recognition software is used to construct databases and to compare data obtained from different gels: in this way duplicate "unknowns" can be identified. Specific small areas showing differences between various histograms can also be isolated and expanded to maximise the differences, thus providing differentiation between closely related bacterial species and the identification of differences within the species to provide new typing schemes. This system should be widely applied in clinical microbiology laboratories in the near future. Images Fig 1 Fig 2 Fig 3 Fig 4 Fig 5 Fig 6 Fig 7 Fig 8 PMID:3312300

Non-invasive detection of candidate pregnancy protein biomarkers in the feces of captive polar bears (Ursus maritimus).

PubMed

Curry, E; Stoops, M A; Roth, T L

2012-07-15

Currently, there is no method of accurately and non-invasively diagnosing pregnancy in polar bears. Specific proteins may exhibit altered profiles in the feces of pregnant bears, but predicting appropriate candidate proteins to investigate is speculative at best. The objective of this study was to identify potential pregnancy biomarker proteins based on their increased abundance in the feces of pregnant polar bears compared to pseudopregnant females (controls) using two-dimensional in-gel electrophoresis (2D-DIGE) and mass spectrometry (MS). Three 2D-DIGE gels were performed to evaluate fecal protein profiles from controls (n=3) and pregnant polar bears (n=3). There were 2224.67±52.39 (mean±SEM) spots resolved per gel. Of these, only five proteins were elevated in the pregnant group (P<0.05), and seven additional spots tended to be higher (0.0599.9% confidence interval. The 11 spots represented seven distinct proteins, five of which were significantly more abundant in the pregnant group: IgGFc-binding protein, filamin-C, carboxypeptidase B, transthyretin, and immunoglobulin heavy chain variable region. To our knowledge, this was the first study that employed 2D-DIGE to identify differentially expressed proteins in fecal samples to characterize a physiological condition other than those related to gastrointestinal disorders. These promising results provided a strong foundation for ensuing efforts to develop a non-invasive pregnancy assay for use in both captive and wild polar bears. Copyright © 2012 Elsevier Inc. All rights reserved.

Comparison of Effectiveness of Betamethasone gel Applied to the Tracheal Tube and IV Dexamethasone on Postoperative Sore Throat: A Randomized Controlled Trial.

PubMed

Tabari, Masumeh; Soltani, Ghasem; Zirak, Nahid; Alipour, Moammad; Khazaeni, Kamran

2013-09-01

Postoperative sore throat is a common complaint in patients with endotracheal intubation and has potentially dangerous complications. This randomized controlled trial study investigated the incidence of postoperative sore throat after general anesthesia when betamethasone gel is applied to a tracheal tube compared with when IV dexamethasone is prescribed. Two hundred and twenty five American Society of Anesthesiologist (ASA)-class I and II patients undergoing elective abdominal surgery with tracheal intubation were randomly divided into three groups: betamethasone gel, intravenous (IV) dexamethasone, and control groups. In the post-anesthesia care unit, a blinded anesthesiologist interviewed all patients regarding postoperative sore throat at 1,6, and 24 hours after surgery. The incidence of sore throat was significantly lower in the betamethasone gel group compared with the IV dexamethasone and control groups, 1, 6, and 24 hours after surgery. In the first day after surgery 10.7% of the betamethasone group had sore throat whereas 26.7% of the IV dexamethasone group and 30.7% of the control group had sore throat. Bucking before extubation was observed in 14(18.4%), 8(10.4%), and 9(12.2%) patients, in the IV dexamethasone, betamethasone gel, and control group, respectively. We concluded that wide spread application of betamethasone gel over tracheal tubes effectively mitigates postoperative sore throat, compared with IV dexamethasone application.

Comparison of polyacrylamide and agarose gel thin-layer isoelectric focusing for the characterization of beta-lactamases.

PubMed

Vecoli, C; Prevost, F E; Ververis, J J; Medeiros, A A; O'Leary, G P

1983-08-01

Plasmid-mediated beta-lactamases from strains of Escherichia coli and Pseudomonas aeruginosa were separated by isoelectric focusing on a 0.8-mm thin-layer agarose gel with a pH gradient of 3.5 to 9.5. Their banding patterns and isoelectric points were compared with those obtained with a 2.0-mm polyacrylamide gel as the support medium. The agarose method produced banding patterns and isoelectric points which corresponded to the polyacrylamide gel data for most samples. Differences were observed for HMS-1 and PSE-1 beta-lactamases. The HMS-1 sample produced two highly resolvable enzyme bands in agarose gels rather than the single faint enzyme band observed on polyacrylamide gels. The PSE-1 sample showed an isoelectric point shift of 0.2 pH unit between polyacrylamide and agarose gel (pI 5.7 and 5.5, respectively). The short focusing time, lack of toxic hazard, and ease of formulation make agarose a practical medium for the characterization of beta-lactamases.

Comparison of polyacrylamide and agarose gel thin-layer isoelectric focusing for the characterization of beta-lactamases.

PubMed Central

Vecoli, C; Prevost, F E; Ververis, J J; Medeiros, A A; O'Leary, G P

1983-01-01

Plasmid-mediated beta-lactamases from strains of Escherichia coli and Pseudomonas aeruginosa were separated by isoelectric focusing on a 0.8-mm thin-layer agarose gel with a pH gradient of 3.5 to 9.5. Their banding patterns and isoelectric points were compared with those obtained with a 2.0-mm polyacrylamide gel as the support medium. The agarose method produced banding patterns and isoelectric points which corresponded to the polyacrylamide gel data for most samples. Differences were observed for HMS-1 and PSE-1 beta-lactamases. The HMS-1 sample produced two highly resolvable enzyme bands in agarose gels rather than the single faint enzyme band observed on polyacrylamide gels. The PSE-1 sample showed an isoelectric point shift of 0.2 pH unit between polyacrylamide and agarose gel (pI 5.7 and 5.5, respectively). The short focusing time, lack of toxic hazard, and ease of formulation make agarose a practical medium for the characterization of beta-lactamases. Images PMID:6605714

Cutaneous wound healing after treatment with plant-derived human recombinant collagen flowable gel.

PubMed

Shilo, Shani; Roth, Sigal; Amzel, Tal; Harel-Adar, Tamar; Tamir, Eran; Grynspan, Frida; Shoseyov, Oded

2013-07-01

Chronic wounds, particularly diabetic ulcers, represent a main public health concern with significant costs. Ulcers often harbor an additional obstacle in the form of tunneled or undermined wounds, requiring treatments that can reach the entire wound tunnel, because bioengineered grafts are typically available only in a sheet form. While collagen is considered a suitable biodegradable scaffold material, it is usually extracted from animal and human cadaveric sources, and accompanied by potential allergic and infectious risks. The purpose of this study was to test the performance of a flowable gel made of human recombinant type I collagen (rhCollagen) produced in transgenic tobacco plants, indicated for the treatment of acute, chronic, and tunneled wounds. The performance of the rhCollagen flowable gel was tested in an acute full-thickness cutaneous wound-healing rat model and compared to saline treatment and two commercial flowable gel control products made of bovine collagen and cadaver human skin collagen. When compared to the three control groups, the rhCollagen-based gel accelerated wound closure and triggered a significant jumpstart to the healing process, accompanied by enhanced re-epithelialization. In a cutaneous full-thickness wound pig model, the rhCollagen-based flowable gel induced accelerated wound healing compared to a commercial product made of bovine tendon collagen. By day 21 post-treatment, 95% wound closure was observed with the rhCollagen product compared to 68% closure in wounds treated with the reference product. Moreover, rhCollagen treatment induced an early angiogenic response and induced a significantly lower inflammatory response than in the control group. In summary, rhCollagen flowable gel proved to be efficacious in animal wound models and is expected to be capable of reducing the healing time of human wounds.

Cutaneous Wound Healing After Treatment with Plant-Derived Human Recombinant Collagen Flowable Gel

PubMed Central

Roth, Sigal; Amzel, Tal; Harel-Adar, Tamar; Tamir, Eran; Grynspan, Frida; Shoseyov, Oded

2013-01-01

Chronic wounds, particularly diabetic ulcers, represent a main public health concern with significant costs. Ulcers often harbor an additional obstacle in the form of tunneled or undermined wounds, requiring treatments that can reach the entire wound tunnel, because bioengineered grafts are typically available only in a sheet form. While collagen is considered a suitable biodegradable scaffold material, it is usually extracted from animal and human cadaveric sources, and accompanied by potential allergic and infectious risks. The purpose of this study was to test the performance of a flowable gel made of human recombinant type I collagen (rhCollagen) produced in transgenic tobacco plants, indicated for the treatment of acute, chronic, and tunneled wounds. The performance of the rhCollagen flowable gel was tested in an acute full-thickness cutaneous wound-healing rat model and compared to saline treatment and two commercial flowable gel control products made of bovine collagen and cadaver human skin collagen. When compared to the three control groups, the rhCollagen-based gel accelerated wound closure and triggered a significant jumpstart to the healing process, accompanied by enhanced re-epithelialization. In a cutaneous full-thickness wound pig model, the rhCollagen-based flowable gel induced accelerated wound healing compared to a commercial product made of bovine tendon collagen. By day 21 post-treatment, 95% wound closure was observed with the rhCollagen product compared to 68% closure in wounds treated with the reference product. Moreover, rhCollagen treatment induced an early angiogenic response and induced a significantly lower inflammatory response than in the control group. In summary, rhCollagen flowable gel proved to be efficacious in animal wound models and is expected to be capable of reducing the healing time of human wounds. PMID:23259631

Cassava flour slurry as a low-cost alternative to commercially available gel for obstetrical ultrasound: a blinded non-inferiority trial comparison of image quality.

PubMed

Aziz, A; Dar, P; Hughes, F; Solorzano, C; Muller, M M; Salmon, C; Salmon, M; Benfield, N

2018-01-12

To evaluate the quality of ultrasound images obtained with cassava flour slurry (CFS) compared with conventional gel in order to determine objectively whether CFS could be a true low-cost alternative. Blinded non-inferiority trial. Obstetrical ultrasound unit in an academic medical centre. Women with a singleton pregnancy, undergoing anatomy ultrasounds. Thirty pregnant women had standard biometry measures obtained with CFS and conventional gel. Images were compared side-by-side in random order by two blinded sonologists and rated for image resolution, detail and total image quality using a 10-cm visual analogue scale. Ratings were compared using paired t-tests. Participant and sonographer experience was measured using five-point Likert scales. Image resolution, detail, and total image quality. Participant experience of gel regarding irritation, messiness, and ease of removal. We found no significant difference between perceived image quality obtained with CFS (mean = 6.2, SD = 1.2) and commercial gel (mean = 6.4, SD = 1.2) [t (28) = -1.1; P = 0.3]. Images were not rated significantly differently for either reviewer in any measure, any standardized image or any view of a specific anatomic structure. All five sonographers rated CFS as easy to obtain clear images and easy for patient and machine cleanup. Only one participant reported itching with CFS. CFS produces comparable image quality to commercial ultrasound gel. The dissemination of these results and the simple CFS recipe could significantly increase access to ultrasound for screening, monitoring and diagnostic purposes in resource-limited settings. This study was internally funded by our department. Low-cost homemade cassava flour slurry creates images equal to commercial ultrasound gel, improving access. © 2018 Royal College of Obstetricians and Gynaecologists.

Proteome of Caulobacter crescentus cell cycle publicly accessible on SWICZ server.

PubMed

Vohradsky, Jiri; Janda, Ivan; Grünenfelder, Björn; Berndt, Peter; Röder, Daniel; Langen, Hanno; Weiser, Jaroslav; Jenal, Urs

2003-10-01

Here we present the Swiss-Czech Proteomics Server (SWICZ), which hosts the proteomic database summarizing information about the cell cycle of the aquatic bacterium Caulobacter crescentus. The database provides a searchable tool for easy access of global protein synthesis and protein stability data as examined during the C. crescentus cell cycle. Protein synthesis data collected from five different cell cycle stages were determined for each protein spot as a relative value of the total amount of [(35)S]methionine incorporation. Protein stability of pulse-labeled extracts were measured during a chase period equivalent to one cell cycle unit. Quantitative information for individual proteins together with descriptive data such as protein identities, apparent molecular masses and isoelectric points, were combined with information on protein function, genomic context, and the cell cycle stage, and were then assembled in a relational database with a world wide web interface (http://proteom.biomed.cas.cz), which allows the database records to be searched and displays the recovered information. A total of 1250 protein spots were reproducibly detected on two-dimensional gel electropherograms, 295 of which were identified by mass spectroscopy. The database is accessible either through clickable two-dimensional gel electrophoretic maps or by means of a set of dedicated search engines. Basic characterization of the experimental procedures, data processing, and a comprehensive description of the web site are presented. In its current state, the SWICZ proteome database provides a platform for the incorporation of new data emerging from extended functional studies on the C. crescentus proteome.

Gene expression in the pulp of ripening bananas. Two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis of in vitro translation products and cDNA cloning of 25 different ripening-related mRNAs.

PubMed Central

Medina-Suárez, R; Manning, K; Fletcher, J; Aked, J; Bird, C R; Seymour, G B

1997-01-01

mRNA was extracted from the pulp and peel of preclimacteric (d 0) bananas (Musa AAA group, cv Grand Nain) and those exposed to ethylene gas for 24 h and stored in air alone for a further 1 (d 2) and 4 d (d 5). Two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis of in vitro translation products from the pulp and peel of these fruits revealed significant up-regulation of numerous transcripts during ripening. The majority of the changes were initiated by d 2, with the level of these messages increasing during the remainder of the ripening period. Pulp tissue from d 2 was used for the construction of a cDNA library. This library was differentially screened for ripening-related clones using cDNA from d-0 and d-2 pulp by a novel microtiter plate method. In the primary screen 250 up- and down-regulated clones were isolated. Of these, 59 differentially expressed clones were obtained from the secondary screen. All of these cDNAs were partially sequenced and grouped into families after database searches. Twenty-five nonredundant groups of pulp clones were identified. These encoded enzymes were involved in ethylene biosynthesis, respiration, starch metabolism, cell wall degradation, and several other key metabolic events. We describe the analysis of these clones and their possible involvement in ripening. PMID:9342865

Connectivity of glass structure. Oxygen number

NASA Astrophysics Data System (ADS)

Medvedev, E. F.; Min'ko, N. I.

2018-03-01

With reference to mathematics, crystal chemistry and chemical technology of synthesis of glass structures in the solution (sol-gel technology), the paper is devoted to the study of the degree of connectivity of a silicon-oxygen backbone (fSi) and the oxygen number (R) [1]. It reveals logical contradictions and uncertainty of mathematical expressions of parameters, since fSi is not similar to the oxygen number. The connectivity of any structure is a result of various types of bonds: ion-covalent, donor-acceptor, hydrogen bonds, etc. Besides, alongside with SiO2, many glass compositions contain other glass-forming elements due to tetrahedral sites thus formed. The connectivity function of a glassy network with any set of glass-forming elements is roughly ensured by connectivity factor Y [2], which has monovalent elements loosening a glassy network. The paper considers the existence of various structural motives in hydrogen-impermeable glasses containing B2O3, Al2O3, PbO, Na2O, K2O and rare-earth elements. Hence, it also describes gradual nucleation, change of crystal forms, and structure consolidation in the process of substance intake from a matrix solution according to sol-gel technology. The crystal form varied from two-dimensional plates to three-dimensional and dendritical ones [3]. Alternative parameters, such as the oxygen number (O) and the structure connectivity factor (Y), were suggested. Functional dependence of Y=f(O) to forecast the generated structures was obtained for two- and multicomponent glass compositions.

Bioinspired Tuning of Hydrogel Permeability-Rigidity Dependency for 3D Cell Culture

NASA Astrophysics Data System (ADS)

Lee, Min Kyung; Rich, Max H.; Baek, Kwanghyun; Lee, Jonghwi; Kong, Hyunjoon

2015-03-01

Hydrogels are being extensively used for three-dimensional immobilization and culture of cells in fundamental biological studies, biochemical processes, and clinical treatments. However, it is still a challenge to support viability and regulate phenotypic activities of cells in a structurally stable gel, because the gel becomes less permeable with increasing rigidity. To resolve this challenge, this study demonstrates a unique method to enhance the permeability of a cell-laden hydrogel while avoiding a significant change in rigidity of the gel. Inspired by the grooved skin textures of marine organisms, a hydrogel is assembled to present computationally optimized micro-sized grooves on the surface. Separately, a gel is engineered to preset aligned microchannels similar to a plant's vascular bundles through a uniaxial freeze-drying process. The resulting gel displays significantly increased water diffusivity with reduced changes of gel stiffness, exclusively when the microgrooves and microchannels are aligned together. No significant enhancement of rehydration is achieved when the microgrooves and microchannels are not aligned. Such material design greatly enhances viability and neural differentiation of stem cells and 3D neural network formation within the gel.

Sol-gel technique for the preparation of beta-cyclodextrin derivative stationary phase in open-tubular capillary electrochromatography.

PubMed

Wang, Y; Zeng, Z; Guan, N; Cheng, J

2001-07-01

A novel open-tubular capillary electrochromatography (OT-CEC) column coated with 2,6-dibutyl-beta-cyclodextrin (DB-beta-CD) was prepared using sol-gel technique. In the sol-gel approach, owing to the three-dimensional network of sol-gel and the strong chemical bond between the stationary phase and the surface of capillary columns, good chromatographic characteristics and unique selectivity in separating isomers were shown. We achieved high efficiencies of 5-14 x 10(4) plates/m for the isomeric nitrophenols using the sol-gel-derived DB-beta-CD columns. The migration time reproducibility of the separation of the isomeric nitrophenols was better than 2.2% over five runs and 4.5% from column to column. These sol-gel-coated DB-beta-CD columns have shown improved separations of isomeric aminophenols, isomeric dihydroxybenzenes and isomeric nitrophenols, in comparison with the sol-gel matrix capillary column. The influences of buffer pH and methanol solvent on separation were investigated. The chiral resolution of enantiomers such as ibuprofen and binaphthol was explored primarily.

RNA-primed complementary-sense DNA synthesis of the geminivirus African cassava mosaic virus.

PubMed Central

Saunders, K; Lucy, A; Stanley, J

1992-01-01

The plant DNA virus African cassava mosaic virus (ACMV) is believed to replicate by a rolling circle mechanism. To investigate complementary-sense DNA (lagging strand) synthesis, we have analysed the heterogenous form of complementary-sense DNA (H3 DNA) from infected Nicotiana benthamiana by two-dimensional agarose gel electrophoresis and blot hybridisation. The presence of an RNA moeity is demonstrated by comparison of results for nucleic acids resolved on neutral/alkaline and neutral/formamide gels, suggesting that complementary-sense DNA synthesis on the virus-sense single-stranded DNA template is preceded by the synthesis of an RNA primer. Hybridisation with probes to specific parts of ACMV DNA A genome indicates that synthesis of the putative RNA primer initiates between nucleotides 2581-221, a region that includes intergenic sequences that have been implicated in geminivirus DNA replication and the control of gene expression. Images PMID:1475192

Non-volatile copolymer compositions for fabricating gel element microarrays

PubMed Central

Golova, Julia B.; Chernov, Boris K.; Perov, Alexander N.; Reynolds, Jennifer; Linger, Yvonne L.; Kukhtin, Alexander; Chandler, Darrell P.

2011-01-01

By modifying polymer compositions and cross-linking reagents, we have developed a simple yet effective manufacturing strategy for copolymerized three-dimensional gel element arrays. A new gel-forming monomer (2-(hydroxyethyl) methacrylamide; HEMAA) was used that possesses low volatility and improves the stability of copolymerized gel element arrays to on-chip thermal cycling procedures relative to previously used monomers. Probe immobilization efficiency within the new polymer was 55%, equivalent to that obtained with acrylamide (AA) and methacrylamide (MA) monomers. Non-specific binding of single stranded targets was equivalent for all monomers. Increasing cross-linker chain length improved hybridization kinetics and end-point signal intensities relative to N,N-methylenebisacrylamide (Bis). The new copolymer formulation was successfully applied to a model orthopox array. Because HEMAA greatly simplifies gel element array manufacture, we expect it (in combination with new cross-linkers described herein) to find widespread application in microarray science. PMID:22033291

Porous alumina scaffold produced by sol-gel combined polymeric sponge method

NASA Astrophysics Data System (ADS)

Hasmaliza, M.; Fazliah, M. N.; Shafinaz, R. J.

2012-09-01

Sol gel is a novel method used to produce high purity alumina with nanometric scale. In this study, three-dimensional porous alumina scaffold was produced using sol-gel polymeric sponge method. Briefly, sol gel alumina was prepared by evaporation and polymeric sponge cut to designated sizes were immersed in the sol gel followed by sintering at 1250 and 1550°C. In order to study the cell interaction, the porous alumina scaffold was sterilized using autoclave prior to Human Mesenchymal Stem Cells (HMSCs) seeding on the scaffold and the cell proliferation was assessed by alamarBlue® assay. SEM results showed that during the 21 day period, HMSCs were able to attach on the scaffold surface and the interconnecting pores while maintaining its proliferation. These findings suggested the potential use of the porous alumina produced as a scaffold for implantation procedure.

Comparative proteomics analysis of oral cancer cell lines: identification of cancer associated proteins

PubMed Central

2014-01-01

Background A limiting factor in performing proteomics analysis on cancerous cells is the difficulty in obtaining sufficient amounts of starting material. Cell lines can be used as a simplified model system for studying changes that accompany tumorigenesis. This study used two-dimensional gel electrophoresis (2DE) to compare the whole cell proteome of oral cancer cell lines vs normal cells in an attempt to identify cancer associated proteins. Results Three primary cell cultures of normal cells with a limited lifespan without hTERT immortalization have been successfully established. 2DE was used to compare the whole cell proteome of these cells with that of three oral cancer cell lines. Twenty four protein spots were found to have changed in abundance. MALDI TOF/TOF was then used to determine the identity of these proteins. Identified proteins were classified into seven functional categories – structural proteins, enzymes, regulatory proteins, chaperones and others. IPA core analysis predicted that 18 proteins were related to cancer with involvements in hyperplasia, metastasis, invasion, growth and tumorigenesis. The mRNA expressions of two proteins – 14-3-3 protein sigma and Stress-induced-phosphoprotein 1 – were found to correlate with the corresponding proteins’ abundance. Conclusions The outcome of this analysis demonstrated that a comparative study of whole cell proteome of cancer versus normal cell lines can be used to identify cancer associated proteins. PMID:24422745

The Two-Component System PhoPR of Clostridium acetobutylicum Is Involved in Phosphate-Dependent Gene Regulation ▿

PubMed Central

Fiedler, Tomas; Mix, Maren; Meyer, Uta; Mikkat, Stefan; Glocker, Michael O.; Bahl, Hubert; Fischer, Ralf-Jörg

2008-01-01

The phoPR gene locus of Clostridium acetobutylicum ATCC 824 comprises two genes, phoP and phoR. Deduced proteins are predicted to represent a response regulator and sensor kinase of a phosphate-dependent two-component regulatory system. We analyzed the expression patterns of phoPR in Pi-limited chemostat cultures and in response to Pi pulses. A basic transcription level under high-phosphate conditions was shown, and a significant increase in mRNA transcript levels was found when external Pi concentrations dropped below 0.3 mM. In two-dimensional gel electrophoresis experiments, a 2.5-fold increase in PhoP was observed under Pi-limiting growth conditions compared to growth with an excess of Pi. At least three different transcription start points for phoP were determined by primer extension analyses. Proteins PhoP and an N-terminally truncated *PhoR were individually expressed heterologously in Escherichia coli and purified. Autophosphorylation of *PhoR and phosphorylation of PhoP were shown in vitro. Electromobility shift assays proved that there was a specific binding of PhoP to the promoter region of the phosphate-regulated pst operon of C. acetobutylicum. PMID:18689481

Dosimetric verification of small fields in the lung using lung-equivalent polymer gel and Monte Carlo simulation.

PubMed

Gharehaghaji, Nahideh; Dadgar, Habib Alah

2018-01-01

The main purpose of this study was evaluate a polymer-gel-dosimeter (PGD) for three-dimensional verification of dose distributions in the lung that is called lung-equivalent gel (LEG) and then to compare its result with Monte Carlo (MC) method. In the present study, to achieve a lung density for PGD, gel is beaten until foam is obtained, and then sodium dodecyl sulfate is added as a surfactant to increase the surface tension of the gel. The foam gel was irradiated with 1 cm × 1 cm field size in the 6 MV photon beams of ONCOR SIEMENS LINAC, along the central axis of the gel. The LEG was then scanned on a 1.5 Tesla magnetic resonance imaging scanner after irradiation using a multiple-spin echo sequence. Least-square fitting the pixel values from 32 consecutive images using a single exponential decay function derived the R2 relaxation rates. Moreover, 6 and 18 MV photon beams of ONCOR SIEMENS LINAC are simulated using MCNPX MC Code. The MC model is used to calculate the depth dose water and low-density water resembling the soft tissue and lung, respectively. Percentages of dose reduction in the lung region relative to homogeneous phantom for 6 MV photon beam were 44.6%, 39%, 13%, and 7% for 0.5 cm × 0.5 cm, 1 cm × 1 cm, 2 cm × 2 cm, and 3 cm × 3 cm fields, respectively. For 18 MV photon beam, the results were found to be 82%, 69%, 46%, and 25.8% for the same field sizes, respectively. Preliminary results show good agreement between depth dose measured with the LEG and the depth dose calculated using MCNP code. Our study showed that the dose reduction with small fields in the lung was very high. Thus, inaccurate prediction of absorbed dose inside the lung and also lung/soft-tissue interfaces with small photon beams may lead to critical consequences for treatment outcome.

A comparison of a refrigerant and a topical anesthetic gel as preinjection anesthetics: a clinical evaluation.

PubMed

Kosaraju, Amar; Vandewalle, Kraig S

2009-01-01

The authors used a split-mouth design to determine the effectiveness of a refrigerant compared with that of a topical anesthetic gel in reducing the pain experienced during a posterior palatal anesthetic injection. Sixteen participants received a five-second application of a refrigerant (1,1,1,3,3-pentafluoropropane/1,1,1,2-tetrafluoroethane) and a two-minute application of a topical anesthetic gel (20 percent benzocaine gel) in the posterior palatal area before an injection of a local anesthetic solution was administered with a 30-gauge needle. Participants rated the pain they experienced after each injection by using a 100-millimeter visual analog scale (VAS) with endpoints of "no pain" and "worst possible pain." The authors calculated VAS scores by measuring the distance in millimeters from the no pain end of the scale. They analyzed data with a paired t test (alpha = .05). The group receiving the refrigerant had a mean VAS score of 17.7 +/- 15.3 mm, and the group receiving the topical anesthetic gel had a VAS score of 26.2 +/- 18.0 mm. The use of the refrigerant compared with the use of topical anesthetic gel significantly reduced the pain experienced during administration of local anesthetic injections (P = .02). The use of a refrigerant as a preinjection anesthetic was more effective compared with the use of a topical anesthetic gel in reducing the pain experienced by participants who received a posterior palatal injection. The potential benefits of using a refrigerant rather than a topical anesthetic gel are pain reduction, decreased application time, ease of application and avoidance of displeasing taste.

Combining strategies to optimize a gel formulation containing miconazole: the influence of modified cyclodextrin on textural properties and drug release.

PubMed

Ribeiro, Andreza Maria; Figueiras, Ana; Freire, Cristina; Santos, Delfim; Veiga, Francisco

2010-06-01

Miconazol, an antimycotic drug, is commonly formulated into semisolid formulations designed to be applied in the oral cavity to treat oral candidiasis. However, given its limited aqueous solubility, permeation through the biological membranes is low and therefore its activity is also limited. Cyclodextrins (CDs) have been widely used to increase the solubility and stability of poorly water-soluble drugs. The aim of this study is to formulate a gel containing an inclusion complex between a modified CD, methyl-beta-cyclodextrin (MbetaCD), and miconazole (MCZ). The influence of the CD on the textural properties of the prepared gel and the drug release from formulation were evaluated. The gels were prepared using two polymers, Carbopol 71G and Pluronic F127, which were selected taking into account their bioadhesiveness and thermal-sensitive gelling properties, respectively. Texture profile analyses were performed at two different temperatures to ascertain the influence of the temperature on the gel texture properties. The in vitro MCZ release profiles from the prepared gel and the commercial gel formulations were evaluated and compared using modified Franz diffusion cells. The addition of MbetaCD to the gel resulted in a decrease of the gel adhesiveness and firmness, and the MCZ release profile through f1 and f2 proved to be similar to the commercial product. A gel comprising miconazol in the form of an inclusion complex with MbetaCD showed suitable textural properties to be applied to the buccal mucosa. The MbetaCD enhanced the solubility of the MCZ in the gel formulation resulting in adequate in vitro drug release profiles.

Further considerations on the thermal stabilization of the nuclear matrix in mouse erythroleukemia cells.

PubMed

Martelli, A M; Falcieri, E; Gobbi, P; Manzoli, L; Cataldi, A; Rana, R A; Cocco, L

1992-04-01

The morphology and the polypeptide composition of the nuclear matrix obtained from 37 degrees C incubated nuclei has been studied in mouse erythroleukemia cells. From a structural point of view, in the absence of heat treatment, the matrix lacked identifiable nucleolar remnants and the internal fibrogranular meshwork whereas a peripheral lamina was seen. On the contrary, the matrix obtained from heat exposed nuclei displayed very electrondense nucleolar remnants and an abundant inner network. These results were obtained irrespective of the type of extracting agent (2M NaCl or 0.2 M (NH4)2SO4) used to remove histones and other soluble proteins. The heat stabilization of the matrix could not be prevented by sulfhydryl blocking chemicals such as iodoacetamide and n-ethylmaleimide, thus suggesting that heat does not stabilize the matrix by inducing the formation of disulfide bonds. Only limited differences in the polypeptide pattern of matrix isolated under different conditions were seen using one-dimensional pore gradient polyacrylamide gels stained with both Coomassie Brilliant Blue and silver despite the fact that the matrix fraction from heat treated nuclei retained about three fold more protein in comparison with controls. The same results were obtained also by means of two-dimensional non-equilibrium gel electrophoresis.

Bioimaging of metals in brain tissue by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) and metallomics.

PubMed

Becker, J Sabine; Matusch, Andreas; Palm, Christoph; Salber, Dagmar; Morton, Kathryn A; Becker, J Susanne

2010-02-01

Laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) has been developed and established as an emerging technique in the generation of quantitative images of metal distributions in thin tissue sections of brain samples (such as human, rat and mouse brain), with applications in research related to neurodegenerative disorders. A new analytical protocol is described which includes sample preparation by cryo-cutting of thin tissue sections and matrix-matched laboratory standards, mass spectrometric measurements, data acquisition, and quantitative analysis. Specific examples of the bioimaging of metal distributions in normal rodent brains are provided. Differences to the normal were assessed in a Parkinson's disease and a stroke brain model. Furthermore, changes during normal aging were studied. Powerful analytical techniques are also required for the determination and characterization of metal-containing proteins within a large pool of proteins, e.g., after denaturing or non-denaturing electrophoretic separation of proteins in one-dimensional and two-dimensional gels. LA-ICP-MS can be employed to detect metalloproteins in protein bands or spots separated after gel electrophoresis. MALDI-MS can then be used to identify specific metal-containing proteins in these bands or spots. The combination of these techniques is described in the second section.

Embryonic stem cells growing in 3-dimensions shift from reliance on the substrate to each other for mechanical support.

PubMed

Teo, Ailing; Lim, Mayasari; Weihs, Daphne

2015-07-16

Embryonic stem cells (ESCs) grow into three-dimensional (3D) spheroid structures en-route to tissue growth. In vitro spheroids can be controllably induced on a two-dimensional (2D) substrate with high viability. Here we use a method for inducing pluripotent embryoid body (EB) formation on flat polyacrylamide gels while simultaneously evaluating the dynamic changes in the mechano-biology of the growing 3D spheroids. During colony growth in 3D, pluripotency is conserved while the spheroid-substrate interactions change significantly. We correlate colony-size, cell-applied traction-forces, and expressions of cell-surface molecules indicating cell-cell and cell-substrate interactions, while verifying pluripotency. We show that as the colony size increases with time, the stresses applied by the spheroid to the gel decrease in the 3D growing EBs; control cells growing in 2D-monolayers maintain unvarying forces. Concurrently, focal-adhesion mediated cell-substrate interactions give way to E-cadherin cell-cell connections, while pluripotency. The mechano-biological changes occurring in the growing embryoid body are required for stabilization of the growing pluripotent 3D-structure, and can affect its potential uses including differentiation. This could enable development of more effective expansion, differentiation, and separation approaches for clinical purposes. Copyright © 2015 Elsevier Ltd. All rights reserved.

Detection of Macromolecular Fractions in HCN Polymers Using Electrophoretic and Ultrafiltration Techniques.

PubMed

Marín-Yaseli, Margarita R; Cid, Cristina; Yagüe, Ana I; Ruiz-Bermejo, Marta

2017-02-01

Elucidating the origin of life involves synthetic as well as analytical challenges. Herein, for the first time, we describe the use of gel electrophoresis and ultrafiltration to fractionate HCN polymers. Since the first prebiotic synthesis of adenine by Oró, HCN polymers have gained much interest in studies on the origins of life due to the identification of biomonomers and related compounds within them. Here, we demonstrate that macromolecular fractions with electrophoretic mobility can also be detected within HCN polymers. The migration of polymers under the influence of an electric field depends not only on their sizes (one-dimensional electrophoresis) but also their different isoelectric points (two-dimensional electrophoresis, 2-DE). The same behaviour was observed for several macromolecular fractions detected in HCN polymers. Macromolecular fractions with apparent molecular weights as high as 250 kDa were detected by tricine-SDS gel electrophoresis. Cationic macromolecular fractions with apparent molecular weights as high as 140 kDa were also detected by 2-DE. The HCN polymers synthesized were fractionated by ultrafiltration. As a result, the molecular weight distributions of the macromolecular fractions detected in the HCN polymers directly depended on the synthetic conditions used to produce these polymers. The implications of these results for prebiotic chemistry will be discussed. © 2017 Wiley-VHCA AG, Zurich, Switzerland.

A novel tantalum-based sol-gel packed microextraction syringe for highly specific enrichment of phosphopeptides in MALDI-MS applications.

PubMed

Çelikbıçak, Ömür; Atakay, Mehmet; Güler, Ülkü; Salih, Bekir

2013-08-07

A new tantalum-based sol-gel material was synthesized using a unique sol-gel synthesis pathway by PEG incorporation into the sol-gel structure without performing a calcination step. This improved its chemical and physical properties for the high capacity and selective enrichment of phosphopeptides from protein digests in complex biological media. The specificity of the tantalum-based sol-gel material for phosphopeptides was evaluated and compared with tantalum(V) oxide (Ta2O5) in different phosphopeptide enrichment applications. The tantalum-based sol-gel and tantalum(V) oxide were characterized in detail using FT-IR spectroscopy, X-ray diffraction (XRD) and scanning electron microscopy (SEM), and also using a surface area and pore size analyzer. In the characterization studies, the surface morphology, pore volume, crystallinity of the materials and PEG incorporation into the sol-gel structure to produce a more hydrophilic material were successfully demonstrated. The X-ray diffractograms of the two different materials were compared and it was noted that the broad signals of the tantalum-based sol-gel clearly represented the amorphous structure of the sol-gel material, which was more likely to create enough surface area and to provide more accessible tantalum atoms for phosphopeptides to be easily adsorbed when compared with the neat and more crystalline structure of Ta2O5. Therefore, the phosphopeptide enrichment performance of the tantalum-based sol-gels was found to be remarkably higher than the more crystalline Ta2O5 in our studies. Phosphopeptides at femtomole levels could be selectively enriched using the tantalum-based sol-gel and detected with a higher signal-to-noise ratio by matrix-assisted laser desorption/ionization-mass spectrometer (MALDI-MS). Moreover, phosphopeptides in a tryptic digest of non-fat bovine milk as a complex real-world biological sample were retained with higher yield using a tantalum-based sol-gel. Additionally, the sol-gel material was packed into a standard syringe (0.5 mL) to enhance the ease of use of the sol-gel material and for the elimination of additional mixing and separation procedures during the adsorption, washing and elution steps of the enrichment procedure. It was found that up to 28 phosphopeptides in milk digest were easily detectable by MALDI-MS at femtomole levels (around 20 fmol) using the microextraction syringe within less than one minute.

Synthesis and Characterization of Types A and B Gelatin Methacryloyl for Bioink Applications

PubMed Central

Lee, Bae Hoon; Lum, Nathaniel; Seow, Li Yuan; Lim, Pei Qi; Tan, Lay Poh

2016-01-01

Gelatin methacryloyl (GelMA) has been increasingly considered as an important bioink material due to its tailorable mechanical properties, good biocompatibility, and ability to be photopolymerized in situ as well as printability. GelMA can be classified into two types: type A GelMA (a product from acid treatment) and type B GelMA (a product from alkali treatment). In current literature, there is little research on the comparison of type A GelMA and type B GelMA in terms of synthesis, rheological properties, and printability for bioink applications. Here, we report the synthesis, rheological properties, and printability of types A and B GelMA. Types A and B GelMA samples with different degrees of substitution (DS) were prepared in a controllable manner by a time-lapse loading method of methacrylic anhydride (MAA) and different feed ratios of MAA to gelatin. Type B GelMA tended to have a slightly higher DS compared to type A GelMA, especially in a lower feed ratio of MAA to gelatin. All the type A and type B GelMA solutions with different DS exhibited shear thinning behaviours at 37 °C. However, only GelMA with a high DS had an easy-to-extrude feature at room temperature. The cell-laden printed constructs of types A and B GelMA at 20% w/v showed around 75% cell viability. PMID:28773918

Anti-hyperglycemic and anti-hypercholesterolemic effects of Aloe vera leaf gel in hyperlipidemic type 2 diabetic patients: a randomized double-blind placebo-controlled clinical trial.

PubMed

Huseini, Hasan Fallah; Kianbakht, Saeed; Hajiaghaee, Reza; Dabaghian, Fataneh Hashem

2012-03-01

Diabetes mellitus type 2 with dyslipidemia is a common disease. Previous studies suggest that aloe (Aloe vera L.) leaf gel may positively affect the blood glucose and lipid levels in dyslipidemic type 2 diabetic patients. Thus, in this randomized double-blind placebo-controlled clinical trial with hyperlipidemic (hypercholesterolemic and/or hypertriglyceridemic) type 2 diabetic patients aged 40 to 60 years not using other anti-hyperlipidemic agents and resistant to daily intake of two 5 mg glyburide tablets and two 500 mg metformin tablets, the efficacy and safety of taking aloe gel (one 300 mg capsule every 12 hours for 2 months) combined with the aforementioned drugs in treatment of 30 patients were evaluated and compared with the placebo group (n = 30). The aloe gel lowered the fasting blood glucose, HbA1c, total cholesterol, and LDL levels significantly (p = 0.036, p = 0.036, p = 0.006, and p = 0.004, respectively) without any significant effects on the other blood lipid levels and liver/kidney function tests (p > 0.05) compared with the placebo at the endpoint. No adverse effects were reported. The results suggest that aloe gel may be a safe anti-hyperglycemic and anti-hypercholesterolemic agent for hyperlipidemic type 2 diabetic patients. © Georg Thieme Verlag KG Stuttgart · New York.