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Sample records for mass labeled 13c14-decabromodiphenylethane

  1. Guided mass spectrum labelling in atom probe tomography.

    PubMed

    Haley, D; Choi, P; Raabe, D

    2015-12-01

    Atom probe tomography (APT) is a valuable near-atomic scale imaging technique, which yields mass spectrographic data. Experimental correctness can often pivot on the identification of peaks within a dataset, this is a manual process where subjectivity and errors can arise. The limitations of manual procedures complicate APT experiments for the operator and furthermore are a barrier to technique standardisation. In this work we explore the capabilities of computer-guided ranging to aid identification and analysis of mass spectra. We propose a fully robust algorithm for enumeration of the possible identities of detected peak positions, which assists labelling. Furthermore, a simple ranking scheme is developed to allow for evaluation of the likelihood of each possible identity being the likely assignment from the enumerated set. We demonstrate a simple, yet complete work-chain that allows for the conversion of mass-spectra to fully identified APT spectra, with the goal of minimising identification errors, and the inter-operator variance within APT experiments. This work chain is compared to current procedures via experimental trials with different APT operators, to determine the relative effectiveness and precision of the two approaches. It is found that there is little loss of precision (and occasionally gain) when participants are given computer assistance. We find that in either case, inter-operator precision for ranging varies between 0 and 2 "significant figures" (2σ confidence in the first n digits of the reported value) when reporting compositions. Intra-operator precision is weakly tested and found to vary between 1 and 3 significant figures, depending upon species composition levels. Finally it is suggested that inconsistencies in inter-operator peak labelling may be the largest source of scatter when reporting composition data in APT.

  2. Label-free peptide profiling of Orbitrap™ full mass spectra

    PubMed Central

    2011-01-01

    Background We developed a new version of the open source software package Peptrix that can yet compare large numbers of Orbitrap™ LC-MS data. The peptide profiling results for Peptrix on MS1 spectra were compared with those obtained from a small selection of open source and commercial software packages: msInspect, Sieve™ and Progenesis™. The properties compared in these packages were speed, total number of detected masses, redundancy of masses, reproducibility in numbers and CV of intensity, overlap of masses, and differences in peptide peak intensities. Reproducibility measurements were taken for the different MS1 software applications by measuring in triplicate a complex peptide mixture of immunoglobulin on the Orbitrap™ mass spectrometer. Values of peptide masses detected from the high intensity peaks of the MS1 spectra by peptide profiling were verified with values of the MS2 fragmented and sequenced masses that resulted in protein identifications with a significant score. Findings Peptrix finds about the same number of peptide features as the other packages, but peptide masses are in some cases approximately 5 to 10 times less redundant present in the peptide profile matrix. The Peptrix profile matrix displays the largest overlap when comparing the number of masses in a pair between two software applications. The overlap of peptide masses between software packages of low intensity peaks in the spectra is remarkably low with about 50% of the detected masses in the individual packages. Peptrix does not differ from the other packages in detecting 96% of the masses that relate to highly abundant sequenced proteins. MS1 peak intensities vary between the applications in a non linear way as they are not processed using the same method. Conclusions Peptrix is capable of peptide profiling using Orbitrap™ files and finding differential expressed peptides in body fluid and tissue samples. The number of peptide masses detected in Orbitrap™ files can be

  3. Probing Protein Structure by Amino Acid-Specific Covalent Labeling and Mass Spectrometry

    PubMed Central

    Mendoza, Vanessa Leah; Vachet, Richard W.

    2009-01-01

    For many years, amino acid-specific covalent labeling has been a valuable tool to study protein structure and protein interactions, especially for systems that are difficult to study by other means. These covalent labeling methods typically map protein structure and interactions by measuring the differential reactivity of amino acid side chains. The reactivity of amino acids in proteins generally depends on the accessibility of the side chain to the reagent, the inherent reactivity of the label and the reactivity of the amino acid side chain. Peptide mass mapping with ESI- or MALDI-MS and peptide sequencing with tandem MS are typically employed to identify modification sites to provide site-specific structural information. In this review, we describe the reagents that are most commonly used in these residue-specific modification reactions, details about the proper use of these covalent labeling reagents, and information about the specific biochemical problems that have been addressed with covalent labeling strategies. PMID:19016300

  4. Genetically encoded protein photocrosslinker with a transferable mass spectrometry-identifiable label

    PubMed Central

    Yang, Yi; Song, Haiping; He, Dan; Zhang, Shuai; Dai, Shizhong; Lin, Shixian; Meng, Rong; Wang, Chu; Chen, Peng R.

    2016-01-01

    Coupling photocrosslinking reagents with mass spectrometry has become a powerful tool for studying protein–protein interactions in living systems, but it still suffers from high rates of false-positive identifications as well as the lack of information on interaction interface due to the challenges in deciphering crosslinking peptides. Here we develop a genetically encoded photo-affinity unnatural amino acid that introduces a mass spectrometry-identifiable label (MS-label) to the captured prey proteins after photocrosslinking and prey–bait separation. This strategy, termed IMAPP (In-situ cleavage and MS-label transfer After Protein Photocrosslinking), enables direct identification of photo-captured substrate peptides that are difficult to uncover by conventional genetically encoded photocrosslinkers. Taking advantage of the MS-label, the IMAPP strategy significantly enhances the confidence for identifying protein–protein interactions and enables simultaneous mapping of the binding interface under living conditions. PMID:27460181

  5. [Nonradioactive iodine-labeled antibodies-inductively coupled plasma mass spectrometry for immunoassay].

    PubMed

    Li, Jing-Xi; Wang, Xiao-Ru; Zhuang, Zhi-Xia; Cui, Wei-Gang

    2010-03-01

    In the present study, the system of nonradioactive iodine-labeled-antibodies linking inductively coupled plasma mass spectrometry for immunoassay was reported. The goat-anti-Escherichia coli and goat anti rabbit were considered as simulant antigen and antibody respectively in order to establish a new method of immunoassay by inductively coupled plasma mass spectrometry which has the advantage of high sensitivity, low detection limit and preferable linearity range. During the experiment, the N-bromosuccinimide, a mild oxidant, was used to oxidize the non-radioactive iodine (127 I) that labeled the protein. The method of nonradioactive iodine labeled protein was established and the best labeling condition was explored. The compound of I was purified by Sephadex G50 column chromatography, then the stability and activity were examined. The results showed that the labeling program was simple, reaction time was within two minutes, the labeling yield achieved 63.12% and none of I shed from the compound after 96 hours. The simulant antigen and antibody reacted on polystyrene microtiter plate and the I was detected by ICP-MS, the detection limit of the method was 0.12 mg x L(-1), relative standard deviation (n = 9) was less than 3% and the linearly dependent coefficient was 0.998 7. This system can also be used in analysis of other protein, nucleic acid and so on.

  6. Effect of tumor mass and antigenic nature on the biodistribution of labeled monoclonal antibodies in mice

    SciTech Connect

    Watanabe, Y.; Endo, K.; Koizumi, M.; Kawamura, Y.; Saga, T.; Sakahara, H.; Kuroki, M.; Matsuoka, Y.; Konishi, J.

    1989-06-01

    The effect of tumor mass and antigenic nature on the biodistribution of 111In- and 125I-labeled monoclonal antibodies (MoAbs) was studied using F(ab')2 fragments of three representative anti-tumor MoAbs and SW1116 human colorectal carcinoma grown in nude mice. The 19-9, F33-104 anti-CEA, and 17-1A MoAbs showed specific binding to SW1116 cells. The former two MoAbs recognize circulating CA 19-9 with molecular weights of more than 5,000,000 and CEA of Mr 170,000-180,000, respectively, whereas 17-1A reacts with a nonshedding antigen. Both percentage injected dose per gram tumor and tumor-to-blood ratios were inversely proportional to the tumor mass in nude mice administered 111In- and 125I-labeled 19-9, but liver uptake increased as tumor size increased. Analysis of serum samples and tumor homogenates demonstrated the presence of a high-molecular-weight species, probably due to the antibody binding to CA 19-9. In the case of 111In-labeled anti-CEA MoAb, tumor uptake also decreased and liver uptake increased with tumor size, but this effect was less obvious than that of 19-9. In contrast, tumor and liver uptake of 125I-labeled anti-CEA MoAb, 111In- and 125I-labeled 17-1A and control antibodies were independent of tumor mass. The absolute tumor uptake and tumor-to-blood ratios of all 125I-labeled antibodies were lower than those of the 111In-labeled ones. And the effect of tumor mass was also weaker with 125I-labeled antibodies, probably due to in vivo dehalogenation. These results indicate that the effect of tumor size on the incorporation of labeled MoAb into tumors is dependent on the antigenic nature to be targeted and/or radionuclides used for labeling and that high concentrations of circulating high molecular weight antigens may limit in vivo use of MoAb conjugates.

  7. Hydroponic isotope labeling of entire plants and high-performance mass spectrometry for quantitative plant proteomics.

    PubMed

    Bindschedler, Laurence V; Mills, Davinia J S; Cramer, Rainer

    2012-01-01

    Hydroponic isotope labeling of entire plants (HILEP) combines hydroponic plant cultivation and metabolic labeling with stable isotopes using (15)N-containing inorganic salts to label whole and mature plants. Employing (15)N salts as the sole nitrogen source for HILEP leads to the production of healthy-looking plants which contain (15)N proteins labeled to nearly 100%. Therefore, HILEP is suitable for quantitative plant proteomic analysis, where plants are grown in either (14)N- or (15)N-hydroponic media and pooled when the biological samples are collected for relative proteome quantitation. The pooled (14)N-/(15)N-protein extracts can be fractionated in any suitable way and digested with a protease for shotgun proteomics, using typically reverse phase liquid chromatography nanoelectrospray ionization tandem mass spectrometry (RPLC-nESI-MS/MS). Best results were obtained with a hybrid ion trap/FT-MS mass spectrometer, combining high mass accuracy and sensitivity for the MS data acquisition with speed and high-throughput MS/MS data acquisition, increasing the number of proteins identified and quantified and improving protein quantitation. Peak processing and picking from raw MS data files, protein identification, and quantitation were performed in a highly automated way using integrated MS data analysis software with minimum manual intervention, thus easing the analytical workflow. In this methodology paper, we describe how to grow Arabidopsis plants hydroponically for isotope labeling using (15)N salts and how to quantitate the resulting proteomes using a convenient workflow that does not require extensive bioinformatics skills.

  8. Increased Protein Structural Resolution from Diethylpyrocarbonate-based Covalent Labeling and Mass Spectrometric Detection

    NASA Astrophysics Data System (ADS)

    Zhou, Yuping; Vachet, Richard W.

    2012-04-01

    Covalent labeling and mass spectrometry are seeing increased use together as a way to obtain insight into the 3-dimensional structure of proteins and protein complexes. Several amino acid specific (e.g., diethylpyrocarbonate) and non-specific (e.g., hydroxyl radicals) labeling reagents are available for this purpose. Diethylpyrocarbonate (DEPC) is a promising labeling reagent because it can potentially probe up to 30% of the residues in the average protein and gives only one reaction product, thereby facilitating mass spectrometric analysis. It was recently reported, though, that DEPC modifications are labile for some amino acids. Here, we show that label loss is more significant and widespread than previously thought, especially for Ser, Thr, Tyr, and His residues, when relatively long protein digestion times are used. Such label loss ultimately decreases the amount of protein structural information that is obtainable with this reagent. We find, however, that the number of DEPC modified residues and, thus, protein structural information, can be significantly increased by decreasing the time between the covalent labeling reaction and the mass spectrometric analysis. This is most effectively accomplished using short (e.g., 2 h) proteolytic digestions with enzymes such as immobilized chymotrypsin or Glu-C rather than using methods (e.g., microwave or ultrasonic irradiation) that accelerate proteolysis in other ways. Using short digestion times, we show that the percentage of solvent accessible residues that can be modified by DEPC increases from 44% to 67% for cytochrome c, 35% to 81% for myoglobin, and 76% to 95% for β-2-microglobulin. In effect, these increased numbers of modified residues improve the protein structural resolution available from this covalent labeling method. Compared with typical overnight digestion conditions, the short digestion times decrease the average distance between modified residues from 11 to 7 Å for myoglobin, 13 to 10 Å for

  9. Dynamic analysis of CO₂ labeling and cell respiration using membrane-inlet mass spectrometry.

    PubMed

    Yang, Tae Hoon

    2014-01-01

    Here, we introduce a mass spectrometry-based analytical method and relevant technical details for dynamic cell respiration and CO2 labeling analysis. Such measurements can be utilized as additional information and constraints for model-based (13)C metabolic flux analysis. Dissolved dynamics of oxygen consumption and CO2 mass isotopomer evolution from (13)C-labeled tracer substrates through different cellular processes can be precisely measured on-line using a miniaturized reactor system equipped with a membrane-inlet mass spectrometer. The corresponding specific rates of physiologically relevant gases and CO2 mass isotopomers can be quantified within a short-term range based on the liquid-phase dynamics of dissolved fermentation gases.

  10. Use of quantum dots as mass and fluorescence labels in microarray biosensing.

    PubMed

    Finetti, Chiara; Plavisch, Lauren; Chiari, Marcella

    2016-01-15

    In this work, we demonstrate the efficacy of a Quantum Dot (QD) mass label strategy to enhance sensitivity in an interferometric technique called interferometric reflectance imaging sensor (IRIS). This biomass detection platform confers the advantage of absolute mass quantification and lower cost, easily implementable equipment. We discuss the advantages of this label when used in parallel with fluorescence detection. QDs represent a unique opportunity to improve sensitivity in both mass-label detection methods due to their large detectable mass, as well as in fluorescence detection, as they fluoresce without quenching. Streptavidin-conjugated QDs (SA-QDs) have been investigated as such a dual-role probe because of their large shape and mass, their 655nm emission peak for fluorescent detection platforms, and their robust insensitivity to photobleaching and quenching. In particular we explored their dual role in a microarrays immunoassay designed to detect antibodies against β-lactoglobulin, a common milk allergen. The SA-QDs formed a large detectable monolayer of 6.2ng/mm(2) in the saturation conditions, a mass signal corroborated by previous studies by Platt et al..

  11. Simplified quantitative glycomics using the stable isotope label Girard's reagent p by electrospray ionization mass spectrometry.

    PubMed

    Wang, Chengjian; Wu, Zhiyu; Yuan, Jiangbei; Wang, Bo; Zhang, Ping; Zhang, Ying; Wang, Zhongfu; Huang, Linjuan

    2014-02-01

    Fast, sensitive, and simple methods for quantitative analysis of disparities in glycan expression between different biological samples are essential for studies of protein glycosylation patterns (glycomics) and the search for disease glycan biomarkers. Relative quantitation of glycans based on stable isotope labeling combined with mass spectrometric detection represents an emerging and promising technique. However, this technique is undermined by the complexity of mass spectra of isotope-labeled glycans caused by the presence of multiple metal ion adduct signals, which result in a decrease of detection sensitivity and an increase of difficulties in data interpretation. Herein we report a simplified quantitative glycomics strategy, which features nonreductive isotopic labeling of reducing glycans with either nondeuterated (d0-) or deuterated (d5-) Girard's reagent P (GP) without salts introduced and simplified mass spectrometric profiles of d0- and d5-GP derivatives of neutral glycans as molecular ions without complex metal ion adducts, allowing rapid and sensitive quantitative comparison between different glycan samples. We have obtained optimized GP-labeling conditions and good quantitation linearity, reproducibility, and accuracy of data by the method. Its excellent applicability was validated by comparatively quantitative analysis of the neutral N-glycans released from bovine and porcine immunoglobulin G as well as of those from mouse and rat sera. Additionally, we have revealed the potential of this strategy for the high-sensitivity analysis of sialylated glycans as GP derivatives, which involves neutralization of the carboxyl group of sialic acid by chemical derivatization.

  12. Medical devices; ovarian adnexal mass assessment score test system; labeling; black box restrictions. Final rule.

    PubMed

    2011-12-30

    The Food and Drug Administration (FDA) is amending the regulation classifying ovarian adnexal mass assessment score test systems to restrict these devices so that a prescribed warning statement that addresses a risk identified in the special controls guidance document must be in a black box and must appear in all labeling, advertising, and promotional material. The black box warning mitigates the risk to health associated with off-label use as a screening test, stand-alone diagnostic test, or as a test to determine whether or not to proceed with surgery.

  13. Label-free imaging, detection, and mass measurement of single viruses by surface plasmon resonance

    PubMed Central

    Wang, Shaopeng; Shan, Xiaonan; Patel, Urmez; Huang, Xinping; Lu, Jin; Li, Jinghong; Tao, Nongjian

    2010-01-01

    We report on label-free imaging, detection, and mass/size measurement of single viral particles in solution by high-resolution surface plasmon resonance microscopy. Diffraction of propagating plasmon waves along a metal surface by the viral particles creates images of the individual particles, which allow us to detect the binding of the viral particles to surfaces functionalized with and without antibodies. We show that the intensity of the particle image is related to the mass of the particle, from which we determine the mass and mass distribution of influenza viral particles with a mass detection limit of approximately 1 ag (or 0.2 fg/mm2). This work demonstrates a multiplexed method to measure the masses of individual viral particles and to study the binding activity of the viral particles. PMID:20798340

  14. Evaluation of chemical labeling methods for identifying functional arginine residues of proteins by mass spectrometry.

    PubMed

    Wanigasekara, Maheshika S K; Chowdhury, Saiful M

    2016-09-01

    Arginine residues undergo several kinds of post-translational modifications (PTMs). These PTMs are associated with several inflammatory diseases, such as rheumatoid arthritis, atherosclerosis, and diabetes. Mass spectrometric studies of arginine modified proteins and peptides are very important, not only to identify the reactive arginine residues but also to understand the tandem mass spectrometry behavior of these peptides for assigning the sequences unambiguously. Herein, we utilize tandem mass spectrometry to report the performance of two widely used arginine labeling reagents, 1,2-cyclohexanedione (CHD) and phenylglyoxal (PG) with several arginine containing peptides and proteins. Time course labeling studies were performed to demonstrate the selectivity of the reagents in proteins or protein digests. Structural studies on the proteins were also explored to better understand the reaction sites and position of arginine residues. We found CHD showed better labeling efficiencies compared to phenylglyoxal. Reactive arginine profiling on a purified albumin protein clearly pointed out the cellular glycation modification site for this protein with high confidence. We believe these detailed mass-spectrometric studies will provide significant input to profile reactive arginine residues in large-scale studies; therefore, targeted proteomics can be performed to the short listed reactive sites for cellular arginine modifications. PMID:27543028

  15. Determination of Phytochelatins in Rice by Stable Isotope Labeling Coupled with Liquid Chromatography-Mass Spectrometry.

    PubMed

    Liu, Ping; Cai, Wen-Jing; Yu, Lei; Yuan, Bi-Feng; Feng, Yu-Qi

    2015-07-01

    A highly sensitive method was developed for the detection of phytochelatins (PCs) in rice by stable isotope labeling coupled with liquid chromatography-electrospray ionization-tandem mass spectrometry (IL-LC-ESI-MS/MS) analysis. A pair of isotope-labeling reagents [ω-bromoacetonylquinolinium bromide (BQB) and BQB-d(7)] were used to label PCs in plant sample and standard PCs, respectively, and then combined prior to LC/MS analysis. The heavy labeled standards were used as the internal standards for quantitation to minimize the matrix and ion suppression effects in MS analysis. In addition, the ionization efficiency of PCs was greatly enhanced through the introduction of a permanent charged moiety of quaternary ammonium of BQB into PCs. The detection sensitivities of PCs upon BQB labeling improved by 14-750-fold, and therefore, PCs can be quantitated using only 5 mg of plant tissue. Furthermore, under cadmium (Cd) stress, we found that the contents of PCs in rice dramatically increased with the increased concentrations and treatment time of Cd. It was worth noting that PC5 was first identified and quantitated in rice tissues under Cd stress in the current study. Taken together, this IL-LC-ESI-MS/MS method demonstrated to be a promising strategy in detection of PCs in plants with high sensitivity and reliability. PMID:26073168

  16. Determination of Phytochelatins in Rice by Stable Isotope Labeling Coupled with Liquid Chromatography-Mass Spectrometry.

    PubMed

    Liu, Ping; Cai, Wen-Jing; Yu, Lei; Yuan, Bi-Feng; Feng, Yu-Qi

    2015-07-01

    A highly sensitive method was developed for the detection of phytochelatins (PCs) in rice by stable isotope labeling coupled with liquid chromatography-electrospray ionization-tandem mass spectrometry (IL-LC-ESI-MS/MS) analysis. A pair of isotope-labeling reagents [ω-bromoacetonylquinolinium bromide (BQB) and BQB-d(7)] were used to label PCs in plant sample and standard PCs, respectively, and then combined prior to LC/MS analysis. The heavy labeled standards were used as the internal standards for quantitation to minimize the matrix and ion suppression effects in MS analysis. In addition, the ionization efficiency of PCs was greatly enhanced through the introduction of a permanent charged moiety of quaternary ammonium of BQB into PCs. The detection sensitivities of PCs upon BQB labeling improved by 14-750-fold, and therefore, PCs can be quantitated using only 5 mg of plant tissue. Furthermore, under cadmium (Cd) stress, we found that the contents of PCs in rice dramatically increased with the increased concentrations and treatment time of Cd. It was worth noting that PC5 was first identified and quantitated in rice tissues under Cd stress in the current study. Taken together, this IL-LC-ESI-MS/MS method demonstrated to be a promising strategy in detection of PCs in plants with high sensitivity and reliability.

  17. Identification of Asp isomerization in proteins by ¹⁸O labeling and tandem mass spectrometry.

    PubMed

    Zhang, Jennifer; Katta, Viswanatham

    2012-01-01

    Isomerization of aspartic acid (Asp) to isoaspartic acid (isoAsp) via succinimide intermediate is a common route of degradation for proteins that can affect their structural integrity. As Asp/isoAsp is isobaric in mass, it is difficult to identify the site of modification by LC-MS/MS peptide mapping. Here, we describe an approach to label the Asp residue involved in isomerization at the protein level by hydrolyzing the succinimide intermediate in H₂¹⁸O. Tryptic digestion of this labeled protein will result in peptides containing the site of isomerization being 2 Da heavier than the ¹⁶O-containing counterparts, due to ¹⁸O incorporation during the hydrolysis process. Comparison of tandem mass spectra of isomerized peptides with and without ¹⁸O incorporation allows easy identification of the Asp residue involved. This method proved to be especially useful in identifying the sites when isomerization occurs in Asp-Asp motifs.

  18. freeQuant: A Mass Spectrometry Label-Free Quantification Software Tool for Complex Proteome Analysis.

    PubMed

    Deng, Ning; Li, Zhenye; Pan, Chao; Duan, Huilong

    2015-01-01

    Study of complex proteome brings forward higher request for the quantification method using mass spectrometry technology. In this paper, we present a mass spectrometry label-free quantification tool for complex proteomes, called freeQuant, which integrated quantification with functional analysis effectively. freeQuant consists of two well-integrated modules: label-free quantification and functional analysis with biomedical knowledge. freeQuant supports label-free quantitative analysis which makes full use of tandem mass spectrometry (MS/MS) spectral count, protein sequence length, shared peptides, and ion intensity. It adopts spectral count for quantitative analysis and builds a new method for shared peptides to accurately evaluate abundance of isoforms. For proteins with low abundance, MS/MS total ion count coupled with spectral count is included to ensure accurate protein quantification. Furthermore, freeQuant supports the large-scale functional annotations for complex proteomes. Mitochondrial proteomes from the mouse heart, the mouse liver, and the human heart were used to evaluate the usability and performance of freeQuant. The evaluation showed that the quantitative algorithms implemented in freeQuant can improve accuracy of quantification with better dynamic range.

  19. Interconversion of Peptide Mass Spectral Libraries Derivatized with iTRAQ or TMT Labels.

    PubMed

    Zhang, Zheng; Yang, Xiaoyu; Mirokhin, Yuri A; Tchekhovskoi, Dmitrii V; Ji, Weihua; Markey, Sanford P; Roth, Jeri; Neta, Pedatsur; Hizal, Deniz Baycin; Bowen, Michael A; Stein, Stephen E

    2016-09-01

    Derivitization of peptides with isobaric tags such as iTRAQ and TMT is widely employed in proteomics due to their compatibility with multiplex quantitative measurements. We recently made publicly available a large peptide library derived from iTRAQ 4-plex labeled spectra. This resource has not been used for identifying peptides labeled with related tags with different masses, because values for virtually all masses of precursor and most product ions would differ for ions containing the different tags as well as containing different tag-specific peaks. We describe a method for interconverting spectra from iTRAQ 4-plex to TMT (6- and 10-plex) and to iTRAQ 8-plex. We interconvert spectra by appropriately mass shifting sequence ions and discarding derivative-specific peaks. After this "cleaning" of search spectra, we demonstrate that the converted libraries perform well in terms of peptide spectral matches. This is demonstrated by comparing results using sequence database searches as well as by comparing search effectiveness using original and converted libraries. At 1% FDR TMT labeled query spectra match 97% as many spectra against a converted iTRAQ library as compared to an original TMT library. Overall this interconversion strategy provides a practical way to extend results from one derivatization method to others that share related chemistry and do not significantly alter fragmentation profiles. PMID:27386737

  20. Interconversion of Peptide Mass Spectral Libraries Derivatized with iTRAQ or TMT Labels.

    PubMed

    Zhang, Zheng; Yang, Xiaoyu; Mirokhin, Yuri A; Tchekhovskoi, Dmitrii V; Ji, Weihua; Markey, Sanford P; Roth, Jeri; Neta, Pedatsur; Hizal, Deniz Baycin; Bowen, Michael A; Stein, Stephen E

    2016-09-01

    Derivitization of peptides with isobaric tags such as iTRAQ and TMT is widely employed in proteomics due to their compatibility with multiplex quantitative measurements. We recently made publicly available a large peptide library derived from iTRAQ 4-plex labeled spectra. This resource has not been used for identifying peptides labeled with related tags with different masses, because values for virtually all masses of precursor and most product ions would differ for ions containing the different tags as well as containing different tag-specific peaks. We describe a method for interconverting spectra from iTRAQ 4-plex to TMT (6- and 10-plex) and to iTRAQ 8-plex. We interconvert spectra by appropriately mass shifting sequence ions and discarding derivative-specific peaks. After this "cleaning" of search spectra, we demonstrate that the converted libraries perform well in terms of peptide spectral matches. This is demonstrated by comparing results using sequence database searches as well as by comparing search effectiveness using original and converted libraries. At 1% FDR TMT labeled query spectra match 97% as many spectra against a converted iTRAQ library as compared to an original TMT library. Overall this interconversion strategy provides a practical way to extend results from one derivatization method to others that share related chemistry and do not significantly alter fragmentation profiles.

  1. Probing adenosine nucleotide-binding proteins with an affinity-labeled nucleotide probe and mass spectrometry.

    PubMed

    Qiu, Haibo; Wang, Yinsheng

    2007-08-01

    Mass spectrometry combined with chemical labeling strategies has become very important in biological analysis. Herein, we described the application of a biotin-conjugated acyl nucleotide for probing adenosine nucleotide-binding proteins. We demonstrated that the probe reacted specifically with the lysine residue at the nucleotide-binding site of two purified adenosine nucleotide-binding proteins, Escherichia coli recombinase A (RecA) and Saccharomyces cerevisiae alcohol dehydrogenase-I (YADH-I). A single conjugate peptide with a specifically labeled lysine residue was identified, by using LC-MS/MS, from the tryptic digestion mixture of the reaction products of the nucleotide analogue with RecA or YADH-I. The strategy, which involved labeling reaction, enzymatic digestion, affinity purification, and LC-MS/MS analysis, was relatively simple, fast, and straightforward. The method should be generally applicable for the identification of lysine residues at the nucleotide-binding site of other proteins. The biotin-conjugated acyl nucleotide probe also allowed for the enrichment and identification of nucleotide-binding proteins from complex protein mixtures; we showed that more than 50 adenosine nucleotide-binding proteins could be identified from the whole-cell lysates of HeLa-S3 and WM-266-4 cells.

  2. Probing adenosine nucleotide-binding proteins with an affinity labeled-nucleotide probe and mass spectrometry

    PubMed Central

    Qiu, Haibo; Wang, Yinsheng

    2008-01-01

    Mass spectrometry combined with chemical labeling strategies has become very important in biological analysis. Herein, we described the application of a biotin-conjugated acyl nucleotide for probing adenosine nucleotide-binding proteins. We demonstrated that the probe reacted specifically with the lysine residue at the nucleotide-binding site of two purified adenosine nucleotide-binding proteins, Escherichia coli RecA and Saccharomyces cerevisiae alcohol dehydrogenase-I (YADH-I). A single conjugate peptide with a specifically labeled lysine residue was identified, by using LC-MS/MS, from the tryptic digestion mixture of the reaction products of the nucleotide analog with RecA or YADH-I. The strategy, which involved labeling reaction, enzymatic digestion, affinity purification and LC-MS/MS analysis, was relatively simple, fast and straightforward. The method should be generally applicable for the identification of lysine residues at the nucleotide-binding site of other proteins. The biotin-conjugated acyl nucleotide probe also allowed for the enrichment and identification of nucleotide-binding proteins from complex protein mixtures; we showed that more than 50 adenosine nucleotide-binding proteins could be identified from the whole cell lysates of HeLa-S3 and WM-266-4 cells. PMID:17602667

  3. Label free targeted detection and quantification of celiac disease immunogenic epitopes by mass spectrometry.

    PubMed

    van den Broeck, Hetty C; Cordewener, Jan H G; Nessen, Merel A; America, Antoine H P; van der Meer, Ingrid M

    2015-04-24

    Celiac disease (CD) is a food-related disease caused by certain gluten peptides containing T-cell stimulating epitopes from wheat, rye, and barley. CD-patients have to maintain a gluten-free diet and are therefore dependent on reliable testing and labeling of gluten-free products. So far, the R5-ELISA is the approved method to detect if food products can be labeled gluten-free. Because the R5-ELISA detects gluten in general, there is a demand for an improved detection method that quantifies specifically CD-epitopes. Therefore, we developed a new method for detection and quantification of CD-epitopes, based on liquid chromatography (LC) coupled to mass spectrometry (MS) in multiple reaction monitoring (MRM) mode. This method enables targeted label free comparative analysis of the gluten proteins present in different wheat varieties and species, and in wheat-based food products. We have tested our method by analyzing several wheat varieties that vary in CD-epitope content, as was shown before using immunoblotting and specific monoclonal antibodies. The results showed that a modern bread wheat variety Toronto contained the highest amounts of CD immunogenic peptides compared with the older bread wheat variety Minaret and the tetraploid wheat variety Dibillik Sinde. Our developed method can detect quantitatively and simultaneously multiple specific CD-epitopes in a high throughput manner. PMID:25795397

  4. Stable Isotope Labeling Strategy for Curcumin Metabolite Study in Human Liver Microsomes by Liquid Chromatography-Tandem Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Gao, Dan; Chen, Xiaowu; Yang, Xiaomei; Wu, Qin; Jin, Feng; Wen, Hongliang; Jiang, Yuyang; Liu, Hongxia

    2015-04-01

    The identification of drug metabolites is very important in drug development. Nowadays, the most widely used methods are isotopes and mass spectrometry. However, the commercial isotopic labeled reagents are usually very expensive, and the rapid and convenient identification of metabolites is still difficult. In this paper, an 18O isotope labeling strategy was developed and the isotopes were used as a tool to identify drug metabolites using mass spectrometry. Curcumin was selected as a model drug to evaluate the established method, and the 18O labeled curcumin was successfully synthesized. The non-labeled and 18O labeled curcumin were simultaneously metabolized in human liver microsomes (HLMs) and analyzed by liquid chromatography/mass spectrometry (LC-MS). The two groups of chromatograms obtained from metabolic reaction mixture with and without cofactors were compared and analyzed using Metabolynx software (Waters Corp., Milford, MA, USA). The mass spectra of the newly appearing chromatographic peaks in the experimental sample were further analyzed to find the metabolite candidates. Their chemical structures were confirmed by tandem mass spectrometry. Three metabolites, including two reduction products and a glucuronide conjugate, were successfully detected under their specific HLMs metabolic conditions, which were in accordance with the literature reported results. The results demonstrated that the developed isotope labeling method, together with post-acquisition data processing using Metabolynx software, could be used for fast identification of new drug metabolites.

  5. Protected amine labels: a versatile molecular scaffold for multiplexed nominal mass and sub-Da isotopologue quantitative proteomic reagents.

    PubMed

    Ficarro, Scott B; Biagi, Jessica M; Wang, Jinhua; Scotcher, Jenna; Koleva, Rositsa I; Card, Joseph D; Adelmant, Guillaume; He, Huan; Askenazi, Manor; Marshall, Alan G; Young, Nicolas L; Gray, Nathanael S; Marto, Jarrod A

    2014-04-01

    We assemble a versatile molecular scaffold from simple building blocks to create binary and multiplexed stable isotope reagents for quantitative mass spectrometry. Termed Protected Amine Labels (PAL), these reagents offer multiple analytical figures of merit including, (1) robust targeting of peptide N-termini and lysyl side chains, (2) optimal mass spectrometry ionization efficiency through regeneration of primary amines on labeled peptides, (3) an amino acid-based mass tag that incorporates heavy isotopes of carbon, nitrogen, and oxygen to ensure matched physicochemical and MS/MS fragmentation behavior among labeled peptides, and (4) a molecularly efficient architecture, in which the majority of hetero-atom centers can be used to synthesize a variety of nominal mass and sub-Da isotopologue stable isotope reagents. We demonstrate the performance of these reagents in well-established strategies whereby up to four channels of peptide isotopomers, each separated by 4 Da, are quantified in MS-level scans with accuracies comparable to current commercial reagents. In addition, we utilize the PAL scaffold to create isotopologue reagents in which labeled peptide analogs differ in mass based on the binding energy in carbon and nitrogen nuclei, thereby allowing quantification based on MS or MS/MS spectra. We demonstrate accurate quantification for reagents that support 6-plex labeling and propose extension of this scheme to 9-channels based on a similar PAL scaffold. Finally, we provide exemplar data that extend the application of isotopologe-based quantification reagents to medium resolution, quadrupole time-of-flight mass spectrometers.

  6. Metabolomic and mass isotopomer analysis of liver gluconeogenesis and citric acid cycle: II. Heterogeneity of metabolite labeling pattern.

    PubMed

    Yang, Lili; Kasumov, Takhar; Kombu, Rajan S; Zhu, Shu-Han; Cendrowski, Andrea V; David, France; Anderson, Vernon E; Kelleher, Joanne K; Brunengraber, Henri

    2008-08-01

    In this second of two companion articles, we compare the mass isotopomer distribution of metabolites of liver gluconeogenesis and citric acid cycle labeled from NaH(13)CO(3) or dimethyl [1,4-(13)C(2)]succinate. The mass isotopomer distribution of intermediates reveals the reversibility of the isocitrate dehydrogenase + aconitase reactions, even in the absence of a source of alpha-ketoglutarate. In addition, in many cases, a number of labeling incompatibilities were found as follows: (i) glucose versus triose phosphates and phosphoenolpyruvate; (ii) differences in the labeling ratios C-4/C-3 of glucose versus (glyceraldehyde 3-phosphate)/(dihydroxyacetone phosphate); and (iii) labeling of citric acid cycle intermediates in tissue versus effluent perfusate. Overall, our data show that gluconeogenic and citric acid cycle intermediates cannot be considered as sets of homogeneously labeled pools. This probably results from the zonation of hepatic metabolism and, in some cases, from differences in the labeling pattern of mitochondrial versus extramitochondrial metabolites. Our data have implications for the use of labeling patterns for the calculation of metabolic rates or fractional syntheses in liver, as well as for modeling liver intermediary metabolism.

  7. Minimizing technical variation during sample preparation prior to label-free quantitative mass spectrometry.

    PubMed

    Scheerlinck, E; Dhaenens, M; Van Soom, A; Peelman, L; De Sutter, P; Van Steendam, K; Deforce, D

    2015-12-01

    Sample preparation is the crucial starting point to obtain high-quality mass spectrometry data and can be divided into two main steps in a bottom-up proteomics approach: cell/tissue lysis with or without detergents and a(n) (in-solution) digest comprising denaturation, reduction, alkylation, and digesting of the proteins. Here, some important considerations, among others, are that the reagents used for sample preparation can inhibit the digestion enzyme (e.g., 0.1% sodium dodecyl sulfate [SDS] and 0.5 M guanidine HCl), give rise to ion suppression (e.g., polyethylene glycol [PEG]), be incompatible with liquid chromatography-tandem mass spectrometry (LC-MS/MS) (e.g., SDS), and can induce additional modifications (e.g., urea). Taken together, all of these irreproducible effects are gradually becoming a problem when label-free quantitation of the samples is envisioned such as during the increasingly popular high-definition mass spectrometry (HDMS(E)) and sequential window acquisition of all theoretical fragment ion spectra (SWATH) data-independent acquisition strategies. Here, we describe the detailed validation of a reproducible method with sufficient protein yield for sample preparation without any known LC-MS/MS interfering substances by using 1% sodium deoxycholate (SDC) during both cell lysis and in-solution digest. PMID:26302362

  8. Mass Spectrometric Quantification of Histone Post-translational Modifications by a Hybrid Chemical Labeling Method

    PubMed Central

    Maile, Tobias M.; Izrael-Tomasevic, Anita; Cheung, Tommy; Guler, Gulfem D.; Tindell, Charles; Masselot, Alexandre; Liang, Jun; Zhao, Feng; Trojer, Patrick; Classon, Marie; Arnott, David

    2015-01-01

    Mass spectrometry is a powerful alternative to antibody-based methods for the analysis of histone post-translational modifications (marks). A key development in this approach was the deliberate propionylation of histones to improve sequence coverage across the lysine-rich and hydrophilic tails that bear most modifications. Several marks continue to be problematic however, particularly di- and tri-methylated lysine 4 of histone H3 which we found to be subject to substantial and selective losses during sample preparation and liquid chromatography-mass spectrometry. We developed a new method employing a “one-pot” hybrid chemical derivatization of histones, whereby an initial conversion of free lysines to their propionylated forms under mild aqueous conditions is followed by trypsin digestion and labeling of new peptide N termini with phenyl isocyanate. High resolution mass spectrometry was used to collect qualitative and quantitative data, and a novel web-based software application (Fishtones) was developed for viewing and quantifying histone marks in the resulting data sets. Recoveries of 53 methyl, acetyl, and phosphoryl marks on histone H3.1 were improved by an average of threefold overall, and over 50-fold for H3K4 di- and tri-methyl marks. The power of this workflow for epigenetic research and drug discovery was demonstrated by measuring quantitative changes in H3K4 trimethylation induced by small molecule inhibitors of lysine demethylases and siRNA knockdown of epigenetic modifiers ASH2L and WDR5. PMID:25680960

  9. Minimizing technical variation during sample preparation prior to label-free quantitative mass spectrometry.

    PubMed

    Scheerlinck, E; Dhaenens, M; Van Soom, A; Peelman, L; De Sutter, P; Van Steendam, K; Deforce, D

    2015-12-01

    Sample preparation is the crucial starting point to obtain high-quality mass spectrometry data and can be divided into two main steps in a bottom-up proteomics approach: cell/tissue lysis with or without detergents and a(n) (in-solution) digest comprising denaturation, reduction, alkylation, and digesting of the proteins. Here, some important considerations, among others, are that the reagents used for sample preparation can inhibit the digestion enzyme (e.g., 0.1% sodium dodecyl sulfate [SDS] and 0.5 M guanidine HCl), give rise to ion suppression (e.g., polyethylene glycol [PEG]), be incompatible with liquid chromatography-tandem mass spectrometry (LC-MS/MS) (e.g., SDS), and can induce additional modifications (e.g., urea). Taken together, all of these irreproducible effects are gradually becoming a problem when label-free quantitation of the samples is envisioned such as during the increasingly popular high-definition mass spectrometry (HDMS(E)) and sequential window acquisition of all theoretical fragment ion spectra (SWATH) data-independent acquisition strategies. Here, we describe the detailed validation of a reproducible method with sufficient protein yield for sample preparation without any known LC-MS/MS interfering substances by using 1% sodium deoxycholate (SDC) during both cell lysis and in-solution digest.

  10. Status of complete proteome analysis by mass spectrometry: SILAC labeled yeast as a model system

    PubMed Central

    de Godoy, Lyris MF; Olsen, Jesper V; de Souza, Gustavo A; Li, Guoqing; Mortensen, Peter; Mann, Matthias

    2006-01-01

    Background Mass spectrometry has become a powerful tool for the analysis of large numbers of proteins in complex samples, enabling much of proteomics. Due to various analytical challenges, so far no proteome has been sequenced completely. O'Shea, Weissman and co-workers have recently determined the copy number of yeast proteins, making this proteome an excellent model system to study factors affecting coverage. Results To probe the yeast proteome in depth and determine factors currently preventing complete analysis, we grew yeast cells, extracted proteins and separated them by one-dimensional gel electrophoresis. Peptides resulting from trypsin digestion were analyzed by liquid chromatography mass spectrometry on a linear ion trap-Fourier transform mass spectrometer with very high mass accuracy and sequencing speed. We achieved unambiguous identification of more than 2,000 proteins, including very low abundant ones. Effective dynamic range was limited to about 1,000 and effective sensitivity to about 500 femtomoles, far from the subfemtomole sensitivity possible with single proteins. We used SILAC (stable isotope labeling by amino acids in cell culture) to generate one-to-one pairs of true peptide signals and investigated if sensitivity, sequencing speed or dynamic range were limiting the analysis. Conclusion Advanced mass spectrometry methods can unambiguously identify more than 2,000 proteins in a single proteome. Complex mixture analysis is not limited by sensitivity but by a combination of dynamic range (high abundance peptides preventing sequencing of low abundance ones) and by effective sequencing speed. Substantially increased coverage of the yeast proteome appears feasible with further development in software and instrumentation. PMID:16784548

  11. Fragmentation reactions of labeled and untabeled Rhodamine B in a high-resolution Fourier transform ion cyclotron resonance mass spectrometer.

    PubMed

    Clemen, Martin; Gernert, Claus; Peters, Jonathan; Grotemeyer, Jürgen

    2013-01-01

    The fragmentation reactions of Rhodamine B have been investigated by the use of electrospray ionization mass spectra in a high mass resolving ion cyclotron resonance mass spectrometer. Using high resolution, it could be shown that the loss of 44 mass units from the molecular ion is due to propane; the measured masses were inconsistent with loss of carbon dioxide. These conclusions are supported using deuterium-labeled Rhodamine B. This sample again only shows the loss of fully-deuterated propane verifying the high-resolution data. These findings illustrate very clearly that the conclusions based solely on low resolution spectra were false. The general implication on fragmentations of aromatic acids is discussed.

  12. Metal-tag labeling coupled with multiple reaction monitoring-mass spectrometry for absolute quantitation of proteins.

    PubMed

    Wang, Xueying; Wang, Xin; Qin, Weijie; Lin, Hongjun; Wang, Jifeng; Wei, Junying; Zhang, Yangjun; Qian, Xiaohong

    2013-09-21

    Mass spectrometry-based quantitative proteomics, consisting of relative and absolute parts, has been used to discover and validate proteins with key functions related to physiological and pathological processes. Currently, stable isotope dilution-multiple reaction monitoring-mass spectrometry (SID-MRM-MS) is the most commonly used method for the absolute determination of proteins in a biological sample. A prerequisite for this method is obtaining internal standards with isotope labels. Although many approaches have been developed for the labeling and preparation of internal peptides, expensive stable isotope labeling coupled with SID-MRM-MS has limited the application and development of an absolute quantitative method. Recently, a low-cost strategy using metal-tag labeling and MS has been developed for relative quantification of peptides or proteins. The introduction of labeling using metal tags has the merits of allowing multiple labeling and enlarging the mass shift to overcome the overlap of adjacent isotope clusters. However, most papers described MRM-MS for protein absolute quantification based on the metal in its peptides labelled with metal by inductively coupled plasma mass spectrometry (ICP MS) but not on its peptides labelled with metal. In this work, a novel approach based on metal-tag labeling coupled with MRM-MS was established for the absolute quantification of peptides or proteins. The principle of the method is that a bifunctional chelator, 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid bearing an N-hydroxysuccinimide ester (DOTA-NHS ester), is used to modify the N-termini of signature peptides from a target protein, and the modified peptides then chelate a certain metal, such as thulium, to form metal-tagged peptides (Tm-DOTA-P). Internal peptides are chemically synthesized and labeled with another metal, such as terbium (Tb-DOTA-P), as the internal standard. Both the Tb-DOTA- and Tm-DOTA-labeled peptides in samples can be analysed via

  13. Raman spectroscopic and mass spectrometric investigations of the hydrogen isotopes and isotopically labelled methane

    SciTech Connect

    Jewett, J.R., Fluor Daniel Hanford

    1997-02-24

    Suitable analytical methods must be tested and developed for monitoring the individual process steps within the fuel cycle of a fusion reactor and for tritium accountability. The utility of laser-Raman spectroscopy accompanied by mass spectrometry with an Omegatron was investigated using the analysis of all hydrogen isotopes and isotopically labeled methanes as an example. The Omegatron is useful for analyzing all hydrogen isotopes mixed with the stable helium isotopes. The application of this mass spectrometer were demonstrated by analyzing mixtures of deuterated methanes. In addition, it was employed to study the radiochemical Witzbach exchange reaction between tritium and methanes. A laser-Raman spectrometer was designed for analysis of tritium-containing gases and was built from individual components. A tritium-compatible, metal-sealed Raman cuvette having windows with good optical properties and additional means for measuring the stray light was first used successfully in this work. The Raman spectra of the hydrogen isotopes were acquired in the pure rotation mode and in the rotation-vibration mode and were used for on. The deuterated methanes were measured by Raman spectroscopy, the wavenumbers determined were assigned to the corresponding vibrations, and the wavenumbers for the rotational fine-structure were summarized in tables. The fundamental Vibrations of the deuterated methanes produced Witzbach reactions were detected and assigned. The fundamental vibrations of the molecules were obtained with Raman spectroscopy for the first time in this work. The @-Raman spectrometer assembled is well suited for the analysis of tritium- containing gases and is practical in combination with mass spectrometry using an Omegatron, for studying gases used in fusion.

  14. Protected Amine Labels: A Versatile Molecular Scaffold for Multiplexed Nominal Mass and Sub-Da Isotopologue Quantitative Proteomic Reagents

    PubMed Central

    Ficarro, Scott B.; Biagi, Jessica M.; Wang, Jinhua; Scotcher, Jenna; Koleva, Rositsa I.; Card, Joseph D.; Adelmant, Guillaume; He, Huan; Askenazi, Manor; Marshall, Alan G.; Young, Nicolas L.; Gray, Nathanael S.; Marto, Jarrod A.

    2014-01-01

    We assemble a versatile molecular scaffold from simple building blocks to create binary and multiplexed stable isotope reagents for quantitative mass spectrometry. Termed Protected Amine Labels (PAL), these reagents offer multiple analytical figures of merit including, (i) robust targeting of peptide N-termini and lysyl side chains, (ii) optimal mass spectrometry ionization efficiency through regeneration of primary amines on labeled peptides, (iii) an amino acid-based mass tag that incorporates heavy isotopes of carbon, nitrogen, and oxygen to ensure matched physicochemical and MS/MS fragmentation behavior among labeled peptides, and (iv) a molecularly efficient architecture, in which the majority of hetero-atom centers can be used to synthesize a variety of nominal mass and sub-Da isotopologue stable isotope reagents. We demonstrate the performance of these reagents in well-established strategies whereby up to four channels of peptide isotopomers, each separated by 4 Da are quantified in MS-level scans with accuracies comparable to current commercial reagents. In addition we utilize the PAL scaffold to create isotopologue reagents in which labeled peptide analogs differ in mass based on the binding energy in carbon and nitrogen nuclei, thereby allowing quantification based on MS or MS/MS spectra. We demonstrate accurate quantification for reagents that support 6-plex labeling and propose extension of this scheme to 9-channels based on a similar PAL scaffold. Finally we provide exemplar data that extends the application of isotopologe-based quantification reagents to medium resolution, quadrupole time-of-flight mass spectrometers. PMID:24496597

  15. Label-Free Quantitative Mass Spectrometry Reveals a Panel of Differentially Expressed Proteins in Colorectal Cancer

    PubMed Central

    Fan, Nai-Jun; Gao, Jiang-Ling; Liu, Yan; Song, Wei; Zhang, Zhan-Yang; Gao, Chun-Fang

    2015-01-01

    To identify potential biomarkers involved in CRC, a shotgun proteomic method was applied to identify soluble proteins in three CRCs and matched normal mucosal tissues using high-performance liquid chromatography and mass spectrometry. Label-free protein profiling of three CRCs and matched normal mucosal tissues were then conducted to quantify and compare proteins. Results showed that 67 of the 784 identified proteins were linked to CRC (28 upregulated and 39 downregulated). Gene Ontology and DAVID databases were searched to identify the location and function of differential proteins that were related to the biological processes of binding, cell structure, signal transduction, cell adhesion, and so on. Among the differentially expressed proteins, tropomyosin-3 (TPM3), endoplasmic reticulum resident protein 29 (ERp29), 18 kDa cationic antimicrobial protein (CAMP), and heat shock 70 kDa protein 8 (HSPA8) were verified to be upregulated in CRC tissue and seven cell lines through western blot analysis. Furthermore, the upregulation of TPM3, ERp29, CAMP, and HSPA8 was validated in 69 CRCs byimmunohistochemistry (IHC) analysis. Combination of TPM3, ERp29, CAMP, and HSPA8 can identify CRC from matched normal mucosal achieving an accuracy of 73.2% using IHC score. These results suggest that TPM3, ERp29, CAMP, and HSPA8 are great potential IHC diagnostic biomarkers for CRC. PMID:25699276

  16. Measuring the dynamics of E. coli ribosome biogenesis using pulse-labeling and quantitative mass spectrometry

    PubMed Central

    Chen, Stephen S.; Sperling, Edit; Silverman, Joshua M.; Davis, Joseph H.; Williamson, James R.

    2012-01-01

    The ribosome is an essential organelle responsible for cellular protein synthesis. Until recently, the study of ribosome assembly has been largely limited to in vitro assays, with few attempts to reconcile these results with the more complex in vivo ribosome biogenesis process. Here, we characterize the ribosome synthesis and assembly pathway for each E. coli ribosomal protein (r-protein) in vivo using a stable isotope pulse-labeling timecourse. Isotope incorporation into assembled ribosomes was measured by quantitative mass spectrometry (qMS) and fit using steady-state flux models. Most r-proteins exhibit precursor pools ranging in size from 0% to 7% of completed ribosomes, and that the sizes of these individual r-protein pools correlate well with the order of r-protein binding in vitro. Additionally, we observe anomalously large precursor pools for specific r-proteins with known extra-ribosomal functions and we have detected three r-proteins with significant turnover during steady-state growth. Taken together, this highly precise, time-dependent proteomic qMS approach should prove useful in future studies of ribosome biogenesis and could be easily extended to explore other complex biological processes in a cellular context. PMID:23090316

  17. Analysis of liposoluble carboxylic acids metabolome in human serum by stable isotope labeling coupled with liquid chromatography-mass spectrometry.

    PubMed

    Zhu, Quan-Fei; Zhang, Zheng; Liu, Ping; Zheng, Shu-Jian; Peng, Ke; Deng, Qian-Yun; Zheng, Fang; Yuan, Bi-Feng; Feng, Yu-Qi

    2016-08-19

    Fatty acids (FAs) are groups of liposoluble carboxylic acids (LCAs) and play important roles in various physiological processes. Abnormal contents or changes of FAs are associated with a series of diseases. Here we developed a strategy with stable isotope labeling combined with liquid chromatography-tandem mass spectrometry (IL-LC-MS) analysis for comprehensive profiling and relative quantitation of LCAs in human serum. In this strategy, a pair of isotope labeling reagents (2-dimethylaminoethylamine (DMED)) and d4-2-dimethylaminoethylamine (d4-DMED) were employed to selectively label carboxyl groups of LCAs. The DMED and d4-DMED labeled products can lose four characteristic neutral fragments of 45 and 49Da or 63 and 67Da in collision-induced dissociation. Therefore, quadruple neutral loss scan (QNLS) mode was established and used for non-targeted profiling of LCAs. The peak pairs of DMED and d4-DMED labeling with the same retention time, intensity and characteristic mass differences were extracted from the two NLS spectra respectively, and assigned as potential LCA candidates. Using this strategy, 241 LCA candidates were discovered in the human serum; 156 carboxylic acid compounds could be determined by searching HMDB and METLIN databases (FAs are over 90%) and 21 of these LCAs were successfully identified by standards. Subsequently, a modified pseudo-targeted method with multiple reaction monitoring (MRM) detection mode was developed and used for relative quantification of LCAs in human serum from type 2 diabetes mellitus (T2DM) patients and healthy controls. As a result, 81 LCAs were found to have significant difference between T2DM patients and healthy controls. Taken together, the isotope labeling combined with tandem mass spectrometry analysis demonstrated to be a powerful strategy for identification and quantification of LCA compounds in serum samples. PMID:27432792

  18. 18O-Labeled Proteome Reference as Global Internal Standards for Targeted Quantification by Selected Reaction Monitoring-Mass Spectrometry

    SciTech Connect

    Kim, Jong Seo; Fillmore, Thomas L.; Liu, Tao; Robinson, Errol W.; Hossain, Mahmud; Champion, Boyd L.; Moore, Ronald J.; Camp, David G.; Smith, Richard D.; Qian, Weijun

    2011-10-11

    Selected reaction monitoring-mass spectrometry (SRM-MS) is an emerging technology for high throughput targeted protein quantification and verification in biological and biomarker discovery studies; however, the cost associated with the use of stable isotope labeled synthetic peptides as internal standards is prohibitive for quantitatively screening large numbers of candidate proteins as often required in the pre-verification phase of biomarker discovery. Herein we present the proof-of-concept experiments of using an 18O-labeled 'universal' reference as comprehensive internal standards for quantitative SRM-MS analysis. With an 18O-labeled whole proteome sample as reference, every peptide of interest will have its own corresponding heavy isotope labeled internal standard, thus providing an ideal approach for quantitative screening of a large number of candidates using SRM-MS. Our results showed that the 18O incorporation efficiency using a recently improved protocol was >99.5% for most peptides investigated, a level comparable to 13C/15N labeled synthetic peptides in terms of heavy isotope incorporation. The accuracy, reproducibility, and linear dynamic range of quantification were further assessed based on known ratios of standard proteins spiked into mouse plasma with an 18O-labeled mouse plasma reference. A dynamic range of four orders of magnitude in relative concentration was obtained with high reproducibility (i.e., coefficient of variance <10%) based on the 16O/18O peak area ratios. Absolute and relative quantification of C-reactive protein and prostate-specific antigen were demonstrated by coupling an 18O-labeled reference with standard additions of protein standards. Collectively, our results demonstrated that the use of 18O-labeled reference provides a convenient and effective strategy for quantitative SRM screening of large number of candidate proteins.

  19. Liquid chromatography tandem mass spectrometry for measuring ¹³C-labeling in intermediates of the glycolysis and pentose phosphate pathway.

    PubMed

    Cocuron, Jean-Christophe; Alonso, Ana Paula

    2014-01-01

    This chapter describes a procedure to analyze (13)C-labeled phosphorylated compounds by liquid chromatography tandem mass spectrometry. Phosphorylated compounds, intermediaries of the glycolysis and pentose phosphate pathway, are separated by anion exchange chromatography and their isotopic labeling is determined by mass spectrometry. A sensitivity in the fmole range is achieved using scheduled multiple reaction monitoring mode.

  20. Combining combing and secondary ion mass spectrometry to study DNA on chips using (13)C and (15)N labeling.

    PubMed

    Cabin-Flaman, Armelle; Monnier, Anne-Francoise; Coffinier, Yannick; Audinot, Jean-Nicolas; Gibouin, David; Wirtz, Tom; Boukherroub, Rabah; Migeon, Henri-Noël; Bensimon, Aaron; Jannière, Laurent; Ripoll, Camille; Norris, Victor

    2016-01-01

    Dynamic secondary ion mass spectrometry ( D-SIMS) imaging of combed DNA - the combing, imaging by SIMS or CIS method - has been developed previously using a standard NanoSIMS 50 to reveal, on the 50 nm scale, individual DNA fibers labeled with different, non-radioactive isotopes in vivo and to quantify these isotopes. This makes CIS especially suitable for determining the times, places and rates of DNA synthesis as well as the detection of the fine-scale re-arrangements of DNA and of molecules associated with combed DNA fibers. Here, we show how CIS may be extended to (13)C-labeling via the detection and quantification of the (13)C (14)N (-) recombinant ion and the use of the (13)C: (12)C ratio, we discuss how CIS might permit three successive labels, and we suggest ideas that might be explored using CIS. PMID:27429742

  1. Combining combing and secondary ion mass spectrometry to study DNA on chips using 13C and 15N labeling

    PubMed Central

    Cabin-Flaman, Armelle; Monnier, Anne-Francoise; Coffinier, Yannick; Audinot, Jean-Nicolas; Gibouin, David; Wirtz, Tom; Boukherroub, Rabah; Migeon, Henri-Noël; Bensimon, Aaron; Jannière, Laurent; Ripoll, Camille; Norris, Victor

    2016-01-01

    Dynamic secondary ion mass spectrometry ( D-SIMS) imaging of combed DNA – the combing, imaging by SIMS or CIS method – has been developed previously using a standard NanoSIMS 50 to reveal, on the 50 nm scale, individual DNA fibers labeled with different, non-radioactive isotopes in vivo and to quantify these isotopes. This makes CIS especially suitable for determining the times, places and rates of DNA synthesis as well as the detection of the fine-scale re-arrangements of DNA and of molecules associated with combed DNA fibers. Here, we show how CIS may be extended to 13C-labeling via the detection and quantification of the 13C 14N - recombinant ion and the use of the 13C: 12C ratio, we discuss how CIS might permit three successive labels, and we suggest ideas that might be explored using CIS. PMID:27429742

  2. Modulating fluorescence anisotropy of dye-labeled DNA without involving mass amplification.

    PubMed

    Pei, Xiaojing; Huang, Hongduan; Chen, Yang; Li, Chenxi; Liu, Feng; Li, Na

    2016-07-01

    Fluorescence anisotropy, known as a simple, homogeneous and cost-effective analytical technology, is an invaluable technique for studying the micro-environmental changes of the dye associated with the molecular interactions. An in-depth understanding of the variables affecting the fluorescence anisotropy signal can facilitate better experimental designs to effectively improve the analytical performance. This work is a follow-up effort in evaluating the factors that can significantly influence fluorescence anisotropy. We systematically studied fluorescence anisotropy of dsDNA with the changing length based on dye-DNA interactions, with the fluorophores in the end-labeling, the middle-site-labeling, and multiple number of labeling manners. The fluorescence anisotropy value and the base-pair response dynamic range could be expanded by labeling the fluorophores in the middle of dsDNA and increasing the number of labels on dsDNA. The C overhang configuration in the end-labeling manner could enhance the fluorescence anisotropy signal but not expand the base-pair response range. Results from all the labeling fluorophores reinforced the leveling-off effect, i.e., the fluorescence anisotropy signal does not response to the increased length of the DNA duplex when the length is larger than a critical number of base pairs. These findings provide perspectives about choosing appropriate fluorescent dyes and labeling sites for simple and universal fluorescence anisotropy designs in various applications. PMID:27154716

  3. In-Gel Stable-Isotope Labeling (ISIL): a strategy for mass spectrometry-based relative quantification.

    PubMed

    Asara, John M; Zhang, Xiang; Zheng, Bin; Christofk, Heather H; Wu, Ning; Cantley, Lewis C

    2006-01-01

    Most proteomics approaches for relative quantification of protein expression use a combination of stable-isotope labeling and mass spectrometry. Traditionally, researchers have used difference gel electrophoresis (DIGE) from stained 1D and 2D gels for relative quantification. While differences in protein staining intensity can often be visualized, abundant proteins can obscure less abundant proteins, and quantification of post-translational modifications is difficult. A method is presented for quantifying changes in the abundance of a specific protein or changes in specific modifications of a protein using In-gel Stable-Isotope Labeling (ISIL). Proteins extracted from any source (tissue, cell line, immunoprecipitate, etc.), treated under two experimental conditions, are resolved in separate lanes by gel electrophoresis. The regions of interest (visualized by staining) are reacted separately with light versus heavy isotope-labeled reagents, and the gel slices are then mixed and digested with proteases. The resulting peptides are then analyzed by LC-MS to determine relative abundance of light/heavy isotope pairs and analyzed by LC-MS/MS for identification of sequence and modifications. The strategy compares well with other relative quantification strategies, and in silico calculations reveal its effectiveness as a global relative quantification strategy. An advantage of ISIL is that visualization of gel differences can be used as a first quantification step followed by accurate and sensitive protein level stable-isotope labeling and mass spectrometry-based relative quantification.

  4. Elemental labelling combined with liquid chromatography inductively coupled plasma mass spectrometry for quantification of biomolecules: A review

    PubMed Central

    Kretschy, Daniela; Koellensperger, Gunda; Hann, Stephan

    2012-01-01

    This article reviews novel quantification concepts where elemental labelling is combined with flow injection inductively coupled plasma mass spectrometry (FI-ICP-MS) or liquid chromatography inductively coupled plasma mass spectrometry (LC–ICP-MS), and employed for quantification of biomolecules such as proteins, peptides and related molecules in challenging sample matrices. In the first sections an overview on general aspects of biomolecule quantification, as well as of labelling will be presented emphasizing the potential, which lies in such methodological approaches. In this context, ICP-MS as detector provides high sensitivity, selectivity and robustness in biological samples and offers the capability for multiplexing and isotope dilution mass spectrometry (IDMS). Fundamental methodology of elemental labelling will be highlighted and analytical, as well as biomedical applications will be presented. A special focus will lie on established applications underlining benefits and bottlenecks of such approaches for the implementation in real life analysis. Key research made in this field will be summarized and a perspective for future developments including sophisticated and innovative applications will given. PMID:23062431

  5. Nano-scale liquid chromatography-mass spectrometry of 2-aminobenzamide-labeled oligosaccharides at low femtomole sensitivity

    NASA Astrophysics Data System (ADS)

    Wuhrer, Manfred; Koeleman, Carolien A. M.; Hokke, Cornelis H.; Deelder, André M.

    2004-03-01

    Conventional normal-phase high performance liquid chromatography of fluorescently labeled oligosaccharides with or without on-line mass spectrometry is an established tool for the structure characterization of protein and lipid derived glycans. Here we describe the miniaturization of such a system to the nano-scale using a 75 [mu]m internal diameter normal-phase amide column on-line with electrospray ionization ion-trap mass spectrometry. 2-Aminobenzamide-labeled oligosaccharides have a predictable retention on this normal-phase column that can be expressed as glucose units by comparison to the retention of a standard 2-aminobenzamide glucose polymer mixture. Isobaric compounds are separated on the basis of their structural differences, and by on-line electrospray ionization ion-trap mass spectrometry, the sequence of the monosaccharides can be deduced. The major improvement of the on-line nano-LC-MS system in comparison to conventional systems is the gain in sensitivity with detection of low femtomole amounts of glycans. This implies that LC-MS of 2-aminobenzamide-labeled oligosaccharides can now be performed at higher sensitivity than their analysis with fluorescence detection.

  6. Metal Oxide-Based Selective Enrichment Combined with Stable Isotope Labeling-Mass Spectrometry Analysis for Profiling of Ribose Conjugates.

    PubMed

    Chu, Jie-Mei; Qi, Chu-Bo; Huang, Yun-Qing; Jiang, Han-Peng; Hao, Yan-Hong; Yuan, Bi-Feng; Feng, Yu-Qi

    2015-07-21

    Some modified ribonucleosides in biological fluids have been evaluated as cancer-related metabolites. Detection of endogenous modified ribonucleosides in biological fluids may serve as a noninvasive cancers diagnostic method. However, determination of modified ribonucleosides is still challenging because of their low abundance and serious matrix interferences in biological fluids. Here, we developed a novel strategy for comprehensive profiling of ribose conjugates from biological fluids using metal oxide-based dispersive solid-phase extraction (DSPE) followed with in vitro stable isotope labeling and double neutral loss scan-mass spectrometry analysis (DSPE-SIL-LC-DNLS-MS). Cerium dioxide (CeO2) was used to selectively recognize and capture ribose conjugates from complex biological samples under basic environment. The enriched ribose conjugates were subsequently labeled with a pair of isotope labeling reagents (acetone and acetone-d6). The glucosidic bond of acetone labeled ribose conjugates is readily ruptured, and the generated ribose that carries an isotope tag can be lost as a neutral fragment under collision induced dissociation (CID). Since the light (acetone) and heavy (acetone-d6) labeled compounds have the same chemical structures and can generate different neutral loss fragments (NL 172 and 178 Da), it is therefore highly convenient to profile ribose conjugates by double neutral loss scan mode in mass spectrometry analysis. In this respect, the light and heavy labeled compounds were ionized at the same condition but recorded separately on MS spectra, which can significantly improve the detection specificity and facilitate the identification of ribose conjugates. Using the developed DSPE-SIL-LC-DNLS-MS strategy, we profiled the ribose conjugates in human urine, and 49 ribose conjugates were readily identified, among which 7 ribose conjugates exhibited significant contents change between healthy controls and lymphoma patients. The DSPE

  7. Proteomic Analysis of Protein Turnover by Metabolic Whole Rodent Pulse-Chase Isotopic Labeling and Shotgun Mass Spectrometry Analysis.

    PubMed

    Savas, Jeffrey N; Park, Sung Kyu; Yates, John R

    2016-01-01

    The analysis of protein half-life and degradation dynamics has proven critically important to our understanding of a broad and diverse set of biological conditions ranging from cancer to neurodegeneration. Historically these protein turnover measures have been performed in cells by monitoring protein levels after "pulse" labeling of newly synthesized proteins and subsequent chase periods. Comparing the level of labeled protein remaining as a function of time to the initial level reveals the protein's half-life. In this method we provide a detailed description of the workflow required for the determination of protein turnover rates on a whole proteome scale in vivo. Our approach starts with the metabolic labeling of whole rodents by restricting all the nitrogen in their diet to exclusively nitrogen-15 in the form of spirulina algae. After near complete organismal labeling with nitrogen-15, the rodents are then switched to a normal nitrogen-14 rich diet for time periods of days to years. Tissues are harvested, the extracts are fractionated, and the proteins are digested to peptides. Peptides are separated by multidimensional liquid chromatography and analyzed by high resolution orbitrap mass spectrometry (MS). The nitrogen-15 containing proteins are then identified and measured by the bioinformatic proteome analysis tools Sequest, DTASelect2, and Census. In this way, our metabolic pulse-chase approach reveals in vivo protein decay rates proteome-wide. PMID:26867752

  8. Quantitative Cross-linking/Mass Spectrometry Using Isotope-labeled Cross-linkers and MaxQuant.

    PubMed

    Chen, Zhuo A; Fischer, Lutz; Cox, Jürgen; Rappsilber, Juri

    2016-08-01

    The conceptually simple step from cross-linking/mass spectrometry (CLMS) to quantitative cross-linking/mass spectrometry (QCLMS) is compounded by technical challenges. Currently, quantitative proteomics software is tightly integrated with the protein identification workflow. This prevents automatically quantifying other m/z features in a targeted manner including those associated with cross-linked peptides. Here we present a new release of MaxQuant that permits starting the quantification process from an m/z feature list. Comparing the automated quantification to a carefully manually curated test set of cross-linked peptides obtained by cross-linking C3 and C3b with BS(3) and isotope-labeled BS(3)-d4 revealed a number of observations: (1) Fully automated process using MaxQuant can quantify cross-links in our reference data set with 68% recall rate and 88% accuracy. (2) Hidden quantification errors can be converted into exposed failures by label-swap replica, which makes label-swap replica an essential part of QCLMS. (3) Cross-links that failed during automated quantification can be recovered by semi-automated re-quantification. The integrated workflow of MaxQuant and semi-automated assessment provides the maximum of quantified cross-links. In contrast, work on larger data sets or by less experienced users will benefit from full automation in MaxQuant. PMID:27302889

  9. Stable isotope labeling tandem mass spectrometry (SILT): integration with peptide identification and extension to data-dependent scans.

    PubMed

    Elbert, Donald L; Mawuenyega, Kwasi G; Scott, Evan A; Wildsmith, Kristin R; Bateman, Randall J

    2008-10-01

    Quantitation of relative or absolute amounts of proteins by mass spectrometry can be prone to large errors. The use of MS/MS ion intensities and stable isotope labeling, which we term stable isotope labeling tandem mass spectrometry (SILT), decreases the effects of contamination from unrelated compounds. We present a software package (SILTmass) that automates protein identification and quantification by the SILT method. SILTmass has the ability to analyze the kinetics of protein turnover, in addition to relative and absolute protein quantitation. Instead of extracting chromatograms to find elution peaks, SILTmass uses only scans in which a peptide is identified and that meet an ion intensity threshold. Using only scans with identified peptides, the accuracy and precision of SILT is shown to be superior to precursor ion intensities, particularly at high or low dilutions of the isotope labeled compounds or with low amounts of protein. Using example scans, we demonstrate likely reasons for the improvements in quantitation by SILT. The appropriate use of variable modifications in peptide identification is described for measurement of protein turnover kinetics. The combination of identification with SILT facilitates quantitation without peak detection and helps to ensure the appropriate use of variable modifications for kinetics experiments.

  10. Quantitative Cross-linking/Mass Spectrometry Using Isotope-labeled Cross-linkers and MaxQuant*

    PubMed Central

    Cox, Jürgen

    2016-01-01

    The conceptually simple step from cross-linking/mass spectrometry (CLMS) to quantitative cross-linking/mass spectrometry (QCLMS) is compounded by technical challenges. Currently, quantitative proteomics software is tightly integrated with the protein identification workflow. This prevents automatically quantifying other m/z features in a targeted manner including those associated with cross-linked peptides. Here we present a new release of MaxQuant that permits starting the quantification process from an m/z feature list. Comparing the automated quantification to a carefully manually curated test set of cross-linked peptides obtained by cross-linking C3 and C3b with BS3 and isotope-labeled BS3-d4 revealed a number of observations: (1) Fully automated process using MaxQuant can quantify cross-links in our reference data set with 68% recall rate and 88% accuracy. (2) Hidden quantification errors can be converted into exposed failures by label-swap replica, which makes label-swap replica an essential part of QCLMS. (3) Cross-links that failed during automated quantification can be recovered by semi-automated re-quantification. The integrated workflow of MaxQuant and semi-automated assessment provides the maximum of quantified cross-links. In contrast, work on larger data sets or by less experienced users will benefit from full automation in MaxQuant. PMID:27302889

  11. Preprocessing significantly improves the peptide/protein identification sensitivity of high-resolution isobarically labeled tandem mass spectrometry data.

    PubMed

    Sheng, Quanhu; Li, Rongxia; Dai, Jie; Li, Qingrun; Su, Zhiduan; Guo, Yan; Li, Chen; Shyr, Yu; Zeng, Rong

    2015-02-01

    Isobaric labeling techniques coupled with high-resolution mass spectrometry have been widely employed in proteomic workflows requiring relative quantification. For each high-resolution tandem mass spectrum (MS/MS), isobaric labeling techniques can be used not only to quantify the peptide from different samples by reporter ions, but also to identify the peptide it is derived from. Because the ions related to isobaric labeling may act as noise in database searching, the MS/MS spectrum should be preprocessed before peptide or protein identification. In this article, we demonstrate that there are a lot of high-frequency, high-abundance isobaric related ions in the MS/MS spectrum, and removing isobaric related ions combined with deisotoping and deconvolution in MS/MS preprocessing procedures significantly improves the peptide/protein identification sensitivity. The user-friendly software package TurboRaw2MGF (v2.0) has been implemented for converting raw TIC data files to mascot generic format files and can be downloaded for free from https://github.com/shengqh/RCPA.Tools/releases as part of the software suite ProteomicsTools. The data have been deposited to the ProteomeXchange with identifier PXD000994. PMID:25435543

  12. Preprocessing significantly improves the peptide/protein identification sensitivity of high-resolution isobarically labeled tandem mass spectrometry data.

    PubMed

    Sheng, Quanhu; Li, Rongxia; Dai, Jie; Li, Qingrun; Su, Zhiduan; Guo, Yan; Li, Chen; Shyr, Yu; Zeng, Rong

    2015-02-01

    Isobaric labeling techniques coupled with high-resolution mass spectrometry have been widely employed in proteomic workflows requiring relative quantification. For each high-resolution tandem mass spectrum (MS/MS), isobaric labeling techniques can be used not only to quantify the peptide from different samples by reporter ions, but also to identify the peptide it is derived from. Because the ions related to isobaric labeling may act as noise in database searching, the MS/MS spectrum should be preprocessed before peptide or protein identification. In this article, we demonstrate that there are a lot of high-frequency, high-abundance isobaric related ions in the MS/MS spectrum, and removing isobaric related ions combined with deisotoping and deconvolution in MS/MS preprocessing procedures significantly improves the peptide/protein identification sensitivity. The user-friendly software package TurboRaw2MGF (v2.0) has been implemented for converting raw TIC data files to mascot generic format files and can be downloaded for free from https://github.com/shengqh/RCPA.Tools/releases as part of the software suite ProteomicsTools. The data have been deposited to the ProteomeXchange with identifier PXD000994.

  13. Stable isotope-labeled vitamin D, metabolites and chemical analogs: Synthesis and use in mass spectrometric studies

    SciTech Connect

    Coldwell, R.D.; Trafford, D.J.; Varley, M.J.; Kirk, D.N.; Makin, H.L. )

    1990-10-01

    Methods for the measurement of vitamin D and its metabolites using stable isotope-labeled internal standards and mass spectrometry are reviewed. The synthesis of both labeled and unlabeled standards is illustrated, and details of the synthesis of (26,26,27,27,27(-2)H5)-25,26-dihydroxyvitamin D3 and (28,28,28(-2)H3)-24,25-dihydroxyvitamin D2 are given. The use of in vitro biologic systems for the production of further metabolites of deuterated 25-hydroxyvitamin D3 is discussed. Use of deuterated 25-hydroxydihydrotachysterol3 as a substrate in the isolated perfused rat kidney has provided valuable data for the assignment of structure to a number of metabolites of 25-hydroxydihydrotachysterol3 formed in this system. 51 refs.

  14. Measuring the Composition and Stable-Isotope Labeling of Algal Biomass Carbohydrates via Gas Chromatography/Mass Spectrometry.

    PubMed

    McConnell, Brian O; Antoniewicz, Maciek R

    2016-05-01

    We have developed a method to measure carbohydrate composition and stable-isotope labeling in algal biomass using gas chromatography/mass spectrometry (GC/MS). The method consists of two-stage hydrochloric acid hydrolysis, followed by chemical derivatization of the released monomer sugars and quantification by GC/MS. Fully (13)C-labeled sugars are used as internal standards for composition analysis. This convenient, reliable, and accurate single-platform workflow offers advantages over existing methods and opens new opportunities to study carbohydrate metabolism of algae under autotrophic, mixotrophic, and heterotrophic conditions using metabolic flux analysis and isotopic tracers such as (2)H2O and (13)C-glucose. PMID:27042946

  15. Stable isotope labeling assisted liquid chromatography-electrospray tandem mass spectrometry for quantitative analysis of endogenous gibberellins.

    PubMed

    Hao, Yan-Hong; Zhang, Zheng; Wang, Lu; Liu, Chao; Lei, Ai-Wen; Yuan, Bi-Feng; Feng, Yu-Qi

    2015-11-01

    In the current study, we developed a stable isotope labeling strategy for the absolute quantification of gibberellins (GAs) by high performance liquid chromatography-electrospray tandem mass spectrometry (HPLC-ESI-MS/MS). N,N-dimethyl ethylenediamine (DMED) and its deuterated counterpart d(4)-DMED were used to derivatize GAs extracted from plant tissue samples and GA standards respectively. The both derivatives of GAs were mixed and then subjected to HPLC-ESI-MS/MS analysis. The absolute quantification of GAs in plant tissues could be achieved by calculating the peak area ratios of DMED labeled GAs/d(4)-DMED labeled GAs. In the proposed strategy, the derivatization reaction of the labeling reagents with GAs could be completed rapidly (within 5 min) with high efficiency (>99%) under mild conditions. The resulting derivatives could produce specific fragments in collision induced dissociation (CID), leading to high selectivity in multiple-reaction monitoring (MRM) mode, thus enhanced the reliability of the LC-MS/MS method. Furthermore, the limits of quantitation (LOQs) of GAs were considerably decreased (2-32 folds) due to incorporating easily ionized moieties into GAs, and the quantification of GAs in plant tissue could be achieved without isotopically labeled GA standards. Good linearity was obtained with correlation coefficients R(2) values of >0.99. The limits of detection (LODs) and quantitation (LOQs) ranged from 0.02 to 0.74 pg and 0.07 to 2.45 pg, respectively. Eleven GAs could be successfully determined in spiked sample with 72-128% recoveries and the relative standard deviations (RSDs) were between 1.0% and 13.9%. Finally, the developed method was successfully applied for the detection of GAs in 50mg (fresh weight) Oryza sativa leaves. PMID:26452831

  16. Stable isotope labeling assisted liquid chromatography-electrospray tandem mass spectrometry for quantitative analysis of endogenous gibberellins.

    PubMed

    Hao, Yan-Hong; Zhang, Zheng; Wang, Lu; Liu, Chao; Lei, Ai-Wen; Yuan, Bi-Feng; Feng, Yu-Qi

    2015-11-01

    In the current study, we developed a stable isotope labeling strategy for the absolute quantification of gibberellins (GAs) by high performance liquid chromatography-electrospray tandem mass spectrometry (HPLC-ESI-MS/MS). N,N-dimethyl ethylenediamine (DMED) and its deuterated counterpart d(4)-DMED were used to derivatize GAs extracted from plant tissue samples and GA standards respectively. The both derivatives of GAs were mixed and then subjected to HPLC-ESI-MS/MS analysis. The absolute quantification of GAs in plant tissues could be achieved by calculating the peak area ratios of DMED labeled GAs/d(4)-DMED labeled GAs. In the proposed strategy, the derivatization reaction of the labeling reagents with GAs could be completed rapidly (within 5 min) with high efficiency (>99%) under mild conditions. The resulting derivatives could produce specific fragments in collision induced dissociation (CID), leading to high selectivity in multiple-reaction monitoring (MRM) mode, thus enhanced the reliability of the LC-MS/MS method. Furthermore, the limits of quantitation (LOQs) of GAs were considerably decreased (2-32 folds) due to incorporating easily ionized moieties into GAs, and the quantification of GAs in plant tissue could be achieved without isotopically labeled GA standards. Good linearity was obtained with correlation coefficients R(2) values of >0.99. The limits of detection (LODs) and quantitation (LOQs) ranged from 0.02 to 0.74 pg and 0.07 to 2.45 pg, respectively. Eleven GAs could be successfully determined in spiked sample with 72-128% recoveries and the relative standard deviations (RSDs) were between 1.0% and 13.9%. Finally, the developed method was successfully applied for the detection of GAs in 50mg (fresh weight) Oryza sativa leaves.

  17. 76 FR 82129 - Medical Devices; Ovarian Adnexal Mass Assessment Score Test System; Labeling; Black Box Restrictions

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-12-30

    ... surgery. In the Federal Register of March 23, 2011 (76 FR 16292 at 12694), FDA published a final rule that... black box warning to address the risk of off-label use. In the Federal Register of March 23, 2011 (76 FR... Federal Register of March 23, 2011 (76 FR 16350 at 16352), FDA published a proposed rule to require...

  18. Structural analysis of alpha-helical proteins from wool using cysteine labelling and mass spectrometry.

    PubMed

    O'Cualain, R D M; Sims, P F G; Carr, C M

    2011-10-01

    A simple reduction/labelling/extraction protocol has been developed to fractionate cortical matrix proteins from filament proteins in wool. Through differential labelling of cysteine residues their relative accessibility in the wool fibre has been investigated. This has allowed the preliminary development of a map of the chemical functionality that is accessible within wool fibres under native conditions. Protein analyses of wool subjected to mechanical action, wet chemical permonosulphate/sulphite treatment and dry argon plasma treatment revealed that none of these detectably improved the accessibility of functional groups at the wool cortex. It is anticipated that this analytical method can be extended to improve the sensitivity and scope with which chemical functionality within native fibres can be mapped and lead to a better understanding of the potential limits/opportunities for fibre modification.

  19. Quantitative Analysis of rRNA Modifications Using Stable Isotope Labeling and Mass Spectrometry

    PubMed Central

    2015-01-01

    Post-transcriptional RNA modifications that are introduced during the multistep ribosome biogenesis process are essential for protein synthesis. The current lack of a comprehensive method for a fast quantitative analysis of rRNA modifications significantly limits our understanding of how individual modification steps are coordinated during biogenesis inside the cell. Here, an LC-MS approach has been developed and successfully applied for quantitative monitoring of 29 out of 36 modified residues in the 16S and 23S rRNA from Escherichia coli. An isotope labeling strategy is described for efficient identification of ribose and base methylations, and a novel metabolic labeling approach is presented to allow identification of MS-silent pseudouridine modifications. The method was used to measure relative abundances of modified residues in incomplete ribosomal subunits compared to a mature 15N-labeled rRNA standard, and a number of modifications in both 16S and 23S rRNA were present in substoichiometric amounts in the preribosomal particles. The RNA modification levels correlate well with previously obtained profiles for the ribosomal proteins, suggesting that RNA is modified in a schedule comparable to the association of the ribosomal proteins. Importantly, this study establishes an efficient workflow for a global monitoring of ribosomal modifications that will contribute to a better understanding of mechanisms of RNA modifications and their impact on intracellular processes in the future. PMID:24422502

  20. Label-free detection of differential protein expression by LC/MALDI mass spectrometry.

    PubMed

    Neubert, Hendrik; Bonnert, Timothy P; Rumpel, Klaus; Hunt, Brandon T; Henle, Ernst S; James, Ian T

    2008-06-01

    Protein abundance changes during disease or experimental perturbation are increasingly analyzed by label-free LC/MS approaches. Here we demonstrate the use of LC/MALDI MS for label-free detection of protein expression differences using Escherichia coli cultures grown on arabinose, fructose or glucose as a carbon source. The advantages of MALDI, such as detection of only singly charged ions, and MALDI plate archiving to facilitate retrospective MS/MS data collection are illustrated. MALDI spectra from RP chromatography of tryptic digests of the E. coli lysates were aligned and quantitated using the Rosetta Elucidator system. Approximately 5000 peptide signals were detected in all LC/MALDI runs spanning over 3 orders of magnitude of signal intensity. The average coefficients of variation for all signals across the entire intensity range in all technical replicates were found to be <25%. Pearson correlation coefficients from 0.93 to 0.98 for pairwise comparisons illustrate high replicate reproducibility. Expression differences determined by Analysis of Variance highlighted over 500 isotope clusters ( p < 0.01), which represented candidates for targeted peptide identification using MS/MS. Biologically interpretable protein identifications that could be derived underpin the general utility of this label-free LC/MALDI strategy.

  1. Stable isotope-labeled collagen: a novel and versatile tool for quantitative collagen analyses using mass spectrometry.

    PubMed

    Taga, Yuki; Kusubata, Masashi; Ogawa-Goto, Kiyoko; Hattori, Shunji

    2014-08-01

    Collagens are the most abundant proteins in animals and are involved in many physiological/pathological events. Although various methods have been used to quantify collagen and its post-translational modifications (PTMs) over the years, it is still difficult to accurately quantify type-specific collagen and minor collagen PTMs. We report a novel quantitative method targeting collagen using stable isotope-labeled collagen named "SI-collagen", which was labeled with isotopically heavy lysine, arginine, and proline in fibroblasts culture. We prepared highly labeled and purified SI-collagen for use as an internal standard in mass spectrometric analysis, particularly for a new approach using amino acid hydrolysis. Our method enabled accurate collagen analyses, including quantification of (1) type-specific collagen (types I and III in this paper), (2) total collagen, and (3) collagen PTMs by LC-MS with high sensitivity. SI-collagen is also applicable to other diverse analyses of collagen and can be a powerful tool for various studies, such as detailed investigation of collagen-related disorders.

  2. Label-free high-throughput screening via mass spectrometry: a single cystathionine quantitative method for multiple applications.

    PubMed

    Holt, Tom G; Choi, Bernard K; Geoghagen, Neil S; Jensen, Kristian K; Luo, Qi; LaMarr, William A; Makara, Gergely M; Malkowitz, Lorraine; Ozbal, Can C; Xiong, Yusheng; Dufresne, Claude; Luo, Ming-Juan

    2009-10-01

    Label-free mass spectrometric (MS) technologies are particularly useful for enzyme assay design for drug discovery screens. MS permits the selective detection of enzyme substrates or products in a wide range of biological matrices without need for derivatization, labeling, or capture technologies. As part of a cardiovascular drug discovery effort aimed at finding modulators of cystathionine beta-synthase (CBS), we used the RapidFire((R)) label-free high-throughput MS (HTMS) technology to develop a high-throughput screening (HTS) assay for CBS activity. The in vitro assay used HTMS to quantify the unlabeled product of the CBS reaction, cystathionine. Cystathionine HTMS analyses were carried out with a throughput of 7 s per sample and quantitation over a linear range of 80-10,000 nM. A compound library of 25,559 samples (or 80 384-well plates) was screened as singlets using the HTMS assay in a period of 8 days. With a hit rate of 0.32%, the actives showed a 90% confirmation rate. The in vitro assay was applied to secondary screens in more complex matrices with no additional analytical development. Our results show that the HTMS method was useful for screening samples containing serum, for cell-based assays, and for liver explants. The novel extension of the in vitro analytical method, without modification, to secondary assays resulted in a significant and advantageous economy of development time for the drug discovery project.

  3. Human Vitamin B12 Absorption and Metabolism are Measured by Accelerator Mass Spectrometry Using Specifically Labeled 14C-Cobalamin

    SciTech Connect

    Carkeet, C; Dueker, S R; Lango, J; Buchholz, B A; Miller, J W; Green, R; Hammock, B D; Roth, J R; Anderson, P J

    2006-01-26

    There is need for an improved test of human ability to assimilate dietary vitamin B{sub 12}. Assaying and understanding absorption and uptake of B{sub 12} is important because defects can lead to hematological and neurological complications. Accelerator mass spectrometry (AMS) is uniquely suited for assessing absorption and kinetics of {sup 14}C-labeled substances after oral ingestion because it is more sensitive than decay counting and can measure levels of carbon-14 ({sup 14}C) in microliter volumes of biological samples, with negligible exposure of subjects to radioactivity. The test we describe employs amounts of B{sub 12} in the range of normal dietary intake. The B{sub 12} used was quantitatively labeled with {sup 14}C at one particular atom of the DMB moiety by exploiting idiosyncrasies of Salmonellametabolism. In order to grow aerobically on ethanolamine, S. entericamust be provided with either pre-formed B{sub 12} or two of its precursors: cobinamide and dimethylbenzimidazole (DMB). When provided with {sup 14}C-DMB specifically labeled in the C2 position, cells produced {sup 14}C-B{sub 12} of high specific activity (2.1 GBq/mmol, 58 mCi/mmol) and no detectable dilution of label from endogenous DMB synthesis. In a human kinetic study, a physiological dose (1.5 mg, 2.2 KBq/59 nCi) of purified {sup 14}C-B{sub 12} was administered and showed plasma appearance and clearance curves consistent with the predicted behavior of the pure vitamin. This method opens new avenues for study of B{sub 12} assimilation.

  4. Probing Protein 3D Structures and Conformational Changes Using Electrochemistry-Assisted Isotope Labeling Cross-Linking Mass Spectrometry.

    PubMed

    Zheng, Qiuling; Zhang, Hao; Wu, Shiyong; Chen, Hao

    2016-05-01

    This study presents a new chemical cross-linking mass spectrometry (MS) method in combination with electrochemistry and isotope labeling strategy for probing both protein three-dimensional (3D) structures and conformational changes. For the former purpose, the target protein/protein complex is cross-linked with equal mole of premixed light and heavy isotope labeled cross-linkers carrying electrochemically reducible disulfide bonds (i.e., DSP-d0 and DSP-d8 in this study, DSP = dithiobis[succinimidyl propionate]), digested and then electrochemically reduced followed with online MS analysis. Cross-links can be quickly identified because of their reduced intensities upon electrolysis and the presence of doublet isotopic peak characteristics. In addition, electroreduction converts cross-links into linear peptides, facilitating MS/MS analysis to gain increased information about their sequences and modification sites. For the latter purpose of probing protein conformational changes, an altered procedure is adopted, in which the protein in two different conformations is cross-linked using DSP-d0 and DSP-d8 separately, and then the two protein samples are mixed in 1:1 molar ratio. The merged sample is subjected to digestion and electrochemical mass spectrometric analysis. In such a comparative cross-linking experiment, cross-links could still be rapidly recognized based on their responses to electrolysis. More importantly, the ion intensity ratios of light and heavy isotope labeled cross-links reveal the conformational changes of the protein, as exemplified by examining the effect of Ca(2+) on calmodulin conformation alternation. This new cross-linking MS method is fast and would have high value in structural biology. Graphical Abstract ᅟ. PMID:26902947

  5. Probing Protein 3D Structures and Conformational Changes Using Electrochemistry-Assisted Isotope Labeling Cross-Linking Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Zheng, Qiuling; Zhang, Hao; Wu, Shiyong; Chen, Hao

    2016-05-01

    This study presents a new chemical cross-linking mass spectrometry (MS) method in combination with electrochemistry and isotope labeling strategy for probing both protein three-dimensional (3D) structures and conformational changes. For the former purpose, the target protein/protein complex is cross-linked with equal mole of premixed light and heavy isotope labeled cross-linkers carrying electrochemically reducible disulfide bonds (i.e., DSP-d0 and DSP-d8 in this study, DSP = dithiobis[succinimidyl propionate]), digested and then electrochemically reduced followed with online MS analysis. Cross-links can be quickly identified because of their reduced intensities upon electrolysis and the presence of doublet isotopic peak characteristics. In addition, electroreduction converts cross-links into linear peptides, facilitating MS/MS analysis to gain increased information about their sequences and modification sites. For the latter purpose of probing protein conformational changes, an altered procedure is adopted, in which the protein in two different conformations is cross-linked using DSP-d0 and DSP-d8 separately, and then the two protein samples are mixed in 1:1 molar ratio. The merged sample is subjected to digestion and electrochemical mass spectrometric analysis. In such a comparative cross-linking experiment, cross-links could still be rapidly recognized based on their responses to electrolysis. More importantly, the ion intensity ratios of light and heavy isotope labeled cross-links reveal the conformational changes of the protein, as exemplified by examining the effect of Ca2+ on calmodulin conformation alternation. This new cross-linking MS method is fast and would have high value in structural biology.

  6. Probing Protein 3D Structures and Conformational Changes Using Electrochemistry-Assisted Isotope Labeling Cross-Linking Mass Spectrometry.

    PubMed

    Zheng, Qiuling; Zhang, Hao; Wu, Shiyong; Chen, Hao

    2016-05-01

    This study presents a new chemical cross-linking mass spectrometry (MS) method in combination with electrochemistry and isotope labeling strategy for probing both protein three-dimensional (3D) structures and conformational changes. For the former purpose, the target protein/protein complex is cross-linked with equal mole of premixed light and heavy isotope labeled cross-linkers carrying electrochemically reducible disulfide bonds (i.e., DSP-d0 and DSP-d8 in this study, DSP = dithiobis[succinimidyl propionate]), digested and then electrochemically reduced followed with online MS analysis. Cross-links can be quickly identified because of their reduced intensities upon electrolysis and the presence of doublet isotopic peak characteristics. In addition, electroreduction converts cross-links into linear peptides, facilitating MS/MS analysis to gain increased information about their sequences and modification sites. For the latter purpose of probing protein conformational changes, an altered procedure is adopted, in which the protein in two different conformations is cross-linked using DSP-d0 and DSP-d8 separately, and then the two protein samples are mixed in 1:1 molar ratio. The merged sample is subjected to digestion and electrochemical mass spectrometric analysis. In such a comparative cross-linking experiment, cross-links could still be rapidly recognized based on their responses to electrolysis. More importantly, the ion intensity ratios of light and heavy isotope labeled cross-links reveal the conformational changes of the protein, as exemplified by examining the effect of Ca(2+) on calmodulin conformation alternation. This new cross-linking MS method is fast and would have high value in structural biology. Graphical Abstract ᅟ.

  7. Determination of thiol metabolites in human urine by stable isotope labeling in combination with pseudo-targeted mass spectrometry analysis

    NASA Astrophysics Data System (ADS)

    Liu, Ping; Qi, Chu-Bo; Zhu, Quan-Fei; Yuan, Bi-Feng; Feng, Yu-Qi

    2016-02-01

    Precursor ion scan and multiple reaction monitoring scan (MRM) are two typical scan modes in mass spectrometry analysis. Here, we developed a strategy by combining stable isotope labeling (IL) with liquid chromatography-mass spectrometry (LC-MS) under double precursor ion scan (DPI) and MRM for analysis of thiols in 5 types of human cancer urine. Firstly, the IL-LC-DPI-MS method was applied for non-targeted profiling of thiols from cancer samples. Compared to traditional full scan mode, the DPI method significantly improved identification selectivity and accuracy. 103 thiol candidates were discovered in all cancers and 6 thiols were identified by their standards. It is worth noting that pantetheine, for the first time, was identified in human urine. Secondly, the IL-LC-MRM-MS method was developed for relative quantification of thiols in cancers compared to healthy controls. All the MRM transitions of light and heavy labeled thiols were acquired from urines by using DPI method. Compared to DPI method, the sensitivity of MRM improved by 2.1-11.3 folds. In addition, the concentration of homocysteine, γ-glutamylcysteine and pantetheine enhanced more than two folds in cancer patients compared to healthy controls. Taken together, the method demonstrated to be a promising strategy for identification and comprehensive quantification of thiols in human urines.

  8. Determination of thiol metabolites in human urine by stable isotope labeling in combination with pseudo-targeted mass spectrometry analysis

    PubMed Central

    Liu, Ping; Qi, Chu-Bo; Zhu, Quan-Fei; Yuan, Bi-Feng; Feng, Yu-Qi

    2016-01-01

    Precursor ion scan and multiple reaction monitoring scan (MRM) are two typical scan modes in mass spectrometry analysis. Here, we developed a strategy by combining stable isotope labeling (IL) with liquid chromatography-mass spectrometry (LC-MS) under double precursor ion scan (DPI) and MRM for analysis of thiols in 5 types of human cancer urine. Firstly, the IL-LC-DPI-MS method was applied for non-targeted profiling of thiols from cancer samples. Compared to traditional full scan mode, the DPI method significantly improved identification selectivity and accuracy. 103 thiol candidates were discovered in all cancers and 6 thiols were identified by their standards. It is worth noting that pantetheine, for the first time, was identified in human urine. Secondly, the IL-LC-MRM-MS method was developed for relative quantification of thiols in cancers compared to healthy controls. All the MRM transitions of light and heavy labeled thiols were acquired from urines by using DPI method. Compared to DPI method, the sensitivity of MRM improved by 2.1–11.3 folds. In addition, the concentration of homocysteine, γ-glutamylcysteine and pantetheine enhanced more than two folds in cancer patients compared to healthy controls. Taken together, the method demonstrated to be a promising strategy for identification and comprehensive quantification of thiols in human urines. PMID:26888486

  9. Conformational changes of recombinant monoclonal antibodies by limited proteolytic digestion, stable isotope labeling, and liquid chromatography-mass spectrometry.

    PubMed

    Ponniah, Gomathinayagam; Nowak, Christine; Kita, Adriana; Cheng, Guilong; Kori, Yekaterina; Liu, Hongcheng

    2016-03-15

    Limited proteolytic digestion is a method with a long history that has been used to study protein domain structures and conformational changes. A method of combining limited proteolytic digestion, stable isotope labeling, and mass spectrometry was established in the current study to investigate protein conformational changes. Recombinant monoclonal antibodies with or without the conserved oligosaccharides, and with or without oxidation of the conserved methionine residues, were used to test the newly proposed method. All of the samples were digested in ammonium bicarbonate buffer prepared in normal water. The oxidized deglycosylated sample was also digested in ammonium bicarbonate buffer prepared in (18)O-labeled water. The sample from the digestion in (18)O-water was spiked into each sample digested in normal water. Each mixed sample was subsequently analyzed by liquid chromatography-mass spectrometry (LC-MS). The molecular weight differences between the peptides digested in normal water versus (18)O-water were used to differentiate peaks from the samples. The relative peak intensities of peptides with or without the C-terminal incorporation of (18)O atoms were used to determine susceptibility of different samples to trypsin and chymotrypsin. The results demonstrated that the method was capable of detecting local conformational changes of the recombinant monoclonal antibodies caused by deglycosylation and oxidation. PMID:26747642

  10. Metabolomics relative quantitation with mass spectrometry using chemical derivatization and isotope labeling

    DOE PAGESBeta

    O'Maille, Grace; Go, Eden P.; Hoang, Linh; Want, Elizabeth J.; Smith, Colin; O'Maille, Paul; NordstrÖm, Anders; Morita, Hirotoshi; Qin, Chuan; Uritboonthai, Wilasinee; et al

    2008-01-01

    Comprehensive detection and quantitation of metabolites from a biological source constitute the major challenges of current metabolomics research. Two chemical derivatization methodologies, butylation and amination, were applied to human serum for ionization enhancement of a broad spectrum of metabolite classes, including steroids and amino acids. LC-ESI-MS analysis of the derivatized serum samples provided a significant signal elevation across the total ion chromatogram to over a 100-fold increase in ionization efficiency. It was also demonstrated that derivatization combined with isotopically labeled reagents facilitated the relative quantitation of derivatized metabolites from individual as well as pooled samples.

  11. Selected reaction monitoring (SRM) mass spectrometry without isotope labeling can be used for rapid protein quantification.

    PubMed

    Zhi, Wenbo; Wang, Meiyao; She, Jin-Xiong

    2011-06-15

    The validation of putative biomarker candidates has become the major bottle-neck in protein biomarker development. Conventional immunoaffinity methods are limited by the availability of antibodies and kits. Here we demonstrate the feasibility of using selected reaction monitoring (SRM) without isotope labeling to achieve fast and reproducible quantification of serum proteins. The SRM/MRM assays for three standard serum proteins, including ceruloplasmin (CP), serum aymloid A (SAA) and sex hormone binding globulin (SHBG), have good linear ranges, generally 10(3) to 10(4) . There are almost perfect correlations between SRM intensities and the loaded peptide amounts (R(2) is usually ~0.99). Our data suggest that SRM/MRM is able to quantify proteins within the range of 0.2-2 fmol, which is comparable to the commercial ELISA/LUMINEX kits for these proteins. Excellent correlations between SRM/MRM and ELISA/LUMINEX assays were observed for SAA and SHBG (R(2)=0.928 and 0.851, respectively). However, the correlation between SRM/MRM and ELISA for CP is less desirable (R(2)=0.565). The reproducibility for SRM/MRM assays is generally very good but may depend on the proteins/peptides being analyzed (R(2)=0.931 and 0.882 for SAA and SHBG, and 0.723 for CP). The SRM/MRM assay without isotope labeling is a rapid and useful method for protein biomarker validation in a modest number of samples and is especially useful when other assays such as ELISA or LUMINEX are not available. PMID:21594933

  12. Mass spectrometry-based, label-free quantitative proteomics of round spermatids in mice.

    PubMed

    Wang, Hailong; Li, Yan; Yang, Lijuan; Yu, Baofeng; Yan, Ping; Pang, Min; Li, Xiaobing; Yang, Hong; Zheng, Guoping; Xie, Jun; Guo, Rui

    2014-10-01

    Round haploid spermatids are formed at the completion of meiosis. These spermatids then undergo morphological and cytological changes during spermiogenesis. Although sperm proteomes have been extensively studied, relatively few studies have specifically investigated the proteome of round spermatids. We developed a label-free quantitative method in combination with 2D-nano-LC-ESI-MS/MS to investigate the proteome of round spermatids in mice. Analysis of the proteomic data identified 2,331 proteins in the round spermatids. Functional classification of the proteins based on Gene Ontology terms and enrichment analysis further revealed the following: 504 of the identified proteins are predicted to be involved in the generation of precursor metabolites and energy; 343 proteins in translation and protein targeting; 298 proteins in nucleotide and nucleic acid metabolism; 275 and 289 proteins in transport and cellular component organization, respectively. A number of the identified proteins were associated with cytoskeleton organization (183), protein degradation (116) and response to stimulus (115). KEGG pathway analysis identified 68 proteins that are annotated as components of the ribosomal pathway and 17 proteins were related to aminoacyl-tRNA biosynthesis. The round spermatids also contained 28 proteins involved in the proteasome pathway and 40 proteins in the lysosome pathway. A total of 60 proteins were annotated as parts of the spliceosome pathway, in which heterogeneous nuclear RNA is converted to mRNA. Approximately 94 proteins were identified as actin‑binding proteins, involved in the regulation of the actin cytoskeleton. In conclusion, using a label-free shotgun proteomic approach, we identified numerous proteins associated with spermiogenesis in round spermatids. PMID:25109358

  13. Stoichiometry of chromatin-associated protein complexes revealed by label-free quantitative mass spectrometry-based proteomics.

    PubMed

    Smits, Arne H; Jansen, Pascal W T C; Poser, Ina; Hyman, Anthony A; Vermeulen, Michiel

    2013-01-01

    Many cellular proteins assemble into macromolecular protein complexes. The identification of protein-protein interactions and quantification of their stoichiometry is therefore crucial to understand the molecular function of protein complexes. Determining the stoichiometry of protein complexes is usually achieved by mass spectrometry-based methods that rely on introducing stable isotope-labeled reference peptides into the sample of interest. However, these approaches are laborious and not suitable for high-throughput screenings. Here, we describe a robust and easy to implement label-free relative quantification approach that combines the detection of high-confidence protein-protein interactions with an accurate determination of the stoichiometry of the identified protein-protein interactions in a single experiment. We applied this method to two chromatin-associated protein complexes for which the stoichiometry thus far remained elusive: the MBD3/NuRD and PRC2 complex. For each of these complexes, we accurately determined the stoichiometry of the core subunits while at the same time identifying novel interactors and their stoichiometry.

  14. Production, Purification, and Characterization of 15N-Labeled DNA Repair Proteins as Internal Standards for Mass Spectrometric Measurements

    PubMed Central

    Jaruga, Pawel; Nelson, Bryant C.; Lowenthal, Mark S.; Jemth, Ann-Sofie; Loseva, Olga; Coskun, Erdem; Helleday, Thomas

    2016-01-01

    Oxidatively induced DNA damage is caused in living organisms by a variety of damaging agents, resulting in the formation of a multiplicity of lesions, which are mutagenic and cytotoxic. Unless repaired by DNA repair mechanisms before DNA replication, DNA lesions can lead to genomic instability, which is one of the hallmarks of cancer. Oxidatively induced DNA damage is mainly repaired by base excision repair pathway with the involvement of a plethora of proteins. Cancer tissues develop greater DNA repair capacity than normal tissues by overexpressing DNA repair proteins. Increased DNA repair in tumors that removes DNA lesions generated by therapeutic agents before they became toxic is a major mechanism in the development of therapy resistance. Evidence suggests that DNA repair capacity may be a predictive biomarker of patient response. Thus, knowledge of DNA–protein expressions in disease-free and cancerous tissues may help predict and guide development of treatments and yield the best therapeutic response. Our laboratory has developed methodologies that use mass spectrometry with isotope dilution for the measurement of expression of DNA repair proteins in human tissues and cultured cells. For this purpose, full-length 15N-labeled analogs of a number of human DNA repair proteins have been produced and purified to be used as internal standards for positive identification and accurate quantification. This chapter describes in detail the protocols of this work. The use of 15N-labeled proteins as internal standards for the measurement of several DNA repair proteins in vivo is also presented. PMID:26791985

  15. Dynamic Changes in Rat Mesenteric Lymph Proteins Following Trauma Using Label-Free Mass Spectrometry

    PubMed Central

    D’Alessandro, Angelo; Dzieciatkowska, Monika; Peltz, Erik D.; Moore, Ernest E.; Jordan, Janeen R.; Silliman, Christopher C.; Banerjee, Anirban; Hansen, Kirk C.

    2014-01-01

    Early events triggered by post-trauma/hemorrhagic shock currently represent a leading cause of morbidity and mortality in these patients. The causative agents of these events have been associated with increased neutrophil priming secondary to shock-dependent alterations of mesenteric lymph. Previous studies have suggested that unknown soluble components of the post-shock mesenteric lymph are main drivers of these events. In the present study, we applied a label free proteomics approach to further delve into the early proteome changes of the mesenteric lymph in response to hemorrhagic shock. Time-course analyses were performed by sampling the lymph every thirty minutes post-shock up until 3h (the time window within which a climax in neutrophil priming was observed). There are novel, transient early post-hemorrhagic shock alterations to the proteome and previously undocumented post-shock protein alterations. These results underlie the triggering of coagulation and pro-inflammatory responses secondary to trauma/hemorrhagic shock, metabolic deregulation and apoptosis, and alterations to proteases/anti-proteases homeostasis, which are suggestive of the potential implication of extracellular matrix proteases in priming neutrophil activation. Finally, there is a likely correlation between early PSML post-shock neutrophil priming and proteomics changes, above all protease/anti-proteases impaired homeostasis (especially of serine proteases and metalloproteases). PMID:25243424

  16. Cell Surface Proteome of Dental Pulp Stem Cells Identified by Label-Free Mass Spectrometry

    PubMed Central

    Niehage, Christian; Karbanová, Jana; Steenblock, Charlotte

    2016-01-01

    Multipotent mesenchymal stromal cells (MSCs) are promising tools for regenerative medicine. They can be isolated from different sources based on their plastic-adherence property. The identification of reliable cell surface markers thus becomes the Holy Grail for their prospective isolation. Here, we determine the cell surface proteomes of human dental pulp-derived MSCs isolated from single donors after culture expansion in low (2%) or high (10%) serum-containing media. Cell surface proteins were tagged on intact cells using cell impermeable, cleavable sulfo-NHS-SS-biotin, which allows their enrichment by streptavidin pull-down. For the proteomic analyses, we first compared label-free methods to analyze cell surface proteomes i.e. composition, enrichment and proteomic differences, and we developed a new mathematical model to determine cell surface protein enrichment using a combinatorial gene ontology query. Using this workflow, we identified 101 cluster of differentiation (CD) markers and 286 non-CD cell surface proteins. Based on this proteome profiling, we identified 14 cell surface proteins, which varied consistently in abundance when cells were cultured under low or high serum conditions. Collectively, our analytical methods provide a basis for identifying the cell surface proteome of dental pulp stem cells isolated from single donors and its evolution during culture or differentiation. Our data provide a comprehensive cell surface proteome for the precise identification of dental pulp-derived MSC populations and their isolation for potential therapeutic intervention. PMID:27490675

  17. Cell Surface Proteome of Dental Pulp Stem Cells Identified by Label-Free Mass Spectrometry.

    PubMed

    Niehage, Christian; Karbanová, Jana; Steenblock, Charlotte; Corbeil, Denis; Hoflack, Bernard

    2016-01-01

    Multipotent mesenchymal stromal cells (MSCs) are promising tools for regenerative medicine. They can be isolated from different sources based on their plastic-adherence property. The identification of reliable cell surface markers thus becomes the Holy Grail for their prospective isolation. Here, we determine the cell surface proteomes of human dental pulp-derived MSCs isolated from single donors after culture expansion in low (2%) or high (10%) serum-containing media. Cell surface proteins were tagged on intact cells using cell impermeable, cleavable sulfo-NHS-SS-biotin, which allows their enrichment by streptavidin pull-down. For the proteomic analyses, we first compared label-free methods to analyze cell surface proteomes i.e. composition, enrichment and proteomic differences, and we developed a new mathematical model to determine cell surface protein enrichment using a combinatorial gene ontology query. Using this workflow, we identified 101 cluster of differentiation (CD) markers and 286 non-CD cell surface proteins. Based on this proteome profiling, we identified 14 cell surface proteins, which varied consistently in abundance when cells were cultured under low or high serum conditions. Collectively, our analytical methods provide a basis for identifying the cell surface proteome of dental pulp stem cells isolated from single donors and its evolution during culture or differentiation. Our data provide a comprehensive cell surface proteome for the precise identification of dental pulp-derived MSC populations and their isolation for potential therapeutic intervention. PMID:27490675

  18. ARiBo pull-down for riboproteomic studies based on label-free quantitative mass spectrometry

    PubMed Central

    Di Tomasso, Geneviève; Miller Jenkins, Lisa M.

    2016-01-01

    As part of their normal life cycle, most RNA molecules associate with several proteins that direct their fate and regulate their function. Here, we describe a novel method for identifying proteins that associate with a target RNA. The procedure is based on the ARiBo method for affinity purification of RNA, which was originally developed to quickly purify RNA with high yields and purity under native conditions. The ARiBo method was further optimized using in vitro transcribed RNA to capture RNA-associating proteins from cellular extracts with high yields and low background protein contamination. For these RNA pull-downs, stem–loops present in the immature forms of let-7 miRNAs (miRNA stem–loops) were used as the target RNAs. Label-free quantitative mass spectrometry analysis allowed for the reliable identification of proteins that are specific to the stem–loops present in the immature forms of two miRNAs, let-7a-1 and let-7g. Several proteins known to bind immature forms of these let-7 miRNAs were identified, but with an improved coverage compared to previous studies. In addition, several novel proteins were identified that better define the protein interactome of the let-7 miRNA stem–loops and further link let-7 biogenesis to important biological processes such as development and tumorigenesis. Thus, combining the ARiBo pull-down method with label-free quantitative mass spectrometry provides an effective proteomic approach for identification of proteins that associate with a target RNA. PMID:27659051

  19. Residual CTAB Ligands as Mass Spectrometry Labels to Monitor Cellular Uptake of Au Nanorods.

    PubMed

    García, Isabel; Henriksen-Lacey, Malou; Sánchez-Iglesias, Ana; Grzelczak, Marek; Penadés, Soledad; Liz-Marzán, Luis M

    2015-06-01

    Gold nanorods have numerous applications in biomedical research, including diagnostics, bioimaging, and photothermal therapy. Even though surfactant removal and surface conjugation with antifouling molecules such as polyethylene glycol (PEG) are required to minimize nonspecific protein binding and cell uptake, the reliable characterization of these processes remains challenging. We propose here the use of laser desorption/ionization mass spectrometry (LDI-MS) to study the ligand exchange efficiency of cetyltrimethylammonium bromide (CTAB)-coated nanorods with different PEG grafting densities and to characterize nanorod internalization in cells. Application of LDI-MS analysis shows that residual CTAB consistently remains adsorbed on PEG-capped Au nanorods. Interestingly, such residual CTAB can be exploited as a mass barcode to discern the presence of nanorods in complex fluids and in vitro cellular systems, even at very low concentrations. PMID:26266492

  20. Phosphonium labeling for increasing metabolomic coverage of neutral lipids using electrospray ionization mass spectrometry†

    PubMed Central

    Woo, Hin-Koon; Go, Eden P.; Hoang, Linh; Trauger, Sunia A.; Bowen, Benjamin; Siuzdak, Gary; Northen, Trent R.

    2011-01-01

    Mass spectrometry has become an indispensable tool for the global study of metabolites (metabolomics), primarily using electrospray ionization mass spectrometry (ESI-MS). However, many important classes of molecules such as neutral lipids do not ionize well by ESI and go undetected. Chemical derivatization of metabolites can enhance ionization for increased sensitivity and metabolomic coverage. Here we describe the use of tris(2,4,6,-trimethoxyphenyl)phosphonium acetic acid (TMPP-AA) to improve liquid chromatography (LC)/ESI-MS detection of hydroxylated metabolites (i.e. lipids) from serum extracts. Cholesterol which is not normally detected from serum using ESI is observed with attomole sensitivity. This approach was applied to identify four endogenous lipids (hexadecanoyl-sn-glycerol, dihydrotachysterol, octadecanol, and alpha-tocopherol) from human serum. Overall, this approach extends the types of metabolites which can be detected using standard ESI-MS instrumentation and demonstrates the potential for targeted metabolomics analysis. PMID:19449318

  1. A neurosteroid analogue photolabeling reagent labels the colchicine binding site on tubulin: A mass spectrometric analysis

    PubMed Central

    Chen, Zi-Wei; Chen, Li-Hai; Akentieva, Natalia; Lichti, Cheryl F.; Darbandi, Ramin; Hastings, Randy; Covey, Douglas F.; Reichert, David E.; Townsend, R. Reid; Evers, Alex S.

    2013-01-01

    Previous studies have shown that the neurosteroid analogue, 6-Azi-pregnanolone (6-AziP), photolabels voltage-dependent anion channels and proteins of approximately 55 kDa in rat brain membranes. The present study used two dimensional electrophoresis and nano-electrospray ionization ion trap mass spectrometry (nano-ESI-MS) to identify the 55 kDa proteins (pI 4.8) as isoforms of β-tubulin. This identification was confirmed by immuno-blot and immunoprecipitation of photolabeled protein with anti-β-tubulin antibody and by the demonstration that 6-AziP photolabels purified bovine brain tubulin in a concentration-dependent pattern. To identify the photolabeling sites, purified bovine brain tubulin was photolabeled with 6-AziP, digested with trypsin, and analyzed by matrix-assisted laser desorption/ionization mass spectrometry (MALDI). A 6-AziP adduct of TAVCDIPPR (m/z=1287.77), a β-tubulin specific peptide, was detected by MALDI. High resolution LC-MS/MS analysis identified that 6-AziP was covalently bound to cysteine 354 (Cys-354), previously identified as a colchicine binding site. 6-AziP photolabeling was inhibited by 2-methoxyestradiol, an endogenous derivative of estradiol thought to bind to the colchicine site. Structural modeling predicted that neurosteroids could dock in this colchicine site at the interface between α- and β-tubulin with the photolabeling group of 6-AziP positioned proximate to Cys-354. PMID:22451060

  2. Label-free mass spectrometry proteome quantification of human embryonic kidney cells following 24 hours of sialic acid overproduction

    PubMed Central

    2013-01-01

    Background Cell surface glycoprotein sialylation is one of the most ubiquitous glycan modifications found on higher eukaryotes. The surface sialylation pattern of cells is influenced by the cellular environment but also by the Golgi sialyltransferase activity and flux of metabolites through sialic acid producing pathways. Altered cell surface sialic acid patterns have been observed in several cancers and other pathological conditions. In this experiment we examined the cellular proteomic changes that occur in human embryonic kidney cells after 24 hours of sialic acid overproduction using N-Acetylmannosamine. We utilized high resolution mass spectrometry and label free protein quantification to characterize the relative changes in protein abundance as well as multiple reaction monitoring to quantify the cellular sialic acid levels. Results Using N-Acetylmannosamine we were able to induce sialic acid production to almost 70-fold compared to non-induced control cells. Mass spectrometric analysis of cellular proteome of control and induced cells identified 1802 proteins of which 105 displayed significant changes in abundance. Functional analysis of the resulting relative changes in protein abundance revealed regulation of several cellular pathways including protein transport, metabolic and signaling pathways and remodeling of epithelial adherens junctions. We also identified several physically interacting co-regulated proteins in the set of changed proteins. Conclusions In this experiment we show that increased metabolic flux through sialic acid producing pathway affects the abundance of several protein transport, epithelial adherens junction, signaling and metabolic pathway related proteins. PMID:23915316

  3. A new approach for the comparative analysis of multiprotein complexes based on 15N metabolic labeling and quantitative mass spectrometry.

    PubMed

    Trompelt, Kerstin; Steinbeck, Janina; Terashima, Mia; Hippler, Michael

    2014-01-01

    The introduced protocol provides a tool for the analysis of multiprotein complexes in the thylakoid membrane, by revealing insights into complex composition under different conditions. In this protocol the approach is demonstrated by comparing the composition of the protein complex responsible for cyclic electron flow (CEF) in Chlamydomonas reinhardtii, isolated from genetically different strains. The procedure comprises the isolation of thylakoid membranes, followed by their separation into multiprotein complexes by sucrose density gradient centrifugation, SDS-PAGE, immunodetection and comparative, quantitative mass spectrometry (MS) based on differential metabolic labeling ((14)N/(15)N) of the analyzed strains. Detergent solubilized thylakoid membranes are loaded on sucrose density gradients at equal chlorophyll concentration. After ultracentrifugation, the gradients are separated into fractions, which are analyzed by mass-spectrometry based on equal volume. This approach allows the investigation of the composition within the gradient fractions and moreover to analyze the migration behavior of different proteins, especially focusing on ANR1, CAS, and PGRL1. Furthermore, this method is demonstrated by confirming the results with immunoblotting and additionally by supporting the findings from previous studies (the identification and PSI-dependent migration of proteins that were previously described to be part of the CEF-supercomplex such as PGRL1, FNR, and cyt f). Notably, this approach is applicable to address a broad range of questions for which this protocol can be adopted and e.g. used for comparative analyses of multiprotein complex composition isolated from distinct environmental conditions. PMID:24686495

  4. Morpheus Spectral Counter: A computational tool for label-free quantitative mass spectrometry using the Morpheus search engine.

    PubMed

    Gemperline, David C; Scalf, Mark; Smith, Lloyd M; Vierstra, Richard D

    2016-03-01

    Label-free quantitative MS based on the Normalized Spectral Abundance Factor (NSAF) has emerged as a straightforward and robust method to determine the relative abundance of individual proteins within complex mixtures. Here, we present Morpheus Spectral Counter (MSpC) as the first computational tool that directly calculates NSAF values from output obtained from Morpheus, a fast, open-source, peptide-MS/MS matching engine compatible with high-resolution accurate-mass instruments. NSAF has distinct advantages over other MS-based quantification methods, including a greater dynamic range as compared to isobaric tags, no requirement to align and re-extract MS1 peaks, and increased speed. MSpC features an easy-to-use graphic user interface that additionally calculates both distributed and unique NSAF values to permit analyses of both protein families and isoforms/proteoforms. MSpC determinations of protein concentration were linear over several orders of magnitude based on the analysis of several high-mass accuracy datasets either obtained from PRIDE or generated with total cell extracts spiked with purified Arabidopsis 20S proteasomes. The MSpC software was developed in C# and is open sourced under a permissive license with the code made available at http://dcgemperline.github.io/Morpheus_SpC/.

  5. Measurement of the isotope enrichment of stable isotope-labeled proteins using high-resolution mass spectra of peptides.

    PubMed

    MacCoss, Michael J; Wu, Christine C; Matthews, Dwight E; Yates, John R

    2005-12-01

    Stable isotope-enriched molecules are used as internal standards and as tracers of in vivo substrate metabolism. The accurate conversion of measured ratios in the mass spectrometer to mole ratios is complicated because a polyatomic molecule containing enriched atoms will result in a combinatorial distribution of isotopomers depending on the enrichment and number of "labeled" atoms. This effect could potentially cause a large error in the mole ratio measurement depending on which isotope peak or peaks were used to determine the ratio. We report a computational method that predicts isotope distributions over a range of enrichments and compares the predicted distributions to experimental peptide isotope distributions obtained by Fourier transform ion cyclotron resonance mass spectrometry. Our approach is accurate with measured enrichments within 1.5% of expected isotope distributions. The method is also precise with 4.9, 2.0, and 0.8% relative standard deviations for peptides containing 59, 79, and 99 atom % excess (15)N, respectively. The approach is automated making isotope enrichment calculations possible for thousands of peptides in a single muLC-FTICR-MS experiment.

  6. Mass Spectrometric Analysis of the Cell Surface N-Glycoproteome by Combining Metabolic Labeling and Click Chemistry

    NASA Astrophysics Data System (ADS)

    Smeekens, Johanna M.; Chen, Weixuan; Wu, Ronghu

    2015-04-01

    Cell surface N-glycoproteins play extraordinarily important roles in cell-cell communication, cell-matrix interactions, and cellular response to environmental cues. Global analysis is exceptionally challenging because many N-glycoproteins are present at low abundances and effective separation is difficult to achieve. Here, we have developed a novel strategy integrating metabolic labeling, copper-free click chemistry, and mass spectrometry (MS)-based proteomics methods to analyze cell surface N-glycoproteins comprehensively and site-specifically. A sugar analog containing an azido group, N-azidoacetylgalactosamine, was fed to cells to label glycoproteins. Glycoproteins with the functional group on the cell surface were then bound to dibenzocyclooctyne-sulfo-biotin via copper-free click chemistry under physiological conditions. After protein extraction and digestion, glycopeptides with the biotin tag were enriched by NeutrAvidin conjugated beads. Enriched glycopeptides were deglycosylated with peptide- N-glycosidase F in heavy-oxygen water, and in the process of glycan removal, asparagine was converted to aspartic acid and tagged with 18O for MS analysis. With this strategy, 144 unique N-glycopeptides containing 152 N-glycosylation sites were identified in 110 proteins in HEK293T cells. As expected, 95% of identified glycoproteins were membrane proteins, which were highly enriched. Many sites were located on important receptors, transporters, and cluster of differentiation proteins. The experimental results demonstrated that the current method is very effective for the comprehensive and site-specific identification of the cell surface N-glycoproteome and can be extensively applied to other cell surface protein studies.

  7. Mass spectrometric analysis of the cell surface N-glycoproteome by combining metabolic labeling and click chemistry.

    PubMed

    Smeekens, Johanna M; Chen, Weixuan; Wu, Ronghu

    2015-04-01

    Cell surface N-glycoproteins play extraordinarily important roles in cell-cell communication, cell-matrix interactions, and cellular response to environmental cues. Global analysis is exceptionally challenging because many N-glycoproteins are present at low abundances and effective separation is difficult to achieve. Here, we have developed a novel strategy integrating metabolic labeling, copper-free click chemistry, and mass spectrometry (MS)-based proteomics methods to analyze cell surface N-glycoproteins comprehensively and site-specifically. A sugar analog containing an azido group, N-azidoacetylgalactosamine, was fed to cells to label glycoproteins. Glycoproteins with the functional group on the cell surface were then bound to dibenzocyclooctyne-sulfo-biotin via copper-free click chemistry under physiological conditions. After protein extraction and digestion, glycopeptides with the biotin tag were enriched by NeutrAvidin conjugated beads. Enriched glycopeptides were deglycosylated with peptide-N-glycosidase F in heavy-oxygen water, and in the process of glycan removal, asparagine was converted to aspartic acid and tagged with 18O for MS analysis. With this strategy, 144 unique N-glycopeptides containing 152 N-glycosylation sites were identified in 110 proteins in HEK293T cells. As expected, 95% of identified glycoproteins were membrane proteins, which were highly enriched. Many sites were located on important receptors, transporters, and cluster of differentiation proteins. The experimental results demonstrated that the current method is very effective for the comprehensive and site-specific identification of the cell surface N-glycoproteome and can be extensively applied to other cell surface protein studies.

  8. New untargeted metabolic profiling combining mass spectrometry and isotopic labeling: application on Aspergillus fumigatus grown on wheat.

    PubMed

    Cano, Patricia M; Jamin, Emilien L; Tadrist, Souria; Bourdaud'hui, Pascal; Péan, Michel; Debrauwer, Laurent; Oswald, Isabelle P; Delaforge, Marcel; Puel, Olivier

    2013-09-01

    Characterization of fungal secondary metabolomes has become a challenge due to the industrial applications of many of these molecules, and also due to the emergence of fungal threats to public health and natural ecosystems. Given that, the aim of the present study was to develop an untargeted method to analyze fungal secondary metabolomes by combining high-accuracy mass spectrometry and double isotopic labeling of fungal metabolomes. The strain NRRL 35693 of Aspergillus fumigatus , an important fungal pathogen, was grown on three wheat grain substrates: (1) naturally enriched grains (99% (12)C), (2) grains enriched 96.8% with (13)C, (3) grains enriched with 53.4% with (13)C and 96.8% with (15)N. Twenty-one secondary metabolites were unambiguously identified by high-performance liquid chromatography-high-resolution mass spectrometry (HPLC-HRMS) analysis. AntiBase 2012 was used to confirm the identity of these metabolites. Additionally, on the basis of tandem mass spectrometry (MS(n)) experiments, it was possible to identify for the first time the formula and the structure of fumigaclavine D, a new member of the fumigaclavines family. Post biosynthesis degradation of tryptoquivaline F by methanol was also identified during HPLC-HRMS analysis by the detection of a carbon atom of nonfungal origin. The interest of this method lies not only on the unambiguous determination of the exact chemical formulas of fungal secondary metabolites but also on the easy discrimination of nonfungal products. Validation of the method was thus successfully achieved in this study, and it can now be applied to other fungal metabolomes, offering great possibilities for the discovery of new drugs or toxins. PMID:23901908

  9. Normalization Approaches for Removing Systematic Biases Associated with Mass Spectrometry and Label-Free Proteomics

    SciTech Connect

    Callister, Stephen J.; Barry, Richard C.; Adkins, Joshua N.; Johnson, Ethan T.; Qian, Weijun; Webb-Robertson, Bobbie-Jo M.; Smith, Richard D.; Lipton, Mary S.

    2006-02-01

    Central tendency, linear regression, locally weighted regression, and quantile techniques were investigated for normalization of peptide abundance measurements obtained from high-throughput liquid chromatography-Fourier transform ion cyclotron resonance mass spectrometry (LC-FTICR MS). Arbitrary abundances of peptides were obtained from three sample sets, including a standard protein sample, two Deinococcus radiodurans samples taken from different growth phases, and two mouse striatum samples from control and methamphetamine-stressed mice (strain C57BL/6). The selected normalization techniques were evaluated in both the absence and presence of biological variability by estimating extraneous variability prior to and following normalization. Prior to normalization, replicate runs from each sample set were observed to be statistically different, while following normalization replicate runs were no longer statistically different. Although all techniques reduced systematic bias, assigned ranks among the techniques revealed significant trends. For most LC-FTICR MS analyses, linear regression normalization ranked either first or second among the four techniques, suggesting that this technique was more generally suitable for reducing systematic biases.

  10. MASS LOSS AND NITROGEN DYNAMICS DURING THE DECOMPOSITION OF A N-LABELED N2-FIXING EPOPHYTIC LICHEN, LOBARIA OREGANA (TUCK.) MULL. ARG.

    EPA Science Inventory

    We studied mass loss and nitrogen dynamics during fall and spring initiated decomposition of an N2-fixing epiphytic lichen, Lobaria oregana (Tuck.) Mull. Arg. using 15N. We developed a method of labeling lichens with 15N that involved spraying lichen material with a nutrient sol...

  11. Comprehensive characterization of glioblastoma tumor tissues for biomarker identification using mass spectrometry-based label-free quantitative proteomics

    PubMed Central

    Heroux, Maxime S.; Chesnik, Marla A.; Halligan, Brian D.; Al-Gizawiy, Mona; Connelly, Jennifer M.; Mueller, Wade M.; Rand, Scott D.; Cochran, Elizabeth J.; LaViolette, Peter S.; Malkin, Mark G.; Schmainda, Kathleen M.

    2014-01-01

    Cancer is a complex disease; glioblastoma (GBM) is no exception. Short survival, poor prognosis, and very limited treatment options make it imperative to unravel the disease pathophysiology. The critically important identification of proteins that mediate various cellular events during disease is made possible with advancements in mass spectrometry (MS)-based proteomics. The objective of our study is to identify and characterize proteins that are differentially expressed in GBM to better understand their interactions and functions that lead to the disease condition. Further identification of upstream regulators will provide new potential therapeutic targets. We analyzed GBM tumors by SDS-PAGE fractionation with internal DNA markers followed by liquid chromatography-tandem mass spectrometry (MS). Brain tissue specimens obtained for clinical purposes during epilepsy surgeries were used as controls, and the quantification of MS data was performed by label-free spectral counting. The differentially expressed proteins were further characterized by Ingenuity Pathway Analysis (IPA) to identify protein interactions, functions, and upstream regulators. Our study identified several important proteins that are involved in GBM progression. The IPA revealed glioma activation with z score 2.236 during unbiased core analysis. Upstream regulators STAT3 and SP1 were activated and CTNNα was inhibited. We verified overexpression of several proteins by immunoblot to complement the MS data. This work represents an important step towards the identification of GBM biomarkers, which could open avenues to identify therapeutic targets for better treatment of GBM patients. The workflow developed represents a powerful and efficient method to identify biomarkers in GBM. PMID:24803679

  12. Chip electrophoresis of active banana ingredients with label-free detection utilizing deep UV native fluorescence and mass spectrometry.

    PubMed

    Ohla, Stefan; Schulze, Philipp; Fritzsche, Stefanie; Belder, Detlev

    2011-02-01

    In the present work, we report on a rapid and straightforward approach for the determination of biologically active compounds in bananas applying microchip electrophoresis (MCE). For this purpose, we applied label-free detection utilizing deep UV fluorescence detection with excitation at 266 nm. Using this approach, we could identify dopamine and serotonin, their precursors tryptophan and tyrosine and also the isoquinoline alkaloid salsolinol in less than 1 min. In bananas, after 10 days of ripening, we additionally found the compound levodopa which is a metabolite of the tyrosine pathway. Quantitative analysis of extracts by external calibration revealed concentrations of serotonin, tryptophan, and tyrosine from 2.7 to 7.6 μg/mL with relative standard deviations of less than 3.5%. The corresponding calibration plots showed good linearity with correlation coefficients higher than 0.985. For reliable peak assignment, the compounds were also analyzed by coupling chip electrophoresis with mass spectrometry. This paper demonstrates exemplarily the applicability of MCE with native fluorescence detection for rapid analysis of natural compounds in fruits and reveals the potential of chip-based separation systems for the analysis of complex mixtures. PMID:21181134

  13. Tracing nitrogenous disinfection byproducts after medium pressure UV water treatment by stable isotope labeling and high resolution mass spectrometry.

    PubMed

    Kolkman, Annemieke; Martijn, Bram J; Vughs, Dennis; Baken, Kirsten A; van Wezel, Annemarie P

    2015-04-01

    Advanced oxidation processes are important barriers for organic micropollutants (e.g., pharmaceuticals, pesticides) in (drinking) water treatment. Studies indicate that medium pressure (MP) UV/H2O2 treatment leads to a positive response in Ames mutagenicity tests, which is then removed after granulated activated carbon (GAC) filtration. The formed potentially mutagenic substances were hitherto not identified and may result from the reaction of photolysis products of nitrate with (photolysis products of) natural organic material (NOM). In this study we present an innovative approach to trace the formation of disinfection byproducts (DBPs) of MP UV water treatment, based on stable isotope labeled nitrate combined with high resolution mass spectrometry. It was shown that after MP UV treatment of artificial water containing NOM and nitrate, multiple nitrogen containing substances were formed. In total 84 N-DBPs were detected at individual concentrations between 1 to 135 ng/L bentazon-d6 equivalents, with a summed concentration of 1.2 μg/L bentazon-d6 equivalents. The chemical structures of three byproducts were confirmed. Screening for the 84 N-DBPs in water samples from a full-scale drinking water treatment plant based on MP UV/H2O2 treatment showed that 22 of the N-DBPs found in artificial water were also detected in real water samples.

  14. Relative Abundance of Integral Plasma Membrane Proteins in Arabidopsis Leaf and Root Tissue Determined by Metabolic Labeling and Mass Spectrometry

    PubMed Central

    Bernfur, Katja; Larsson, Olaf; Larsson, Christer; Gustavsson, Niklas

    2013-01-01

    Metabolic labeling of proteins with a stable isotope (15N) in intact Arabidopsis plants was used for accurate determination by mass spectrometry of differences in protein abundance between plasma membranes isolated from leaves and roots. In total, 703 proteins were identified, of which 188 were predicted to be integral membrane proteins. Major classes were transporters, receptors, proteins involved in membrane trafficking and cell wall-related proteins. Forty-one of the integral proteins, including nine of the 13 isoforms of the PIP (plasma membrane intrinsic protein) aquaporin subfamily, could be identified by peptides unique to these proteins, which made it possible to determine their relative abundance in leaf and root tissue. In addition, peptides shared between isoforms gave information on the proportions of these isoforms. A comparison between our data for protein levels and corresponding data for mRNA levels in the widely used database Genevestigator showed an agreement for only about two thirds of the proteins. By contrast, localization data available in the literature for 21 of the 41 proteins show a much better agreement with our data, in particular data based on immunostaining of proteins and GUS-staining of promoter activity. Thus, although mRNA levels may provide a useful approximation for protein levels, detection and quantification of isoform-specific peptides by proteomics should generate the most reliable data for the proteome. PMID:23990937

  15. Mass-tag labeling reveals site-specific and endogenous levels of protein S-fatty acylation.

    PubMed

    Percher, Avital; Ramakrishnan, Srinivasan; Thinon, Emmanuelle; Yuan, Xiaoqiu; Yount, Jacob S; Hang, Howard C

    2016-04-19

    Fatty acylation of cysteine residues provides spatial and temporal control of protein function in cells and regulates important biological pathways in eukaryotes. Although recent methods have improved the detection and proteomic analysis of cysteine fatty (S-fatty) acylated proteins, understanding how specific sites and quantitative levels of this posttranslational modification modulate cellular pathways are still challenging. To analyze the endogenous levels of protein S-fatty acylation in cells, we developed a mass-tag labeling method based on hydroxylamine-sensitivity of thioesters and selective maleimide-modification of cysteines, termed acyl-PEG exchange (APE). We demonstrate that APE enables sensitive detection of protein S-acylation levels and is broadly applicable to different classes of S-palmitoylated membrane proteins. Using APE, we show that endogenous interferon-induced transmembrane protein 3 is S-fatty acylated on three cysteine residues and site-specific modification of highly conserved cysteines are crucial for the antiviral activity of this IFN-stimulated immune effector. APE therefore provides a general and sensitive method for analyzing the endogenous levels of protein S-fatty acylation and should facilitate quantitative studies of this regulated and dynamic lipid modification in biological systems.

  16. Mass-tag labeling reveals site-specific and endogenous levels of protein S-fatty acylation.

    PubMed

    Percher, Avital; Ramakrishnan, Srinivasan; Thinon, Emmanuelle; Yuan, Xiaoqiu; Yount, Jacob S; Hang, Howard C

    2016-04-19

    Fatty acylation of cysteine residues provides spatial and temporal control of protein function in cells and regulates important biological pathways in eukaryotes. Although recent methods have improved the detection and proteomic analysis of cysteine fatty (S-fatty) acylated proteins, understanding how specific sites and quantitative levels of this posttranslational modification modulate cellular pathways are still challenging. To analyze the endogenous levels of protein S-fatty acylation in cells, we developed a mass-tag labeling method based on hydroxylamine-sensitivity of thioesters and selective maleimide-modification of cysteines, termed acyl-PEG exchange (APE). We demonstrate that APE enables sensitive detection of protein S-acylation levels and is broadly applicable to different classes of S-palmitoylated membrane proteins. Using APE, we show that endogenous interferon-induced transmembrane protein 3 is S-fatty acylated on three cysteine residues and site-specific modification of highly conserved cysteines are crucial for the antiviral activity of this IFN-stimulated immune effector. APE therefore provides a general and sensitive method for analyzing the endogenous levels of protein S-fatty acylation and should facilitate quantitative studies of this regulated and dynamic lipid modification in biological systems. PMID:27044110

  17. Use of an intravenous microdose of 14C-labeled drug and accelerator mass spectrometry to measure absolute oral bioavailability in dogs; cross-comparison of assay methods by accelerator mass spectrometry and liquid chromatography-tandem mass spectrometry.

    PubMed

    Miyaji, Yoshihiro; Ishizuka, Tomoko; Kawai, Kenji; Hamabe, Yoshimi; Miyaoka, Teiji; Oh-hara, Toshinari; Ikeda, Toshihiko; Kurihara, Atsushi

    2009-01-01

    A technique utilizing simultaneous intravenous microdosing of (14)C-labeled drug with oral dosing of non-labeled drug for measurement of absolute bioavailability was evaluated using R-142086 in male dogs. Plasma concentrations of R-142086 were measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and those of (14)C-R-142086 were measured by accelerator mass spectrometry (AMS). The absence of metabolites in the plasma and urine was confirmed by a single radioactive peak of the parent compound in the chromatogram after intravenous microdosing of (14)C-R-142086 (1.5 microg/kg). Although plasma concentrations of R-142086 determined by LC-MS/MS were approximately 20% higher than those of (14)C-R-142086 as determined by AMS, there was excellent correlation (r=0.994) between both concentrations after intravenous dosing of (14)C-R-142086 (0.3 mg/kg). The oral bioavailability of R-142086 at 1 mg/kg obtained by simultaneous intravenous microdosing of (14)C-R-142086 was 16.1%, this being slightly higher than the value (12.5%) obtained by separate intravenous dosing of R-142086 (0.3 mg/kg). In conclusion, on utilizing simultaneous intravenous microdosing of (14)C-labeled drug in conjunction with AMS analysis, absolute bioavailability could be approximately measured in dogs, but without total accuracy. Bioavailability in humans may possibly be approximately measured at an earlier stage and at a lower cost. PMID:19430168

  18. A New Strategy of Using O18-Labeled Iodoacetic Acid for Mass Spectrometry-Based Protein Quantitation

    NASA Astrophysics Data System (ADS)

    Wang, Shunhai; Kaltashov, Igor A.

    2012-07-01

    A new O18 labeling protocol is designed to assist quantitation of cysteine-containing proteins using LC/MS. Unlike other O18 labeling strategies, the labeling is carried out at the intact protein level (prior to its digestion) during reduction/alkylation of cysteine side chains using O18-labeled iodoacetic acid (IAA). The latter can be easily prepared by exchanging carboxylic oxygen atoms of commercially available IAA in O18-enriched water at low pH. Since incorporation of the O18 label in the protein occurs at the whole protein, rather than peptide level, the quantitation results are not peptide-dependent. The excellent stability of the label in mild pH conditions provides flexibility and robustness needed of sample processing steps following the labeling. In contrast to generally costly isotope labeling reagents, this approach uses only two relatively inexpensive commercially available reagents (IAA and H2O18). The feasibility of the new method is demonstrated using an 80 kDa human serum transferrin (hTf) as a model, where linear quantitation is achieved across a dynamic range spanning three orders of magnitude. The new approach can be used in quantitative proteomics applications and is particularly suitable for a variety of tasks in the biopharmaceutical sector, ranging from pharmacokinetic studies to quality control of protein therapeutics.

  19. Release of PCDD/PCDF to air and land during open burning of sugarcane and forest litter over soil fortified with mass labelled PCDD/PCDF

    NASA Astrophysics Data System (ADS)

    Black, Robert R.; (Mick) Meyer, Carl P.; Yates, Alan; Van Zwieten, Lukas; Chittim, Brock G.; Mueller, Jochen F.

    2012-11-01

    The contribution of PCDD/PCDF emissions from soil during open burning of biomass was examined. Mass labelled PCDD/PCDF was added to soil containing native PCDD/PCDF and biomass was laid out on this soil and burnt, simulating sugarcane trash and forest fires. Smoke samples were collected using a high volume portable field sampler. After each fire the concentration of all mass labelled PCDD/PCDF congeners in the surface soil decreased, however, the concentration of some native 2,3,7,8 substituted congeners increased, indicating that formation was occurring. Mass labelled PCDD/PCDF congeners were detected in all ash samples, mean 2.8 pg g-1 (range 0.5-8 pg g-1), demonstrating release from the soil. Additionally, mass labelled PCDD/PCDF congeners were detected in all air samples mean 1.2 μg (t fuel)-1 (range 0.2-2.0 μg (t fuel)-1), again demonstrating release from the soil. Native 2,3,7,8 substituted congeners detected in the air samples were dominated (in terms of contribution to total congener mass) by Cl8DD (90% for forest litter and 77% for sugarcane). The major contributor to TEQ of emissions from both forest litter and sugarcane was 1, 2, 3, 7, 8-Cl5DD (40-64% and 57-75%, respectively). These results demonstrate that release of PCDD/PCDF from soil to air and land occurs during open burning of biomass when soil temperatures are sufficiently elevated.

  20. Characterization of uniformly and atom-specifically 13C-labeled heparin and heparan sulfate polysaccharide precursors using 13C NMR spectroscopy and ESI mass spectrometry

    PubMed Central

    Nguyen, Thao K. N.; Tran, Vy M.; Victor, Xylophone V.; Skalicky, Jack J.; Kuberan, Balagurunathan

    2010-01-01

    The biological actions of heparin and heparan sulfate, two structurally related glycosaminoglycans, depend on the organization of the complex heparanome. Due to the structural complexity of the heparanome, the sequence of variably sulfonated uronic acid and glucosamine residues is usually characterized by the analysis of smaller oligosaccharide and disaccharide fragments. Even characterization of smaller heparin/heparan sulfate oligosaccharide or disaccharide fragments using simple 1D 1H NMR spectroscopy is often complicated by the extensive signal overlap. 13C NMR signals, on the other hand, overlap less and therefore, 13C NMR spectroscopy can greatly facilitate the structural elucidation of the complex heparanome and provide finer insights into the structural basis for biological functions. This is the first report of the preparation of anomeric carbon-specific 13C-labeled heparin/heparan sulfate precursors from the Escherichia coli K5 strain. Uniformly 13C- and 15N-labeled precursors were also produced and characterized by 13C NMR spectroscopy. Mass spectrometric analysis of enzymatically fragmented disaccharides revealed that anomeric carbon-specific labeling efforts resulted in a minor loss/scrambling of 13C in the precursor backbone, whereas uniform labeling efforts resulted in greater than 95% 13C isotope enrichment in the precursor backbone. These labeled precursors provided high-resolution NMR signals with great sensitivity and set the stage for studying the heparanome–proteome interactions. PMID:20832774

  1. Synthesis of deuterium-labeled 17-hydroxyprogesterone suitable as an internal standard for isotope dilution mass spectrometry

    SciTech Connect

    Shimizu, K.; Yamaga, N.; Kohara, H.

    1988-03-01

    A synthesis is reported of 17-hydroxyprogesterone, labeled with four atoms of deuterium at ring C and suitable for use as an internal standard for isotope dilution mass spectrometry. Base-catalyzed equilibration of methyl 3 alpha-acetoxy-12-oxo-cholanate (III) with /sup 2/H/sub 2/O, followed by reduction of the 12-oxo group by the modified Wolff-Kisher method using (/sup 2/H)diethylene glycol and (/sup 2/H)hydrazine hydrate afforded (11,11,12,12,23,23(-2)H)lithocholic acid (V). The Meystre-Miescher degradation of the side chain of V yielded 3 alpha-hydroxy-5 beta-(11,11,12,12(-2)H)pregnan-20-one (X). Oxidation of the 3,20-enol-diacetate of X with perbenzoic acid followed by saponification afforded 3 alpha,17-dihydroxy-5 beta-(11,11,12,12(-2)H)pregnan-20-one (XI). Oxidation of XI with N-bromoacetamide yielded 17-hydroxy-5 beta-(11,11,12,12(-2)H)pregnane-3,20-dione (XII). Bromination of XII followed by dehydrobromination yielded 17-hydroxy-(11,11,12,12(-2)H) progesterone (XIV), consisting of 0.3% /sup 2/H0-, 1.1% /sup 2/H/sub 1/-, 8.6% /sup 2/H/sub 2/-, 37.1% /sup 2/H/sub 3/-, 52.1% /sup 2/H/sub 4/-, and 0.8% /sup 2/H/sub 5/-species.

  2. Altered Retinoic Acid Metabolism in Diabetic Mouse Kidney Identified by 18O Isotopic Labeling and 2D Mass Spectrometry

    PubMed Central

    Starkey, Jonathan M.; Zhao, Yingxin; Sadygov, Rovshan G.; Haidacher, Sigmund J.; LeJeune, Wanda S.; Dey, Nilay; Luxon, Bruce A.; Kane, Maureen A.; Napoli, Joseph L.; Denner, Larry; Tilton, Ronald G.

    2010-01-01

    Background Numerous metabolic pathways have been implicated in diabetes-induced renal injury, yet few studies have utilized unbiased systems biology approaches for mapping the interconnectivity of diabetes-dysregulated proteins that are involved. We utilized a global, quantitative, differential proteomic approach to identify a novel retinoic acid hub in renal cortical protein networks dysregulated by type 2 diabetes. Methodology/Principal Findings Total proteins were extracted from renal cortex of control and db/db mice at 20 weeks of age (after 12 weeks of hyperglycemia in the diabetic mice). Following trypsinization, 18O- and 16O-labeled control and diabetic peptides, respectively, were pooled and separated by two dimensional liquid chromatography (strong cation exchange creating 60 fractions further separated by nano-HPLC), followed by peptide identification and quantification using mass spectrometry. Proteomic analysis identified 53 proteins with fold change ≥1.5 and p≤0.05 after Benjamini-Hochberg adjustment (out of 1,806 proteins identified), including alcohol dehydrogenase (ADH) and retinaldehyde dehydrogenase (RALDH1/ALDH1A1). Ingenuity Pathway Analysis identified altered retinoic acid as a key signaling hub that was altered in the diabetic renal cortical proteome. Western blotting and real-time PCR confirmed diabetes-induced upregulation of RALDH1, which was localized by immunofluorescence predominantly to the proximal tubule in the diabetic renal cortex, while PCR confirmed the downregulation of ADH identified with mass spectrometry. Despite increased renal cortical tissue levels of retinol and RALDH1 in db/db versus control mice, all-trans-retinoic acid was significantly decreased in association with a significant decrease in PPARβ/δ mRNA. Conclusions/Significance Our results indicate that retinoic acid metabolism is significantly dysregulated in diabetic kidneys, and suggest that a shift in all-trans-retinoic acid metabolism is a novel feature in

  3. Application of 13C isotope labeling using liquid chromatography mass spectrometry (LC-MS) to determining phosphate-containing metabolic incorporation

    PubMed Central

    Bhowmik, Salil Kumar; Putluri, Vasanta; Kommagani, Ramakrishna; Konde, Sai Aparna; Lydon, John P.; Sreekumar, Arun; Putluri, Nagireddy

    2016-01-01

    Here, we describe an approach wherein negative electrospray ionization mass spectrometry has used to understand the relative flux through phosphate containing metabolic intermediates associated with central carbon metabolism after administering cells with 13C-labeled substrates. The method was applied to examine the 13C incorporation through glycolysis in T47D breast cancer cells and showed reduction of glycolytic relative flux upon treatment with 2-Deoxyglucose. PMID:24338880

  4. Analysis of cytochrome P450 metabolites of arachidonic acid by stable isotope probe labeling coupled with ultra high-performance liquid chromatography/mass spectrometry.

    PubMed

    Zhu, Quan-Fei; Hao, Yan-Hong; Liu, Ming-Zhou; Yue, Jiang; Ni, Jian; Yuan, Bi-Feng; Feng, Yu-Qi

    2015-09-01

    Cytochrome P450 metabolites of arachidonic acid (AA) belong to eicosanoids and are potent lipid mediators of inflammation. It is well-known that eicosanoids play an important role in numerous pathophysiological processes. Therefore, quantitative analysis of cytochrome P450 metabolites of AA, including hydroxyeicosatetraenoic acids (HETEs), epoxyeicosatreinoic acids (EETs), and dihydroxyeicosatrienoic acids (DHETs) can provide crucial information to uncover underlying mechanisms of cytochrome P450 metabolites of AA related diseases. Herein, we developed a highly sensitive method to identify and quantify HETEs, EETs, and DHETs in lipid extracts of biological samples based on stable isotope probe labeling coupled with ultra high-performance liquid chromatography/mass spectrometry. To this end, a pair of stable isotope probes, 2-dimethylaminoethylamine (DMED) and d4-2-dimethylaminoethylamine (d4-DMED), were utilized to facilely label eicosanoids. The heavy labeled eicosanoid standards were prepared and used as internal standards for quantification to minimize the matrix and ion suppression effects in mass spectrometry analysis. In addition, the detection sensitivities of DMED labeled eicosanoids improved by 3-104 folds in standard solution and 5-138 folds in serum matrix compared with unlabeled analytes. Moreover, a good separation of eicosanoids isomers was achieved upon DMED labeling. The established method provided substantial sensitivity (limit of quantification at sub-picogram), high specificity, and broad linear dynamics range (3 orders of magnitude). We further quantified cytochrome P450 metabolites of AA in rat liver, heart, brain tissues and human serum using the developed method. The results showed that 19 eicosanoids could be distinctly detected and the contents of 11-, 15-, 16-, 20-HETE, 5,6-EET, and 14,15-EET in type 2 diabetes mellitus patients and 5-, 11-, 12-, 15-, 16-, 20-HETE, 8,9-EET, and 5,6-DHET in myeloid leukemia patients had significant changes

  5. Quantitative Proteomic Analysis of Wheat Seeds during Artificial Ageing and Priming Using the Isobaric Tandem Mass Tag Labeling.

    PubMed

    Lv, Yangyong; Zhang, Shuaibing; Wang, Jinshui; Hu, Yuansen

    2016-01-01

    Wheat (Triticum aestivum L.) is an important crop worldwide. The physiological deterioration of seeds during storage and seed priming is closely associated with germination, and thus contributes to plant growth and subsequent grain yields. In this study, wheat seeds during different stages of artificial ageing (45°C; 50% relative humidity; 98%, 50%, 20%, and 1% Germination rates) and priming (hydro-priming treatment) were subjected to proteomics analysis through a proteomic approach based on the isobaric tandem mass tag labeling. A total of 162 differentially expressed proteins (DEPs) mainly involved in metabolism, energy supply, and defense/stress responses, were identified during artificial ageing and thus validated previous physiological and biochemical studies. These DEPs indicated that the inability to protect against ageing leads to the incremental decomposition of the stored substance, impairment of metabolism and energy supply, and ultimately resulted in seed deterioration. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that the up-regulated proteins involved in seed ageing were mainly enriched in ribosome, whereas the down-regulated proteins were mainly accumulated in energy supply (starch and sucrose metabolism) and stress defense (ascorbate and aldarate metabolism). Proteins, including hemoglobin 1, oleosin, agglutinin, and non-specific lipid-transfer proteins, were first identified in aged seeds and might be regarded as new markers of seed deterioration. Of the identified proteins, 531 DEPs were recognized during seed priming compared with unprimed seeds. In contrast to the up-regulated DEPs in seed ageing, several up-regulated DEPs in priming were involved in energy supply (tricarboxylic acid cycle, glycolysis, and fatty acid oxidation), anabolism (amino acids, and fatty acid synthesis), and cell growth/division. KEGG and protein-protein interaction analysis indicated that the up-regulated proteins in seed priming were mainly

  6. Quantitative Proteomic Analysis of Wheat Seeds during Artificial Ageing and Priming Using the Isobaric Tandem Mass Tag Labeling.

    PubMed

    Lv, Yangyong; Zhang, Shuaibing; Wang, Jinshui; Hu, Yuansen

    2016-01-01

    Wheat (Triticum aestivum L.) is an important crop worldwide. The physiological deterioration of seeds during storage and seed priming is closely associated with germination, and thus contributes to plant growth and subsequent grain yields. In this study, wheat seeds during different stages of artificial ageing (45°C; 50% relative humidity; 98%, 50%, 20%, and 1% Germination rates) and priming (hydro-priming treatment) were subjected to proteomics analysis through a proteomic approach based on the isobaric tandem mass tag labeling. A total of 162 differentially expressed proteins (DEPs) mainly involved in metabolism, energy supply, and defense/stress responses, were identified during artificial ageing and thus validated previous physiological and biochemical studies. These DEPs indicated that the inability to protect against ageing leads to the incremental decomposition of the stored substance, impairment of metabolism and energy supply, and ultimately resulted in seed deterioration. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that the up-regulated proteins involved in seed ageing were mainly enriched in ribosome, whereas the down-regulated proteins were mainly accumulated in energy supply (starch and sucrose metabolism) and stress defense (ascorbate and aldarate metabolism). Proteins, including hemoglobin 1, oleosin, agglutinin, and non-specific lipid-transfer proteins, were first identified in aged seeds and might be regarded as new markers of seed deterioration. Of the identified proteins, 531 DEPs were recognized during seed priming compared with unprimed seeds. In contrast to the up-regulated DEPs in seed ageing, several up-regulated DEPs in priming were involved in energy supply (tricarboxylic acid cycle, glycolysis, and fatty acid oxidation), anabolism (amino acids, and fatty acid synthesis), and cell growth/division. KEGG and protein-protein interaction analysis indicated that the up-regulated proteins in seed priming were mainly

  7. Quantitative Proteomic Analysis of Wheat Seeds during Artificial Ageing and Priming Using the Isobaric Tandem Mass Tag Labeling

    PubMed Central

    Lv, Yangyong; Zhang, Shuaibing; Wang, Jinshui; Hu, Yuansen

    2016-01-01

    Wheat (Triticum aestivum L.) is an important crop worldwide. The physiological deterioration of seeds during storage and seed priming is closely associated with germination, and thus contributes to plant growth and subsequent grain yields. In this study, wheat seeds during different stages of artificial ageing (45°C; 50% relative humidity; 98%, 50%, 20%, and 1% Germination rates) and priming (hydro-priming treatment) were subjected to proteomics analysis through a proteomic approach based on the isobaric tandem mass tag labeling. A total of 162 differentially expressed proteins (DEPs) mainly involved in metabolism, energy supply, and defense/stress responses, were identified during artificial ageing and thus validated previous physiological and biochemical studies. These DEPs indicated that the inability to protect against ageing leads to the incremental decomposition of the stored substance, impairment of metabolism and energy supply, and ultimately resulted in seed deterioration. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that the up-regulated proteins involved in seed ageing were mainly enriched in ribosome, whereas the down-regulated proteins were mainly accumulated in energy supply (starch and sucrose metabolism) and stress defense (ascorbate and aldarate metabolism). Proteins, including hemoglobin 1, oleosin, agglutinin, and non-specific lipid-transfer proteins, were first identified in aged seeds and might be regarded as new markers of seed deterioration. Of the identified proteins, 531 DEPs were recognized during seed priming compared with unprimed seeds. In contrast to the up-regulated DEPs in seed ageing, several up-regulated DEPs in priming were involved in energy supply (tricarboxylic acid cycle, glycolysis, and fatty acid oxidation), anabolism (amino acids, and fatty acid synthesis), and cell growth/division. KEGG and protein-protein interaction analysis indicated that the up-regulated proteins in seed priming were mainly

  8. 18O-labeled proteome reference as global internal standards for targeted quantification by selected reaction monitoring-mass spectrometry.

    PubMed

    Kim, Jong-Seo; Fillmore, Thomas L; Liu, Tao; Robinson, Errol; Hossain, Mahmud; Champion, Boyd L; Moore, Ronald J; Camp, David G; Smith, Richard D; Qian, Wei-Jun

    2011-12-01

    Selected reaction monitoring (SRM)-MS is an emerging technology for high throughput targeted protein quantification and verification in biomarker discovery studies; however, the cost associated with the application of stable isotope-labeled synthetic peptides as internal standards can be prohibitive for screening a large number of candidate proteins as often required in the preverification phase of discovery studies. Herein we present a proof of concept study using an (18)O-labeled proteome reference as global internal standards (GIS) for SRM-based relative quantification. The (18)O-labeled proteome reference (or GIS) can be readily prepared and contains a heavy isotope ((18)O)-labeled internal standard for every possible tryptic peptide. Our results showed that the percentage of heavy isotope ((18)O) incorporation applying an improved protocol was >99.5% for most peptides investigated. The accuracy, reproducibility, and linear dynamic range of quantification were further assessed based on known ratios of standard proteins spiked into the labeled mouse plasma reference. Reliable quantification was observed with high reproducibility (i.e. coefficient of variance <10%) for analyte concentrations that were set at 100-fold higher or lower than those of the GIS based on the light ((16)O)/heavy ((18)O) peak area ratios. The utility of (18)O-labeled GIS was further illustrated by accurate relative quantification of 45 major human plasma proteins. Moreover, quantification of the concentrations of C-reactive protein and prostate-specific antigen was illustrated by coupling the GIS with standard additions of purified protein standards. Collectively, our results demonstrated that the use of (18)O-labeled proteome reference as GIS provides a convenient, low cost, and effective strategy for relative quantification of a large number of candidate proteins in biological or clinical samples using SRM.

  9. Qualitative Metabolome Analysis of Human Cerebrospinal Fluid by 13C-/12C-Isotope Dansylation Labeling Combined with Liquid Chromatography Fourier Transform Ion Cyclotron Resonance Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Guo, Kevin; Bamforth, Fiona; Li, Liang

    2011-02-01

    Metabolome analysis of human cerebrospinal fluid (CSF) is challenging because of low abundance of metabolites present in a small volume of sample. We describe and apply a sensitive isotope labeling LC-MS technique for qualitative analysis of the CSF metabolome. After a CSF sample is divided into two aliquots, they are labeled by 13C-dansyl and 12C-dansyl chloride, respectively. The differentially labeled aliquots are then mixed and subjected to LC-MS using Fourier-transform ion cyclotron resonance mass spectrometry (FTICR MS). Dansylation offers significant improvement in the performance of chromatography separation and detection sensitivity. Moreover, peaks detected in the mass spectra can be readily analyzed for ion pair recognition and database search based on accurate mass and/or retention time information. It is shown that about 14,000 features can be detected in a 25-min LC-FTICR MS run of a dansyl-labeled CSF sample, from which about 500 metabolites can be profiled. Results from four CSF samples are compared to gauge the detectability of metabolites by this method. About 261 metabolites are commonly detected in replicate runs of four samples. In total, 1132 unique metabolite ion pairs are detected and 347 pairs (31%) matched with at least one metabolite in the Human Metabolome Database. We also report a dansylation library of 220 standard compounds and, using this library, about 85 metabolites can be positively identified. Among them, 21 metabolites have never been reported to be associated with CSF. These results illustrate that the dansylation LC-FTICR MS method can be used to analyze the CSF metabolome in a more comprehensive manner.

  10. Multi-Isotope Secondary Ion Mass Spectrometry Combining Heavy Water 2H with 15N Labeling As Complementary Tracers for Metabolic Heterogeneity at the Single-Cell Level

    NASA Astrophysics Data System (ADS)

    Kopf, S.; McGlynn, S.; Cowley, E.; Green, A.; Newman, D. K.; Orphan, V. J.

    2014-12-01

    Metabolic rates of microbial communities constitute a key physiological parameter for understanding the in situ growth constraints for life in any environment. Isotope labeling techniques provide a powerful approach for measuring such biological activity, due to the use of isotopically enriched substrate tracers whose incorporation into biological materials can be detected with high sensitivity by isotope-ratio mass spectrometry. Nano-meter scale secondary ion mass spectrometry (NanoSIMS) combined with stable isotope labeling provides a unique tool for studying the spatiometabolic activity of microbial populations at the single cell level in order to assess both community structure and population diversity. However, assessing the distribution and range of microbial activity in complex environmental systems with slow-growing organisms, diverse carbon and nitrogen sources, or heterotrophic subpopulations poses a tremendous technical challenge because the introduction of isotopically labeled substrates frequently changes the nutrient availability and can inflate or bias measures of activity. Here, we present the use of hydrogen isotope labeling with deuterated water as an important new addition to the isotopic toolkit and apply it for the determination of single cell microbial activities by NanoSIMS imaging. This tool provides a labeling technique that minimally alters any aquatic chemical environment, can be administered with strong labels even in minimal addition (natural background is very low), is an equally universal substrate for all forms of life even in complex, carbon and nitrogen saturated systems, and can be combined with other isotopic tracers. The combination of heavy water labeling with the most commonly used NanoSIMS tracer, 15N, is technically challenging but opens up a powerful new set of multi-tracer experiments for the study of microbial activity in complex communities. We present the first truly simultaneous single cell triple isotope system

  11. Combining Capillary Electrophoresis Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry and Stable Isotopic Labeling Techniques for Comparative Crustacean Peptidomics

    PubMed Central

    Wang, Junhua; Zhang, Yuzhuo; Xiang, Feng; Zhang, Zichuan; Li, Lingjun

    2010-01-01

    Herein we describe a sensitive and straightforward off-line capillary electrophoresis (CE) matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) interface in conjunction with stable isotopic labeling (SIL) technique for comparative neuropeptidomic analysis in crustacean model organisms. Two SIL schemes, including a binary H/D formaldehyde labeling technique and novel, laboratory-developed multiplexed dimethylated leucine-based isobaric tagging reagents, have been evaluated in these proof-of-concept experiments. We employ these isotopic labeling techniques in conjunction with CE-MALDI MS for quantitative peptidomic analyses of the pericardial organs isolated from two crustacean species, the European green crab Carcinus maenas and the blue crab Callinectes sapidus. Isotopically labeled peptide pairs are found to co-migrate in CE fractions and quantitative changes in relative abundances of peptide pairs are obtained by comparing peak intensities of respective peptide pairs. Several neuropeptide families exhibit changes in response to salinity stress, suggesting potential physiological functions of these signaling peptides. PMID:20334868

  12. Automation of dimethylation after guanidination labeling chemistry and its compatibility with common buffers and surfactants for mass spectrometry-based shotgun quantitative proteome analysis.

    PubMed

    Lo, Andy; Tang, Yanan; Chen, Lu; Li, Liang

    2013-07-25

    Isotope labeling liquid chromatography-mass spectrometry (LC-MS) is a major analytical platform for quantitative proteome analysis. Incorporation of isotopes used to distinguish samples plays a critical role in the success of this strategy. In this work, we optimized and automated a chemical derivatization protocol (dimethylation after guanidination, 2MEGA) to increase the labeling reproducibility and reduce human intervention. We also evaluated the reagent compatibility of this protocol to handle biological samples in different types of buffers and surfactants. A commercially available liquid handler was used for reagent dispensation to minimize analyst intervention and at least twenty protein digest samples could be prepared in a single run. Different front-end sample preparation methods for protein solubilization (SDS, urea, Rapigest™, and ProteaseMAX™) and two commercially available cell lysis buffers were evaluated for compatibility with the automated protocol. It was found that better than 94% desired labeling could be obtained in all conditions studied except urea, where the rate was reduced to about 92% due to carbamylation on the peptide amines. This work illustrates the automated 2MEGA labeling process can be used to handle a wide range of protein samples containing various reagents that are often encountered in protein sample preparation for quantitative proteome analysis.

  13. Expression, purification, and mass spectrometric analysis of 15N, 13C-labeled RGD-hirudin, expressed in Pichia pastoris, for NMR studies.

    PubMed

    Huang, Yinong; Zhang, Yanling; Wu, Yi; Wang, Jue; Liu, Xingang; Dai, Linsen; Wang, Longsheng; Yu, Min; Mo, Wei

    2012-01-01

    A novel recombinant hirudin, RGD-hirudin, inhibits the activity of thrombin and the aggregation of platelets. Here, we successfully expressed (15)N, (13)C-labeled RGD-hirudin in Pichia pastoris in a fermenter. The protein was subsequently purified to yield sufficient quantities for structural and functional studies. The purified protein was characterized by HPLC and MALDI-TOF mass spectroscopy. Analysis revealed that the protein was pure and uniformly labeled with (15)N and (13)C. A bioassay showed that the anti-thrombin activity and the anti-platelet aggregation ability of the labeled protein were the same as those of unlabeled RGD-hirudin. Multidimensional heteronuclear NMR spectroscopy has been used to determine almost complete backbone (15)N, (13)C and (1)H resonance assignments of the r-RGD-Hirudin. The (15)N-(1)H HSQC spectrum of uniformly (15)N, (13)C-labeled RGD-hirudin allowed successful assignment of the signals. Examples of the quality of the data are provided for the (15)N-(l)H correlation spectrum, and by selected planes of the CBCA(CO)NH, CBCANH, and HNCO experiments. These results provide a basis for further studies on the structure-function relationship of RGD-hirudin with thrombin and platelets. PMID:22879918

  14. The use of label-free mass spectrometry for relative quantification of sarcoplasmic proteins during the processing of dry-cured ham.

    PubMed

    Gallego, Marta; Mora, Leticia; Concepción Aristoy, M; Toldrá, Fidel

    2016-04-01

    The aim of this work was to quantify changes in the abundance of the major sarcoplasmic proteins throughout the ham dry-curing process by using a label-free mass spectrometry methodology based on the measurement of mass spectral peak intensities obtained from the extracted ion chromatogram. For this purpose, extraction of sarcoplasmic proteins was followed by trypsin digestion and analysis by nanoliquid chromatography coupled to tandem mass spectrometry (Q/TOF) for the identification and relative quantification of sarcoplasmic proteins through individual quantification of trypsinised peptides. In total, 20 proteins, including 12 glycolytic enzymes, were identified and quantified. The accuracy of the protocol was based on MS/MS replicates, and beta-lactoglobulin protein was used to normalise data and correct possible variations during sample preparation or LC-MS/MS analysis. Mass spectrometry-based proteomics provides precise identification and quantification of proteins in comparison with traditional methodologies based on gel electrophoresis, especially in the case of overlapping proteins. Moreover, the label-free approach used in this study proved to be a simple, fast, reliable method for evaluating proteolytic degradation of sarcoplasmic proteins during the processing of dry-cured ham.

  15. High-Performance Chemical Isotope Labeling Liquid Chromatography-Mass Spectrometry for Profiling the Metabolomic Reprogramming Elicited by Ammonium Limitation in Yeast.

    PubMed

    Luo, Xian; Zhao, Shuang; Huan, Tao; Sun, Difei; Friis, R Magnus N; Schultz, Michael C; Li, Liang

    2016-05-01

    Information about how yeast metabolism is rewired in response to internal and external cues can inform the development of metabolic engineering strategies for food, fuel, and chemical production in this organism. We report a new metabolomics workflow for the characterization of such metabolic rewiring. The workflow combines efficient cell lysis without using chemicals that may interfere with downstream sample analysis and differential chemical isotope labeling liquid chromatography mass spectrometry (CIL LC-MS) for in-depth yeast metabolome profiling. Using (12)C- and (13)C-dansylation (Dns) labeling to analyze the amine/phenol submetabolome, we detected and quantified a total of 5719 peak pairs or metabolites. Among them, 120 metabolites were positively identified using a library of 275 Dns-metabolite standards, and 2980 metabolites were putatively identified based on accurate mass matches to metabolome databases. We also applied (12)C- and (13)C-dimethylaminophenacyl (DmPA) labeling to profile the carboxylic acid submetabolome and detected over 2286 peak pairs, from which 33 metabolites were positively identified using a library of 188 DmPA-metabolite standards, and 1595 metabolites were putatively identified. Using this workflow for metabolomic profiling of cells challenged by ammonium limitation revealed unexpected links between ammonium assimilation and pantothenate accumulation that might be amenable to engineering for better acetyl-CoA production in yeast. We anticipate that efforts to improve other schemes of metabolic engineering will benefit from application of this workflow to multiple cell types. PMID:26947805

  16. Statistical Model to Analyze Quantitative Proteomics Data Obtained by 18O/16O Labeling and Linear Ion Trap Mass Spectrometry

    PubMed Central

    Jorge, Inmaculada; Navarro, Pedro; Martínez-Acedo, Pablo; Núñez, Estefanía; Serrano, Horacio; Alfranca, Arántzazu; Redondo, Juan Miguel; Vázquez, Jesús

    2009-01-01

    Statistical models for the analysis of protein expression changes by stable isotope labeling are still poorly developed, particularly for data obtained by 16O/18O labeling. Besides large scale test experiments to validate the null hypothesis are lacking. Although the study of mechanisms underlying biological actions promoted by vascular endothelial growth factor (VEGF) on endothelial cells is of considerable interest, quantitative proteomics studies on this subject are scarce and have been performed after exposing cells to the factor for long periods of time. In this work we present the largest quantitative proteomics study to date on the short term effects of VEGF on human umbilical vein endothelial cells by 18O/16O labeling. Current statistical models based on normality and variance homogeneity were found unsuitable to describe the null hypothesis in a large scale test experiment performed on these cells, producing false expression changes. A random effects model was developed including four different sources of variance at the spectrum-fitting, scan, peptide, and protein levels. With the new model the number of outliers at scan and peptide levels was negligible in three large scale experiments, and only one false protein expression change was observed in the test experiment among more than 1000 proteins. The new model allowed the detection of significant protein expression changes upon VEGF stimulation for 4 and 8 h. The consistency of the changes observed at 4 h was confirmed by a replica at a smaller scale and further validated by Western blot analysis of some proteins. Most of the observed changes have not been described previously and are consistent with a pattern of protein expression that dynamically changes over time following the evolution of the angiogenic response. With this statistical model the 18O labeling approach emerges as a very promising and robust alternative to perform quantitative proteomics studies at a depth of several thousand proteins

  17. Tc-99m-labeled red blood cells for the measurement of red cell mass in newborn infants: concise communication

    SciTech Connect

    Linderkamp, O.; Betke, K.; Fendel, H.; Klemm, J.; Lorenzen, K.; Riegel, K.P.

    1980-07-01

    In vitro and in vivo investigations were performed to examine the binding of Tc-99m to neonatal red blood cells (RBC). Labeling efficiency was about 90%, and unbound Tc-99m less than 3% after one washing, in premature and full-term newborns and in children. Thus presence of high percentages of fetal hemoglobin (Hb F) did not influence the labeling of RBCs with Tc-99m. RBCs of 11 newborns were hemolysed and the distribution of Tc-99m on RBC components was analyzed. Although Hb F percentage averaged (60.0 +- 8.10)% (s.d.), only (11.9 +- 3.7)% of Tc-99m was bound by Hb F, whereas (45.0 +- 6.1)% was associated with Hb A. RBC membranes bound (13.7 +- 4.3)% and (29.3 +- 4.0)% were found unbound in hemolysates. These results indicate that Tc-99m preferentially binds to beta chains. In vivo equilibration of Tc-99m RBCs and of albumin labeled with Evans blue was investigated in five newborn infants. Tc-99m RBCs were stable in each case during the first hour after injection. Elution of Tc-99m from RBCs was (3.4 +- 1.5)% per h. Body-to-venous hematocrit ratio averaged 0.86 +- 0.03.

  18. Analysis of 13C labeling enrichment in microbial culture applying metabolic tracer experiments using gas chromatography-combustion-isotope ratio mass spectrometry.

    PubMed

    Heinzle, Elmar; Yuan, Yongbo; Kumar, Sathish; Wittmann, Christoph; Gehre, Matthias; Richnow, Hans-Herrmann; Wehrung, Patrick; Adam, Pierre; Albrecht, Pierre

    2008-09-15

    The applicability of gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS) for the quantification of 13C enrichment of proteinogenic amino acids in metabolic tracer experiments was evaluated. Measurement of the 13C enrichment of proteinogenic amino acids from cell hydrolyzates of Corynebacterium glutamicum growing on different mixtures containing between 0.5 and 10% [1-13C]glucose shows the significance of kinetic isotope effects in metabolic flux studies at low degree of labeling. We developed a method to calculate the 13C enrichment. The approach to correct for these effects in metabolic flux studies using delta13C measurement by GC-C-IRMS uses two parallel experiments applying substrate with natural abundance and 13C-enriched tracer substrate, respectively. The fractional enrichment obtained in natural substrate is subtracted from that of the enriched one. Tracer studies with C. glutamicum resulted in a statistically identical relative fractional enrichment of 13C in proteinogenic amino acids over the whole range of applied concentrations of [1-13C]glucose. The current findings indicate a great potential of GC-C-IRMS for labeling quantification in 13C metabolic flux analysis with low labeling degree of tracer substrate directly in larger scale bioreactors.

  19. Gold nanoparticles labeling with hybridization chain reaction amplification strategy for the sensitive detection of HepG2 cells by inductively coupled plasma mass spectrometry.

    PubMed

    Zhang, Xing; Chen, Beibei; He, Man; Wang, Han; Hu, Bin

    2016-12-15

    Sensitive detection of circulating tumor cells (CTCs) is of great significance in the early detection of cancer and cancer metastasis. This work reported an efficient, specific, and sensitive immunoassay protocol for detection of tumor cells by using inductively coupled plasma mass spectrometry (ICP-MS) with gold nanoparticles (AuNPs) labeling and hybridization chain reaction (HCR) amplification. In the established approach, antibodies against epithelial cell adhesion molecule (anti-EpCAM) conjugated magnetic beads (MBs) were used for selective capture of tumor cells from peripheral blood, aptamer was applied for the recognition of captured tumor cells, and AuNPs labeled DNA concatamer was used as the signal probe for tumor cell labeling and ICP-MS detection. Due to the dual amplification effect of AuNPs and HCR, the limit of detection of this ICP-MS based method for HepG2 cells was as low as 15 cells, and the linear range was 40-8000 cells with the relative standard deviation for seven replicate detections of 200 HepG2 cells was 8.7%. Furthermore, the applicability of the method for the analysis of peripheral blood samples was demonstrated by the spiking tests. The established method was highly specific and sensitive for the detection of HepG2 cells, and has a good application potential in clinical diagnosis.

  20. Differential quantitative proteomics of Porphyromonas gingivalis by linear ion trap mass spectrometry: Non-label methods comparison, q-values and LOWESS curve fitting

    NASA Astrophysics Data System (ADS)

    Xia, Qiangwei; Wang, Tiansong; Park, Yoonsuk; Lamont, Richard J.; Hackett, Murray

    2007-01-01

    Differential analysis of whole cell proteomes by mass spectrometry has largely been applied using various forms of stable isotope labeling. While metabolic stable isotope labeling has been the method of choice, it is often not possible to apply such an approach. Four different label free ways of calculating expression ratios in a classic "two-state" experiment are compared: signal intensity at the peptide level, signal intensity at the protein level, spectral counting at the peptide level, and spectral counting at the protein level. The quantitative data were mined from a dataset of 1245 qualitatively identified proteins, about 56% of the protein encoding open reading frames from Porphyromonas gingivalis, a Gram-negative intracellular pathogen being studied under extracellular and intracellular conditions. Two different control populations were compared against P. gingivalis internalized within a model human target cell line. The q-value statistic, a measure of false discovery rate previously applied to transcription microarrays, was applied to proteomics data. For spectral counting, the most logically consistent estimate of random error came from applying the locally weighted scatter plot smoothing procedure (LOWESS) to the most extreme ratios generated from a control technical replicate, thus setting upper and lower bounds for the region of experimentally observed random error.

  1. Quantitative isomer-specific N-glycan fingerprinting using isotope coded labeling and high performance liquid chromatography-electrospray ionization-mass spectrometry with graphitic carbon stationary phase.

    PubMed

    Michael, Claudia; Rizzi, Andreas M

    2015-02-27

    Glycan reductive isotope labeling (GRIL) using (12)C6-/(13)C6-aniline as labeling reagent is reported with the aim of quantitative N-glycan fingerprinting. Porous graphitized carbon (PGC) as stationary phase in capillary scale HPLC coupled to electrospray mass spectrometry with time of flight analyzer was applied for the determination of labeled N-glycans released from glycoproteins. The main benefit of using stable isotope-coding in the context of comparative glycomics lies in the improved accuracy and precision of the quantitative analysis in combined samples and in the potential of correcting for structure-dependent incomplete enzymatic release of oligosaccharides when comparing identical target proteins. The method was validated with respect to mobile phase parameters, reproducibility, accuracy, linearity and limit of detection/quantification (LOD/LOQ) using test glycoproteins. It is shown that the developed method is capable of determining relative amounts of N-glycans (including isomers) comparing two samples in one single HPLC-MS run. The analytical potential and usefulness of GRIL in combination with PGC-ESI-TOF-MS is demonstrated comparing glycosylation in human monoclonal antibodies produced in Chinese hamster ovary cells (CHO) and hybridoma cell lines.

  2. Development of isotope labeling liquid chromatography mass spectrometry for mouse urine metabolomics: quantitative metabolomic study of transgenic mice related to Alzheimer's disease.

    PubMed

    Peng, Jun; Guo, Kevin; Xia, Jianguo; Zhou, Jianjun; Yang, Jing; Westaway, David; Wishart, David S; Li, Liang

    2014-10-01

    Because of a limited volume of urine that can be collected from a mouse, it is very difficult to apply the common strategy of using multiple analytical techniques to analyze the metabolites to increase the metabolome coverage for mouse urine metabolomics. We report an enabling method based on differential isotope labeling liquid chromatography mass spectrometry (LC-MS) for relative quantification of over 950 putative metabolites using 20 μL of urine as the starting material. The workflow involves aliquoting 10 μL of an individual urine sample for ¹²C-dansylation labeling that target amines and phenols. Another 10 μL of aliquot was taken from each sample to generate a pooled sample that was subjected to ¹³C-dansylation labeling. The ¹²C-labeled individual sample was mixed with an equal volume of the ¹³C-labeled pooled sample. The mixture was then analyzed by LC-MS to generate information on metabolite concentration differences among different individual samples. The interday repeatability for the LC-MS runs was assessed, and the median relative standard deviation over 4 days was 5.0%. This workflow was then applied to a metabolomic biomarker discovery study using urine samples obtained from the TgCRND8 mouse model of early onset familial Alzheimer's disease (FAD) throughout the course of their pathological deposition of beta amyloid (Aβ). It was showed that there was a distinct metabolomic separation between the AD prone mice and the wild type (control) group. As early as 15-17 weeks of age (presymptomatic), metabolomic differences were observed between the two groups, and after the age of 25 weeks the metabolomic alterations became more pronounced. The metabolomic changes at different ages corroborated well with the phenotype changes in this transgenic mice model. Several useful candidate biomarkers including methionine, desaminotyrosine, taurine, N1-acetylspermidine, and 5-hydroxyindoleacetic acid were identified. Some of them were found in previous

  3. Quantification of peptide m/z distributions from 13C-labeled cultures with high resolution mass spectrometry

    Technology Transfer Automated Retrieval System (TEKTRAN)

    With the introduction of orbital trap mass spectrometers molecular masses can be determined with great precision and accuracy. In addition, orbital trap spectrometers (Orbitraps) are sensitive and possess a linear dynamic range of multiple orders of magnitude. These qualities make the Orbitrap well-...

  4. Balancing the (carbon) budget: Using linear inverse models to estimate carbon flows and mass-balance 13C:15N labelling experiments in low oxygen sediments.

    NASA Astrophysics Data System (ADS)

    Hunter, William Ross; Van Oevelen, Dick; Witte, Ursula

    2013-04-01

    Over 1 million km2 of seafloor experience permanent low-oxygen conditions within oxygen minimum zones (OMZs). OMZs are predicted to grow as a consequence of climate change, potentially affecting oceanic biogeochemical cycles. The Arabian Sea OMZ impinges upon the western Indian continental margin at bathyal depths (150 - 1500m) producing a strong depth dependent oxygen gradient at the sea floor. The influence of the OMZ upon the short term processing of organic matter by sediment ecosystems was investigated using in situ stable isotope pulse chase experiments. These deployed doses of 13C:15N labeled organic matter onto the sediment surface at four stations from across the OMZ (water depth 540 - 1100 m; [O2] = 0.35 - 15 μM). In order to prevent experimentally anoxia, the mesocosms were not sealed. 13C and 15N labels were traced into sediment, bacteria, fauna and 13C into sediment porewater DIC and DOC. However, the DIC and DOC flux to the water column could not be measured, limiting our capacity to obtain mass-balance for C in each experimental mesocosm. Linear Inverse Modeling (LIM) provides a method to obtain a mass-balanced model of carbon flow that integrates stable-isotope tracer data with community biomass and biogeochemical flux data from a range of sources. Here we present an adaptation of the LIM methodology used to investigate how ecosystem structure influenced carbon flow across the Indian margin OMZ. We demonstrate how oxygen conditions affect food-web complexity, affecting the linkages between the bacteria, foraminifera and metazoan fauna, and their contributions to benthic respiration. The food-web models demonstrate how changes in ecosystem complexity are associated with oxygen availability across the OMZ and allow us to obtain a complete carbon budget for the stationa where stable-isotope labelling experiments were conducted.

  5. Ag+ labeling: a convenient new tool for the characterization of hydrogen-bonded supramolecular assemblies by MALDI-TOF mass spectrometry.

    PubMed

    Timmerman, P; Jolliffe, K A; Calama, M C; Weidmann, J L; Prins, L J; Cardullo, F; Snellink-Ruël, B H; Fokkens, R H; Nibbering, N M; Shinkai, S; Reinhoudt, D N

    2000-11-17

    Herein we describe our results on the characterization of a wide variety of different hydrogen-bonded assemblies by means of a novel matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) technique with Ag+ labeling. The labeling technique with Ag+ ions is extremely mild and provides a nondestructive way to generate charged assemblies that can be detected by mass spectrometry. Up to now more than 25 different single (1(3).2(3)), double (3(3).2(6)), and tetrarosettes (4(3).2(12)) have been successfully characterized by the use of this method. The success of the method entirely depends on the presence of a suitable binding site for the Ag+ ion. A variety of functionalities has been identified that provide strong binding sites for Ag+, either acting in a cooperative way (pi-arene and pi-alkene donor functionalities) or individually (cyano and crown ether functionalities). The method works well for assemblies with molecular weights between 2,000 and 8,000 Da, and most likely far beyond this limit.

  6. Determination of sup 13 C labeling pattern of citric acid cycle intermediates by gas chromatography-mass spectrometry

    SciTech Connect

    Di Donato, L.; Montgomery, J.A.; Des Rosiers, C.; David, F.; Garneau, M.; Brunengraber, H. )

    1990-02-26

    Investigations of the regulation of the citric acid cycle require determination of labeling patterns of cycle intermediates. These were assayed to date, using infusion of: (i) ({sup 14}C)tracer followed by chemical degradation of intermediates and (ii) ({sup 13}C)tracer followed by NMR analysis of intermediates. The authors developed a strategy to analyze by GC-MS the ({sup 13}C) labeling pattern of {mu}mole samples of citrate (CIT), isocitrate (ICIT), 2-ketoglutarate (2-KG), glutamate (GLU) and glutamine (GLN). These are enzymatically or chemically converted to 2-KG, ICIT, 4-aminobutyrate (GABA) and 2-hydroxyglutarate (2-OHG). GC-MS analyses of TMS or TBDMS derivatives of these compounds yield the enrichment of each carbon. The authors confirmed the identity of each fragment using the spectra of (1-{sup 13}C), (5-{sup 13}C), (2,3,3,4,4-{sup 2}H{sub 5})glutamate and (1-{sup 13}C), (1,4-{sup 13}C)GABA.

  7. Laccase-catalyzed reactions of 17β-estradiol in the presence of humic acid: Resolved by high-resolution mass spectrometry in combination with (13)C labeling.

    PubMed

    Sun, Kai; Luo, Qi; Gao, Yanzheng; Huang, Qingguo

    2016-02-01

    The widespread presence of estrogens in natural waters poses potential threats to the aquatic organisms and human health. It is known that estrogens undergo enzyme-catalyzed oxidative coupling (ECOC) reactions, which may impact their environmental fate and can be used in wastewater treatment to remove estrogens, but little information is available on how natural organic matter (NOM) may influence 17β-estradiol (E2) transformation in ECOC processes. A series of experiments were conducted to examine the transformation of E2 in aqueous solution containing humic acid (HA) as model NOM by laccase-mediated ECOC reactions. The impact of HA on the reaction behaviors and product distribution is systematically characterized. The presence of HA inhibited the extent of E2 self-coupling in laccase-mediated systems, while promoted cross-coupling between E2 and HA. Reconfiguration of humic molecules was also observed and characterized by changes in absorbance at 275 nm and the ratios between A250 nm/A365 nm. In particular, experiments were conducted with un-labeled E2 mixed with (13)C3-labeled E2 at a set ratio, with the products probed using high-resolution mass spectrometry (HRMS). The high m/z accuracy of HRMS enabled the use of isotope ratio as a tracer to identify possible cross-coupling products between E2 and HA. Such a method combining HRMS and isotope labeling provides a novel means for identification of products in a reaction system involving NOM or other complex matrices. These findings provide a basis for optimization of ECOC reactions for estrogen removal, and also help to understand the environmental transformation of estrogens. PMID:26692517

  8. A label-free differential quantitative mass spectrometry method for the characterization and identification of protein changes during citrus fruit development

    PubMed Central

    2010-01-01

    Background Citrus is one of the most important and widely grown commodity fruit crops. In this study a label-free LC-MS/MS based shot-gun proteomics approach was taken to explore three main stages of citrus fruit development. These approaches were used to identify and evaluate changes occurring in juice sac cells in various metabolic pathways affecting citrus fruit development and quality. Results Protein changes in citrus juice sac cells were identified and quantified using label-free shotgun methodologies. Two alternative methods, differential mass-spectrometry (dMS) and spectral counting (SC) were used to analyze protein changes occurring during earlier and late stages of fruit development. Both methods were compared in order to develop a proteomics workflow that could be used in a non-model plant lacking a sequenced genome. In order to resolve the bioinformatics limitations of EST databases from species that lack a full sequenced genome, we established iCitrus. iCitrus is a comprehensive sequence database created by merging three major sources of sequences (HarvEST:citrus, NCBI/citrus/unigenes, NCBI/citrus/proteins) and improving the annotation of existing unigenes. iCitrus provided a useful bioinformatics tool for the high-throughput identification of citrus proteins. We have identified approximately 1500 citrus proteins expressed in fruit juice sac cells and quantified the changes of their expression during fruit development. Our results showed that both dMS and SC provided significant information on protein changes, with dMS providing a higher accuracy. Conclusion Our data supports the notion of the complementary use of dMS and SC for label-free comparative proteomics, broadening the identification spectrum and strengthening the identification of trends in protein expression changes during the particular processes being compared. PMID:21162737

  9. Differential binding of ppGpp and pppGpp to E. coli RNA polymerase: photo-labeling and mass spectral studies.

    PubMed

    Syal, Kirtimaan; Chatterji, Dipankar

    2015-12-01

    (p)ppGpp, a secondary messenger, is induced under stress and shows pleiotropic response. It binds to RNA polymerase and regulates transcription in Escherichia coli. More than 25 years have passed since the first discovery was made on the direct interaction of ppGpp with E. coli RNA polymerase. Several lines of evidence suggest different modes of ppGpp binding to the enzyme. Earlier cross-linking experiments suggested that the β-subunit of RNA polymerase is the preferred site for ppGpp, whereas recent crystallographic studies pinpoint the interface of β'/ω-subunits as the site of action. With an aim to validate the binding domain and to follow whether tetra- and pentaphosphate guanosines have different location on RNA polymerase, this work was initiated. RNA polymerase was photo-labeled with 8-azido-ppGpp/8-azido-pppGpp, and the product was digested with trypsin and subjected to mass spectrometry analysis. We observed three new peptides in the trypsin digest of the RNA polymerase labeled with 8-azido-ppGpp, of which two peptides correspond to the same pocket on β'-subunit as predicted by X-ray structural analysis, whereas the third peptide was mapped on the β-subunit. In the case of 8-azido-pppGpp-labeled RNA polymerase, we have found only one cross-linked peptide from the β'-subunit. However, we were unable to identify any binding site of pppGpp on the β-subunit. Interestingly, we observed that pppGpp at high concentration competes out ppGpp bound to RNA polymerase more efficiently, whereas ppGpp cannot titrate out pppGpp. The competition between tetraphosphate guanosine and pentaphosphate guanosine for E. coli RNA polymerase was followed by gel-based assay as well as by a new method known as DRaCALA assay.

  10. Spatially tracking 13C labeled substrate (bicarbonate) accumulation in microbial communities using laser ablation isotope ratio mass spectrometry

    SciTech Connect

    Moran, James J.; Doll, Charles G.; Bernstein, Hans C.; Renslow, Ryan S.; Cory, Alexandra B.; Hutchison, Janine R.; Lindemann, Stephen R.; Fredrickson, Jim K.

    2014-08-25

    This is a manuscript we would like to submit for publication in Environmental Microbiology Reports. This manuscript contains a description of a laser ablation isotope ratio mass spectrometry methodology developed at PNNL and applied to a microbial system at a PNNL project location – Hot Lake, Washington. I will submit a word document containing the entire manuscript with this Erica input request form.

  11. Production, Purification, and Characterization of ¹⁵N-Labeled DNA Repair Proteins as Internal Standards for Mass Spectrometric Measurements.

    PubMed

    Reddy, Prasad T; Jaruga, Pawel; Nelson, Bryant C; Lowenthal, Mark S; Jemth, Ann-Sofie; Loseva, Olga; Coskun, Erdem; Helleday, Thomas; Dizdaroglu, Miral

    2016-01-01

    Oxidatively induced DNA damage is caused in living organisms by a variety of damaging agents, resulting in the formation of a multiplicity of lesions, which are mutagenic and cytotoxic. Unless repaired by DNA repair mechanisms before DNA replication, DNA lesions can lead to genomic instability, which is one of the hallmarks of cancer. Oxidatively induced DNA damage is mainly repaired by base excision repair pathway with the involvement of a plethora of proteins. Cancer tissues develop greater DNA repair capacity than normal tissues by overexpressing DNA repair proteins. Increased DNA repair in tumors that removes DNA lesions generated by therapeutic agents before they became toxic is a major mechanism in the development of therapy resistance. Evidence suggests that DNA repair capacity may be a predictive biomarker of patient response. Thus, knowledge of DNA-protein expressions in disease-free and cancerous tissues may help predict and guide development of treatments and yield the best therapeutic response. Our laboratory has developed methodologies that use mass spectrometry with isotope dilution for the measurement of expression of DNA repair proteins in human tissues and cultured cells. For this purpose, full-length (15)N-labeled analogs of a number of human DNA repair proteins have been produced and purified to be used as internal standards for positive identification and accurate quantification. This chapter describes in detail the protocols of this work. The use of (15)N-labeled proteins as internal standards for the measurement of several DNA repair proteins in vivo is also presented.

  12. High performance liquid chromatography-tandem mass spectrometry method for ex vivo metabolic studies of a rhenium-labeled radiopharmaceutical for liver cancer.

    PubMed

    Chen, Wei-Hsi; Liao, Chen-Wei; Luo, Tsai-Yueh; Chang, Yu; Men, Lee-Chung; Hsieh, Yi-Cheng

    2014-01-01

    The radio-isotope rhenium-labeled N-[2-(triphenylmethyl)thioethyl]-3-aza-19-ethyloxycarbonyl-3-[2-(triphenylmethyl)thioethyl] octadecanoate) ligand (188Re-MN-16ET) is a novel therapeutic agent under preclinical evaluation for hepatoma. A reversed-phase high performance liquid chromatography coupled with a tandem mass spectrometric analysis method and diode array detector (DAD) involving a T type splitter was developed to characterize this pharmaceutical in rat liver tissue solution and determine its biotransformation rate. The separation was accomplished on a C18 column (chromolith silica, 4.6 mm x 100 mm) using an acetonitrile-ammonium acetate buffer gradient as the mobile phase. The detection was achieved by DAD set at 250nm and tandem mass spectrometry using electrospray ionization in the positive ion mode. Re-MN-16ET displayed a retention time of 23.2 min and a transition ion pair corresponding to m/z677 --> 631 for multiple reaction monitoring. Its biotransformation reaction in rat liver homogenate proceeded for 90 min in a 37°C water bath. The characterization was conducted using aliquots that were extracted and concentrated from the reaction mixture for various incubation times. Re-MN-16ET exhibited a biotransformation half-life (t1/2) of 8-9 min in liver tissue solution and was almost completely exhausted after 90 min. Two of its metabolites, consisting of the Re-labeled carboxylic acid derivative, predominately, and its corresponding demetallized disulfide ligand were found in the liver homogenate, providing a metabolism pathway for the radio-pharmaceutical.

  13. Mass spectrometric analysis of pharmaceutical adulterants in products labeled as botanical dietary supplements or herbal remedies: a review.

    PubMed

    Vaclavik, Lukas; Krynitsky, Alexander J; Rader, Jeanne I

    2014-11-01

    The increased availability and use of botanical dietary supplements and herbal remedies among consumers has been accompanied by an increased frequency of adulteration of these products with synthetic pharmaceuticals. Unscrupulous producers may add drugs and analogues of various classes, such as phosphodiesterase type 5 (PDE-5) inhibitors, weight loss, hypoglycemic, antihypertensive and anti-inflammatory agents, or anabolic steroids, to develop or intensify biological effects of dietary supplements or herbal remedies. The presence of such adulterated products in the marketplace is a worldwide problem and their consumption poses health risks to consumers. Analytical methods that allow rapid and reliable testing of dietary supplements for the presence of synthetic drugs are needed to address such fraudulent practices. Mass spectrometry (MS) and hyphenated techniques such as liquid chromatography-mass spectrometry (LC-MS) and gas chromatography-mass spectrometry (GC-MS) have become primary tools in this endeavor. The present review critically assesses the role and summarizes the applications of MS in the analysis of pharmaceutical adulterants in botanical dietary supplements and herbal remedies. The uses of MS techniques in detection, confirmation, and quantification of known pharmaceutical adulterants as well as in screening for and structure elucidation of unexpected adulterants and novel designer drugs are discussed. PMID:25270866

  14. Characterization of TATP gas phase product ion chemistry via isotope labeling experiments using ion mobility spectrometry interfaced with a triple quadrupole mass spectrometer.

    PubMed

    Tomlinson-Phillips, Jill; Wooten, Alfred; Kozole, Joseph; Deline, James; Beresford, Pamela; Stairs, Jason

    2014-09-01

    Identification of the fragment ion species associated with the ion reaction mechanism of triacetone triperoxide (TATP), a homemade peroxide-based explosive, is presented. Ion mobility spectrometry (IMS) has proven to be a key analytical technique in the detection of trace explosive material. Unfortunately, IMS alone does not provide chemical identification of the ions detected; therefore, it is unknown what ion species are actually formed and separated by the IMS. In IMS, ions are primarily characterized by their drift time, which is dependent on the ion׳s mass and molecular cross-section; thus, IMS as a standalone technique does not provide structural signatures, which is in sharp contrast to the chemical and molecular information that is generally obtained from other customary analytical techniques, such as NMR, Raman and IR spectroscopy and mass spectrometry. To help study the ion chemistry that gives rise to the peaks observed in IMS, the hardware of two different commercial IMS instruments has been directly coupled to triple quadrupole (QQQ) mass spectrometers, in order to ascertain each ion׳s corresponding mass/charge (m/z) ratios with different dopants at two temperatures. Isotope labeling was then used to help identify and confirm the molecular identity of the explosive fragment and adduct ions of TATP. The m/z values and isotope labeling experiments were used to help propose probable molecular formulas for the ion fragments. In this report, the fragment and adduct ions m/z 58 and 240 of TATP have been confirmed to be [C3H6NH·H](+) and [TATP·NH4](+), respectively; while the fragment ions m/z 73 and 89 of TATP are identified as having the molecular formulas [C4H9NH2](+) and [C4H9O2](+), respectively. It is anticipated that the work in this area will not only help to facilitate improvements in mobility-based detection (IMS and MS), but also aid in the development and optimization of MS-based detection algorithms for TATP.

  15. Characterization of TATP gas phase product ion chemistry via isotope labeling experiments using ion mobility spectrometry interfaced with a triple quadrupole mass spectrometer.

    PubMed

    Tomlinson-Phillips, Jill; Wooten, Alfred; Kozole, Joseph; Deline, James; Beresford, Pamela; Stairs, Jason

    2014-09-01

    Identification of the fragment ion species associated with the ion reaction mechanism of triacetone triperoxide (TATP), a homemade peroxide-based explosive, is presented. Ion mobility spectrometry (IMS) has proven to be a key analytical technique in the detection of trace explosive material. Unfortunately, IMS alone does not provide chemical identification of the ions detected; therefore, it is unknown what ion species are actually formed and separated by the IMS. In IMS, ions are primarily characterized by their drift time, which is dependent on the ion׳s mass and molecular cross-section; thus, IMS as a standalone technique does not provide structural signatures, which is in sharp contrast to the chemical and molecular information that is generally obtained from other customary analytical techniques, such as NMR, Raman and IR spectroscopy and mass spectrometry. To help study the ion chemistry that gives rise to the peaks observed in IMS, the hardware of two different commercial IMS instruments has been directly coupled to triple quadrupole (QQQ) mass spectrometers, in order to ascertain each ion׳s corresponding mass/charge (m/z) ratios with different dopants at two temperatures. Isotope labeling was then used to help identify and confirm the molecular identity of the explosive fragment and adduct ions of TATP. The m/z values and isotope labeling experiments were used to help propose probable molecular formulas for the ion fragments. In this report, the fragment and adduct ions m/z 58 and 240 of TATP have been confirmed to be [C3H6NH·H](+) and [TATP·NH4](+), respectively; while the fragment ions m/z 73 and 89 of TATP are identified as having the molecular formulas [C4H9NH2](+) and [C4H9O2](+), respectively. It is anticipated that the work in this area will not only help to facilitate improvements in mobility-based detection (IMS and MS), but also aid in the development and optimization of MS-based detection algorithms for TATP. PMID:24913870

  16. Acetylation and glycation of fibrinogen in vitro occur at specific lysine residues in a concentration dependent manner: A mass spectrometric and isotope labeling study

    SciTech Connect

    Svensson, Jan; Bergman, Ann-Charlotte; Adamson, Ulf; Blombaeck, Margareta; Wallen, Hakan; Joerneskog, Gun

    2012-05-04

    Highlights: Black-Right-Pointing-Pointer Fibrinogen was incubated in vitro with glucose or aspirin. Black-Right-Pointing-Pointer Acetylations and glycations were found at twelve lysine sites by mass spectrometry. Black-Right-Pointing-Pointer The labeling by aspirin and glucose occurred dose-dependently. Black-Right-Pointing-Pointer No competition between glucose and aspirin for binding to fibrinogen was found. -- Abstract: Aspirin may exert part of its antithrombotic effects through platelet-independent mechanisms. Diabetes is a condition in which the beneficial effects of aspirin are less prominent or absent - a phenomenon called 'aspirin resistance'. We investigated whether acetylation and glycation occur at specific sites in fibrinogen and if competition between glucose and aspirin in binding to fibrinogen occurs. Our hypothesis was that such competition might be one explanation to 'aspirin resistance' in diabetes. After incubation of fibrinogen in vitro with aspirin (0.8 mM, 24 h) or glucose (100 mM, 5-10 days), we found 12 modified sites with mass spectrometric techniques. Acetylations in the {alpha}-chain: {alpha}K191, {alpha}K208, {alpha}K224, {alpha}K429, {alpha}K457, {alpha}K539, {alpha}K562, in the {beta}-chain: {beta}K233, and in the {gamma}-chain: {gamma}K170 and {gamma}K273. Glycations were found at {beta}K133 and {gamma}K75, alternatively {gamma}K85. Notably, the lysine 539 is a site involved in FXIII-mediated cross-linking of fibrin. With isotope labeling in vitro, using [{sup 14}C-acetyl]salicylic acid and [{sup 14}C]glucose, a labeling of 0.013-0.084 and 0.12-0.5 mol of acetylated and glycated adduct/mol fibrinogen, respectively, was found for clinically (12.9-100 {mu}M aspirin) and physiologically (2-8 mM glucose) relevant plasma concentrations. No competition between acetylation and glycation could be demonstrated. Thus, fibrinogen is acetylated at several lysine residues, some of which are involved in the cross-linking of fibrinogen. This may

  17. Quantitative determination of free and total bisphenol A in human urine using labeled BPA glucuronide and isotope dilution mass spectrometry.

    PubMed

    Kubwabo, Cariton; Kosarac, Ivana; Lalonde, Kaela; Foster, Warren G

    2014-07-01

    Bisphenol A (BPA) is a widely used industrial chemical in the manufacturing of polycarbonate plastic bottles, food and beverage can linings, thermal receipts, and dental sealants. Animal and human studies suggest that BPA may disrupt normal hormonal function and hence, potentially, have negative effects on the human health. While total BPA is frequently reported, it is recognized that free BPA is the biologically active form and is rarely reported in the literature. The objective of this study was to develop a sensitive and improved method for the measurement of free and total BPA in human urine. Use of a labeled conjugated BPA (bisphenol A-d6 β-D-glucuronide) allowed for the optimization of the enzymatic reaction and permitted an accurate determination of the conjugated BPA concentration in urine samples. In addition, a (13)C12-BPA internal standard was used to account for the analytical recoveries and performance of the isotope dilution method. Solid-phase extraction (SPE) combined with derivatization and analysis using a triple quadrupole GC-EI/MS/MS system achieved very low method detection limit of 0.027 ng/mL. BPA concentrations were measured in urine samples collected during the second and third trimesters of pregnancy in 36 Canadian women. Total maternal BPA concentrations in urine samples ranged from not detected to 9.40 ng/mL (median, 1.21 ng/mL), and free BPA concentrations ranged from not detected to 0.950 ng/mL (median, 0.185 ng/mL). Eighty-six percent of the women had detectable levels of conjugated BPA, whereas only 22 % had detectable levels of free BPA in their urine. BPA levels measured in this study agreed well with data reported internationally.

  18. Variation in airborne 137Cs peak levels with altitude from high-altitude locations across Europe after the arrival of Fukushima-labeled air masses

    NASA Astrophysics Data System (ADS)

    Masson, Olivier; Bieringer, Jacqueline; Dalheimer, Axel; Estier, Sybille; Evrard, Olivier; Penev, Ilia; Ringer, Wolfgang; Schlosser, Clemens; Steinkopff, Thomas; Tositti, Laura; de Vismes-Ott, Anne

    2015-04-01

    During the Fukushima Daiichi nuclear power plant (FDNPP) accident, a dozen of high-altitude aerosol sampling stations, located between 850 and 3,454 m above sea level (a.s.l.), provided airborne activity levels across Europe (Fig. 1). This represents at most 5% of the total number of aerosol sampling locations that delivered airborne activity levels (at least one result) in Europe, in connection with this nuclear accident. High altitude stations are typically equipped with a high volume sampler that collects aerosols on filters. The Fukushima-labeled air mass arrival and the peak of airborne cesium-137 (137Cs) activity levels were registered in Europe at different dates depending on the location, with differences up to a factor of six on a regional scale. Besides this statement related to lowland areas, we have compared the maximum airborne levels registered at high-altitude European locations (850 m < altitudes < 3450 m) with what was observed at the closest lowland location. The vertical distribution of 137Cs peak level was not uniform even after a long travel time/distance from Japan. This being true at least in the atmospheric boundary layer and in the lower free troposphere. Moreover the relation '137Csmax vs. altitude' shows a decreasing trend (Fig. 2). Results and discussion : Comparison of 137Cs and 7Be levels shows simultaneous increases at least when the 137Cs airborne level rose for the first time (Fig. 3). Zugspitze and Jungfraujoch stations attest of a time shift between 7Be and 137Cs peak that can be due to the particular dynamic of air movements at such high altitudes. After the 137Cs peak value, the plume concentration decreased whatever the 7Be level. Due to the cosmogenic origin of 7Be, its increase in the ground-level air is usually associated with downwind air movements, i.e. stratospheric air intrusions or at least air from high-tropospheric levels, into lower atmospheric layers. This means that Fukushima-labeled air masses registered at ground

  19. Comparative proteomic analysis of the aging soleus and extensor digitorum longus rat muscles using TMT labeling and mass spectrometry.

    PubMed

    Chaves, Daniela F S; Carvalho, Paulo C; Lima, Diogo B; Nicastro, Humberto; Lorenzeti, Fábio M; Siqueira-Filho, Mário; Hirabara, Sandro M; Alves, Paulo H M; Moresco, James J; Yates, John R; Lancha, Antonio H

    2013-10-01

    Sarcopenia describes an age-related decline in skeletal muscle mass, strength, and function that ultimately impairs metabolism and leads to poor balance, frequent falling, limited mobility, and a reduction in quality of life. Here we investigate the pathogenesis of sarcopenia through a proteomic shotgun approach. In brief, we employed tandem mass tags to quantitate and compare the protein profiles obtained from young versus old rat slow-twitch type of muscle (soleus) and a fast-twitch type of muscle (extensor digitorum longus, EDL). Our results disclose 3452 and 1848 proteins identified from soleus and EDL muscles samples, of which 78 and 174 were found to be differentially expressed, respectively. In general, most of the proteins were structural related and involved in energy metabolism, oxidative stress, detoxification, or transport. Aging affected soleus and EDL muscles differently, and several proteins were regulated in opposite ways. For example, pyruvate kinase had its expression and activity different in both soleus and EDL muscles. We were able to verify with existing literature many of our differentially expressed proteins as candidate aging biomarkers and, most importantly, disclose several new candidate biomarkers such as the glioblastoma amplified sequence, zero β-globin, and prolargin. PMID:24001182

  20. Characterization of Breast Cancer Interstitial Fluids by TmT Labeling, LTQ-Orbitrap Velos Mass Spectrometry and Pathway Analysis

    PubMed Central

    Cinzia, Raso; Carlo, Cosentino; Marco, Gaspari; Natalia, Malara; Xuemei, Han; Daniel, McClatchy; Kyu, Park Sung; Maria, Renne; Nuria, Vadalà; Ubaldo, Prati; Giovanni, Cuda; Vincenzo, Mollace; Francesco, Amato; Yates, John R.

    2012-01-01

    Cancer is currently considered as the end point of numerous genomic and epigenomic mutations and as the result of the interaction of transformed cells within the stromal microenvironment. The present work focuses on breast cancer, one of the most common malignancies affecting the female population in industrialized countries. In this study we perform a proteomic analysis of bioptic samples from human breast cancer, namely interstitial fluids and primary cells, normal vs disease tissues, using Tandem mass Tags (TmT) quantitative mass spectrometry combined with the MudPIT technique. To the best of our knowledge this work, with over 1700 proteins identified, represents the most comprehensive characterization of the breast cancer interstitial fluid proteome to date. Network analysis was used to identify functionally active networks in the breast cancer associated samples. From the list of differentially expressed genes we have retrieved the associated functional interaction networks. Many different signaling pathways were found activated, strongly linked to invasion, metastasis development, proliferation and with a significant cross-talking rate. This pilot study presents evidence that the proposed quantitative proteomic approach can be applied to discriminate between normal and tumoral samples and for the discovery of yet unknown carcinogenesis mechanisms and therapeutic strategies. PMID:22563702

  1. Mass

    SciTech Connect

    Quigg, Chris

    2007-12-05

    In the classical physics we inherited from Isaac Newton, mass does not arise, it simply is. The mass of a classical object is the sum of the masses of its parts. Albert Einstein showed that the mass of a body is a measure of its energy content, inviting us to consider the origins of mass. The protons we accelerate at Fermilab are prime examples of Einsteinian matter: nearly all of their mass arises from stored energy. Missing mass led to the discovery of the noble gases, and a new form of missing mass leads us to the notion of dark matter. Starting with a brief guided tour of the meanings of mass, the colloquium will explore the multiple origins of mass. We will see how far we have come toward understanding mass, and survey the issues that guide our research today.

  2. Selective Analysis of Sulfur-Containing Species in a Heavy Crude Oil by Deuterium Labeling Reactions and Ultrahigh Resolution Mass Spectrometry.

    PubMed

    Wang, Xuxiao; Schrader, Wolfgang

    2015-12-17

    A heavy crude oil has been treated with deuterated alkylating reagents (CD₃I and C₂D₅I) and directly analyzed without any prior fractionation and chromatographic separation by high-field Orbitrap Fourier Transform Mass Spectrometry (FTMS) and Fourier Transform Ion Cyclotron Resonance Mass Spectrometry (FT-ICR MS) using electrospray ionization (ESI). The reaction of a polycyclic aromatic sulfur heterocycles (PASHs) dibenzothiophene (DBT), in the presence of silver tetrafluoroborate (AgBF₄) with ethyl iodide (C₂H₅I) in anhydrous dichloroethane (DCE) was optimized as a sample reaction to study heavy crude oil mixtures, and the reaction yield was monitored and determined by proton nuclear magnetic resonance spectroscopy (¹H-NMR). The obtained conditions were then applied to a mixture of standard aromatic CH-, N-, O- and S-containing compounds and then a heavy crude oil, and only sulfur-containing compounds were selectively alkylated. The deuterium labeled alkylating reagents, iodomethane-d₃ (CD₃I) and iodoethane-d₅ (C₂D₅I), were employed to the alkylation of heavy crude oil to selectively differentiate the tagged sulfur species from the original crude oil.

  3. Selective Analysis of Sulfur-Containing Species in a Heavy Crude Oil by Deuterium Labeling Reactions and Ultrahigh Resolution Mass Spectrometry

    PubMed Central

    Wang, Xuxiao; Schrader, Wolfgang

    2015-01-01

    A heavy crude oil has been treated with deuterated alkylating reagents (CD3I and C2D5I) and directly analyzed without any prior fractionation and chromatographic separation by high-field Orbitrap Fourier Transform Mass Spectrometry (FTMS) and Fourier Transform Ion Cyclotron Resonance Mass Spectrometry (FT-ICR MS) using electrospray ionization (ESI). The reaction of a polycyclic aromatic sulfur heterocycles (PASHs) dibenzothiophene (DBT), in the presence of silver tetrafluoroborate (AgBF4) with ethyl iodide (C2H5I) in anhydrous dichloroethane (DCE) was optimized as a sample reaction to study heavy crude oil mixtures, and the reaction yield was monitored and determined by proton nuclear magnetic resonance spectroscopy (1H-NMR). The obtained conditions were then applied to a mixture of standard aromatic CH-, N-, O- and S-containing compounds and then a heavy crude oil, and only sulfur-containing compounds were selectively alkylated. The deuterium labeled alkylating reagents, iodomethane-d3 (CD3I) and iodoethane-d5 (C2D5I), were employed to the alkylation of heavy crude oil to selectively differentiate the tagged sulfur species from the original crude oil. PMID:26694374

  4. Stable-isotope-labeled Histone Peptide Library for Histone Post-translational Modification and Variant Quantification by Mass Spectrometry *

    PubMed Central

    Lin, Shu; Wein, Samuel; Gonzales-Cope, Michelle; Otte, Gabriel L.; Yuan, Zuo-Fei; Afjehi-Sadat, Leila; Maile, Tobias; Berger, Shelley L.; Rush, John; Lill, Jennie R.; Arnott, David; Garcia, Benjamin A.

    2014-01-01

    To facilitate accurate histone variant and post-translational modification (PTM) quantification via mass spectrometry, we present a library of 93 synthetic peptides using Protein-Aqua™ technology. The library contains 55 peptides representing different modified forms from histone H3 peptides, 23 peptides representing H4 peptides, 5 peptides representing canonical H2A peptides, 8 peptides representing H2A.Z peptides, and peptides for both macroH2A and H2A.X. The PTMs on these peptides include lysine mono- (me1), di- (me2), and tri-methylation (me3); lysine acetylation; arginine me1; serine/threonine phosphorylation; and N-terminal acetylation. The library was subjected to chemical derivatization with propionic anhydride, a widely employed protocol for histone peptide quantification. Subsequently, the detection efficiencies were quantified using mass spectrometry extracted ion chromatograms. The library yields a wide spectrum of detection efficiencies, with more than 1700-fold difference between the peptides with the lowest and highest efficiencies. In this paper, we describe the impact of different modifications on peptide detection efficiencies and provide a resource to correct for detection biases among the 93 histone peptides. In brief, there is no correlation between detection efficiency and molecular weight, hydrophobicity, basicity, or modification type. The same types of modifications may have very different effects on detection efficiencies depending on their positions within a peptide. We also observed antagonistic effects between modifications. In a study of mouse trophoblast stem cells, we utilized the detection efficiencies of the peptide library to correct for histone PTM/variant quantification. For most histone peptides examined, the corrected data did not change the biological conclusions but did alter the relative abundance of these peptides. For a low-abundant histone H2A variant, macroH2A, the corrected data led to a different conclusion than the

  5. Determination of 5-methyltetrahydrofolate (13C-labeled and unlabeled) in human plasma and urine by combined liquid chromatography mass spectrometry.

    PubMed

    Hart, David J; Finglas, Paul M; Wolfe, Caroline A; Mellon, Fred; Wright, Anthony J A; Southon, Susan

    2002-06-15

    The association of folates with the prevention of neural tube defects and reduced risk of other chronic diseases has stimulated interest in the development of techniques for the study of their bioavailability in humans. Stable isotope protocols differentiate between oral and/or intravenous test doses of folate and natural levels of folate already present in the body. An liquid chromatography/mass spectrometry (LC/MS) procedure is described that has been validated for the determination of [13C]5-methyltetrahydropteroyl monoglutamic acid ([13C]5-CH3H4PteGlu) in plasma and urine, following oral dosing of volunteers with different labeled folates. Folate binding protein affinity columns were used for sample purification prior to LC/MS determination. Chromatographic separation was achieved using a Superspher 100RP18 (4 microm) column and mobile phase of 0.1 mol/L acetic acid (pH 3.3):acetonitrile (90:10; 250 microL/min). Selected ion monitoring was conducted on the [M-H](-) ion: m/z 458 and 459 for analyzing 5-CH3H4PteGlu; m/z 464 [M+6-H](-) to determine 5-CH3H4PteGlu derived from the label dose; m/z 444 for analysis of 2H4PteGlu internal standard, and m/z 446 and 478 to confirm that there was no direct absorption of unmetabolized compounds. Calibration was linear over the range 0-9 x 10(-9) mol/L; the limits of detection and quantification were 0.2 x 10(-9) and 0.55 x 10(-9) mol/L, respectively. The mean coefficient of variation of the ratios (m/z 463/458) was 7.4%. The method has potential applications for other key folates involved in one-carbon metabolism.

  6. Simultaneous quantification of labeled (2)H5-glycerol, (13)C6-glucose, and endogenous D-glucose in mouse plasma using liquid chromatography tandem mass spectrometry.

    PubMed

    Jahouh, Farid; Wang, Rong

    2015-11-01

    Monitoring the level of glucose and glycerol or their labeled derivatives in biological fluid for kinetic studies has always been challenging, especially in mice, because of the limited volume in addition to the complexity of plasma. For such application, we developed a simple, fast, and sensitive method for the simultaneous measurement of absolute concentrations of labeled (2)H5-glycerol and (13)C6-glucose as well as endogenous D-glucose using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). In our study, 15.0 μL of mouse plasma was processed by a one-step protein precipitation, followed by LC-MS/MS analysis. The quantification of the analytes was carried out by monitoring the product ion scan of their corresponding deprotonated molecular ions and constructing the extracted ion fragmentogram by choosing a specific product ion for each analyte (equivalent to precursor ion to product ion transitions). The limit of detection (LOD) was evaluated to be 1.0 μM for both (2)H5-glycerol and (13)C6-glucose, and the limit of quantitation (LOQ) was observed to be 5.0 μM for both (2)H5-glycerol and (13)C6-glucose in diluted mice plasma that corresponds to 50 μM in plasma or 4.60 and 9.01 mg/dL of glycerol and glucose in plasma, respectively. The extraction recoveries are 81.9 % (CV = 8.1 %) for (2)H5-glycerol and 26.2 % (CV = 13.6 %) for (13)C6-glucose. PMID:26362155

  7. Determination of 5-methyltetrahydrofolate (13C-labeled and unlabeled) in human plasma and urine by combined liquid chromatography mass spectrometry.

    PubMed

    Hart, David J; Finglas, Paul M; Wolfe, Caroline A; Mellon, Fred; Wright, Anthony J A; Southon, Susan

    2002-06-15

    The association of folates with the prevention of neural tube defects and reduced risk of other chronic diseases has stimulated interest in the development of techniques for the study of their bioavailability in humans. Stable isotope protocols differentiate between oral and/or intravenous test doses of folate and natural levels of folate already present in the body. An liquid chromatography/mass spectrometry (LC/MS) procedure is described that has been validated for the determination of [13C]5-methyltetrahydropteroyl monoglutamic acid ([13C]5-CH3H4PteGlu) in plasma and urine, following oral dosing of volunteers with different labeled folates. Folate binding protein affinity columns were used for sample purification prior to LC/MS determination. Chromatographic separation was achieved using a Superspher 100RP18 (4 microm) column and mobile phase of 0.1 mol/L acetic acid (pH 3.3):acetonitrile (90:10; 250 microL/min). Selected ion monitoring was conducted on the [M-H](-) ion: m/z 458 and 459 for analyzing 5-CH3H4PteGlu; m/z 464 [M+6-H](-) to determine 5-CH3H4PteGlu derived from the label dose; m/z 444 for analysis of 2H4PteGlu internal standard, and m/z 446 and 478 to confirm that there was no direct absorption of unmetabolized compounds. Calibration was linear over the range 0-9 x 10(-9) mol/L; the limits of detection and quantification were 0.2 x 10(-9) and 0.55 x 10(-9) mol/L, respectively. The mean coefficient of variation of the ratios (m/z 463/458) was 7.4%. The method has potential applications for other key folates involved in one-carbon metabolism. PMID:12054449

  8. High-resolution twin-ion metabolite extraction (HiTIME) mass spectrometry: nontargeted detection of unknown drug metabolites by isotope labeling, liquid chromatography mass spectrometry, and automated high-performance computing.

    PubMed

    Leeming, Michael G; Isaac, Andrew P; Pope, Bernard J; Cranswick, Noel; Wright, Christine E; Ziogas, James; O'Hair, Richard A J; Donald, William A

    2015-04-21

    The metabolic fate of a compound can often determine the success of a new drug lead. Thus, significant effort is directed toward identifying the metabolites formed from a given molecule. Here, an automated and nontargeted procedure is introduced for detecting drug metabolites without authentic metabolite standards via the use of stable isotope labeling, liquid chromatography mass spectrometry (LC/MS), and high-performance computing. LC/MS of blood plasma extracts from rats that were administered a 1:1 mixture of acetaminophen (APAP) and (13)C6-APAP resulted in mass spectra that contained "twin" ions for drug metabolites that were not detected in control spectra (i.e., no APAP administered). Because of the development of a program (high-resolution twin-ion metabolite extraction; HiTIME) that can identify twin-ions in high-resolution mass spectra without centroiding (i.e., reduction of mass spectral peaks to single data points), 9 doublets corresponding to APAP metabolites were identified. This is nearly twice that obtained by use of existing programs that make use of centroiding to reduce computational cost under these conditions with a quadrupole time-of-flight mass spectrometer. By a manual search for all reported APAP metabolite ions, no additional twin-ion signals were assigned. These data indicate that all the major metabolites of APAP and multiple low-abundance metabolites (e.g., acetaminophen hydroxy- and methoxysulfate) that are rarely reported were detected. This methodology can be used to detect drug metabolites without prior knowledge of their identity. HiTIME is freely available from https://github.com/bjpop/HiTIME .

  9. Point-of-Purchase Calorie Labeling Has Little Influence on Calories Ordered Regardless of Body Mass Index.

    PubMed

    Rendell, Sarah Litman; Swencionis, Charles

    2014-09-01

    The obesity epidemic has incited legislation aimed to inform consumers of the nutritional value of food items available in restaurants and fast food establishments, with the presumption that knowing the caloric content in a meal might enable patrons to make healthier choices when ordering. However, available research shows mixed results regarding consumers' use of calorie information to promote healthier purchases. The aim of this study was to determine whether menu type, specifically having viewed a menu with calorie disclosures or not, would have an impact on how many calories were in a lunch meal ordered by a patron. Additionally, we sought to identify body mass index (BMI) as a moderator of the relationship between viewing a menu with or without calorie information and the number of calories an individual orders for lunch. Two hundred forty-five adults participated in the study and completed the questionnaire. Results indicated neither menu type, nor reporting having seen calorie information, was significantly related to the number of calories in the foods that participants ordered, even after controlling demographic variables age, sex, income, education, race/ethnicity, and BMI. BMI did not serve as a moderator in the relationship between menu type and food calories ordered. Implications for policy change and clinical work with overweight and obese patients are discussed.

  10. 13C- and 15N-Labeling Strategies Combined with Mass Spectrometry Comprehensively Quantify Phospholipid Dynamics in C. elegans

    PubMed Central

    Drechsler, Robin; Gafken, Philip R.; Olsen, Carissa Perez

    2015-01-01

    Membranes define cellular and organelle boundaries, a function that is critical to all living systems. Like other biomolecules, membrane lipids are dynamically maintained, but current methods are extremely limited for monitoring lipid dynamics in living animals. We developed novel strategies in C. elegans combining 13C and 15N stable isotopes with mass spectrometry to directly quantify the replenishment rates of the individual fatty acids and intact phospholipids of the membrane. Using multiple measurements of phospholipid dynamics, we found that the phospholipid pools are replaced rapidly and at rates nearly double the turnover measured for neutral lipid populations. In fact, our analysis shows that the majority of membrane lipids are replaced each day. Furthermore, we found that stearoyl-CoA desaturases (SCDs), critical enzymes in polyunsaturated fatty acid production, play an unexpected role in influencing the overall rates of membrane maintenance as SCD depletion affected the turnover of nearly all membrane lipids. Additionally, the compromised membrane maintenance as defined by LC-MS/MS with SCD RNAi resulted in active phospholipid remodeling that we predict is critical to alleviate the impact of reduced membrane maintenance in these animals. Not only have these combined methodologies identified new facets of the impact of SCDs on the membrane, but they also have great potential to reveal many undiscovered regulators of phospholipid metabolism. PMID:26528916

  11. Comprehensive and highly sensitive urinary steroid hormone profiling method based on stable isotope-labeling liquid chromatography-mass spectrometry.

    PubMed

    Dai, Weidong; Huang, Qiang; Yin, Peiyuan; Li, Jia; Zhou, Jia; Kong, Hongwei; Zhao, Chunxia; Lu, Xin; Xu, Guowang

    2012-12-01

    Steroid hormones are crucial substances that mediate a wide range of vital physiological functions of the body. Determination of the levels of steroid hormones plays an important role in understanding the mechanism of the steroid hormone-related diseases. In this study, we present a novel targeted metabolic profiling method based on the introduction of an easily protonated stable isotope tag to a hydroxyl-containing steroid hormone with a synthesized derivatization reagent, deuterium 4-(dimethylamino)-benzoic acid (d(4)-DMBA), and liquid chromatography-mass spectrometry (LC-MS). Different from other reported derivatization reagents that have been used to enhance the sensitivities for estrogens or androgens, our method is comprehensive with the capability of covering hydroxyl-containing androgens, estrogens, corticoids, and progestogens. Furthermore, the nonderivatized steroid hormones (e.g., 17α-hydroxyprogesterone, progesterone, and androstenedione) were not destroyed during the derivatization process, and their levels could still be obtained in one LC-MS run. We were able to detect 24 steroid hormones at subng/mL levels (the lower limit of detection could reach 5 pg/mL for estrone and 16α-hydroxy estrone, which is equivalent to 0.1 pg on column) with maximum sensitivity enhancement factors of more than 10(3)- to 10(4)-fold after derivatization. The method was successfully applied to the measurement of free (unconjugated) steroid hormones in urine samples of males, females, and pregnant women. Because the significant role the steroid hormone pathway plays in humans, a comprehensive, sensitive, specific, and accurate method for profiling the steroid hormone metabolome shall offer new insights into hormone-related diseases. PMID:23110480

  12. Towards absolute quantification of therapeutic monoclonal antibody in serum by LC-MS/MS using isotope-labeled antibody standard and protein cleavage isotope dilution mass spectrometry.

    PubMed

    Heudi, Olivier; Barteau, Samuel; Zimmer, Dieter; Schmidt, Joerg; Bill, Kurt; Lehmann, Natalie; Bauer, Christian; Kretz, Olivier

    2008-06-01

    Although LC-MS methods are increasingly used for the absolute quantification of proteins, the lack of appropriate internal standard (IS) hinders the development of rapid and standardized analytical methods for both in vitro and in vivo studies. Here, we have developed a novel method for the absolute quantification of a therapeutic protein, which is monoclonal antibody (mAb). The method combines liquid chromatography tandem mass spectrometry (LC-MS/MS) and protein cleavage isotope dilution mass spectrometry with the isotope-labeled mAb as IS. The latter was identical to the analyzed mAb with the exception that each threonine contains four (13)C atoms and one (15)N atom. Serum samples were spiked with IS prior to the overnight trypsin digestion and subsequent sample cleanup. Sample extracts were analyzed on a C18 ACE column (150 mm x 4.6 mm) using an LC gradient time of 11 min. Endogenous mAb concentrations were determined by calculating the peak height ratio of its signature peptide to the corresponding isotope-labeled peptide. The linear dynamic range was established between 5.00 and 1000 microg/mL mAb with accuracy and precision within +/-15% at all concentrations and below +/-20% at the LLOQ (lower limit of quantification). The overall method recovery in terms of mAb was 14%. The losses due to sample preparation (digestion and purification) were 72% from which about 32% was due to the first step of the method, the sample digestion. This huge loss during sample preparation strongly emphasizes the necessity to employ an IS right from the beginning. Our method was successfully applied to the mAb quantification in marmoset serum study samples, and the precision obtained on duplicate samples was, in most cases, below 20%. The comparison with enzyme-linked immunosorbent assay (ELISA) showed higher exposure in terms of AUC and Cmax with the LC-MS/MS method. Possible reasons for this discrepancy are discussed in this study. The results of this study indicate that our LC

  13. Principles of protein labeling techniques.

    PubMed

    Obermaier, Christian; Griebel, Anja; Westermeier, Reiner

    2015-01-01

    Protein labeling methods prior to separation and analysis have become indispensable approaches for proteomic profiling. Basically, three different types of tags are employed: stable isotopes, mass tags, and fluorophores. While proteins labeled with stable isotopes and mass tags are measured and differentiated by mass spectrometry, fluorescent labels are detected with fluorescence imagers. The major purposes for protein labeling are monitoring of biological processes, reliable quantification of compounds and specific detection of protein modifications and isoforms in multiplexed samples, enhancement of detection sensitivity, and simplification of detection workflows. Proteins can be labeled during cell growth by incorporation of amino acids containing different isotopes, or in biological fluids, cells or tissue samples by attaching specific groups to the ε-amino group of lysine, the N-terminus, or the cysteine residues. The principles and the modifications of the different labeling approaches on the protein level are described; benefits and shortcomings of the methods are discussed.

  14. Comparative Label-Free Mass Spectrometric Analysis of Mildly versus Severely Affected mdx Mouse Skeletal Muscles Identifies Annexin, Lamin, and Vimentin as Universal Dystrophic Markers.

    PubMed

    Holland, Ashling; Henry, Michael; Meleady, Paula; Winkler, Claudia K; Krautwald, Mirjam; Brinkmeier, Heinrich; Ohlendieck, Kay

    2015-01-01

    The primary deficiency in the membrane cytoskeletal protein dystrophin results in complex changes in dystrophic muscles. In order to compare the degree of secondary alterations in differently affected subtypes of skeletal muscles, we have conducted a global analysis of proteome-wide changes in various dystrophin-deficient muscles. In contrast to the highly degenerative mdx diaphragm muscle, which showed considerable alterations in 35 distinct proteins, the spectrum of mildly to moderately dystrophic skeletal muscles, including interosseus, flexor digitorum brevis, soleus, and extensor digitorum longus muscle, exhibited a smaller number of changed proteins. Compensatory mechanisms and/or cellular variances may be responsible for differing secondary changes in individual mdx muscles. Label-free mass spectrometry established altered expression levels for diaphragm proteins associated with contraction, energy metabolism, the cytoskeleton, the extracellular matrix and the cellular stress response. Comparative immunoblotting verified the differences in the degree of secondary changes in dystrophin-deficient muscles and showed that the up-regulation of molecular chaperones, the compensatory increase in proteins of the intermediate filaments, the fibrosis-related increase in collagen levels and the pathophysiological decrease in calcium binding proteins is more pronounced in mdx diaphragm as compared to the less severely affected mdx leg muscles. Annexin, lamin, and vimentin were identified as universal dystrophic markers. PMID:26102067

  15. Comparison of copper labeling followed by liquid chromatography-inductively coupled plasma mass spectrometry and immunochemical assays for serum hepcidin-25 determination.

    PubMed

    Konz, Tobias; Alonso-García, Javier; Montes-Bayón, María; Sanz-Medel, Alfredo

    2013-10-17

    Hepcidin-25 has been defined as the key biomarker in iron metabolism. This peptide binds to the iron transporter ferroportin to cause its degradation. Therefore, the need for specific, accurate and precise methods for the quantification of hepcidin-25 in biological fluids is dramatically increasing. In this regard, the use of rapid immunochemical methods that provide low limit of quantification is desired for routine clinical use. However, such fast methodologies should be first analytically evaluated and compared with alternative strategies to check for their advantages and limitations. Here we compare the use of a commercial immunochemical assay for hepcidin determination with a novel analytical approach based on Cu-labeling of the peptide followed by Cu determination using liquid chromatography (HPLC) and plasma mass spectrometry (ICP-MS). The figures of merit of both systems reveal similar analytical characteristics and both seem to be adequate for the determination of the peptide at biologically relevant concentrations in human serum samples. The analysis of a larger number of samples (n=50) by both techniques showed a good agreement in the concentrations found. Such finding permits to address the hepcidin recovery in the sample preparation procedure necessary for the HPLC-ICP-MS analysis in human serum that turn out to be 76-85%. Additionally, limitations due to cross-reactivity issues of the ELISA method could be addressed in some of the samples by using LC-ICP-MS and were confirmed by LC-Electrospray-MS.

  16. Differential mass spectrometry: a label-free LC-MS method for finding significant differences in complex peptide and protein mixtures.

    PubMed

    Wiener, Matthew C; Sachs, Jeffrey R; Deyanova, Ekaterina G; Yates, Nathan A

    2004-10-15

    Efficiently identifying and quantifying disease- or treatment-related changes in the abundance of proteins is an important area of research for the pharmaceutical industry. Here we describe an automated, label-free method for finding differences in complex mixtures using complete LC-MS data sets, rather than subsets of extracted peaks or features. The method selectively finds statistically significant differences in the intensity of both high-abundance and low-abundance ions, accounting for the variability of measured intensities and the fact that true differences will persist in time. The method was used to compare two complex peptide mixtures with known peptide differences. This controlled experiment allowed us to assess the validity of each difference found and so to analyze the method's sensitivity and specificity. The method detects both presence versus absence and a 2-fold change in peptide concentration near the limit of detection of the instrument used, where chromatographic peaks may not be sufficiently well defined to be detected in individual samples. The method is more sensitive and gives fewer false positives than subtractive methods that ignore signal variability. Differential mass spectrometry combined with targeted MS/MS analysis of only identified differences may save both computation time and human effort compared to shotgun proteomics approaches. PMID:15481957

  17. Label-free determination of protein-ligand binding constants using mass spectrometry and validation using surface plasmon resonance and isothermal titration calorimetry.

    PubMed

    Jecklin, Matthias C; Schauer, Stefan; Dumelin, Christoph E; Zenobi, Renato

    2009-01-01

    We performed a systematic comparison of three label-free methods for quantitative assessment of binding strengths of proteins interacting with small molecule ligands. The performance of (1) nanoelectrospray ionization mass spectrometry (nESI-MS), (2) surface plasmon resonance (SPR), and (3) isothermal titration calorimetry (ITC) was compared for the determination of dissociation constants (K(D)). The model system studied for this purpose was the human carbonic anhydrase I (hCAI) with eight known and well characterized sulfonamide inhibitors (Krishnamurthy et al., Chem. Rev. 2008, 108: 946-1051). The binding affinities of the inhibitors chosen vary by more than four orders of magnitude e.g., the K(D) value determined for ethoxzolamide by nESI-MS was 5 +/- 1 nM and the K(D) value for sulfanilamide was 145.7 +/- 10.0 microM. The agreement of the determined K(D) values by the three methods investigated was excellent for ethoxzolamide and benzenesulfonamide (variation with experimental error), good for acetazolamide and 4-carboxybenzenesulfonamide (variation by approximately one order of magnitude), but poor for others e.g., sulpiride. The accuracies of the K(D) values are determined, and advantages and drawbacks of the individual methods are discussed. Moreover, we critically evaluate the three examined methods in terms of ease of the measurement, sample consumption, time requirement, and discuss their limitations. PMID:19373858

  18. Review, evaluation, and discussion of the challenges of missing value imputation for mass spectrometry-based label-free global proteomics

    SciTech Connect

    Webb-Robertson, Bobbie-Jo M.; Wiberg, Holli K.; Matzke, Melissa M.; Brown, Joseph N.; Wang, Jing; McDermott, Jason E.; Smith, Richard D.; Rodland, Karin D.; Metz, Thomas O.; Pounds, Joel G.; Waters, Katrina M.

    2015-04-09

    In this review, we apply selected imputation strategies to label-free liquid chromatography–mass spectrometry (LC–MS) proteomics datasets to evaluate the accuracy with respect to metrics of variance and classification. We evaluate several commonly used imputation approaches for individual merits and discuss the caveats of each approach with respect to the example LC–MS proteomics data. In general, local similarity-based approaches, such as the regularized expectation maximization and least-squares adaptive algorithms, yield the best overall performances with respect to metrics of accuracy and robustness. However, no single algorithm consistently outperforms the remaining approaches, and in some cases, performing classification without imputation sometimes yielded the most accurate classification. Thus, because of the complex mechanisms of missing data in proteomics, which also vary from peptide to protein, no individual method is a single solution for imputation. In summary, on the basis of the observations in this review, the goal for imputation in the field of computational proteomics should be to develop new approaches that work generically for this data type and new strategies to guide users in the selection of the best imputation for their dataset and analysis objectives.

  19. CVD Prevention Through Policy: a Review of Mass Media, Food/Menu Labeling, Taxation/Subsidies, Built Environment, School Procurement, Worksite Wellness, and Marketing Standards to Improve Diet.

    PubMed

    Afshin, Ashkan; Penalvo, Jose; Del Gobbo, Liana; Kashaf, Michael; Micha, Renata; Morrish, Kurtis; Pearson-Stuttard, Jonathan; Rehm, Colin; Shangguan, Siyi; Smith, Jessica D; Mozaffarian, Dariush

    2015-11-01

    Poor diet is the leading cause of cardiovascular disease in the USA and globally. Evidence-based policies are crucial to improve diet and population health. We reviewed the effectiveness for a range of policy levers to alter diet and diet-related risk factors. We identified evidence to support benefits of focused mass media campaigns (especially for fruits, vegetables, salt), food pricing strategies (both subsidies and taxation, with stronger effects at lower income levels), school procurement policies (for increasing healthful or reducing unhealthful choices), and worksite wellness programs (especially when comprehensive and multicomponent). Evidence was inconclusive for food and menu labeling (for consumer or industry behavior) and changes in local built environment (e.g., availability or accessibility of supermarkets, fast food outlets). We found little empiric evidence evaluating marketing restrictions, although broad principles and large resources spent on marketing suggest utility. Widespread implementation and evaluation of evidence-based policy strategies, with further research on other strategies with mixed/limited evidence, are essential "population medicine" to reduce health and economic burdens and inequities of diet-related illness worldwide. PMID:26370554

  20. Review, evaluation, and discussion of the challenges of missing value imputation for mass spectrometry-based label-free global proteomics

    DOE PAGESBeta

    Webb-Robertson, Bobbie-Jo M.; Wiberg, Holli K.; Matzke, Melissa M.; Brown, Joseph N.; Wang, Jing; McDermott, Jason E.; Smith, Richard D.; Rodland, Karin D.; Metz, Thomas O.; Pounds, Joel G.; et al

    2015-04-09

    In this review, we apply selected imputation strategies to label-free liquid chromatography–mass spectrometry (LC–MS) proteomics datasets to evaluate the accuracy with respect to metrics of variance and classification. We evaluate several commonly used imputation approaches for individual merits and discuss the caveats of each approach with respect to the example LC–MS proteomics data. In general, local similarity-based approaches, such as the regularized expectation maximization and least-squares adaptive algorithms, yield the best overall performances with respect to metrics of accuracy and robustness. However, no single algorithm consistently outperforms the remaining approaches, and in some cases, performing classification without imputation sometimes yieldedmore » the most accurate classification. Thus, because of the complex mechanisms of missing data in proteomics, which also vary from peptide to protein, no individual method is a single solution for imputation. In summary, on the basis of the observations in this review, the goal for imputation in the field of computational proteomics should be to develop new approaches that work generically for this data type and new strategies to guide users in the selection of the best imputation for their dataset and analysis objectives.« less

  1. CVD Prevention Through Policy: a Review of Mass Media, Food/Menu Labeling, Taxation/Subsidies, Built Environment, School Procurement, Worksite Wellness, and Marketing Standards to Improve Diet.

    PubMed

    Afshin, Ashkan; Penalvo, Jose; Del Gobbo, Liana; Kashaf, Michael; Micha, Renata; Morrish, Kurtis; Pearson-Stuttard, Jonathan; Rehm, Colin; Shangguan, Siyi; Smith, Jessica D; Mozaffarian, Dariush

    2015-11-01

    Poor diet is the leading cause of cardiovascular disease in the USA and globally. Evidence-based policies are crucial to improve diet and population health. We reviewed the effectiveness for a range of policy levers to alter diet and diet-related risk factors. We identified evidence to support benefits of focused mass media campaigns (especially for fruits, vegetables, salt), food pricing strategies (both subsidies and taxation, with stronger effects at lower income levels), school procurement policies (for increasing healthful or reducing unhealthful choices), and worksite wellness programs (especially when comprehensive and multicomponent). Evidence was inconclusive for food and menu labeling (for consumer or industry behavior) and changes in local built environment (e.g., availability or accessibility of supermarkets, fast food outlets). We found little empiric evidence evaluating marketing restrictions, although broad principles and large resources spent on marketing suggest utility. Widespread implementation and evaluation of evidence-based policy strategies, with further research on other strategies with mixed/limited evidence, are essential "population medicine" to reduce health and economic burdens and inequities of diet-related illness worldwide.

  2. A study of the elimination of water from lithium-cationized tripeptide methyl esters by means of tandem mass spectrometry and isotope labeling.

    PubMed

    Talaty, Erach R; Cooper, Travis J; Piland, Debra L; Bateman, David J; Syed, Adeel; Stevenson, William; Van Stipdonk, Michael J

    2006-01-01

    Extensive isotope labeling (2H, 13C and 15N), collision-induced dissociation (CID) and multiple-stage tandem mass spectrometry were used to investigate the elimination of H2O from a series of model, metal-cationized tripeptide methyl esters. The present results corroborate our earlier suggestion that loss of water from lithiated peptides is initiated by a nucleophilic attack from the N-terminal side upon an amide carbonyl carbon atom to form a five-membered ring as an intermediate followed by 1,2-elimination of water. We show that the nucleophilic atom is the oxygen atom of the N-terminal amide group in the fragmentation of [AcGGGOMe+Li]+ as well as [GGGOMe+Li]+. However, the subsequent fragmentation is markedly different in the two cases as a result of the absence and presence of a free amino group. In particular, extensive scrambling of protons in the alpha-positions of GGGOMe is observed, presumably as a consequence of intervention of the basic amino group.

  3. Nutrition Labeling

    NASA Astrophysics Data System (ADS)

    Metzger, Lloyd E.

    Nutrition labeling regulations differ in countries around the world. The focus of this chapter is on nutrition labeling regulations in the USA, as specified by the Food and Drug Administration (FDA) and the Food Safety and Inspection Service (FSIS) of the United States Department of Agriculture (USDA). A major reason for analyzing the chemical components of foods in the USA is nutrition labeling regulations. Nutrition label information is not only legally required in many countries, but also is of increasing importance to consumers as they focus more on health and wellness.

  4. Quantitative analysis of 15N labeled positional isomers of glutamine and citrulline via electrospray ionization tandem mass spectrometry of their dansyl derivatives

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The enteral metabolism of glutamine and citrulline are intertwined because, while glutamine is one of the main fuel sources for the enterocyte, citrulline is one of its products. It has been shown that the administration of 15N labeled glutamine results in the incorporation of the 15N label into cit...

  5. Linkage and Branch Analysis of High-Mannose Oligosaccharides Using Closed-Ring Labeling of 8-Aminopyrene-1,3,6-Trisulfonate and P-Aminobenzoic Ethyl Ester and Negative Ion Trap Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Chen, Shu-Ting; Her, Guor-Rong

    2012-08-01

    A strategy based on negative ion electrospray ionization tandem mass spectrometry and closed-ring labeling with both 8-aminopyrene-1,3,6-trisulfonate (APTS) and p-aminobenzoic acid ethyl ester (ABEE) was developed for linkage and branch determination of high-mannose oligosaccharides. X-type cross-ring fragment ions obtained from APTS-labeled oligosaccharides by charge remote fragmentation provided information on linkages near the non-reducing terminus. In contrast, A-type cross-ring fragment ions observed from ABEE-labeled oligosaccharides yielded information on linkages near the reducing terminus. This complementary information provided by APTS- and ABEE-labeled oligosaccharides was utilized to delineate the structures of the high-mannose oligosaccharides. As a demonstration of this approach, the linkages and branches of high-mannose oligosaccharides Man5GlcNAc2, Man6GlcNAc2, Man8GlcNAc2, and Man9GlcNAc2 cleaved from the ribonuclease B were assigned from MS2 spectra of ABEE- and APTS-labeled derivatives.

  6. Fish proteins as targets of ferrous-catalyzed oxidation: identification of protein carbonyls by fluorescent labeling on two-dimensional gels and MALDI-TOF/TOF mass spectrometry.

    PubMed

    Pazos, Manuel; da Rocha, Angela Pereira; Roepstorff, Peter; Rogowska-Wrzesinska, Adelina

    2011-07-27

    Protein oxidation in fish meat is considered to affect negatively the muscle texture. An important source of free radicals taking part in this process is Fenton's reaction dependent on ferrous ions present in the tissue. The aim of this study was to investigate the susceptibility of cod muscle proteins in sarcoplasmic and myofibril fractions to in vitro metal-catalyzed oxidation and to point out protein candidates that might play a major role in the deterioration of fish quality. Extracted control proteins and proteins subjected to free radicals generated by Fe(II)/ascorbate mixture were labeled with fluorescein-5-thiosemicarbazide (FTSC) to tag carbonyl groups and separated by two-dimensional gel electrophoresis. Consecutive visualization of protein carbonyl levels by capturing the FTSC signal and total protein levels by capturing the SyproRuby staining signal allowed us to quantify the relative change in protein carbonyl levels corrected for changes in protein content. Proteins were identified using MALDI-TOF/TOF mass spectrometry and homology-based searches. The results show that freshly extracted cod muscle proteins exhibit a detectable carbonylation background and that the incubation with Fe(II)/ascorbate triggers a further oxidation of both sarcoplasmic and myofibril proteins. Different proteins exhibited various degrees of sensitivity to oxidation processes. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH), nucleoside diphosphate kinase B (NDK), triosephosphate isomerase, phosphoglycerate mutase, lactate dehydrogenase, creatine kinase, and enolase were the sarcoplasmic proteins most vulnerable to ferrous-catalyzed oxidation. Moreover, NDK, phosphoglycerate mutase, and GAPDH were identified in several spots differing by their pI, and those forms showed different susceptibilities to metal-catalyzed oxidation, indicating that post-translational modifications may change the resistance of proteins to oxidative damage. The Fe(II)/ascorbate treatment significantly

  7. LC-quadrupole/Orbitrap high-resolution mass spectrometry enables stable isotope-resolved simultaneous quantification and ¹³C-isotopic labeling of acyl-coenzyme A thioesters.

    PubMed

    Frey, Alexander J; Feldman, Daniel R; Trefely, Sophie; Worth, Andrew J; Basu, Sankha S; Snyder, Nathaniel W

    2016-05-01

    Acyl-coenzyme A (acyl-CoA) thioesters are evolutionarily conserved, compartmentalized, and energetically activated substrates for biochemical reactions. The ubiquitous involvement of acyl-CoA thioesters in metabolism, including the tricarboxylic acid cycle, fatty acid metabolism, amino acid degradation, and cholesterol metabolism highlights the broad applicability of applied measurements of acyl-CoA thioesters. However, quantitation of acyl-CoA levels provides only one dimension of metabolic information and a more complete description of metabolism requires the relative contribution of different precursors to individual substrates and pathways. Using two distinct stable isotope labeling approaches, acyl-CoA thioesters can be labeled with either a fixed [(13)C3(15)N1] label derived from pantothenate into the CoA moiety or via variable [(13)C] labeling into the acyl chain from metabolic precursors. Liquid chromatography-hybrid quadrupole/Orbitrap high-resolution mass spectrometry using parallel reaction monitoring, but not single ion monitoring, allowed the simultaneous quantitation of acyl-CoA thioesters by stable isotope dilution using the [(13)C3(15)N1] label and measurement of the incorporation of labeled carbon atoms derived from [(13)C6]-glucose, [(13)C5(15)N2]-glutamine, and [(13)C3]-propionate. As a proof of principle, we applied this method to human B cell lymphoma (WSU-DLCL2) cells in culture to precisely describe the relative pool size and enrichment of isotopic tracers into acetyl-, succinyl-, and propionyl-CoA. This method will allow highly precise, multiplexed, and stable isotope-resolved determination of metabolism to refine metabolic models, characterize novel metabolism, and test modulators of metabolic pathways involving acyl-CoA thioesters. PMID:26968563

  8. Transformation of 17β-estradiol in humic acid solution by ε-MnO2 nanorods as probed by high-resolution mass spectrometry combined with (13)C labeling.

    PubMed

    Sun, Kai; Liang, Shangtao; Kang, Fuxing; Gao, Yanzheng; Huang, Qingguo

    2016-07-01

    Steroidal estrogens (SEs), widespread in aquatic systems, have a potential to disrupt the endocrine system of wildlife species and humans. In our experiments, the performance of ε-MnO2 nanorods in transforming 17β-estradiol (E2) was investigated, and the effect of humic acid (HA) on the reaction behaviors was systematically characterized. Reconfiguration of humic molecules was also investigated by high-performance size exclusion chromatography (HPSEC). Results indicated that ε-MnO2 nanomaterials ensured efficient removal of E2 from the aqueous solution. The presence of HA hindered the transformation of E2, while enhanced the cross-coupling between E2 and humic molecules. In particular, we used a mixture of un-labeled E2 and (13)C3-labeled E2 at a 1: 1 set ratio (w/w) to probe the reaction products via high-resolution mass spectrometry (HRMS). The combination of HRMS and (13)C3-labeling revealed the intermediate products including estrone (E1), and hydroxylated, quinone-like, and ring-opened species, as well as E2 dimer and trimer. More importantly, possible cross-coupling products between E2 and HA were also identified. A reaction mechanism including two-electron oxidation and single-electron oxidation was proposed. The applied analytical approach using HRMS along with (13)C3-labeling for reaction-product identification is crucial to understanding the role of HA in the transformation of SEs.

  9. Transformation of 17β-estradiol in humic acid solution by ε-MnO2 nanorods as probed by high-resolution mass spectrometry combined with (13)C labeling.

    PubMed

    Sun, Kai; Liang, Shangtao; Kang, Fuxing; Gao, Yanzheng; Huang, Qingguo

    2016-07-01

    Steroidal estrogens (SEs), widespread in aquatic systems, have a potential to disrupt the endocrine system of wildlife species and humans. In our experiments, the performance of ε-MnO2 nanorods in transforming 17β-estradiol (E2) was investigated, and the effect of humic acid (HA) on the reaction behaviors was systematically characterized. Reconfiguration of humic molecules was also investigated by high-performance size exclusion chromatography (HPSEC). Results indicated that ε-MnO2 nanomaterials ensured efficient removal of E2 from the aqueous solution. The presence of HA hindered the transformation of E2, while enhanced the cross-coupling between E2 and humic molecules. In particular, we used a mixture of un-labeled E2 and (13)C3-labeled E2 at a 1: 1 set ratio (w/w) to probe the reaction products via high-resolution mass spectrometry (HRMS). The combination of HRMS and (13)C3-labeling revealed the intermediate products including estrone (E1), and hydroxylated, quinone-like, and ring-opened species, as well as E2 dimer and trimer. More importantly, possible cross-coupling products between E2 and HA were also identified. A reaction mechanism including two-electron oxidation and single-electron oxidation was proposed. The applied analytical approach using HRMS along with (13)C3-labeling for reaction-product identification is crucial to understanding the role of HA in the transformation of SEs. PMID:27086077

  10. μPET imaging of the pharmacokinetic behavior of medium and high molar mass (89)Zr-labeled poly(2-ethyl-2-oxazoline) in comparison to poly(ethylene glycol).

    PubMed

    Wyffels, Leonie; Verbrugghen, Thomas; Monnery, Bryn D; Glassner, Mathias; Stroobants, Sigrid; Hoogenboom, Richard; Staelens, Steven

    2016-08-10

    Poly(2-oxazoline)s are a promising class of polymers for biomedical applications and a versatile alternative to poly(ethylene glycol)s (PEG). In this work, the pharmacokinetic behavior of well defined (89)Zr-labeled poly(2-ethyl-2-oxazoline)s (PEtOx) was evaluated and compared to that of (89)Zr-labeled PEG, both with varying molar mass. Amine-terminated PEtOx of low dispersity in a molar mass range of 20 to 110kDa and PEG of 20 and 40kDa were functionalized with a desferrioxamine chelator and radiolabeled with (89)Zr. The tissue distribution of both radiolabeled PEtOx and PEG polymers was studied by means of micro Positron Emission Tomography (μPET) molecular imaging in mice longitudinally up to 1week post injection, followed by ex vivo biodistribution. As previously described for other classes of non-ionic polymers, the blood clearance of PEtOx decreased with molar mass. The cut off for glomerular filtration of PEtOx is likely to be around 40kDa. The head to head comparison of PEG and PEtOx revealed that the biodistribution is mostly dominated by polymer chain length and not polymer molar mass. This study constitutes an important addition to further establishing PEtOx as a promising polymer in biomedical applications. PMID:27235979

  11. Atmospheric pressure gas chromatography coupled to quadrupole-time of flight mass spectrometry as a tool for identification of volatile migrants from autoadhesive labels used for direct food contact.

    PubMed

    Canellas, Elena; Vera, Paula; Nerín, Cristina

    2014-11-01

    Pressure-sensitive adhesives (PSA) are used to manufacture labels that are applied directly on the food. These adhesives could contain not only intentionally added compounds (IAS) to the adhesive formula but also non-intentionally added substances (NIAS), due to the impurities from the raw materials used, decomposition of the initial components or from chemical interactions between them. These compounds could migrate to the food and contaminate it. In this study, gas chromatography coupled with mass spectrometry (GC-MS/Q) and atmospheric pressure gas chromatography coupled to a quadrupole hyphenated to a time of flight mass spectrometer (APGC-MS/Q-TOF) have been used for identification of unknown compounds and NIAS coming from a PSA. Seven compounds were identified by GC-MS/Q, and other eight compounds remained initially unknown. The structure of these eight new compounds was elucidated by working with the spectra obtained by APGC-MS/Q-TOF. Finally, two different migration studies were carried out. The first one with Tenax as solid food simulant in contact with the paper label containing the adhesive and the second one with isooctane filled in a natural pork intestine where the label containing the adhesive was applied on the external side. The results are shown and discussed. PMID:25395134

  12. Atmospheric pressure gas chromatography coupled to quadrupole-time of flight mass spectrometry as a tool for identification of volatile migrants from autoadhesive labels used for direct food contact.

    PubMed

    Canellas, Elena; Vera, Paula; Nerín, Cristina

    2014-11-01

    Pressure-sensitive adhesives (PSA) are used to manufacture labels that are applied directly on the food. These adhesives could contain not only intentionally added compounds (IAS) to the adhesive formula but also non-intentionally added substances (NIAS), due to the impurities from the raw materials used, decomposition of the initial components or from chemical interactions between them. These compounds could migrate to the food and contaminate it. In this study, gas chromatography coupled with mass spectrometry (GC-MS/Q) and atmospheric pressure gas chromatography coupled to a quadrupole hyphenated to a time of flight mass spectrometer (APGC-MS/Q-TOF) have been used for identification of unknown compounds and NIAS coming from a PSA. Seven compounds were identified by GC-MS/Q, and other eight compounds remained initially unknown. The structure of these eight new compounds was elucidated by working with the spectra obtained by APGC-MS/Q-TOF. Finally, two different migration studies were carried out. The first one with Tenax as solid food simulant in contact with the paper label containing the adhesive and the second one with isooctane filled in a natural pork intestine where the label containing the adhesive was applied on the external side. The results are shown and discussed.

  13. Data set of the protein expression profiles of Luminal A, Claudin-low and overexpressing HER2(+) breast cancer cell lines by iTRAQ labelling and tandem mass spectrometry.

    PubMed

    Calderón-González, Karla Grisel; Valero Rustarazo, Ma Luz; Labra-Barrios, Maria Luisa; Bazán-Méndez, César Isaac; Tavera-Tapia, Alejandra; Herrera-Aguirre, Marí aEsther; Sánchez Del Pino, Manuel M; Gallegos-Pérez, José Luis; González-Márquez, Humberto; Hernández-Hernández, Jose Manuel; León-Ávila, Gloria; Rodríguez-Cuevas, Sergio; Guisa-Hohenstein, Fernando; Luna-Arias, Juan Pedro

    2015-09-01

    Breast cancer is the most common and the leading cause of mortality in women worldwide. There is a dire necessity of the identification of novel molecules useful in diagnosis and prognosis. In this work we determined the differentially expression profiles of four breast cancer cell lines compared to a control cell line. We identified 1020 polypeptides labelled with iTRAQ with more than 95% in confidence. We analysed the common proteins in all breast cancer cell lines through IPA software (IPA core and Biomarkers). In addition, we selected the specific overexpressed and subexpressed proteins of the different molecular classes of breast cancer cell lines, and classified them according to protein class and biological process. Data in this article is related to the research article "Determination of the protein expression profiles of breast cancer cell lines by Quantitative Proteomics using iTRAQ Labelling and Tandem Mass Spectrometry" (Calderón-González et al. [1] in press). PMID:26217805

  14. Characterization of a benzyladenine binding-site peptide isolated from a wheat cytokinin-binding protein: Sequence analysis and identification of a single affinity-labeled histidine residue by mass spectrometry

    SciTech Connect

    Brinegar, A.C.; Cooper, G.; Stevens, A.; Hauer, C.R.; Shabanowitz, J.; Hunt, D.F.; Fox, J.E. )

    1988-08-01

    A wheat embryo cytokinin-binding protein was covalently modified with the radiolabeled photoaffinity ligand 2-azido-N{sup 6}-({sup 14}C)benzyladenine. A single labeled peptide was obtained after proteolytic digestion and isolation by reversed-phase and anion-exchange HPLC. Sequencing by classical Edman degradation identified 11 of the 12 residues but failed to identify the labeled amino acid. Analysis by laser photodissociation Fourier-transform mass spectrometry of 10 pmol of the peptide independently confirmed the Edman data and also demonstrated that the histidine residue nearest the C terminus (underlined) was modified by the reagent in the sequence Ala-Phe-Leu-Gln-Pro-Ser-His-His{und His}-Asp-Ala-Asp-Glu.

  15. The use of stable isotope labeling and liquid chromatography/tandem mass spectrometry techniques to study the pharmacokinetics and bioavailability of the antimigraine drug, MK-0462 (rizatriptan) in dogs.

    PubMed

    Barrish, A; Olah, T V; Gatto, G J; Michel, K B; Dobrinska, M R; Gilbert, J D

    1996-01-01

    MK-0462 (rizatriptan) is a 5HT1D agonist being developed for the treatment of migraine. The assay for this substance in plasma and urine is based on HPLC with tandem mass spectrometry (MS/MS) detection. The procedure has been modified to include the simultaneous determination of the [triazole-13C2, 15N3-] stable-isotope-labelled analogue for which the lower quantifiable limit was 0.1 ng mL-1. The assay has been applied to study the pharmacokinetics of MK-0462 after simultaneous oral and intravenous administration of the drug and its stable-isotope-labelled analogue to dogs. The experiment afforded an estimate of plasma clearance concomitant with a precise measurement of the drug's oral bioavailability. The increasing use of LC-MS/MS in quantitative experiments may renew interest in stable isotopes as tools for pharmaceutical research. PMID:8755236

  16. Data set of the protein expression profiles of Luminal A, Claudin-low and overexpressing HER2(+) breast cancer cell lines by iTRAQ labelling and tandem mass spectrometry.

    PubMed

    Calderón-González, Karla Grisel; Valero Rustarazo, Ma Luz; Labra-Barrios, Maria Luisa; Bazán-Méndez, César Isaac; Tavera-Tapia, Alejandra; Herrera-Aguirre, Marí aEsther; Sánchez Del Pino, Manuel M; Gallegos-Pérez, José Luis; González-Márquez, Humberto; Hernández-Hernández, Jose Manuel; León-Ávila, Gloria; Rodríguez-Cuevas, Sergio; Guisa-Hohenstein, Fernando; Luna-Arias, Juan Pedro

    2015-09-01

    Breast cancer is the most common and the leading cause of mortality in women worldwide. There is a dire necessity of the identification of novel molecules useful in diagnosis and prognosis. In this work we determined the differentially expression profiles of four breast cancer cell lines compared to a control cell line. We identified 1020 polypeptides labelled with iTRAQ with more than 95% in confidence. We analysed the common proteins in all breast cancer cell lines through IPA software (IPA core and Biomarkers). In addition, we selected the specific overexpressed and subexpressed proteins of the different molecular classes of breast cancer cell lines, and classified them according to protein class and biological process. Data in this article is related to the research article "Determination of the protein expression profiles of breast cancer cell lines by Quantitative Proteomics using iTRAQ Labelling and Tandem Mass Spectrometry" (Calderón-González et al. [1] in press).

  17. Method for evaluating the potential of C labeled plant polyphenols to cross the blood-brain barrier using accelerator mass spectrometry.

    PubMed

    Janle, Elsa M; Lila, Mary Ann; Grannan, Michael; Wood, Lauren; Higgins, Aine; Yousef, Gad G; Rogers, Randy B; Kim, Helen; Jackson, George S; Weaver, Connie M

    2010-04-01

    Bioactive compounds in botanicals may be beneficial in preventing age-related neurodegenerative diseases, but for many compounds conventional methods may be inadequate to detect if these compounds cross the blood brain barrier or to track the pharmacokinetics in the brain. By combining a number of unique technologies it has been possible to utilize the power of AMS to study the pharmacokinetics of bioactive compounds in the brain at very low concentrations. (14)C-labeled compounds can be biosynthesized by plant cell suspension cultures co-incubated with radioisotopically-labeled sucrose and isolated and separated into a series of bioactive fractions.To study the pharmacokinetics and tissue distribution of (14)C labeled plant polyphenols, rats were implanted with jugular catheters, subcutaneous ultrafiltration probes and brain microdialysis probes. Labeled fractions were dosed orally. Interstitial fluid (ISF) and brain microdialysate samples were taken in tandem with blood samples. It was often possible to determine (14)C in blood and ISF with a β-counter. However, brain microdialysate samples (14)C levels on the order of 10(7) atoms/sample required AMS technology. The Brain Microdialysate(AUC)/Serum(AUC) ranged from .021- to .029, with the higher values for the glycoside fractions. By using AMS in combination with traditional methods, it is possible to study uptake by blood, distribution to ISF and determine the amount of a dose which can reach the brain and follow the pharmacokinetics in the brain. PMID:20419067

  18. Method for evaluating the potential of 14C labeled plant polyphenols to cross the blood-brain barrier using accelerator mass spectrometry

    NASA Astrophysics Data System (ADS)

    Janle, Elsa M.; Lila, Mary Ann; Grannan, Michael; Wood, Lauren; Higgins, Aine; Yousef, Gad G.; Rogers, Randy B.; Kim, Helen; Jackson, George S.; Weaver, Connie M.

    2010-04-01

    Bioactive compounds in botanicals may be beneficial in preventing age-related neurodegenerative diseases, but for many compounds conventional methods may be inadequate to detect if these compounds cross the blood-brain barrier or to track the pharmacokinetics in the brain. By combining a number of unique technologies it has been possible to utilize the power of AMS to study the pharmacokinetics of bioactive compounds in the brain at very low concentrations. 14C labeled compounds can be biosynthesized by plant cell suspension cultures co-incubated with radioisotopically-labeled sucrose and isolated and separated into a series of bioactive fractions. To study the pharmacokinetics and tissue distribution of 14C labeled plant polyphenols, rats were implanted with jugular catheters, subcutaneous ultrafiltration probes and brain microdialysis probes. Labeled fractions were dosed orally. Interstitial fluid (ISF) and brain microdialysate samples were taken in tandem with blood samples. It was often possible to determine 14C in blood and ISF with a β-counter. However, brain microdialysate samples 14C levels on the order of 10 7 atoms/sample required AMS technology. The Brain Microdialysate AUC/Serum AUC ranged from .021- to .029, with the higher values for the glycoside fractions. By using AMS in combination with traditional methods, it is possible to study uptake by blood, distribution to ISF and determine the amount of a dose which can reach the brain and follow the pharmacokinetics in the brain.

  19. Investigation of low-abundant in vitro metabolites of stable isotope-labelled BAL4815 by accurate mass capillary-LC-ESI-qTof-MS and MS/MS.

    PubMed

    Wind, Mathias; Spickermann, Jochen; Schleimer, Michael; Donzelli, Massimiliano; Gebhardt, Klaus; Sturm-Haurany, Rima; Klauer, Dominique; Fullhardt, Pascal; Schmitt-Hoffmann, Anne

    2006-07-01

    The metabolic profile of BAL4815, an antifungal azole drug, was determined using in vitro rat hepatocyte incubations and subsequent analysis by capillary LC-qTof-MS and MS/MS including accurate mass determination. For the detection of the metabolites, a mixture of the drug and its deuterium-labelled analogue was used for incubations. Metabolic stability of BAL4815 was high in cultured rat hepatocytes. However, several low-abundant metabolites were detected by the use of capillary LC-qTof-MS and manual investigation of the data. The peak intensity of the most abundant metabolite was close to the limit of detection. Except for an apparent oxidation product, the masses of the other detected metabolites could not be assigned to a single and frequently occurring biotransformation. Accurate mass determination and possible elemental compositions suggested that metabolism occurred through a combination of glutathionylation and defluorination. This was verified using accurate mass MS/MS. The use of accurate mass measurements and the derived suggestions for the elemental compositions were essential to elucidate this atypical metabolic pathway. A mass accuracy better than 8 ppm could be achieved for most assigned MS and MS/MS signals with intensities less than 6 cps in the spectra.

  20. Whole proteome analysis of the protozoan parasite Trypanosoma brucei using stable isotope labeling by amino acids in cell culture and mass spectrometry.

    PubMed

    Cirovic, Olivera; Ochsenreiter, Torsten

    2014-01-01

    The single-celled protozoan Trypanosoma brucei spp. is the causative agent of human African trypanosomiasis and nagana in cattle. Quantitative proteomics for the first time has allowed for the characterization of the proteome from several different life stages of the parasite (Butter et al., Mol Cell Proteomics 12:172-179, 2013; Gunasekera et al., BMC Genomics 13:556, 2012; Urbaniak et al., PloS One 7(5):e36619, 2012). To achieve this, stable isotope labeling by amino acids in cell culture (SILAC) (Ong et al., Mol Cell Proteomics 1:376-386, 2002) was adapted to T. brucei spp. cultures. T. brucei cells grown in standard media with dialyzed fetal calf serum containing heavy isotope-labeled amino acids (arginine and lysine) show efficient incorporation of the labeled amino acids into the whole cell proteome (8-12 divisions) and no detectable amino acid conversions. The method can be applied to both of the major life stages of the parasite and in combination with RNAi or gene knockout approaches.

  1. Studies of peptide a- and b-type fragment ions using stable isotope labeling and integrated ion mobility/tandem mass spectrometry.

    PubMed

    Riba Garcia, Isabel; Giles, Kevin; Bateman, Robert H; Gaskell, Simon J

    2008-12-01

    The structures of peptide a- and b-type fragment ions were studied using synthetic peptides including a set of isomeric peptides, differing in the sequence location of an alanine residue labeled with (15)N and uniformly with (13)C. The pattern of isotope labeling of second-generation fragment ions derived via a(n) and b(n) ions (where n = 4 or 5) suggested that these intermediates existed in part as macrocyclic structures, where alternative sites of ring opening gave rise to different linear forms whose simple cleavage might give rise to the observed final products. Similar conclusions were derived from combined ion mobility/tandem MS analyses where different fragmentation patterns were observed for isomeric a- or b-type ions that display different ion mobilities. These analyses were facilitated by a new approach to the processing of ion mobility/tandem MS data, from which distinct and separate product ion spectra are derived from ions that are incompletely separated by ion mobility. Finally, an example is provided of evidence for a macrocyclic structure for b(n) ions where n = 8 or 9.

  2. Detection of reactive metabolites using isotope-labeled glutathione trapping and simultaneous neutral loss and precursor ion scanning with ultra-high-pressure liquid chromatography triple quadruple mass spectrometry.

    PubMed

    Huang, Ke; Huang, Lingyi; van Breemen, Richard B

    2015-04-01

    Metabolic activation of drugs to electrophilic species is responsible for over 60% of black box warnings and drug withdrawals from the market place in the United States. Reactive metabolite trapping using glutathione (GSH) and analysis using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) or HPLC with high resolution mass spectrometry (mass defect filtering) have enabled screening for metabolic activation to become routine during drug development. However, current MS-based approaches cannot detect all GSH conjugates present in complex mixtures, especially those present in extracts of botanical dietary supplements. To overcome these limitations, a fast triple quadrupole mass spectrometer-based approach was developed that can detect positively and negatively charged GSH conjugates in a single analysis without the need for advanced knowledge of the elemental compositions of potential conjugates and while avoiding false positives. This approach utilized UHPLC instead of HPLC to shorten separation time and enhance sensitivity, incorporated stable-isotope labeled GSH to avoid false positives, and used fast polarity switching electrospray MS/MS to detect GSH conjugates that form positive and/or negative ions. The general new method was then used to test the licorice dietary supplement Glycyrrhiza glabra, which was found to form multiple GSH conjugates upon metabolic activation. Among the GSH conjugates found in the licorice assay were conjugates with isoliquiritigenin and glabridin, which is an irreversible inhibitor of cytochrome P450 enzymes.

  3. Spatial and temporal distribution of 13C labelled plant residues in soil aggregates and Lumbricus terrestris surface casts: A combination of Transmission Electron Microscopy and Nanoscale Secondary Ion Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Vidal, Alix; Remusat, Laurent; Watteau, Françoise; Derenne, Sylvie; Quenea, Katell

    2016-04-01

    Earthworms play a central role in litter decomposition, soil structuration and carbon cycling. They ingest both organic and mineral compounds which are mixed, complexed with mucus and dejected in form of casts at the soil surface and along burrows. Bulk isotopic or biochemical technics have often been used to study the incorporation of litter in soil and casts, but they could not reflect the complex interaction between soil, plant and microorganisms at the microscale. However, the heterogeneous distribution of organic carbon in soil structures induces contrasted microbial activity areas. Nano-scale secondary ion mass spectrometry (NanoSIMS), which is a high spatial resolution method providing elemental and isotopic maps of organic and mineral materials, has recently been applied in soil science (Herrmann et al., 2007; Vogel et al., 2014). The combination of Nano-scale secondary ion mass spectrometry (NanoSIMS) and Transmission Electron Microscopy (TEM) has proven its potential to investigate labelled residues incorporation in earthworm casts (Vidal et al., 2016). In line of this work, we studied the spatial and temporal distribution of plant residues in soil aggregates and earthworm surface casts. This study aimed to (1) identify the decomposition states of labelled plant residues incorporated at different time steps, in casts and soil, (2) identify the microorganisms implied in this decomposition (3) relate the organic matter states of decomposition with their 13C signature. A one year mesocosm experiment was set up to follow the incorporation of 13C labelled Ryegrass (Lolium multiflorum) litter in a soil in the presence of anecic earthworms (Lumbricus terrestris). Soil and surface cast samples were collected after 8 and 54 weeks, embedded in epoxy resin and cut into ultra-thin sections. Soil was fractionated and all and analyzed with TEM and NanoSIMS, obtaining secondary ion images of 12C, 16O, 12C14N, 13C14N and 28Si. The δ13C maps were obtained using the 13C14

  4. 4-Phenylaminomethyl-Benzeneboric Acid Modified Tip Extraction for Determination of Brassinosteroids in Plant Tissues by Stable Isotope Labeling-Liquid Chromatography-Mass Spectrometry.

    PubMed

    Yu, Lei; Ding, Jun; Wang, Ya-Lan; Liu, Ping; Feng, Yu-Qi

    2016-01-19

    Monitoring brassinosteroids (BRs) has been of major interest of researchers as these substances play a crucial role in a variety of phytological processes in plants. However, the determination of endogenous BRs in plant tissue is still a challenging task due to their low abundance and the complex matrix of plant tissues. In this study, a single step strategy by combining tip extraction and in situ derivatization was proposed for BR analysis. In the proposed strategy, a mixed mode sorbent (C8-SO3H) in tip was modified with 4-phenylaminomethyl-benzeneboric acid (4-PAMBA) through cation exchange and hydrophobic interactions, and then used as a boronate affinity media to selectively capture and purify BRs from plant extract through the reaction of boric acid groups of 4-PAMBA and cis-diol on BRs. The BRs-4-PAMBA derivatives formed were easily eluted from the C8-SO3H tip by nullifying the ion exchange and hydrophobic interactions using ammonia acetonitrile, followed by LC-MS/MS analysis. BR standards, isotopically labeled with d5-4-phenylaminomethyl-benzeneboric acid (4-PAMBA-d5) were introduced to improve the assay precision of LC-MS/MS. Under the optimized conditions, the overall process could be completed within 1 h, which is greatly improved in speed compared with previously reported protocols. In addition, the detection sensitivities of labeled BRs were improved by over 2000-fold compared with unlabeled BRs, thus the consumption of plant materials was reduced to 50 mg. Finally, the proposed method was applied for the investigation of BRs response in rice toward Cd stress. PMID:26650986

  5. 4-Phenylaminomethyl-Benzeneboric Acid Modified Tip Extraction for Determination of Brassinosteroids in Plant Tissues by Stable Isotope Labeling-Liquid Chromatography-Mass Spectrometry.

    PubMed

    Yu, Lei; Ding, Jun; Wang, Ya-Lan; Liu, Ping; Feng, Yu-Qi

    2016-01-19

    Monitoring brassinosteroids (BRs) has been of major interest of researchers as these substances play a crucial role in a variety of phytological processes in plants. However, the determination of endogenous BRs in plant tissue is still a challenging task due to their low abundance and the complex matrix of plant tissues. In this study, a single step strategy by combining tip extraction and in situ derivatization was proposed for BR analysis. In the proposed strategy, a mixed mode sorbent (C8-SO3H) in tip was modified with 4-phenylaminomethyl-benzeneboric acid (4-PAMBA) through cation exchange and hydrophobic interactions, and then used as a boronate affinity media to selectively capture and purify BRs from plant extract through the reaction of boric acid groups of 4-PAMBA and cis-diol on BRs. The BRs-4-PAMBA derivatives formed were easily eluted from the C8-SO3H tip by nullifying the ion exchange and hydrophobic interactions using ammonia acetonitrile, followed by LC-MS/MS analysis. BR standards, isotopically labeled with d5-4-phenylaminomethyl-benzeneboric acid (4-PAMBA-d5) were introduced to improve the assay precision of LC-MS/MS. Under the optimized conditions, the overall process could be completed within 1 h, which is greatly improved in speed compared with previously reported protocols. In addition, the detection sensitivities of labeled BRs were improved by over 2000-fold compared with unlabeled BRs, thus the consumption of plant materials was reduced to 50 mg. Finally, the proposed method was applied for the investigation of BRs response in rice toward Cd stress.

  6. Data set of the protein expression profiles of Luminal A, Claudin-low and overexpressing HER2+ breast cancer cell lines by iTRAQ labelling and tandem mass spectrometry

    PubMed Central

    Calderón-González, Karla Grisel; Valero Rustarazo, Ma Luz; Labra-Barrios, Maria Luisa; Bazán-Méndez, César Isaac; Tavera-Tapia, Alejandra; Herrera-Aguirre, Marí;aEsther; Sánchez del Pino, Manuel M.; Gallegos-Pérez, José Luis; González-Márquez, Humberto; Hernández-Hernández, Jose Manuel; León-Ávila, Gloria; Rodríguez-Cuevas, Sergio; Guisa-Hohenstein, Fernando; Luna-Arias, Juan Pedro

    2015-01-01

    Breast cancer is the most common and the leading cause of mortality in women worldwide. There is a dire necessity of the identification of novel molecules useful in diagnosis and prognosis. In this work we determined the differentially expression profiles of four breast cancer cell lines compared to a control cell line. We identified 1020 polypeptides labelled with iTRAQ with more than 95% in confidence. We analysed the common proteins in all breast cancer cell lines through IPA software (IPA core and Biomarkers). In addition, we selected the specific overexpressed and subexpressed proteins of the different molecular classes of breast cancer cell lines, and classified them according to protein class and biological process. Data in this article is related to the research article “Determination of the protein expression profiles of breast cancer cell lines by Quantitative Proteomics using iTRAQ Labelling and Tandem Mass Spectrometry” (Calderón-González et al. [1] in press). PMID:26217805

  7. Label-free mass spectrometric analysis of the mdx-4cv diaphragm identifies the matricellular protein periostin as a potential factor involved in dystrophinopathy-related fibrosis.

    PubMed

    Holland, Ashling; Dowling, Paul; Meleady, Paula; Henry, Michael; Zweyer, Margit; Mundegar, Rustam R; Swandulla, Dieter; Ohlendieck, Kay

    2015-07-01

    Proteomic profiling plays a decisive role in the identification of novel biomarkers of muscular dystrophy and the elucidation of new pathobiochemical mechanisms that underlie progressive muscle wasting. Building on the findings of recent comparative analyses of tissue samples and body fluids from dystrophic animals and patients afflicted with Duchenne muscular dystrophy, we have used here label-free MS to study the severely dystrophic diaphragm from the not extensively characterized mdx-4cv mouse. This animal model of progressive muscle wasting exhibits less dystrophin-positive revertant fibers than the conventional mdx mouse, making it ideal for the future monitoring of experimental therapies. The pathoproteomic signature of the mdx-4cv diaphragm included a significant increase in the fibrosis marker collagen and related extracellular matrix proteins (asporin, decorin, dermatopontin, prolargin) and cytoskeletal proteins (desmin, filamin, obscurin, plectin, spectrin, tubulin, vimentin, vinculin), as well as decreases in proteins of ion homeostasis (parvalbumin) and the contractile apparatus (myosin-binding protein). Importantly, one of the most substantially increased proteins was identified as periostin, a matricellular component and apparent marker of fibrosis and tissue damage. Immunoblotting confirmed a considerable increase of periostin in the dystrophin-deficient diaphragm from both mdx and mdx-4cv mice, suggesting an involvement of this matricellular protein in dystrophinopathy-related fibrosis.

  8. Measurement of 13C and 15N isotope labeling by gas chromatography/combustion/isotope ratio mass spectrometry to study amino acid fluxes in a plant-microbe symbiotic association.

    PubMed

    Molero, Gemma; Aranjuelo, Iker; Teixidor, Pilar; Araus, José Luis; Nogués, Salvador

    2011-03-15

    We have developed a method based on a double labeling with stable isotopes and gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) analyses to study amino acid exchange in a symbiotic plant-microbe association. Isotopic precision was studied for 21 standards including 15 amino acid derivatives, three N-protected amino acid methyl esters, three amines and one international standard. High correlations were observed between the δ(13)C and δ(15)N values obtained by GC/C/IRMS and those obtained by an elemental analyzer (EA) coupled to an isotope ratio mass spectrometer (R(2) = 0.9868 and 0.9992, respectively). The mean precision measured was 0.04‰ for δ(13)C and 0.28‰ for δ(15)N (n = 15). This method was applied in vivo to the symbiotic relationship between alfalfa (Medicago sativa L.) and N(2)-fixing bacteria. Plants were simultaneously labeled over 10 days with (13)C-depleted CO(2) ((12)CO(2)), which was assimilated through photosynthesis by leaves, and (15)N(2) fixed via nodules. Subsequently, the C and N isotope compositions (i.e. δ(13)C and δ(15)N) of free amino acids were analyzed in leaves and nodules by GC/C/IRMS. The method revealed the pattern of C and N exchange between leaves and nodules, highlighting that γ-aminobutanoic acid and glycine may represent an important form of C transport from leaves to the nodules. The results confirmed the validity, reliability and accuracy of the method for assessing C and N fluxes between plants and symbiotic bacteria and support the use of this technique in a broad range of metabolic and fluxomic studies.

  9. Determination of bovine lactoferrin in dairy products by ultra-high performance liquid chromatography-tandem mass spectrometry based on tryptic signature peptides employing an isotope-labeled winged peptide as internal standard.

    PubMed

    Zhang, Jingshun; Lai, Shiyun; Cai, Zengxuan; Chen, Qi; Huang, Baifen; Ren, Yiping

    2014-06-01

    A new and sensitive determination method was developed for bovine lactoferrin in dairy products including infant formulas based on the signature peptide by ultra high-performance liquid chromatography and triple-quadrupole tandem mass spectrometry under the multiple reaction monitoring mode. The simple pretreatment procedures included the addition of a winged peptide containing the isotope-labeled signature peptide as internal standard, followed by an enzymatic digestion with trypsin. The signature peptide was chosen and identified from the tryptic hydrolyzates of bovine lactoferrin by ultra high-performance liquid chromatography and quadrupole-time-of-flight tandem mass spectrometry based on sequence database search. Analytes were separated on an ACQUITY UPLC BEH 300 C18 column and monitored by MS/MS in seven minutes. Quantitative result bias due to matrix effect and tryptic efficiency was corrected through the use of synthetic isotope-labeled standards. The limit of detection and limit of quantification were 0.3 mg/100 g and 1.0 mg/100 g, respectively. Bovine lactoferrin within the concentration range of 10-1000 nmol L(-1) showed a strong linear relationship with a linear correlation coefficient (r) of >0.998. The intra- and inter-day precision of the method were RSD<6.5% and RSD<7.1%, respectively. Excellent repeatability (RSD<6.4%) substantially supported the application of this method for the determination of bovine lactoferrin in dairy samples. The present method was successfully validated and applied to determination of bovine lactoferrin in dairy products including infant formulas.

  10. Proteomic analysis of rat plasma with experimental autoimmune uveitis based on label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS).

    PubMed

    Guo, Dadong; Gu, Peiming; Liu, Zhengfeng; Tang, Kai; Du, Yuxiang; Bi, Hongsheng

    2015-01-22

    Uveitis is a severe autoimmune eye disease that can cause intraocular inflammation even lead to severe vision loss, and the occurrence of uveitis can be closely associated with abnormal expression of proteins. However, the abnormally expressed proteins involved in uveitis are not well identified. Using liquid chromatography-tandem mass spectrometry technique, we examined the alterations in proteomic expression profiling in rat plasma specimens related to experimental autoimmune uveitis (EAU) versus normal samples. In addition, the experimental verification was further performed using enzyme-linked immunosorbent assay (ELISA) for abnormally expressed proteins in EAU rat plasma. The results indicate that 62 proteins were upregulated and 106 proteins were downregulated in plasma from EAU rats compared with those in saline-treated samples. In the meantime, we observed that the plasma level of complement component 3 in EAU rats was upregulated versus saline-treated rats (from 92.32μg/mL to 168.92μg/mL), whereas the level of interleukin-1 receptor accessory protein was downregulated (from 1120.97pg/mL to 798.39pg/mL), and these results were highly in agreement with those of mass spectrometry determination. Taken together, our results indicate that liquid chromatography-tandem mass spectrometry analysis possesses a good resolution for peptides in plasma, and the findings will provide the baseline plasma dataset for EAU rats and the relevant information can contribute to future studies on the understanding the mechanism of uveitis.

  11. Method development for the redox speciation analysis of iron by ion chromatography-inductively coupled plasma mass spectrometry and carryover assessment using isotopically labeled analyte analogues.

    PubMed

    Wolle, Mesay Mulugeta; Fahrenholz, Timothy; Rahman, G M Mizanur; Pamuku, Matt; Kingston, H M 'Skip'; Browne, Damien

    2014-06-20

    An ion chromatography-inductively coupled plasma mass spectrometry (IC-ICP-MS) method was developed for the redox speciation analysis of iron (Fe) based on in-column complexation of Fe(2+) and Fe(3+) by dipicolinic acid (DPA). The effects of column type, mobile phase composition and molecular ion interference were studied in the method optimization. The carryover of the target species in the IC-ICP-MS method was uniquely and effectively evaluated using isotopically enriched analogues of the analytes ((54)Fe(2+) and (57)Fe(3+)). Standard solutions of the enriched standards were injected into the system following analysis of a sample, and the ratios of the isotopes of iron in the enriched standards were calculated based on the chromatographic peak areas. The concentrations of the analytes carried over from the sample to the enriched standards were determined using the quantitative relationship in isotope dilution mass spectrometry (IDMS). In contrast to the routine way of evaluating carryover effect by injecting a blank solution after sample analysis, the use of isotopically enriched standards identified significant analyte carryover in the present method. Extensive experiments were carried out to systematically identify the source of the carryover and to eliminate the problem; the separation column was found to be the exclusive source. More than 95% of the analyte carryover was eliminated by reducing the length of the column. The detection limit of the IC-ICP-MS method (MDL) for the iron species was 2ngg(-1). The method was used to determine Fe(2+) and Fe(3+) in synthetic aqueous standard solutions and a beverage sample.

  12. Popular computational methods to assess multiprotein complexes derived from label-free affinity purification and mass spectrometry (AP-MS) experiments.

    PubMed

    Armean, Irina M; Lilley, Kathryn S; Trotter, Matthew W B

    2013-01-01

    Advances in sensitivity, resolution, mass accuracy, and throughput have considerably increased the number of protein identifications made via mass spectrometry. Despite these advances, state-of-the-art experimental methods for the study of protein-protein interactions yield more candidate interactions than may be expected biologically owing to biases and limitations in the experimental methodology. In silico methods, which distinguish between true and false interactions, have been developed and applied successfully to reduce the number of false positive results yielded by physical interaction assays. Such methods may be grouped according to: (1) the type of data used: methods based on experiment-specific measurements (e.g., spectral counts or identification scores) versus methods that extract knowledge encoded in external annotations (e.g., public interaction and functional categorisation databases); (2) the type of algorithm applied: the statistical description and estimation of physical protein properties versus predictive supervised machine learning or text-mining algorithms; (3) the type of protein relation evaluated: direct (binary) interaction of two proteins in a cocomplex versus probability of any functional relationship between two proteins (e.g., co-occurrence in a pathway, sub cellular compartment); and (4) initial motivation: elucidation of experimental data by evaluation versus prediction of novel protein-protein interaction, to be experimentally validated a posteriori. This work reviews several popular computational scoring methods and software platforms for protein-protein interactions evaluation according to their methodology, comparative strengths and weaknesses, data representation, accessibility, and availability. The scoring methods and platforms described include: CompPASS, SAINT, Decontaminator, MINT, IntAct, STRING, and FunCoup. References to related work are provided throughout in order to provide a concise but thorough introduction to a

  13. Application of Cassette Ultracentrifugation Using Non-labeled Compounds and Liquid Chromatography-Tandem Mass Spectrometry Analysis for High-Throughput Protein Binding Determination.

    PubMed

    Kieltyka, Kasia; McAuliffe, Brian; Cianci, Christopher; Drexler, Dieter M; Shou, Wilson; Zhang, Jun

    2016-03-01

    Membrane-based devices typically used for serum protein binding determination are not fully applicable to highly lipophilic compounds because of nonspecific binding to the device membrane. Ultracentrifugation, however, completely eliminates the issue by using a membrane-free approach, although its wide application has been limited. This lack of utilization is mainly attributed to 2 factors: the high cost in acquiring and handling of radiolabeled compounds and low assay throughput owing to the difficulties in process automation. To overcome these challenges, we report a high-throughput workflow by cassette ultracentrifugation of nonradiolabeled compounds followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. Twenty compounds with diverse physicochemical and protein binding properties were selected for the evaluation of the workflow. To streamline the working process, approaches of matrix balancing for all the samples for LC-MS/MS analysis and determining free fraction without analytical calibration curves were adopted. Both the discrete ultracentrifugation of individual compounds and cassette ultracentrifugation of all the test compounds followed by simultaneous LC-MS/MS analysis exhibited a linear correlation with literature values, demonstrating respectively the validity of the ultracentrifugation process and the cassette approach. The cassette ultracentrifugation using nonradiolabeled compounds followed by LC-MS/MS analysis has greatly facilitated its application for high-throughput protein binding screening in drug discovery.

  14. Mass spectrometry.

    NASA Technical Reports Server (NTRS)

    Burlingame, A. L.; Johanson, G. A.

    1972-01-01

    Review of the current state of mass spectrometry, indicating its unique importance for advanced scientific research. Mass spectrometry applications in computer techniques, gas chromatography, ion cyclotron resonance, molecular fragmentation and ionization, and isotope labeling are covered. Details are given on mass spectrometry applications in bio-organic chemistry and biomedical research. As the subjects of these applications are indicated alkaloids, carbohydrates, lipids, terpenes, quinones, nucleic acid components, peptides, antibiotics, and human and animal metabolisms. Particular attention is given to the mass spectra of organo-inorganic compounds, inorganic mass spectrometry, surface phenomena such as secondary ion and electron emission, and elemental and isotope analysis. Further topics include mass spectrometry in organic geochemistry, applications in geochronology and cosmochemistry, and organic mass spectrometry.

  15. Mass spectrometry-based microassay of (2)H and (13)C plasma glucose labeling to quantify liver metabolic fluxes in vivo.

    PubMed

    Hasenour, Clinton M; Wall, Martha L; Ridley, D Emerson; Hughey, Curtis C; James, Freyja D; Wasserman, David H; Young, Jamey D

    2015-07-15

    Mouse models designed to examine hepatic metabolism are critical to diabetes and obesity research. Thus, a microscale method to quantitatively assess hepatic glucose and intermediary metabolism in conscious, unrestrained mice was developed. [(13)C3]propionate, [(2)H2]water, and [6,6-(2)H2]glucose isotopes were delivered intravenously in short- (9 h) and long-term-fasted (19 h) C57BL/6J mice. GC-MS and mass isotopomer distribution (MID) analysis were performed on three 40-μl arterial plasma glucose samples obtained during the euglycemic isotopic steady state. Model-based regression of hepatic glucose and citric acid cycle (CAC)-related fluxes was performed using a comprehensive isotopomer model to track carbon and hydrogen atom transitions through the network and thereby simulate the MIDs of measured fragment ions. Glucose-6-phosphate production from glycogen diminished, and endogenous glucose production was exclusively gluconeogenic with prolonged fasting. Gluconeogenic flux from phosphoenolpyruvate (PEP) remained stable, whereas that from glycerol modestly increased from short- to long-term fasting. CAC flux [i.e., citrate synthase (VCS)] was reduced with long-term fasting. Interestingly, anaplerosis and cataplerosis increased with fast duration; accordingly, pyruvate carboxylation and the conversion of oxaloacetate to PEP were severalfold higher than VCS in long-term fasted mice. This method utilizes state-of-the-art in vivo methodology and comprehensive isotopomer modeling to quantify hepatic glucose and intermediary fluxes during physiological stress in mice. The small plasma requirements permit serial sampling without stress and the affirmation of steady-state glucose kinetics. Furthermore, the approach can accommodate a broad range of modeling assumptions, isotope tracers, and measurement inputs without the need to introduce ad hoc mathematical approximations.

  16. Biomimetic oxidative treatment of spruce wood studied by pyrolysis-molecular beam mass spectrometry coupled with multivariate analysis and 13C-labeled tetramethylammonium hydroxide thermochemolysis: implications for fungal degradation of wood.

    PubMed

    Arantes, Valdeir; Qian, Yuhui; Kelley, Stephen S; Milagres, Adriane M F; Filley, Timothy R; Jellison, Jody; Goodell, Barry

    2009-11-01

    In this work, pyrolysis-molecular beam mass spectrometry analysis coupled with principal components analysis and (13)C-labeled tetramethylammonium hydroxide thermochemolysis were used to study lignin oxidation, depolymerization, and demethylation of spruce wood treated by biomimetic oxidative systems. Neat Fenton and chelator-mediated Fenton reaction (CMFR) systems as well as cellulosic enzyme treatments were used to mimic the nonenzymatic process involved in wood brown-rot biodegradation. The results suggest that compared with enzymatic processes, Fenton-based treatment more readily opens the structure of the lignocellulosic matrix, freeing cellulose fibrils from the matrix. The results demonstrate that, under the current treatment conditions, Fenton and CMFR treatment cause limited demethoxylation of lignin in the insoluble wood residue. However, analysis of a water-extractable fraction revealed considerable soluble lignin residue structures that had undergone side chain oxidation as well as demethoxylation upon CMFR treatment. This research has implications for our understanding of nonenzymatic degradation of wood and the diffusion of CMFR agents in the wood cell wall during fungal degradation processes.

  17. To tag or not to tag: a comparative evaluation of immunoaffinity-labeling and tandem mass spectrometry for the identification and localization of posttranslational protein carbonylation by 4-hydroxy-2-nonenal, an end-product of lipid peroxidation.

    PubMed

    Guo, Jia; Prokai, Laszlo

    2011-10-19

    Posttranslational carbonylation of proteins by the covalent attachment of the lipid peroxidation product 4-hydroxy-2-nonenal (HNE) is a biomarker of oxidative stress. Tandem mass spectrometry (MS/MS) has become an essential tool for characterization of this modification. Chemical tagging methods have been used to facilitate the immunoaffinity-based enrichment or even quantification of HNE-modified peptides and proteins. With MS/MS spectra of the untagged modified peptides considered as references, a comparative evaluation is presented focusing on the impact of affinity-tagging with four carbonyl-specific reagents (2,4-dinitrophenyl hydrazine, biotin hydrazide, biotinamidohexanoic acid hydrazide and N'-aminooxymethylcarbonyl-hydrazino D-biotin) on collision-induced dissociation of the tagged HNE-carbonylated peptides. Our study has shown that chemical labeling may not be carried out successfully for all the peptides and with all the reagents. The attachment of a tag usually cannot circumvent the occurrence of strong neutral losses observed with untagged species and, in addition, fragmentation of the introduced tag may also happen. Chemical tagging of certain peptides may, nevertheless, afford more sequence ions upon MS/MS than the untagged carbonylated peptide, especially when Michael addition of the lipid peroxidation product occurs on cysteine residues. Therefore, tagging may increase the confidence of identifications of HNE-modified peptides by database searches.

  18. Stable isotope dimethyl labelling for quantitative proteomics and beyond.

    PubMed

    Hsu, Jue-Liang; Chen, Shu-Hui

    2016-10-28

    Stable-isotope reductive dimethylation, a cost-effective, simple, robust, reliable and easy-to- multiplex labelling method, is widely applied to quantitative proteomics using liquid chromatography-mass spectrometry. This review focuses on biological applications of stable-isotope dimethyl labelling for a large-scale comparative analysis of protein expression and post-translational modifications based on its unique properties of the labelling chemistry. Some other applications of the labelling method for sample preparation and mass spectrometry-based protein identification and characterization are also summarized.This article is part of the themed issue 'Quantitative mass spectrometry'. PMID:27644970

  19. Evidence of the chemical reaction of (18)O-labelled nitrite with CO2 in aqueous buffer of neutral pH and the formation of (18)OCO by isotope ratio mass spectrometry.

    PubMed

    Tsikas, Dimitrios; Böhmer, Anke; Gros, Gerolf; Endeward, Volker

    2016-05-01

    Inorganic nitrite (NO2(-), ON-O(-) ←→ (-)O-NO) is the autoxidation product of nitric oxide (NO). Nitrite can also be formed from inorganic nitrate (ONO2(-)), the major oxidation product of NO in erythrocytes, by the catalytic action of bacterial nitrate reductase in gut and oral microflora. Nitrite can be reduced to NO by certain cellular proteins and enzymes, as well as in the gastric juice under acidic conditions. Hemoglobin, xanthine oxidoreductase and carbonic anhydrase (CA) have been reported to convert nitrite to NO. Renal CA isoforms are involved in the reabsorption of nitrite and may, therefore, play an important role in NO homeostasis. Yet, the mechanisms underlying the action of CA on nitrite are incompletely understood. The nitrate/nitrite system is regarded as a reservoir of NO. We have recently shown that nitrite reacts chemically with carbon dioxide (CO2), the regular substrate of CA. The present communication reports a stable isotope ratio mass spectrometry (IRMS) study on the reaction of NO2(-) and CO2 performed in 50 mM HEPES buffer of pH 7.4 at 37 °C. By using (18)O-labelled nitrite ((18)ON-O(-)/(-18)O-NO) and CO2 we observed formation of (18)O-labelled CO2. This finding is an unequivocal evidence of the chemical reaction of (18)ON-O(-)/(-18)O-NO with CO2. The reaction is rapid and involves nucleophilic attack of the negatively charged nitrite via one of its oxygen atoms on the partially positively charged CO2 molecule to form the putative intermediate (18)ON-O-CO2(-)/(-)O2C-(18)O-NO. The by far largest fraction of this intermediate decomposes back to (18)ON-O(-)/(-18)O-NO and CO2. A very small fraction of the intermediate, however, rearranges and finally decomposes to form (18)OCO and nitrite. This reaction is slower in the presence of an isolated erythrocytic CA isoform II. In summary, NO2(-), CO2 and CA are ubiquitous. The chemical reaction of NO2(-) with CO2 and its modulation by CA isoforms may play important roles in the transport of

  20. Mental Labels and Tattoos

    ERIC Educational Resources Information Center

    Hyatt, I. Ralph

    1977-01-01

    Discusses the ease with which mental labels become imprinted in our system, six basic axioms for maintaining negative mental tattoos, and psychological processes for eliminating mental tattoos and labels. (RK)

  1. Rapid, sensitive and simultaneous determination of fluorescence-labeled designated substances controlled by the Pharmaceutical Affairs Law in Japan by ultra-performance liquid chromatography coupled with electrospray-ionization time-of-flight mass spectrometry.

    PubMed

    Min, Jun Zhe; Hatanaka, Suguru; Toyo'oka, Toshimasa; Inagaki, Shinsuke; Kikura-Hanajiri, Ruri; Goda, Yukihiro

    2009-11-01

    A simultaneous determination method based on ultra-performance liquid chromatography (UPLC) with fluorescence (FL) detection and electrospray-ionization time-of-flight mass spectrometry (ESI-TOF-MS) was developed for 16 "designated substances" (Shitei-Yakubutsu) controlled by the Pharmaceutical Affairs Law in Japan. These substances were first labeled with 4-(N,N-dimethylaminosulfonyl)-7-fluoro-2,1,3-benzoxadiazole at 60 degrees C for 2 h in 0.1 M borax (pH 9.3). The resulting fluorophores were well separated by reversed-phase chromatography using an Acquity UPLC BEH C(18) column (1.7 microm, 100 mm x 2.1 mm i.d.) by isocratic elution with a mixture of water and acetonitrile-methanol (20:80) containing 0.1% formic acid. The separated derivatives were sensitively detected by both FL and TOF-MS. However, the determination of several designated substances by FL detection showed interference from endogenous substances in biological samples. Therefore, the determination in real samples was carried out by a combination of UPLC separation and ESI-TOF-MS detection. The structures of the designated substances were identified from the protonated-molecular ions [M+H](+) obtained from the TOF-MS measurement. The calibration curves obtained from the peak area ratios of the internal standard (I.S.), i.e., 3-phenyl-1-propylamine, and the designated substances versus the injection amounts showed good linearity. The limits of detection (S/N = 3) and the limits of quantification (S/N = 10) in 0.1 mL of human plasma and urine for the present method were 0.30-150 pmol and 1.0-500 pmol, respectively. Good accuracy and precision (according to intraday and interday assays) were also obtained with the present procedure. This method was applied to analyses of human plasma, urine and real products.

  2. Trypsin-catalyzed oxygen-18 labeling for quantitative proteomics

    SciTech Connect

    Qian, Weijun; Petritis, Brianne O.; Nicora, Carrie D.; Smith, Richard D.

    2011-07-01

    Stable isotope labeling based on relative peptide/protein abundance measurements is commonly applied for quantitative proteomics. Recently, trypsin-catalyzed oxygen-18 labeling has grown in popularity due to its simplicity, cost-effectiveness, and its ability to universally label peptides with high sample recovery. In (18)O labeling, both C-terminal carboxyl group atoms of tryptic peptides can be enzymatically exchanged with (18)O, thus providing the labeled peptide with a 4 Da mass shift from the (16)O-labeled sample. Peptide (18)O labeling is ideally suited for generating a labeled "universal" reference sample used for obtaining accurate and reproducible quantitative measurements across large number of samples in quantitative discovery proteomics.

  3. Bar Code Labels

    NASA Technical Reports Server (NTRS)

    1988-01-01

    American Bar Codes, Inc. developed special bar code labels for inventory control of space shuttle parts and other space system components. ABC labels are made in a company-developed anodizing aluminum process and consecutively marketed with bar code symbology and human readable numbers. They offer extreme abrasion resistance and indefinite resistance to ultraviolet radiation, capable of withstanding 700 degree temperatures without deterioration and up to 1400 degrees with special designs. They offer high resistance to salt spray, cleaning fluids and mild acids. ABC is now producing these bar code labels commercially or industrial customers who also need labels to resist harsh environments.

  4. Discovery and Verification of Head-and-neck Cancer Biomarkers by Differential Protein Expression Analysis Using iTRAQ Labeling, Multidimensional Liquid Chromatography, and Tandem Mass Spectrometry*S⃞

    PubMed Central

    Ralhan, Ranju; DeSouza, Leroi V.; Matta, Ajay; Chandra Tripathi, Satyendra; Ghanny, Shaun; Datta Gupta, Siddartha; Bahadur, Sudhir; Siu, K. W. Michael

    2008-01-01

    Multidimensional LC-MS/MS has been used for the analysis of biological samples labeled with isobaric mass tags for relative and absolute quantitation (iTRAQ) to identify proteins that are differentially expressed in human head-and-neck squamous cell carcinomas (HNSCCs) in relation to non-cancerous head-and-neck tissues (controls) for cancer biomarker discovery. Fifteen individual samples (cancer and non-cancerous tissues) were compared against a pooled non-cancerous control (prepared by pooling equal amounts of proteins from six non-cancerous tissues) in five sets by on-line and off-line separation. We identified 811 non-redundant proteins in HNSCCs, including structural proteins, signaling components, enzymes, receptors, transcription factors, and chaperones. A panel of proteins showing consistent differential expression in HNSCC relative to the non-cancerous controls was discovered. Some of the proteins include stratifin (14-3-3σ); YWHAZ (14-3-3ζ); three calcium-binding proteins of the S100 family, S100-A2, S100-A7 (psoriasin), and S100-A11 (calgizarrin); prothymosin α (PTHA); l-lactate dehydrogenase A chain; glutathione S-transferase Pi; APC-binding protein EB1; and fascin. Peroxiredoxin2, carbonic anhydrase I, flavin reductase, histone H3, and polybromo-1D (BAF180) were underexpressed in HNSCCs. A panel of the three best performing biomarkers, YWHAZ, stratifin, and S100-A7, achieved a sensitivity of 0.92 and a specificity of 0.91 in discriminating cancerous from non-cancerous head-and-neck tissues. Verification of differential expression of YWHAZ, stratifin, and S100-A7 proteins in clinical samples of HNSCCs and paired and non-paired non-cancerous tissues by immunohistochemistry, immunoblotting, and RT-PCR confirmed their overexpression in head-and-neck cancer. Verification of YWHAZ, stratifin, and S100-A7 in an independent set of HNSCCs achieved a sensitivity of 0.92 and a specificity of 0.87 in discriminating cancerous from non-cancerous head

  5. Spanish labeling guide.

    PubMed

    Juliá, A M; García, S V; Breckinridge, M F

    1983-01-01

    A systematic reference of English-Spanish prescription label translations is presented. The purpose of the reference list (which is the most comprehensive published to date) is to enable a pharmacist to write precise, accurate label directions in Spanish for any patient who cannot read English.

  6. Spin-labeled polyribonucleotides.

    PubMed Central

    Petrov, A I; Sukhorukov, B I

    1980-01-01

    Poly (U), poly (C) and poly (A) were spin labeled with N-(2,2,5,5-tetramethyl-3-carbonylpyrroline-1-oxyl)-imidazole. This spin label interacts selectively with 2' OH ribose groups of polynucleotides and does not modify the nucleic acid bases. The extent of spin labeling is not dependent upon the nature of the base and is entirely determined by rigidity of the secondary structure of the polynucleotide. The extent of modification for poly (U), poly (C) and poly (A) was 4.2, 1.7 and 1.5 per cent, respectively, the secondary structure of the polynucleotides being practically unchanged. Some physico-chemical properties of the spin-labeled polynucleotides were investigated by ESR spectroscopy. Rotational correlation times of the spin label and activation energy of its motion were calculated. PMID:6253911

  7. Label fusion strategy selection.

    PubMed

    Robitaille, Nicolas; Duchesne, Simon

    2012-01-01

    Label fusion is used in medical image segmentation to combine several different labels of the same entity into a single discrete label, potentially more accurate, with respect to the exact, sought segmentation, than the best input element. Using simulated data, we compared three existing label fusion techniques-STAPLE, Voting, and Shape-Based Averaging (SBA)-and observed that none could be considered superior depending on the dissimilarity between the input elements. We thus developed an empirical, hybrid technique called SVS, which selects the most appropriate technique to apply based on this dissimilarity. We evaluated the label fusion strategies on two- and three-dimensional simulated data and showed that SVS is superior to any of the three existing methods examined. On real data, we used SVS to perform fusions of 10 segmentations of the hippocampus and amygdala in 78 subjects from the ICBM dataset. SVS selected SBA in almost all cases, which was the most appropriate method overall. PMID:22518113

  8. How to Read Drug Labels

    MedlinePlus

    ... and alternative medicine Healthy Aging How to read drug labels Printer-friendly version How to Read Drug ... read drug labels How to read a prescription drug label View a text version of this picture. ...

  9. Capacitive label reader

    DOEpatents

    Arlowe, H.D.

    1983-07-15

    A capacitive label reader includes an outer ring transmitting portion, an inner ring transmitting portion, and a plurality of insulated receiving portions. A label is the mirror-image of the reader except that identifying portions corresponding to the receiving portions are insulated from only one of two coupling elements. Positive and negative pulses applied, respectively, to the two transmitting rings biased a CMOS shift register positively to either a 1 or 0 condition. The output of the CMOS may be read as an indication of the label.

  10. High resolution quantitative proteomics of HeLa cells protein species using stable isotope labeling with amino acids in cell culture(SILAC), two-dimensional gel electrophoresis(2DE) and nano-liquid chromatograpohy coupled to an LTQ-OrbitrapMass spectrometer.

    PubMed

    Thiede, Bernd; Koehler, Christian J; Strozynski, Margarita; Treumann, Achim; Stein, Robert; Zimny-Arndt, Ursula; Schmid, Monika; Jungblut, Peter R

    2013-02-01

    The proteomics field has shifted over recent years from two-dimensional gel electrophoresis (2-DE)-based approaches to SDS-PAGE or gel-free workflows because of the tremendous developments in isotopic labeling techniques, nano-liquid chromatography, and high-resolution mass spectrometry. However, 2-DE still offers the highest resolution in protein separation. Therefore, we combined stable isotope labeling with amino acids in cell culture of controls and apoptotic HeLa cells with 2-DE and the subsequent analysis of tryptic peptides via nano-liquid chromatography coupled to an LTQ-Orbitrap mass spectrometer to obtain quantitative data using the methods with the highest resolving power on all levels of the proteomics workflow. More than 1,200 proteins with more than 2,700 protein species were identified and quantified from 816 Coomassie Brilliant Blue G-250 stained 2-DE spots. About half of the proteins were identified and quantified only in single 2-DE spots. The majority of spots revealed one to five proteins; however, in one 2-DE spot, up to 23 proteins were identified. Only half of the 2-DE spots represented a dominant protein with more than 90% of the whole protein amount. Consequently, quantification based on staining intensities in 2-DE gels would in approximately half of the spots be imprecise, and minor components could not be quantified. These problems are circumvented by quantification using stable isotope labeling with amino acids in cell culture. Despite challenges, as shown in detail for lamin A/C and vimentin, the quantitative changes of protein species can be detected. The combination of 2-DE with high-resolution nano-liquid chromatography-mass spectrometry allowed us to identify proteomic changes in apoptotic cells that would be unobservable using any of the other previously employed proteomic workflows.

  11. High Resolution Quantitative Proteomics of HeLa Cells Protein Species Using Stable Isotope Labeling with Amino Acids in Cell Culture(SILAC), Two-Dimensional Gel Electrophoresis(2DE) and Nano-Liquid Chromatograpohy Coupled to an LTQ-OrbitrapMass Spectrometer*

    PubMed Central

    Thiede, Bernd; Koehler, Christian J.; Strozynski, Margarita; Treumann, Achim; Stein, Robert; Zimny-Arndt, Ursula; Schmid, Monika; Jungblut, Peter R.

    2013-01-01

    The proteomics field has shifted over recent years from two-dimensional gel electrophoresis (2-DE)-based approaches to SDS-PAGE or gel-free workflows because of the tremendous developments in isotopic labeling techniques, nano-liquid chromatography, and high-resolution mass spectrometry. However, 2-DE still offers the highest resolution in protein separation. Therefore, we combined stable isotope labeling with amino acids in cell culture of controls and apoptotic HeLa cells with 2-DE and the subsequent analysis of tryptic peptides via nano-liquid chromatography coupled to an LTQ-Orbitrap mass spectrometer to obtain quantitative data using the methods with the highest resolving power on all levels of the proteomics workflow. More than 1,200 proteins with more than 2,700 protein species were identified and quantified from 816 Coomassie Brilliant Blue G-250 stained 2-DE spots. About half of the proteins were identified and quantified only in single 2-DE spots. The majority of spots revealed one to five proteins; however, in one 2-DE spot, up to 23 proteins were identified. Only half of the 2-DE spots represented a dominant protein with more than 90% of the whole protein amount. Consequently, quantification based on staining intensities in 2-DE gels would in approximately half of the spots be imprecise, and minor components could not be quantified. These problems are circumvented by quantification using stable isotope labeling with amino acids in cell culture. Despite challenges, as shown in detail for lamin A/C and vimentin, the quantitative changes of protein species can be detected. The combination of 2-DE with high-resolution nano-liquid chromatography-mass spectrometry allowed us to identify proteomic changes in apoptotic cells that would be unobservable using any of the other previously employed proteomic workflows. PMID:23033477

  12. Behind the Label "Alcoholic."

    ERIC Educational Resources Information Center

    Wright, Deborah M.

    1989-01-01

    Relates individual's personal story of her childhood influenced by her parent's alcoholism, her own alcoholism as a young adult, and her experiences with counseling. Asks others not to reject her because of the label "alcoholic." (ABL)

  13. Like your labels?

    PubMed

    Field, Michele

    2010-01-01

    The descriptive “conventions” used on food labels are always evolving. Today, however, the changes are so complicated (partly driven by legislation requiring disclosures about environmental impacts, health issues, and geographical provenance) that these labels more often baffle buyers than enlighten them. In a light-handed manner, the article points to how sometimes reading label language can be like deciphering runes—and how if we are familiar with the technical terms, we can find a literal meaning, but still not see the implications. The article could be ten times longer because food labels vary according to cultures—but all food-exporting cultures now take advantage of our short attention-span when faced with these texts. The question is whether less is more—and if so, in this contest for our attention, what “contestant” is voted off. PMID:21539053

  14. Analysis of proteome dynamics in mice by isotopic labeling.

    PubMed

    Price, John C; Ghaemmaghami, Sina

    2014-01-01

    Recent advances in mass spectrometry and in vivo isotopic labeling have enabled proteome-wide analyses of protein turnover in complex organisms. Here, we describe a protocol for analyzing protein turnover rates in mouse tissues by comprehensive (15)N labeling. The procedure involves the complete isotopic labeling of blue green algae (Spirulina platensis) with (15)N and utilizing it as a source of dietary nitrogen for mice. We outline a detailed protocol for in-house production of (15)N-labeled algae, labeling of mice, and analysis of isotope incorporation kinetics by mass spectrometry. The methodology can be adapted to analyze proteome dynamics in most murine tissues and may be particularly useful in the analysis of proteostatic disruptions in mouse models of disease. PMID:24791984

  15. Routing and Label Space Reduction in Label Switching Networks

    NASA Astrophysics Data System (ADS)

    Solano, Fernando; Caro, Luis Fernando; Stidsen, Thomas; Papadimitriou, Dimitri

    This chapter is devoted to the analysis and modeling of some problems related to the optimal usage of the label space in label switching networks. Label space problems concerning three different technologies and architectures - namely Multi-protocol Label Switching (MPLS), Ethernet VLAN-Label Switching (ELS) and All-Optical Label Switching (AOLS) - are discussed in this chapter. Each of these cases yields to different constraints of the general label space reduction problem. We propose a generic optimization model and, then, we describe some adaptations aiming at modeling each particular case. Simulation results are briefly discussed at the end of this chapter.

  16. Robust labeling methods for copy protection of images

    NASA Astrophysics Data System (ADS)

    Langelaar, Gerhard C.; van der Lubbe, Jan C. A.; Lagendijk, Reginald L.

    1997-01-01

    In the European project SMASH a mass multimedia storage device for home usage is being developed. The success of such a storage system depends not only on technical advances, but also on the existence of an adequate copy protection method. Copy protection for visual data requires fast and robust labeling techniques. In this paper, two new labeling techniques are proposed. The first method extends an existing spatial labeling technique. This technique divides the image into blocks and searches an optimal label- embedding level for each block instead of using a fixed embedding-level for the complete image. The embedding-level for each block is dependent on a lower quality JPEG compressed version of the labeled block. The second method removes high frequency DCT-coefficients in some areas to embed a label. A JPEG quality factor and the local image structure determine how many coefficients are discarded during the labeling process. Using both methods a perceptually invisible label of a few hundred bits was embedded in a set of true color images. The label added by the spatial method is very robust against JPEG compression. However, this method is not suitable for real-time applications. Although the second DCT-based method is slightly less resistant to JPEG compression, it is more resistant to line-shifting and cropping than the first one and is suitable for real-time labeling.

  17. Off-Label Drug Use

    MedlinePlus

    ... Your Local Offices Close + - Text Size Off-label Drug Use What is off-label drug use? In the United States new drugs are ... unapproved use of a drug. Is off-label drug use legal? The off-label use of FDA- ...

  18. A Straightforward and Highly Efficient Precipitation/On-pellet Digestion Procedure Coupled to a Long Gradient Nano-LC Separation and Orbitrap Mass Spectrometry for Label-free Expression Profiling of the Swine Heart Mitochondrial Proteome

    PubMed Central

    Duan, Xiaotao; Young, Rebecca; Straubinger, Robert M.; Page, Brian J.; Cao, Jin; Wang, Hao; Yu, Haoying; Canty, John M.; Qu, Jun

    2009-01-01

    For label-free expression profiling of tissue proteomes, efficient protein extraction, thorough and quantitative sample cleanup and digestion procedures, as well as sufficient and reproducible chromatographic separation, are highly desirable but remain challenging. However, optimal methodology has remained elusive, especially for proteomes that are rich in membrane proteins, such as the mitochondria. Here we describe a straightforward and reproducible sample preparation procedure, coupled with a highly selective and sensitive nano-LC/Orbitrap analysis, which enables reliable and comprehensive expression profiling of tissue mitochondria. The mitochondrial proteome of swine heart was selected as a test system. Efficient protein extraction was accomplished using a strong buffer containing both ionic and non-ionic detergents. Overnight precipitation was used for cleanup of the extract, and the sample was subjected to an optimized 2-step, on-pellet digestion approach. In the first step, the protein pellet was dissolved via a 4 h tryptic digestion under vigorous agitation, which nano-LC/LTQ/ETD showed to produce large and incompletely cleaved tryptic peptides. The mixture was then reduced, alkylated, and digested into its full complement of tryptic peptides with additional trypsin. This solvent precipitation/on-pellet digestion procedure achieved significantly higher and more reproducible peptide recovery of the mitochondrial preparation, than observed using a prevalent alternative procedure for label-free expression profiling, SDS-PAGE/in-gel digestion (87% vs. 54%). Furthermore, uneven peptide losses were lower than observed with SDS-PAGE/in-gel digestion. The resulting peptides were sufficiently resolved by a 5 h gradient using a nano-LC configuration that features a low-void-volume, high chromatographic reproducibility, and an LTQ/Orbitrap analyzer for protein identification and quantification. The developed method was employed for label-free comparison of the

  19. Labeling lake water with tritium

    USGS Publications Warehouse

    Frederick, B.J.

    1963-01-01

    A method of packaging tritiated water in a manner that facilitates safe handling in environmental labeling operations, and procedures followed in labeling a large body of water with a small volume of tritiated water are described. ?? 1963.

  20. 99m tc labeled liposomes

    SciTech Connect

    Phillips, W.T.; Klipper, R.W.; Timmons, J.H.; Rudolph, A.S.

    1992-10-27

    This patent describes a method of preparing stable gamma-emitting radionuclide-labeled alkyleneamine oxime, the incubating being for a period of time sufficient to form labeled liposome-encapsulated protein.

  1. Decode the Sodium Label Lingo

    MedlinePlus

    ... For Preschooler For Gradeschooler For Teen Decode the Sodium Label Lingo Published January 24, 2013 Print Email Reading food labels can help you slash sodium. Here's how to decipher them. "Sodium free" or " ...

  2. Learning with imperfectly labeled patterns

    NASA Technical Reports Server (NTRS)

    Chittineni, C. B.

    1979-01-01

    The problem of learning in pattern recognition using imperfectly labeled patterns is considered. The performance of the Bayes and nearest neighbor classifiers with imperfect labels is discussed using a probabilistic model for the mislabeling of the training patterns. Schemes for training the classifier using both parametric and non parametric techniques are presented. Methods for the correction of imperfect labels were developed. To gain an understanding of the learning process, expressions are derived for success probability as a function of training time for a one dimensional increment error correction classifier with imperfect labels. Feature selection with imperfectly labeled patterns is described.

  3. Glycan labeling strategies and their use in identification and quantification

    PubMed Central

    Ruhaak, L. R.; Zauner, G.; Huhn, C.; Bruggink, C.; Deelder, A. M.

    2010-01-01

    Most methods for the analysis of oligosaccharides from biological sources require a glycan derivatization step: glycans may be derivatized to introduce a chromophore or fluorophore, facilitating detection after chromatographic or electrophoretic separation. Derivatization can also be applied to link charged or hydrophobic groups at the reducing end to enhance glycan separation and mass-spectrometric detection. Moreover, derivatization steps such as permethylation aim at stabilizing sialic acid residues, enhancing mass-spectrometric sensitivity, and supporting detailed structural characterization by (tandem) mass spectrometry. Finally, many glycan labels serve as a linker for oligosaccharide attachment to surfaces or carrier proteins, thereby allowing interaction studies with carbohydrate-binding proteins. In this review, various aspects of glycan labeling, separation, and detection strategies are discussed. Figure MALDI-FTICR-MS of 2AA-labeled total plasma N-glycans PMID:20225063

  4. Environmental Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Lebedev, Albert T.

    2013-06-01

    Environmental mass spectrometry is an important branch of science because it provides many of the data that underlie policy decisions that can directly influence the health of people and ecosystems. Environmental mass spectrometry is currently undergoing rapid development. Among the most relevant directions are a significant broadening of the lists of formally targeted compounds; a parallel interest in nontarget chemicals; an increase in the reliability of analyses involving accurate mass measurements, tandem mass spectrometry, and isotopically labeled standards; and a shift toward faster high-throughput analysis, with minimal sample preparation, involving various approaches, including ambient ionization techniques and miniature instruments. A real revolution in analytical chemistry could be triggered with the appearance of robust, simple, and sensitive portable mass spectrometers that can utilize ambient ionization techniques. If the cost of such instruments is reduced to a reasonable level, mass spectrometers could become valuable household devices.

  5. Label and Label-Free Detection Techniques for Protein Microarrays

    PubMed Central

    Syahir, Amir; Usui, Kenji; Tomizaki, Kin-ya; Kajikawa, Kotaro; Mihara, Hisakazu

    2015-01-01

    Protein microarray technology has gone through numerous innovative developments in recent decades. In this review, we focus on the development of protein detection methods embedded in the technology. Early microarrays utilized useful chromophores and versatile biochemical techniques dominated by high-throughput illumination. Recently, the realization of label-free techniques has been greatly advanced by the combination of knowledge in material sciences, computational design and nanofabrication. These rapidly advancing techniques aim to provide data without the intervention of label molecules. Here, we present a brief overview of this remarkable innovation from the perspectives of label and label-free techniques in transducing nano-biological events.

  6. Production of isotopically-labeled standards from a uniformly labeled precursor for quantitative volatile metabolomic studies

    PubMed Central

    Gómez-Cortés, Pilar; Brenna, J. Thomas; Sacks, Gavin L.

    2012-01-01

    Optimal accuracy and precision in small molecule profiling by mass spectrometry generally requires isotopically labeled standards chemically representative of all compounds of interest. However, preparation of mixed standards from commercially available pure compounds is often prohibitively expensive and time consuming, and many labeled compounds are not available in pure form. We used a single prototype uniformly labeled [U-13C]-compound to generate [U-13C]-volatile standards for use in subsequent experimental profiling studies. [U-13C]-α-linolenic acid (C18:3n-3, ALA) was thermally oxidized to produce labeled lipid degradation volatiles which were subsequently characterized qualitatively and quantitatively. Twenty-five [U-13C]-labeled volatiles were identified by headspace solid-phase microextraction-gas chromatography-time of flight-mass spectrometry (HS-SPME-GC-TOF-MS) by comparison of spectra with unlabeled volatiles. Using 250 μL starting sample, labeled volatiles were quantified by a reverse isotope dilution procedure. Using the [U-13C]-labeled standards, limits of detection comparable to or better than previous HS-SPME reports were achieved, 0.010–1.04 ng/g. The performance of the [U-13C]-volatile standards was evaluated using a commodity soybean oil (CSO) oxidized at 60°C from 0 to 15 d. Relative responses of n-decane, an unlabeled internal standard otherwise absent from the mixture, and [U-13C]-oxidation products changed by up to 8-fold as the CSO matrix was oxidized, demonstrating that reliance on a single standard in volatile profiling studies yields inaccurate results due to changing matrix effects. The [U-13C]-standard mixture was used to quantify 25 volatiles in oxidized CSO and low-ALA soybean oil with an average relative standard deviation of 8.5%. Extension of this approach to other labeled substrates, e.g., [U-13C]-sugars and amino acids, for profiling studies should be feasible and can dramatically improve quantitative results compared to

  7. Comments on "innovative method for carbon dioxide determination in human postmortem cardiac gas samples using headspace-gas chromatography-mass spectrometry and stable labeled isotope as internal standard" by Varlet et al.

    PubMed

    Saffaj, T; Ihssane, B

    2014-01-31

    Varlet et al. recently proposed a headspace-gas chromatography-mass spectrometry (HS-GC-MS) method applicable for the routine determination of CO2 in gaseous biologic matrices. This developed bioanalytical method was fully validated according to the SFSTP 1997 guidelines using the accuracy profile as a graphical decision-making tool.In this letter, we discuss the validity of HS-GC-MS method based on the newest SFSTP guideline. In particular, we demonstrate by the estimation of the β-expectation tolerance interval that the error total exceeds the acceptance limits (30%) for the second concentration level (0.5μmol mL(-1) vial HS). Furthermore, we show through the risk profile that the probability to have future results inside the ±30% acceptance limits is smaller than 95%.

  8. Appliance energy labeling takes effect

    SciTech Connect

    Not Available

    1980-06-01

    Consumers buying household appliances will be helped by energy-efficiency labels and minimum efficiency standards required for refrigerators and refrigerator/freezers, freezers, dishwashers, water heaters, clothes washers, room air conditioners, and furnaces. The ENERGYGUIDE labels must be displayed in the store and in catalogs. Two voluntary efficiency programs were combined in the Energy Policy and Conservation Act (EPCA) requiring labels by 1980. Shoppers may compare the efficiencies of appliances and compute the actual cost differential over the lifetime of the equipment. Manufacturers have responded with more-efficient models, but the impact of efficient appliances on energy consumption will be small. A sample label with the required information is illustrated. (DCK)

  9. Labeled Cocaine Analogs

    SciTech Connect

    Goodman, Mark M.; Shi, Bing Zhi; Keil, Robert N.

    1999-03-30

    Novel methods for positron emission tomography or single photon emission spectroscopy using tracer compounds having the structure: ##STR1## where X in .beta. configuration is phenyl, naphthyl; 2,3 or 4-iodophenyl; 2,3 or 4-(trimethylsilyl)phenyl; 3,4,5 or 6-iodonaphthyl; 3,4,5 or 6-(trimethylsilyl)naphthyl; 2,3 or 4-(trialkylstannyl)phenyl; or 3,4,5 or 6-(trialkylstannyl)napthyl Y in .beta. configuration is 2-fluoroethoxy, 3-fluoropropoxy, 4-fluorobutoxy, 2-fluorocyclopropoxy, 2 or 3-fluorocyclobutoxy, R,S 1'-fluoroisopropoxy, R 1'-fluoroisopropoxy, S 1'-fluoroisopropoxy, 1',3'-difluoroisopropoxy, R,S 1'-fluoroisobutoxy, R 1'-fluoroisobutoxy, S 1'-fluoroisobutoxy, R,S 4'-fluoroisobutoxy, R 4'-fluoroisobutoxy, S 4'-fluoroisobutoxy, or 1',1'-di(fluoromethyl)isobutoxy, The compounds bind dopamine transporter protein and can be labeled with .sup.18 F or .sup.123 I for imaging.

  10. 78 FR 66826 - Prior Label Approval System: Generic Label Approval

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-11-07

    ... the Agency (76 FR 75809). FSIS also proposed to combine the regulations that provide for the approval... preamble (76 FR 75814), FSIS wrote: . . . statements on labels that are defined in FSIS's regulations or... ``Product Labeling: Definition of the Term ``Natural'' and related materials (71 FR 70503, Dec. 5, 2006)...

  11. Laser labeling, a safe technology to label produce

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Labeling of the produce has gained marked attention in recent years. Laser labeling technology involves the etching of required information on the surface using a low energy CO2 laser beam. The etching forms alphanumerical characters by pinhole dot matrix depressions. These openings can lead to wat...

  12. Laser labeling, a safe technology to label produce

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Laser labeling of fruits and vegetables is an alternative means to label produce. Low energy CO2 laser beams etch the surface showing the contrasting underlying layer. These etched surfaces can promote water loss and potentially allow for entry of decay organisms. The long-term effects of laser labe...

  13. 76 FR 75809 - Prior Label Approval System: Generic Label Approval

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-12-05

    ... poultry products will take effect January 1, 2012 (75 FR 82148, Dec. 29, 2010). These mandatory features... limited types of labels (e.g., labels for raw, single ingredient meat and poultry products) (48 FR 11410... Agency. On March 25, 1992, FSIS published an Advance Notice of Proposed Rulemaking (ANPRM) (57 FR...

  14. Robust method for investigating nitrogen metabolism of 15N labeled amino acids using AccQ•Tag ultra performance liquid chromatography-photodiode array-electrospray ionization-mass spectrometry: application to a parasitic plant-plant interaction.

    PubMed

    Gaudin, Zachary; Cerveau, Delphine; Marnet, Nathalie; Bouchereau, Alain; Delavault, Philippe; Simier, Philippe; Pouvreau, Jean-Bernard

    2014-01-21

    An AccQ•Tag ultra performance liquid chromatography-photodiode array-electrospray ionization-mass spectrometry (AccQ•Tag-UPLC-PDA-ESI-MS) method is presented here for the fast, robust, and sensitive quantification of (15)N isotopologue enrichment of amino acids in biological samples, as for example in the special biotic interaction between the cultivated specie Brassica napus (rapeseed) and the parasitic weed Phelipanche ramosa (broomrape). This method was developed and validated using amino acid standard solutions containing (15)N amino acid isotopologues and/or biological unlabeled extracts. Apparatus optimization, limits of detection and quantification, quantification reproducibility, and calculation method of (15)N isotopologue enrichment are presented. Using this method, we could demonstrate that young parasite tubercles assimilate inorganic nitrogen as (15)N-ammonium when supplied directly through batch incubation but not when supplied by translocation from host root phloem, contrary to (15)N2-glutamine. (15)N2-glutamine mobility from host roots to parasite tubercles followed by its low metabolism in tubercles suggests that the host-derived glutamine acts as an important nitrogen containing storage compound in the young tubercle of Phelipanche ramosa. PMID:24359440

  15. Nutrition Marketing on Food Labels

    ERIC Educational Resources Information Center

    Colby, Sarah E.; Johnson, LuAnn; Scheett, Angela; Hoverson, Bonita

    2010-01-01

    Objective: This research sought to determine how often nutrition marketing is used on labels of foods that are high in saturated fat, sodium, and/or sugar. Design and Setting: All items packaged with food labels (N = 56,900) in all 6 grocery stores in Grand Forks, ND were surveyed. Main Outcome Measure(s): Marketing strategy, nutrient label…

  16. Meat and Poultry Labeling Terms

    MedlinePlus

    ... Food Standards and Labels: The Facts Labeling and Marketing Information [ Top of Page ] OVEN PREPARED: Product is fully cooked and ready to eat. [ Top of Page ] YOUNG TURKEY: Turkeys of either sex that are less than 8 months of age according to present regulations. [ Top of Page ] Last ...

  17. The Influence of Literacy Labeling (Literacy Learning Outside the Classroom).

    ERIC Educational Resources Information Center

    Hayden, Ruth, Ed.

    1996-01-01

    Outlines perspectives on literacy evident in the mass media: the business perspective, the deficiency perspective, and the combat perspective. Reflects on other literacy metaphors, and argues that, as knowledgeable participants in the public debate, teachers should use literacy labels that reflect more authentic and legitimate viewpoints. (SR)

  18. 21 CFR 201.72 - Potassium labeling.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 4 2014-04-01 2014-04-01 false Potassium labeling. 201.72 Section 201.72 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.72 Potassium labeling. (a) The labeling of over-the-counter (OTC) drug products intended for oral ingestion shall contain the potassium...

  19. 21 CFR 201.72 - Potassium labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 4 2012-04-01 2012-04-01 false Potassium labeling. 201.72 Section 201.72 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.72 Potassium labeling. (a) The labeling of over-the-counter (OTC) drug products intended for oral ingestion shall contain the potassium...

  20. 21 CFR 201.72 - Potassium labeling.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 4 2013-04-01 2013-04-01 false Potassium labeling. 201.72 Section 201.72 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.72 Potassium labeling. (a) The labeling of over-the-counter (OTC) drug products intended for oral ingestion shall contain the potassium...

  1. 21 CFR 610.60 - Container label.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 7 2010-04-01 2010-04-01 false Container label. 610.60 Section 610.60 Food and... GENERAL BIOLOGICAL PRODUCTS STANDARDS Labeling Standards § 610.60 Container label. (a) Full label. The following items shall appear on the label affixed to each container of a product capable of bearing a...

  2. 16 CFR 460.12 - Labels.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 16 Commercial Practices 1 2012-01-01 2012-01-01 false Labels. 460.12 Section 460.12 Commercial Practices FEDERAL TRADE COMMISSION TRADE REGULATION RULES LABELING AND ADVERTISING OF HOME INSULATION § 460.12 Labels. If you are a manufacturer, you must label all packages of your insulation. The labels...

  3. 16 CFR 460.12 - Labels.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 16 Commercial Practices 1 2014-01-01 2014-01-01 false Labels. 460.12 Section 460.12 Commercial Practices FEDERAL TRADE COMMISSION TRADE REGULATION RULES LABELING AND ADVERTISING OF HOME INSULATION § 460.12 Labels. If you are a manufacturer, you must label all packages of your insulation. The labels...

  4. 16 CFR 460.12 - Labels.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 16 Commercial Practices 1 2011-01-01 2011-01-01 false Labels. 460.12 Section 460.12 Commercial Practices FEDERAL TRADE COMMISSION TRADE REGULATION RULES LABELING AND ADVERTISING OF HOME INSULATION § 460.12 Labels. If you are a manufacturer, you must label all packages of your insulation. The labels...

  5. 16 CFR 460.12 - Labels.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 16 Commercial Practices 1 2010-01-01 2010-01-01 false Labels. 460.12 Section 460.12 Commercial Practices FEDERAL TRADE COMMISSION TRADE REGULATION RULES LABELING AND ADVERTISING OF HOME INSULATION § 460.12 Labels. If you are a manufacturer, you must label all packages of your insulation. The labels...

  6. 16 CFR 460.12 - Labels.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 16 Commercial Practices 1 2013-01-01 2013-01-01 false Labels. 460.12 Section 460.12 Commercial Practices FEDERAL TRADE COMMISSION TRADE REGULATION RULES LABELING AND ADVERTISING OF HOME INSULATION § 460.12 Labels. If you are a manufacturer, you must label all packages of your insulation. The labels...

  7. 21 CFR 201.71 - Magnesium labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 4 2012-04-01 2012-04-01 false Magnesium labeling. 201.71 Section 201.71 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.71 Magnesium labeling. (a) The labeling of over-the-counter (OTC) drug products intended for oral ingestion shall contain the magnesium...

  8. 21 CFR 201.71 - Magnesium labeling.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 4 2013-04-01 2013-04-01 false Magnesium labeling. 201.71 Section 201.71 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.71 Magnesium labeling. (a) The labeling of over-the-counter (OTC) drug products intended for oral ingestion shall contain the magnesium...

  9. 21 CFR 201.71 - Magnesium labeling.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 4 2014-04-01 2014-04-01 false Magnesium labeling. 201.71 Section 201.71 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.71 Magnesium labeling. (a) The labeling of over-the-counter (OTC) drug products intended for oral ingestion shall contain the magnesium...

  10. 9 CFR 317.4 - Labeling approval.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... labeling of such final labeling has been submitted for approval to the Food Labeling Division, Regulatory... Secretary upon request. (b) The Food Labeling Division shall permit submission for approval of only sketch... Food Labeling Division, Regulatory Programs, Food Safety and Inspection Service, U.S. Department...

  11. 21 CFR 201.72 - Potassium labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 4 2010-04-01 2010-04-01 false Potassium labeling. 201.72 Section 201.72 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.72 Potassium labeling. (a) The labeling of over-the-counter (OTC) drug products intended for oral ingestion shall contain the potassium...

  12. 21 CFR 201.64 - Sodium labeling.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... contains sodium bicarbonate, sodium phosphate, or sodium biphosphate as an active ingredient for oral... 21 Food and Drugs 4 2011-04-01 2011-04-01 false Sodium labeling. 201.64 Section 201.64 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.64 Sodium labeling. (a) The labeling...

  13. 21 CFR 201.64 - Sodium labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... contains sodium bicarbonate, sodium phosphate, or sodium biphosphate as an active ingredient for oral... 21 Food and Drugs 4 2012-04-01 2012-04-01 false Sodium labeling. 201.64 Section 201.64 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.64 Sodium labeling. (a) The labeling...

  14. 21 CFR 201.64 - Sodium labeling.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... contains sodium bicarbonate, sodium phosphate, or sodium biphosphate as an active ingredient for oral... 21 Food and Drugs 4 2013-04-01 2013-04-01 false Sodium labeling. 201.64 Section 201.64 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.64 Sodium labeling. (a) The labeling...

  15. 21 CFR 201.64 - Sodium labeling.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... contains sodium bicarbonate, sodium phosphate, or sodium biphosphate as an active ingredient for oral... 21 Food and Drugs 4 2014-04-01 2014-04-01 false Sodium labeling. 201.64 Section 201.64 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.64 Sodium labeling. (a) The labeling...

  16. 21 CFR 201.64 - Sodium labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... contains sodium bicarbonate, sodium phosphate, or sodium biphosphate as an active ingredient for oral... 21 Food and Drugs 4 2010-04-01 2010-04-01 false Sodium labeling. 201.64 Section 201.64 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.64 Sodium labeling. (a) The labeling...

  17. Hydrogen Exchange Mass Spectrometry.

    PubMed

    Mayne, Leland

    2016-01-01

    Hydrogen exchange (HX) methods can reveal much about the structure, energetics, and dynamics of proteins. The addition of mass spectrometry (MS) to an earlier fragmentation-separation HX analysis now extends HX studies to larger proteins at high structural resolution and can provide information not available before. This chapter discusses experimental aspects of HX labeling, especially with respect to the use of MS and the analysis of MS data.

  18. Isotopic Labeling of Red Cabbage Anthocyanins with Atmospheric 13-CO2

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Isotopic labeling of plants provides a unique opportunity for understanding metabolic processes. A significant challenge of isotopic labeling during plant growth is that isotopes must be administered without disrupting plant development and at sufficient levels for mass spectral analysis. We describ...

  19. Measurement of deuterium-labeled phylloquinone in plasma by LC-APCI-MS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Deuterium-labeled vegetables were fed to humans for the measurement of both unlabeled and deuterium-labeled phylloquinone in plasma. We developed a technique to determine the quantities of these compounds using liquid chromatography/mass spectrometry with atmospheric pressure chemical ionization (LC...

  20. Labeled Cocaine Analogs

    SciTech Connect

    Goodman, Mark M.; Shi, Bing Zhi; Keil, Robert N.

    1999-01-26

    Novel compounds having the structure: ##STR1## where X in .beta. configuration is phenyl, naphthyl; 2,3 or 4-iodophenyl; 2,3 or 4-(trimethylsilyl)phenyl; 3,4,5 or 6-iodonaphthyl; 3,4,5 or 6-(trimethylsilyl)naphthyl; 2,3 or 4-(trialkylstannyl)phenyl; or 3,4,5 or 6-(trialkylstannyl)naphthyl Y in .beta. configuration is Y.sub.1 or Y.sub.2, where Y.sub.1 is 2-fluoroethoxy, 3-fluoropropoxy, 4-fluorobutoxy, 2-fluorocyclopropoxy, 2 or 3-fluorocyclobutoxy, R,S 1'-fluoroisopropoxy, R 1'-fluoroisopropoxy, S 1'-fluoroisopropoxy, 1',3'-difluoroisopropoxy, R,S 1'-fluoroisobutoxy, R 1'-fluoroisobutoxy, S 1'-fluoroisobutoxy, R,S 4'-fluoroisobutoxy, R 4'-fluoroisobutoxy, S 4'-fluoroisobutoxy, or 1',1'-di(fluoromethyl)isobutoxy, and Y.sub.2 is 2-methanesulfonyloxy ethoxy, 3-methanesulfonyloxy propoxy, 4-methanesulfonyloxy butoxy, 2-methanesulfonyloxy cyclopropoxy, 2 or 3-methanesulfonyloxy cyclobutoxy, 1'methanesulfonyloxy isopropoxy, 1'-fluoro, 3'-methanesulfonyloxy isopropoxy, 1'-methanesulfonyloxy, 3'-fluoro isopropoxy, 1'-methanesulfonyloxy isobutoxy, or 4'-methanesulfonyloxy isobutoxy bind dopamine transporter protein and can be labeled with .sup.18 F or .sup.123 I for imaging.

  1. Synthesis Of Labeled Metabolites

    DOEpatents

    Martinez, Rodolfo A.; Silks, III, Louis A.; Unkefer, Clifford J.; Atcher, Robert

    2004-03-23

    The present invention is directed to labeled compounds, for example, isotopically enriched mustard gas metabolites including: [1,1',2,2'-.sup.13 C.sub.4 ]ethane, 1,1'-sulfonylbis[2-(methylthio); [1,1',2,2'-.sup.13 C.sub.4 ]ethane, 1-[[2-(methylsulfinyl)ethyl]sulfonyl]-2-(methylthio); [1,1',2,2'-.sup.13 C.sub.4 ]ethane, 1,1'-sulfonylbis[2-(methylsulfinyl)]; and, 2,2'-sulfinylbis([1,2-.sup.13 C.sub.2 ]ethanol of the general formula ##STR1## where Q.sup.1 is selected from the group consisting of sulfide (--S--), sulfone (--S(O)--), sulfoxide (--S(O.sub.2)--) and oxide (--O--), at least one C* is .sup.13 C, X is selected from the group consisting of hydrogen and deuterium, and Z is selected from the group consisting of hydroxide (--OH), and --Q.sup.2 --R where Q.sup.2 is selected from the group consisting of sulfide (--S--), sulfone(--S(O)--), sulfoxide (--S(O.sub.2)--) and oxide (--O--), and R is selected from the group consisting of hydrogen, a C.sub.1 to C.sub.4 lower alkyl, and amino acid moieties, with the proviso that when Z is a hydroxide and Q.sup.1 is a sulfide, then at least one X is deuterium.

  2. 21 CFR 1302.04 - Location and size of symbol on label and labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 9 2012-04-01 2012-04-01 false Location and size of symbol on label and labeling. 1302.04 Section 1302.04 Food and Drugs DRUG ENFORCEMENT ADMINISTRATION, DEPARTMENT OF JUSTICE LABELING... and labeling. The symbol shall be prominently located on the label or the labeling of the...

  3. 21 CFR 1302.04 - Location and size of symbol on label and labeling.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 9 2013-04-01 2013-04-01 false Location and size of symbol on label and labeling. 1302.04 Section 1302.04 Food and Drugs DRUG ENFORCEMENT ADMINISTRATION, DEPARTMENT OF JUSTICE LABELING... and labeling. The symbol shall be prominently located on the label or the labeling of the...

  4. Photoactivatable protein labeling by singlet oxygen mediated reactions.

    PubMed

    To, Tsz-Leung; Medzihradszky, Katalin F; Burlingame, Alma L; DeGrado, William F; Jo, Hyunil; Shu, Xiaokun

    2016-07-15

    Protein-protein interactions regulate many biological processes. Identification of interacting proteins is thus an important step toward molecular understanding of cell signaling. The aim of this study was to investigate the use of photo-generated singlet oxygen and a small molecule for proximity labeling of interacting proteins in cellular environment. The protein of interest (POI) was fused with a small singlet oxygen photosensitizer (miniSOG), which generates singlet oxygen ((1)O2) upon irradiation. The locally generated singlet oxygen then activated a biotin-conjugated thiol molecule to form a covalent bond with the proteins nearby. The labeled proteins can then be separated and subsequently identified by mass spectrometry. To demonstrate the applicability of this labeling technology, we fused the miniSOG to Skp2, an F-box protein of the SCF ubiquitin ligase, and expressed the fusion protein in mammalian cells and identified that the surface cysteine of its interacting partner Skp1 was labeled by the biotin-thiol molecule. This photoactivatable protein labeling method may find important applications including identification of weak and transient protein-protein interactions in the native cellular context, as well as spatial and temporal control of protein labeling.

  5. SU-E-I-14: Comparison of Iodine-Labeled and Indium-Labeled Antibody Biodistributions

    SciTech Connect

    Williams, L

    2014-06-01

    Purpose: It is often assumed that animal biodistributions of novel proteins are not dependent upon the radiolabel used in their determination. In units of percent injected dose per gram of tissue (%ID/g), organ uptake results (u) may be obtained using either iodine or metal as radioactive labels. Iodination is preferred as it is a one-step process whereas metal labeling requires two chemical procedures and therefore more protein material. It is important to test whether the radioactive tag leads to variation in the uptake value. Methods: Uptakes of 3antibodies to Carcinoembryonic Antigen (CEA) were evaluated in a nude mouse model bearing 150 to 300 mg LS174T human colon cancer xenografts. Antibodies included diabody (56 kDa), minibody (80kDa) and intact M5A (150 kDa) anti-CEA cognates. Both radioiodine and indium-111 labels were used with uptakes evaluated at 7 time(t) points out to 96 h. Ratios (R) of u(iodine-label)/u(indium-label) were determined for liver, spleen, kidneys, lung and tumor. Results: Hepatic loss was rapid for diabody and minibody; by 24 h their R values were only 2%; i.e., uptake of iodine was 2% of that of indium for these 2 antibodies. By contrast, R for the intact cognate was 50% at that time point. Splenic results were similar. Tumor uptake ratios did not depend upon the antibody type and were 50% at 24 h. Conclusions: Relatively rapid loss of iodine relative to indium in liver and spleen was observed in lower mass antibodies. Tumor ratios were larger and independent of antibody type. Aside from tumor, the R ratio of uptakes depended on the antibody type. R values decreased monotonically with time in all tissues and for all cognates. Using this ratio, one can possibly correct iodine-based u (t) results so that they resemble radiometal-derived biodistributions.

  6. Homosexual Labeling by University Youths

    ERIC Educational Resources Information Center

    Nyberg, Kenneth L.; Alston, Jon P.

    1977-01-01

    Details the responses of young, urban, college-educated people on their attitudes toward homosexuals, specifically focusing on issues of public identification and negative labeling as it effects homosexual persons and their behaviors. (Author/RK)

  7. How to read food labels

    MedlinePlus

    ... 1 serving. You should also pay attention to trans fats on any food label. These fats raise "bad" ... foods and desserts. Many fast food restaurants use trans fats for frying. If a food has these fats, ...

  8. Dietary Supplement Label Database (DSLD)

    MedlinePlus

    ... Print Report Error T he Dietary Supplement Label Database (DSLD) is a joint project of the National ... participants in the latest survey in the DSLD database (NHANES): The search options: Quick Search, Browse Dietary ...

  9. Food Labels Tell the Story!

    MedlinePlus

    ... Environment Kids Health Topics Environment & Health Healthy Living Pollution Reduce, Reuse, Recycle Science – How It Works The ... Pay close attention to serving sizes. Products labeled "light" or "lite" must have 1/3 fewer calories ...

  10. Electrothermal branding for embryo labeling.

    PubMed

    Wang, L; Beebe, D J; Williams, A R; Easley, K D

    1997-11-01

    A novel embryo labeling technique based on electrothermal branding is developed. Two types of micro branding irons are fabricated and tested. One utilizes 25 microns tungsten wire as the heating element. The other utilizes surface micromachining techniques to fabricate polysilicon branding irons. The thermal behavior of the branding irons and the heat distributions in the embryos are analytically modeled. Micron-scale labels on unfertilized bovine embryos are achieved.

  11. Multi-focus cluster labeling.

    PubMed

    Eikvil, Line; Jenssen, Tor-Kristian; Holden, Marit

    2015-06-01

    Document collections resulting from searches in the biomedical literature, for instance, in PubMed, are often so large that some organization of the returned information is necessary. Clustering is an efficient tool for organizing search results. To help the user to decide how to continue the search for relevant documents, the content of each cluster can be characterized by a set of representative keywords or cluster labels. As different users may have different interests, it can be desirable with solutions that make it possible to produce labels from a selection of different topical categories. We therefore introduce the concept of multi-focus cluster labeling to give users the possibility to get an overview of the contents through labels from multiple viewpoints. The concept for multi-focus cluster labeling has been established and has been demonstrated on three different document collections. We illustrate that multi-focus visualizations can give an overview of clusters along axes that general labels are not able to convey. The approach is generic and should be applicable to any biomedical (or other) domain with any selection of foci where appropriate focus vocabularies can be established. A user evaluation also indicates that such a multi-focus concept is useful.

  12. The impact of nutritional labels and socioeconomic status on energy intake. An experimental field study.

    PubMed

    Crockett, Rachel A; Jebb, Susan A; Hankins, Matthew; Marteau, Theresa M

    2014-10-01

    There is some evidence for paradoxical effects of nutritional labelling on energy intake particularly amongst restrained eaters and those with a higher body mass index (BMI) resulting in greater consumption of energy from foods with a positive health message (e.g. "low-fat") compared with the same foods, unlabelled. This study aimed to investigate, in a UK general population sample, the likelihood of paradoxical effects of nutritional labelling on energy intake. Participants (n = 287) attended a London cinema and were offered a large tub of salted or toffee popcorn. Participants were randomised to receive their selected flavour with one of three labels: a green low-fat label, a red high-fat label or no label. Participants watched two film clips while completing measures of demographic characteristics, emotional state and taste of the popcorn. Following the experiment, popcorn consumption was measured. There were no main effects of nutritional labelling on consumption. Contrary to predictions neither BMI nor weight concern moderated the effect of label on consumption. There was a three-way interaction between low-fat label, weight concern and socioeconomic status (SES) such that weight-concerned participants of higher SES who saw a low-fat label consumed more than weight unconcerned participants of similar SES (t = -2.7, P = .04). By contrast, weight-concerned participants of lower SES seeing either type of label, consumed less than those seeing no label (t = -2.04, P = .04). Nutritional labelling may have different effects in different socioeconomic groups. Further studies are required to understand fully the possible contribution of food labelling to health inequalities. PMID:24879885

  13. The impact of nutritional labels and socioeconomic status on energy intake. An experimental field study.

    PubMed

    Crockett, Rachel A; Jebb, Susan A; Hankins, Matthew; Marteau, Theresa M

    2014-10-01

    There is some evidence for paradoxical effects of nutritional labelling on energy intake particularly amongst restrained eaters and those with a higher body mass index (BMI) resulting in greater consumption of energy from foods with a positive health message (e.g. "low-fat") compared with the same foods, unlabelled. This study aimed to investigate, in a UK general population sample, the likelihood of paradoxical effects of nutritional labelling on energy intake. Participants (n = 287) attended a London cinema and were offered a large tub of salted or toffee popcorn. Participants were randomised to receive their selected flavour with one of three labels: a green low-fat label, a red high-fat label or no label. Participants watched two film clips while completing measures of demographic characteristics, emotional state and taste of the popcorn. Following the experiment, popcorn consumption was measured. There were no main effects of nutritional labelling on consumption. Contrary to predictions neither BMI nor weight concern moderated the effect of label on consumption. There was a three-way interaction between low-fat label, weight concern and socioeconomic status (SES) such that weight-concerned participants of higher SES who saw a low-fat label consumed more than weight unconcerned participants of similar SES (t = -2.7, P = .04). By contrast, weight-concerned participants of lower SES seeing either type of label, consumed less than those seeing no label (t = -2.04, P = .04). Nutritional labelling may have different effects in different socioeconomic groups. Further studies are required to understand fully the possible contribution of food labelling to health inequalities.

  14. Molecular and mass spectroscopic analysis of isotopically labeled organic residues

    NASA Technical Reports Server (NTRS)

    Mendoza-Gomez, Celia X.; Greenberg, J. Mayo; Mccain, P.; Ferris, J. P.; Briggs, R.; Degroot, M. S.; Schutte, Willem A.

    1989-01-01

    Experimental studies aimed at understanding the evolution of complex organic molecules on interstellar grains were performed. The photolysis of frozen gas mixtures of various compositions containing H2O, CO, NH3, and CH4 was studied. These species were chosen because of their astrophysical importance as deducted from observational as well as theoretical studies of ice mantles on interstellar grains. These ultraviolet photolyzed ices were warmed up in order to produce refractory organic molecules like the ones formed in molecular clouds when the icy mantles are being irradiated and warmed up either by a nearby stellar source or impulsive heating. The laboratory studies give estimates of the efficiency of production of such organic material under interstellar conditions. It is shown that the gradual carbonization of organic mantles in the diffuse cloud phase leads to higher and higher visual absorptivity - yellow residues become brown in the laboratory. The obtained results can be applied to explaining the organic components of comets and their relevance to the origin of life.

  15. 21 CFR 610.60 - Container label.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 7 2011-04-01 2010-04-01 true Container label. 610.60 Section 610.60 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) BIOLOGICS GENERAL BIOLOGICAL PRODUCTS STANDARDS Labeling Standards § 610.60 Container label. (a) Full label....

  16. 21 CFR 225.180 - Labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 4 2010-04-01 2010-04-01 false Labeling. 225.180 Section 225.180 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS: GENERAL CURRENT GOOD MANUFACTURING PRACTICE FOR MEDICATED FEEDS Labeling § 225.180 Labeling. Labels shall...

  17. 21 CFR 225.180 - Labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 4 2012-04-01 2012-04-01 false Labeling. 225.180 Section 225.180 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS: GENERAL CURRENT GOOD MANUFACTURING PRACTICE FOR MEDICATED FEEDS Labeling § 225.180 Labeling. Labels shall...

  18. 40 CFR 211.108 - Sample label.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 25 2011-07-01 2011-07-01 false Sample label. 211.108 Section 211.108 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) NOISE ABATEMENT PROGRAMS PRODUCT NOISE LABELING General Provisions § 211.108 Sample label. Examples of labels conforming to the requirements...

  19. 40 CFR 211.108 - Sample label.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 25 2014-07-01 2014-07-01 false Sample label. 211.108 Section 211.108 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) NOISE ABATEMENT PROGRAMS PRODUCT NOISE LABELING General Provisions § 211.108 Sample label. Examples of labels conforming to the requirements...

  20. 40 CFR 211.108 - Sample label.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 26 2013-07-01 2013-07-01 false Sample label. 211.108 Section 211.108 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) NOISE ABATEMENT PROGRAMS PRODUCT NOISE LABELING General Provisions § 211.108 Sample label. Examples of labels conforming to the requirements...

  1. 40 CFR 211.108 - Sample label.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 26 2012-07-01 2011-07-01 true Sample label. 211.108 Section 211.108 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) NOISE ABATEMENT PROGRAMS PRODUCT NOISE LABELING General Provisions § 211.108 Sample label. Examples of labels conforming to the requirements...

  2. 9 CFR 381.132 - Labeling approval.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... been submitted for approval to the Food Labeling Division, Regulatory Programs, Food Safety and... request. (b) The Food Labeling Division shall permit submission for approval of only sketch labeling, as... Labeling Division, Regulatory Programs, Food Safety and Inspection Service, U.S. Department of...

  3. 9 CFR 381.132 - Labeling approval.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... product unless the sketch labeling of such final labeling has been submitted for approval to the Food Labeling Division, Regulatory Programs, Food Safety and Inspection Service, and approved by such division... authorized representative of the Secretary upon request. (b) The Food Labeling Division shall...

  4. 9 CFR 381.132 - Labeling approval.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... been submitted for approval to the Food Labeling Division, Regulatory Programs, Food Safety and... request. (b) The Food Labeling Division shall permit submission for approval of only sketch labeling, as... Labeling Division, Regulatory Programs, Food Safety and Inspection Service, U.S. Department of...

  5. 9 CFR 381.132 - Labeling approval.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... been submitted for approval to the Food Labeling Division, Regulatory Programs, Food Safety and... request. (b) The Food Labeling Division shall permit submission for approval of only sketch labeling, as... Labeling Division, Regulatory Programs, Food Safety and Inspection Service, U.S. Department of...

  6. 9 CFR 381.132 - Labeling approval.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... been submitted for approval to the Food Labeling Division, Regulatory Programs, Food Safety and... request. (b) The Food Labeling Division shall permit submission for approval of only sketch labeling, as... Labeling Division, Regulatory Programs, Food Safety and Inspection Service, U.S. Department of...

  7. 40 CFR 211.108 - Sample label.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 24 2010-07-01 2010-07-01 false Sample label. 211.108 Section 211.108 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) NOISE ABATEMENT PROGRAMS PRODUCT NOISE LABELING General Provisions § 211.108 Sample label. Examples of labels conforming to the requirements...

  8. 21 CFR 895.25 - Labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... eliminated by labeling or a change in labeling, or change in advertising if the device is a restricted device... person(s) responsible for the labeling or advertising of the device specifying: (1) The deception or risk... labeling, or change in advertising if the device is a restricted device, necessary to correct the...

  9. 16 CFR 309.17 - Labels.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... edges of the label. If you wish to change the format of this single component label, you must petition... no closer than 3/16″ (.48 cm) from the side edges of the label. If you wish to change the format of.... All labels must be capable of withstanding extremes of weather conditions for a period of at least...

  10. 16 CFR 309.17 - Labels.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... edges of the label. If you wish to change the format of this single component label, you must petition... no closer than 3/16″ (.48 cm) from the side edges of the label. If you wish to change the format of.... All labels must be capable of withstanding extremes of weather conditions for a period of at least...

  11. 16 CFR 309.17 - Labels.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... edges of the label. If you wish to change the format of this single component label, you must petition... no closer than 3/16″ (.48 cm) from the side edges of the label. If you wish to change the format of.... All labels must be capable of withstanding extremes of weather conditions for a period of at least...

  12. Parallel Reaction Monitoring: A Targeted Experiment Performed Using High Resolution and High Mass Accuracy Mass Spectrometry

    PubMed Central

    Rauniyar, Navin

    2015-01-01

    The parallel reaction monitoring (PRM) assay has emerged as an alternative method of targeted quantification. The PRM assay is performed in a high resolution and high mass accuracy mode on a mass spectrometer. This review presents the features that make PRM a highly specific and selective method for targeted quantification using quadrupole-Orbitrap hybrid instruments. In addition, this review discusses the label-based and label-free methods of quantification that can be performed with the targeted approach. PMID:26633379

  13. Optimal policy for labeling training samples

    NASA Astrophysics Data System (ADS)

    Lipsky, Lester; Lopresti, Daniel; Nagy, George

    2013-01-01

    Confirming the labels of automatically classified patterns is generally faster than entering new labels or correcting incorrect labels. Most labels assigned by a classifier, even if trained only on relatively few pre-labeled patterns, are correct. Therefore the overall cost of human labeling can be decreased by interspersing labeling and classification. Given a parameterized model of the error rate as an inverse power law function of the size of the training set, the optimal splits can be computed rapidly. Projected savings in operator time are over 60% for a range of empirical error functions for hand-printed digit classification with ten different classifiers.

  14. 21 CFR 201.70 - Calcium labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... product is more than 3.2 grams: “Ask a doctor before use if you have 1 kidney stones a calcium-restricted... 21 Food and Drugs 4 2012-04-01 2012-04-01 false Calcium labeling. 201.70 Section 201.70 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.70 Calcium labeling. (a) The labeling...

  15. 21 CFR 201.70 - Calcium labeling.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... product is more than 3.2 grams: “Ask a doctor before use if you have 1 kidney stones a calcium-restricted... 21 Food and Drugs 4 2013-04-01 2013-04-01 false Calcium labeling. 201.70 Section 201.70 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.70 Calcium labeling. (a) The labeling...

  16. 21 CFR 201.70 - Calcium labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... product is more than 3.2 grams: “Ask a doctor before use if you have 1 kidney stones a calcium-restricted... 21 Food and Drugs 4 2010-04-01 2010-04-01 false Calcium labeling. 201.70 Section 201.70 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.70 Calcium labeling. (a) The labeling...

  17. 21 CFR 201.70 - Calcium labeling.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... product is more than 3.2 grams: “Ask a doctor before use if you have 1 kidney stones a calcium-restricted... 21 Food and Drugs 4 2014-04-01 2014-04-01 false Calcium labeling. 201.70 Section 201.70 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.70 Calcium labeling. (a) The labeling...

  18. 21 CFR 201.70 - Calcium labeling.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... product is more than 3.2 grams: “Ask a doctor before use if you have 1 kidney stones a calcium-restricted... 21 Food and Drugs 4 2011-04-01 2011-04-01 false Calcium labeling. 201.70 Section 201.70 Food and... LABELING Labeling Requirements for Over-the-Counter Drugs § 201.70 Calcium labeling. (a) The labeling...

  19. Nutrition Labeling Using a Computer Program

    NASA Astrophysics Data System (ADS)

    Metzger, Lloyd E.

    The 1990 Nutrition Labeling and Education Act mandated nutritional labeling of most foods. As a result, a large portion of food analysis is performed for nutritional labeling purposes. A food labeling guide and links to the complete nutritional labeling regulations are available online at http://vm.cfsan.fda.gov/˜dms/flg-toc.html. However, interpretation of these regulations and the appropriate usage of rounding rules, available nutrient content claims, reference amounts, and serving size can be difficult.

  20. 49 CFR 172.430 - POISON label.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 49 Transportation 2 2010-10-01 2010-10-01 false POISON label. 172.430 Section 172.430... SECURITY PLANS Labeling § 172.430 POISON label. (a) Except for size and color, the POISON label must be as follows: EC02MR91.029 (b) In addition to complying with § 172.407, the background on the POISON label...

  1. 49 CFR 172.430 - POISON label.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 49 Transportation 2 2011-10-01 2011-10-01 false POISON label. 172.430 Section 172.430... SECURITY PLANS Labeling § 172.430 POISON label. (a) Except for size and color, the POISON label must be as follows: EC02MR91.029 (b) In addition to complying with § 172.407, the background on the POISON label...

  2. 49 CFR 172.430 - POISON label.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 49 Transportation 2 2013-10-01 2013-10-01 false POISON label. 172.430 Section 172.430... SECURITY PLANS Labeling § 172.430 POISON label. (a) Except for size and color, the POISON label must be as follows: EC02MR91.029 (b) In addition to complying with § 172.407, the background on the POISON label...

  3. 49 CFR 172.430 - POISON label.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 49 Transportation 2 2012-10-01 2012-10-01 false POISON label. 172.430 Section 172.430... SECURITY PLANS Labeling § 172.430 POISON label. (a) Except for size and color, the POISON label must be as follows: EC02MR91.029 (b) In addition to complying with § 172.407, the background on the POISON label...

  4. 49 CFR 172.430 - POISON label.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 49 Transportation 2 2014-10-01 2014-10-01 false POISON label. 172.430 Section 172.430... SECURITY PLANS Labeling § 172.430 POISON label. (a) Except for size and color, the POISON label must be as follows: EC02MR91.029 (b) In addition to complying with § 172.407, the background on the POISON label...

  5. Label-free functional selectivity assays.

    PubMed

    Ferrie, Ann M; Goral, Vasiliy; Wang, Chaoming; Fang, Ye

    2015-01-01

    G protein-coupled receptors (GPCRs) represent the largest class of drug targets. Ligand-directed functional selectivity or biased agonism opens new possibility for discovering GPCR drugs with better efficacy and safety profiles. However, quantification of ligand bias is challenging. Herein, we present five different label-free dynamic mass redistribution (DMR) approaches to assess ligand bias acting at the β2-adrenergic receptor (β2AR). Multiparametric analysis of the DMR agonist profiles reveals divergent pharmacology of a panel of β2AR agonists. DMR profiling using catechol as a conformational probe detects the presence of multiple conformations of the β2AR. DMR assays under microfluidics, together with chemical biology tools, discover ligand-directed desensitization of the receptor. DMR antagonist reverse assays manifest biased antagonism. DMR profiling using distinct probe-modulated cells detects the biased agonism in the context of self-referenced pharmacological activity map. PMID:25563188

  6. Metrics for Labeled Markov Systems

    NASA Technical Reports Server (NTRS)

    Desharnais, Josee; Jagadeesan, Radha; Gupta, Vineet; Panangaden, Prakash

    1999-01-01

    Partial Labeled Markov Chains are simultaneously generalizations of process algebra and of traditional Markov chains. They provide a foundation for interacting discrete probabilistic systems, the interaction being synchronization on labels as in process algebra. Existing notions of process equivalence are too sensitive to the exact probabilities of various transitions. This paper addresses contextual reasoning principles for reasoning about more robust notions of "approximate" equivalence between concurrent interacting probabilistic systems. The present results indicate that:We develop a family of metrics between partial labeled Markov chains to formalize the notion of distance between processes. We show that processes at distance zero are bisimilar. We describe a decision procedure to compute the distance between two processes. We show that reasoning about approximate equivalence can be done compositionally by showing that process combinators do not increase distance. We introduce an asymptotic metric to capture asymptotic properties of Markov chains; and show that parallel composition does not increase asymptotic distance.

  7. Positron emitter labeled enzyme inhibitors

    SciTech Connect

    Fowler, J.S.; MacGregor, R.R.; Wolf, A.P.; Langstrom, B.

    1990-04-03

    This invention involves a new strategy for imaging and mapping enzyme activity in the living human and animal body using positron emitter-labeled suicide enzyme inactivators or inhibitors which become covalently bound to the enzyme as a result of enzymatic catalysis. Two such suicide inactivators for monoamine oxidase have been labeled with carbon-11 and used to map the enzyme subtypes in the living human and animal body using PET. By using positron emission tomography to image the distribution of radioactivity produced by the body penetrating radiation emitted by carbon-11, a map of functionally active monoamine oxidase activity is obtained. Clorgyline and L-deprenyl are suicide enzyme inhibitors and irreversibly inhibit monoamine oxidase. When these inhibitors are labeled with carbon-11 they provide selective probes for monoamine oxidase localization and reactivity in vivo using positron emission tomography.

  8. Positron emitter labeled enzyme inhibitors

    DOEpatents

    Fowler, J.S.; MacGregor, R.R.; Wolf, A.P.

    1987-05-22

    This invention involved a new strategy for imaging and mapping enzyme activity in the living human and animal body using positron emitter-labeled suicide enzyme inactivators or inhibitors which become covalently bound to the enzyme as a result of enzymatic catalysis. Two such suicide in activators for monoamine oxidase have been labeled with carbon-11 and used to map the enzyme subtypes in the living human and animal body using PET. By using positron emission tomography to image the distribution of radioactivity produced by the body penetrating radiation emitted by carbon-11, a map of functionally active monoamine oxidase activity is obtained. Clorgyline and L-deprenyl are suicide enzyme inhibitors and irreversibly inhibit monoamine oxidase. When these inhibitors are labeled with carbon-11 they provide selective probes for monoamine oxidase localization and reactivity in vivo using positron emission tomography. 2 figs.

  9. Positron emitter labeled enzyme inhibitors

    DOEpatents

    Fowler, Joanna S.; MacGregor, Robert R.; Wolf, Alfred P.; Langstrom, Bengt

    1990-01-01

    This invention involves a new strategy for imaging and mapping enzyme activity in the living human and animal body using positron emitter-labeled suicide enzyme inactivators or inhibitors which become covalently bound to the enzyme as a result of enzymatic catalysis. Two such suicide inactivators for monoamine oxidase have been labeled with carbon-11 and used to map the enzyme subtypes in the living human and animal body using PET. By using positron emission tomography to image the distribution of radioactivity produced by the body penetrating radiation emitted by carbon-11, a map of functionally active monoamine oxidase activity is obtained. Clorgyline and L-deprenyl are suicide enzyme inhibitors and irreversibly inhibit monoamine oxidase. When these inhibitors are labeled with carbon-11 they provide selective probes for monoamine oxidase localization and reactivity in vivo using positron emission tomography.

  10. Labeling Feral Spruce Budworm (Lepidoptera: Tortricidae) Populations With Rubidium.

    PubMed

    MacKinnon, Wayne; Eveleigh, Eldon; Silk, Peter; Forbes, Glen

    2016-04-01

    Rubidium (Rb) is a trace element that occurs naturally in low concentrations and is easily absorbed by plants, making it a useful tool for labeling insect defoliators, such as spruce budworm (Choristoneura fumiferana Clemens). Balsam fir trees (Abies balsamea (L.) Miller) injected with either 8 or 16 g per tree of rubidium chloride (RbCl) showed quick uptake and distribution throughout the crown, with no negative effects on tree shoot growth or spruce budworm survival and development. Adult spruce budworm that fed as larvae on trees injected with RbCl were clearly labeled, with significantly higher Rb concentrations than the background levels found in adults that fed as larvae on control trees. Rb concentrations in feral spruce budworm adults for both the 8 g (9 µg/g) and 16 g (25 µg/g) per tree treatments were at least five times lower than those in laboratory-reared adults on 1,000 µg/g RbCl diet (125 µg/g); survival, development, pupal weight, sex ratio, and mating status of spruce budworm were not adversely affected by Rb treatment. Egg masses laid by feral females that fed as larvae on Rb-labeled trees were also labeled with Rb. Injecting trees with RbCl is a viable technique for labeling feral spruce budworm populations to help distinguish local populations from immigrants to better evaluate the success of early intervention strategies such as mating disruption. PMID:26920559

  11. Denture labeling: A new approach.

    PubMed

    Bansal, Pardeep K; Sharma, Akshey; Bhanot, Rajesh

    2011-04-01

    The need for denture labeling is important for forensic and social reasons in case patients need to be identified individually. The importance of denture marking has long been acknowledged by the dental profession. Over the years, various denture marking systems have been reported in the literature, but none till date fulfills all the prescribed ADA specifications. A simple, easy, inexpensive procedure for marking accurate identification marks on dentures with a lead foil is described here. The label caring the patient information is incorporated in the acrylic resin during the denture processing.

  12. Rock Music Gets a Label.

    ERIC Educational Resources Information Center

    Cutietta, Robert

    1986-01-01

    A group called Parents Music Resource Center (PMRC) has captured the media spotlight with a proposal to have warning labels placed on music albums containing sexually explicit or violent lyrics. Major record companies have agreed to a version of the PMRC's demands for a one-year trial period, beginning in 1986. (RM)

  13. Nutrition Marketing on Food Labels

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Nutrition marketing may influence purchasing behavior and thereby be a factor in the obesity epidemic. Very little peer-reviewed research has been published which investigates the relationship between nutrition marketing on food labels and consumer behavior. The purpose of this paper was to give an ...

  14. Revisiting Labels: "Hearing" or Not?

    ERIC Educational Resources Information Center

    Rhoades, Ellen A.

    2010-01-01

    This position paper briefly presents evidence-based findings pertaining to the language of labels for people with hearing loss that relate to stigma, expectation levels, stereotypes, and self-fulfilling prophecies. These constructs are important for auditory-based practitioners, administrators, policymakers, students, families, and persons with…

  15. Psychological effectiveness of carbon labelling

    NASA Astrophysics Data System (ADS)

    Beattie, Geoffrey

    2012-04-01

    Despite the decision by supermarket-giant Tesco to delay its plan to add carbon-footprint information onto all of its 70,000 products, carbon labelling, if carefully designed, could yet change consumer behaviour. However, it requires a new type of thinking about consumers and much additional work.

  16. The Labelling Approach to Deviance.

    ERIC Educational Resources Information Center

    Rains, Prudence M.; Kitsuse, John L.; Duster, Troy; Freidson, Eliot

    2003-01-01

    This reprint of one chapter from the 1975 text, "Issues in the Classification of Children" by Nicholas Hobbs and others, addresses the theoretical, methodological, and empirical issues involved in the "labeling" approach to the sociology of deviance. It examines the social process of classification, the use of classification in social agencies,…

  17. Radionuclide-labeled red blood cells: current status and future prospects

    SciTech Connect

    Srivastava, S.C.; Chervu, L.R.

    1984-04-01

    Radiolabeling of red cells and their clinical and research application in nuclear medicine constitute an area of continued interest and steady growth during the past two decades. Technetium-/sup 99/m-labeled red cells in particular have revolutionized the field of cardiovascular nuclear medicine by making possible the external evaluation of various heart parameters with minimum radiation dose or trauma to the patient. Among other areas of study that use /sup 99/mTc -RBC are blood pool imaging, detection of vascular malformations, red cell mass determination, detection of gastrointestinal bleeding, and of hemangiomas. Heat-damaged /sup 99/mTc -RBC find application in spleen imaging, accessory spleen localization, detection of GI bleeding, and in other areas. A critical evaluation is presented of the various in vitro and in vivo labeling techniques that are currently available for red cell labeling. Even though the presently used procedures provide satisfactory labeled preparations, ideal radioisotopic RBC labels remain to be developed. Intermediate (2-3 days) as well as long-lived (approximately 30 days) radionuclidic labels are highly desirable for a number of clinical procedures where /sup 99/mTc is not useful due to its short half-life. New approaches such as the use of radiolabeled antibodies to red cell antigens, or labeling specific receptor sites in the cell may lead to substantial improvements in the labeling methodology and could yield labeled cells with the least damage and maximum in vivo stability.

  18. Establishing Drug Resistance in Microorganisms by Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Demirev, Plamen A.; Hagan, Nathan S.; Antoine, Miquel D.; Lin, Jeffrey S.; Feldman, Andrew B.

    2013-08-01

    A rapid method to determine drug resistance in bacteria based on mass spectrometry is presented. In it, a mass spectrum of an intact microorganism grown in drug-containing stable isotope-labeled media is compared with a mass spectrum of the intact microorganism grown in non-labeled media without the drug present. Drug resistance is determined by predicting characteristic mass shifts of one or more microorganism biomarkers using bioinformatics algorithms. Observing such characteristic mass shifts indicates that the microorganism is viable even in the presence of the drug, thus incorporating the isotopic label into characteristic biomarker molecules. The performance of the method is illustrated on the example of intact E. coli, grown in control (unlabeled) and 13C-labeled media, and analyzed by MALDI TOF MS. Algorithms for data analysis are presented as well.

  19. The labeling debate in the United States.

    PubMed

    Marchant, Gary E; Cardineau, Guy A

    2013-01-01

    The mandatory labeling of genetically modified (GM) food has become the predominant policy issue concerning biotechnology in the United States. The controversy over GM labeling is being debated at several different levels and branches of government. At the federal level, the Food and Drug Administration, which has primary jurisdiction over food safety and labeling, has steadfastly refused to require labeling of GM foods since 1992 based on its conclusion that GM foods as a category present no unique or higher risks than other foods. Proposed legislation has been repeatedly introduced in the US. Congress over the years to mandate GM labeling, but has made very little progress. With federal labeling requirements apparently stalled, the main activity has switched to the state level, where numerous individual states are considering mandatory GM labeling, either through legislation or proposition. The debate over GM labeling, at both the federal and state levels, has focused on five issues: (1) public opinion; (2) the legality of labeling requirements; (3) the risks and benefits of GM foods; (4) the costs and burdens of GM labeling; and (5) consumer choice. While the pro-labeling forces argue that all of these factors weigh in favor of mandatory GM labeling, a more careful evaluation of the evidence finds that all five factors weigh decisively against mandatory GM labeling requirements.

  20. Improved proteome coverage by using iTRAQ labelling and peptide OFFGEL fractionation

    PubMed Central

    Ernoult, Emilie; Gamelin, Erick; Guette, Catherine

    2008-01-01

    Background The development of mass spectrometric techniques and fractionation methods now allows the investigation of very complex protein mixtures ranging from subcellular structures to tissues. Nevertheless, this work is particularly difficult due to the wide dynamic range of protein concentration in eukaryotic tissues. In this paper, we present a shotgun method whereby the peptides are fractionated using OFFGEL electrophoresis after iTRAQ labelling. Results We demonstrated that iTRAQ peptide labelling enhances MALDI ionisation and that the OFFGEL fractionation of the labelled peptides introduces a supplementary criterion (pI) useful for validation and identification of proteins. We showed that iTRAQ samples allowed lower-concentrated proteins identification in comparison with free-labelled samples. Conclusion The combined use of iTRAQ labelling and OFFGEL fractionation allows a considerable increase in proteome coverage of very complex samples prepared from total cell extracts and supports the low-concentrated protein identification. PMID:18851748

  1. Relative quantification of biomarkers using mixed-isotope labeling coupled with MS

    PubMed Central

    Chapman, Heidi M; Schutt, Katherine L; Dieter, Emily M; Lamos, Shane M

    2013-01-01

    The identification and quantification of important biomarkers is a critical first step in the elucidation of biological systems. Biomarkers take many forms as cellular responses to stimuli and can be manifested during transcription, translation, and/or metabolic processing. Increasingly, researchers have relied upon mixed-isotope labeling (MIL) coupled with MS to perform relative quantification of biomarkers between two or more biological samples. MIL effectively tags biomarkers of interest for ease of identification and quantification within the mass spectrometer by using isotopic labels that introduce a heavy and light form of the tag. In addition to MIL coupled with MS, a number of other approaches have been used to quantify biomarkers including protein gel staining, enzymatic labeling, metabolic labeling, and several label-free approaches that generate quantitative data from the MS signal response. This review focuses on MIL techniques coupled with MS for the quantification of protein and small-molecule biomarkers. PMID:23157360

  2. 21 CFR 211.125 - Labeling issuance.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... identity and conformity to the labeling specified in the master or batch production records. (c) Procedures... discrepancies shall be investigated in accordance with § 211.192. Labeling reconciliation is waived for cut...

  3. 21 CFR 211.125 - Labeling issuance.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... identity and conformity to the labeling specified in the master or batch production records. (c) Procedures... discrepancies shall be investigated in accordance with § 211.192. Labeling reconciliation is waived for cut...

  4. 21 CFR 211.125 - Labeling issuance.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... identity and conformity to the labeling specified in the master or batch production records. (c) Procedures... discrepancies shall be investigated in accordance with § 211.192. Labeling reconciliation is waived for cut...

  5. 21 CFR 211.125 - Labeling issuance.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... identity and conformity to the labeling specified in the master or batch production records. (c) Procedures... discrepancies shall be investigated in accordance with § 211.192. Labeling reconciliation is waived for cut...

  6. 21 CFR 211.125 - Labeling issuance.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... identity and conformity to the labeling specified in the master or batch production records. (c) Procedures... discrepancies shall be investigated in accordance with § 211.192. Labeling reconciliation is waived for cut...

  7. 40 CFR 205.158 - Labeling requirements.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... TRANSPORTATION EQUIPMENT NOISE EMISSION CONTROLS Motorcycles § 205.158 Labeling requirements. (a)(1) The... information: (i) The label heading: Motorcycle Noise Emission Control Information; (ii) The statement: This ___ (model year) ___ (model specific code) motorcycle, ___ (serial number), meets EPA noise...

  8. 40 CFR 1033.135 - Labeling.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... THAT IT IS REMANUFACTURED, EXCEPT AS ALLOWED BY 40 CFR 1033.750.” (3) Label diesel-fueled locomotives... locomotives certified for use with both LSD and ULSD. (c) Engine labels. (1) For engines not...

  9. 21 CFR 640.84 - Labeling.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Albumin (Human) § 640.84 Labeling. In addition to the labeling... percent albumin is administered to a patient with marked dehydration; (d) The protein...

  10. 21 CFR 640.84 - Labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Albumin (Human) § 640.84 Labeling. In addition to the labeling... percent albumin is administered to a patient with marked dehydration; (d) The protein...

  11. 21 CFR 640.84 - Labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Albumin (Human) § 640.84 Labeling. In addition to the labeling... percent albumin is administered to a patient with marked dehydration; (d) The protein...

  12. 21 CFR 640.84 - Labeling.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Albumin (Human) § 640.84 Labeling. In addition to the labeling... percent albumin is administered to a patient with marked dehydration; (d) The protein...

  13. 21 CFR 640.84 - Labeling.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Albumin (Human) § 640.84 Labeling. In addition to the labeling... percent albumin is administered to a patient with marked dehydration; (d) The protein...

  14. 21 CFR 640.94 - Labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Plasma Protein Fraction (Human) § 640.94 Labeling. In addition... package labels shall contain the following information: (a) The osmotic equivalent in terms of plasma,...

  15. 21 CFR 640.94 - Labeling.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Plasma Protein Fraction (Human) § 640.94 Labeling. In addition... package labels shall contain the following information: (a) The osmotic equivalent in terms of plasma,...

  16. 21 CFR 640.94 - Labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Plasma Protein Fraction (Human) § 640.94 Labeling. In addition... package labels shall contain the following information: (a) The osmotic equivalent in terms of plasma,...

  17. 21 CFR 640.94 - Labeling.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Plasma Protein Fraction (Human) § 640.94 Labeling. In addition... package labels shall contain the following information: (a) The osmotic equivalent in terms of plasma,...

  18. 21 CFR 640.94 - Labeling.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Plasma Protein Fraction (Human) § 640.94 Labeling. In addition... package labels shall contain the following information: (a) The osmotic equivalent in terms of plasma,...

  19. 27 CFR 19.437 - Labels.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... affix a label showing the following information: (1) The proprietor's name and plant number; (2) The... paragraph (a) of this section is not required when the sample container bears a label approved under part...

  20. 27 CFR 19.704 - Labels.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... containers bear an approved label pursuant to 27 CFR Part 5 and subpart S of this part and the sample is... spirits to be withdrawn under the provisions of § 19.701, the proprietor shall affix a label showing...

  1. 27 CFR 19.437 - Labels.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... affix a label showing the following information: (1) The proprietor's name and plant number; (2) The... paragraph (a) of this section is not required when the sample container bears a label approved under part...

  2. 27 CFR 19.437 - Labels.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... affix a label showing the following information: (1) The proprietor's name and plant number; (2) The... paragraph (a) of this section is not required when the sample container bears a label approved under part...

  3. 27 CFR 19.437 - Labels.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... affix a label showing the following information: (1) The proprietor's name and plant number; (2) The... paragraph (a) of this section is not required when the sample container bears a label approved under part...

  4. 49 CFR 172.407 - Label specifications.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... Register citations affecting § 172.407, see the List of CFR Sections Affected, which appears in the Finding... the CORROSIVE label. (iii) White may be used for the symbol for the ORGANIC PEROXIDE label. (3)...

  5. 21 CFR 225.80 - Labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... labeling, including placards, upon receipt from the printer shall be proofread against the Master Record... responsible individual, and kept for 1 year after all the labels from that batch have been used. (3) In...

  6. 21 CFR 225.80 - Labeling.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... labeling, including placards, upon receipt from the printer shall be proofread against the Master Record... responsible individual, and kept for 1 year after all the labels from that batch have been used. (3) In...

  7. 99mTc: Labeling Chemistry and Labeled Compounds

    NASA Astrophysics Data System (ADS)

    Alberto, R.; Abram, U.

    This chapter reviews the radiopharmaceutical chemistry of technetium related to the synthesis of perfusion agents and to the labeling of receptor-binding biomolecules. To understand the limitations of technetium chemistry imposed by future application of the complexes in nuclear medicine, an introductory section analyzes the compulsory requirements to be considered when facing the incentive of introducing a novel radiopharmaceutical into the market. Requirements from chemistry, routine application, and market are discussed. In a subsequent section, commercially available 99mTc-based radiopharmaceuticals are treated. It covers the complexes in use for imaging the most important target organs such as heart, brain, or kidney. The commercially available radiopharmaceuticals fulfill the requirements outlined earlier and are discussed with this background. In a following section, the properties and perspectives of the different generations of radiopharmaceuticals are described in a general way, covering characteristics for perfusion agents and for receptor-specific molecules. Technetium chemistry for the synthesis of perfusion agents and the different labeling approaches for target-specific biomolecules are summarized. The review comprises a general introduction to the common approaches currently in use, employing the N x S4-x , [3+1] and 2-hydrazino-nicotinicacid (HYNIC) method as well as more recent strategies such as the carbonyl and the TcN approach. Direct labeling without the need of a bifunctional chelator is briefly reviewed as well. More particularly, recent developments in the labeling of concrete targeting molecules, the second generation of radiopharmaceuticals, is then discussed and prominent examples with antibodies/peptides, neuroreceptor targeting small molecules, myocardial imaging agents, vitamins, thymidine, and complexes relevant to multidrug resistance are given. In addition, a new approach toward peptide drug development is described. The section

  8. Automated labeling in document images

    NASA Astrophysics Data System (ADS)

    Kim, Jongwoo; Le, Daniel X.; Thoma, George R.

    2000-12-01

    The National Library of Medicine (NLM) is developing an automated system to produce bibliographic records for its MEDLINER database. This system, named Medical Article Record System (MARS), employs document image analysis and understanding techniques and optical character recognition (OCR). This paper describes a key module in MARS called the Automated Labeling (AL) module, which labels all zones of interest (title, author, affiliation, and abstract) automatically. The AL algorithm is based on 120 rules that are derived from an analysis of journal page layouts and features extracted from OCR output. Experiments carried out on more than 11,000 articles in over 1,000 biomedical journals show the accuracy of this rule-based algorithm to exceed 96%.

  9. Homosexual Labeling and the Male Role.

    ERIC Educational Resources Information Center

    Karr, Rodney G.

    1978-01-01

    In this study, men were perceived to be less masculine and less preferred as fellow participants if they were labeled homosexual. The man responsible for the primary labeling of the homosexual was perceived as more masculine and more sociable when he labeled the homosexual than when he did not. (Author/WI)

  10. 30 CFR 47.42 - Label contents.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 30 Mineral Resources 1 2012-07-01 2012-07-01 false Label contents. 47.42 Section 47.42 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING HAZARD COMMUNICATION (HazCom) Container Labels and Other Forms of Warning § 47.42 Label contents. When an operator...

  11. 30 CFR 47.42 - Label contents.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 30 Mineral Resources 1 2013-07-01 2013-07-01 false Label contents. 47.42 Section 47.42 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING HAZARD COMMUNICATION (HazCom) Container Labels and Other Forms of Warning § 47.42 Label contents. When an operator...

  12. 40 CFR 600.301 - Labeling requirements.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... ECONOMY AND GREENHOUSE GAS EXHAUST EMISSIONS OF MOTOR VEHICLES Fuel Economy Labeling § 600.301 Labeling... each dealer shall maintain or cause to be maintained on each automobile: (1) A general fuel economy... vehicle for which a specific label is requested which has a combined FTP/HFET-based fuel economy value,...

  13. 40 CFR 600.301 - Labeling requirements.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... ECONOMY AND GREENHOUSE GAS EXHAUST EMISSIONS OF MOTOR VEHICLES Fuel Economy Labeling § 600.301 Labeling... each dealer shall maintain or cause to be maintained on each automobile: (1) A general fuel economy... vehicle for which a specific label is requested which has a combined FTP/HFET-based fuel economy value,...

  14. 40 CFR 600.301 - Labeling requirements.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... ECONOMY AND GREENHOUSE GAS EXHAUST EMISSIONS OF MOTOR VEHICLES Fuel Economy Labeling § 600.301 Labeling... each dealer shall maintain or cause to be maintained on each automobile: (1) A general fuel economy... vehicle for which a specific label is requested which has a combined FTP/HFET-based fuel economy value,...

  15. 21 CFR 660.35 - Labeling.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... STANDARDS FOR DIAGNOSTIC SUBSTANCES FOR LABORATORY TESTS Reagent Red Blood Cells § 660.35 Labeling. In... or end of the label, oustide of the main panel. (2) If washing the cells is required by the manufacturer, the container label shall include appropriate instructions; if the cells should not be...

  16. 21 CFR 660.35 - Labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... STANDARDS FOR DIAGNOSTIC SUBSTANCES FOR LABORATORY TESTS Reagent Red Blood Cells § 660.35 Labeling. In... or end of the label, oustide of the main panel. (2) If washing the cells is required by the manufacturer, the container label shall include appropriate instructions; if the cells should not be...

  17. 30 CFR 47.42 - Label contents.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Label contents. 47.42 Section 47.42 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING HAZARD COMMUNICATION (HazCom) Container Labels and Other Forms of Warning § 47.42 Label contents. When an operator...

  18. 40 CFR 211.104 - Label content.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... Required by U.S. EPA regulation 40 CFR part 211, subpart ___.” EC01FE92.055 ... 40 Protection of Environment 24 2010-07-01 2010-07-01 false Label content. 211.104 Section 211.104... LABELING General Provisions § 211.104 Label content. The following data and information must be on...

  19. 40 CFR 1036.135 - Labeling.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ....135 Labeling. Label your engines as described in 40 CFR 86.007-35(a)(3), with the following additional information: (a) (b) Identify the emission control system. Use terms and abbreviations as described in 40 CFR... labeling requirement to be consistent with the intent of 40 CFR part 1037....

  20. 40 CFR 1036.135 - Labeling.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ....135 Labeling. Label your engines as described in 40 CFR 86.007-35(a)(3), with the following additional information: (a) (b) Identify the emission control system. Use terms and abbreviations as described in 40 CFR... labeling requirement to be consistent with the intent of 40 CFR part 1037....

  1. 16 CFR 306.12 - Labels.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... biodiesel, biomass-based diesel, biodiesel blends, and biomass-based diesel blends. The label is 3 inches (7... the black band. Directly underneath the black band, the label shall read “contains biomass-based... the side edges of the label. (5) For biomass-based diesel blends containing more than 5 percent and...

  2. 21 CFR 820.120 - Device labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Device labeling. 820.120 Section 820.120 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES QUALITY SYSTEM REGULATION Labeling and Packaging Control § 820.120 Device labeling. Each...

  3. 27 CFR 18.55 - Label.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... TREASURY ALCOHOL PRODUCTION OF VOLATILE FRUIT-FLAVOR CONCENTRATE Operations § 18.55 Label. Each container of concentrate will have affixed thereto, before transfer, a label identifying the product and... 27 Alcohol, Tobacco Products and Firearms 1 2014-04-01 2014-04-01 false Label. 18.55 Section...

  4. 21 CFR 610.60 - Container label.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... following items shall appear on the label affixed to each container of a product capable of bearing a full label: (1) The proper name of the product; (2) The name, address, and license number of manufacturer; (3... 21 Food and Drugs 7 2013-04-01 2013-04-01 false Container label. 610.60 Section 610.60 Food...

  5. 27 CFR 19.604 - Caution label.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Caution label. 19.604... OF THE TREASURY LIQUORS DISTILLED SPIRITS PLANTS Containers and Marks Marks § 19.604 Caution label... denaturer may be printed on such label, but no other extraneous matter will be permitted thereon without...

  6. 40 CFR 763.171 - Labeling requirements.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... cannot be removed without defacing or destroying them. Product labels shall appear as in paragraph (d)(2... packaging, the label must be attached to the innermost layer adjacent to the product. If the innermost layer... product's innermost layer of product wrapping or packaging, or a label must be attached to the next...

  7. 40 CFR 211.105 - Label format.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 25 2014-07-01 2014-07-01 false Label format. 211.105 Section 211.105 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) NOISE ABATEMENT PROGRAMS PRODUCT NOISE LABELING General Provisions § 211.105 Label format. (a) Unless specified otherwise in other...

  8. 27 CFR 18.55 - Label.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... TREASURY ALCOHOL PRODUCTION OF VOLATILE FRUIT-FLAVOR CONCENTRATE Operations § 18.55 Label. Each container of concentrate will have affixed thereto, before transfer, a label identifying the product and... 27 Alcohol, Tobacco Products and Firearms 1 2013-04-01 2013-04-01 false Label. 18.55 Section...

  9. 27 CFR 18.55 - Label.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... TREASURY LIQUORS PRODUCTION OF VOLATILE FRUIT-FLAVOR CONCENTRATE Operations § 18.55 Label. Each container of concentrate will have affixed thereto, before transfer, a label identifying the product and... 27 Alcohol, Tobacco Products and Firearms 1 2012-04-01 2012-04-01 false Label. 18.55 Section...

  10. 21 CFR 610.60 - Container label.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... following items shall appear on the label affixed to each container of a product capable of bearing a full label: (1) The proper name of the product; (2) The name, address, and license number of manufacturer; (3... 21 Food and Drugs 7 2014-04-01 2014-04-01 false Container label. 610.60 Section 610.60 Food...

  11. 9 CFR 116.3 - Label records.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    .... Each label shall be identified as to: (1) Name and product code number as it appears on the product... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Label records. 116.3 Section 116.3..., SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS RECORDS AND REPORTS § 116.3 Label...

  12. 40 CFR 211.105 - Label format.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 24 2010-07-01 2010-07-01 false Label format. 211.105 Section 211.105 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) NOISE ABATEMENT PROGRAMS PRODUCT NOISE LABELING General Provisions § 211.105 Label format. (a) Unless specified otherwise in other...

  13. 40 CFR 211.105 - Label format.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 26 2012-07-01 2011-07-01 true Label format. 211.105 Section 211.105 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) NOISE ABATEMENT PROGRAMS PRODUCT NOISE LABELING General Provisions § 211.105 Label format. (a) Unless specified otherwise in other...

  14. 9 CFR 116.3 - Label records.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    .... Each label shall be identified as to: (1) Name and product code number as it appears on the product... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Label records. 116.3 Section 116.3..., SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS RECORDS AND REPORTS § 116.3 Label...

  15. 9 CFR 116.3 - Label records.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    .... Each label shall be identified as to: (1) Name and product code number as it appears on the product... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Label records. 116.3 Section 116.3..., SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS RECORDS AND REPORTS § 116.3 Label...

  16. 21 CFR 610.60 - Container label.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... following items shall appear on the label affixed to each container of a product capable of bearing a full label: (1) The proper name of the product; (2) The name, address, and license number of manufacturer; (3... 21 Food and Drugs 7 2012-04-01 2012-04-01 false Container label. 610.60 Section 610.60 Food...

  17. 9 CFR 116.3 - Label records.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    .... Each label shall be identified as to: (1) Name and product code number as it appears on the product... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Label records. 116.3 Section 116.3..., SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS RECORDS AND REPORTS § 116.3 Label...

  18. 21 CFR 1271.250 - Labeling controls.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Labeling controls. 1271.250 Section 1271.250 Food..., AND CELLULAR AND TISSUE-BASED PRODUCTS Current Good Tissue Practice § 1271.250 Labeling controls. (a) General. You must establish and maintain procedures to control the labeling of HCT/Ps. You must...

  19. 10 CFR 61.57 - Labeling.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 10 Energy 2 2010-01-01 2010-01-01 false Labeling. 61.57 Section 61.57 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) LICENSING REQUIREMENTS FOR LAND DISPOSAL OF RADIOACTIVE WASTE Technical Requirements for Land Disposal Facilities § 61.57 Labeling. Each package of waste must be clearly labeled...

  20. 10 CFR 61.57 - Labeling.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 10 Energy 2 2012-01-01 2012-01-01 false Labeling. 61.57 Section 61.57 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) LICENSING REQUIREMENTS FOR LAND DISPOSAL OF RADIOACTIVE WASTE Technical Requirements for Land Disposal Facilities § 61.57 Labeling. Each package of waste must be clearly labeled...

  1. 10 CFR 61.57 - Labeling.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 10 Energy 2 2011-01-01 2011-01-01 false Labeling. 61.57 Section 61.57 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) LICENSING REQUIREMENTS FOR LAND DISPOSAL OF RADIOACTIVE WASTE Technical Requirements for Land Disposal Facilities § 61.57 Labeling. Each package of waste must be clearly labeled...

  2. 10 CFR 61.57 - Labeling.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 10 Energy 2 2014-01-01 2014-01-01 false Labeling. 61.57 Section 61.57 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) LICENSING REQUIREMENTS FOR LAND DISPOSAL OF RADIOACTIVE WASTE Technical Requirements for Land Disposal Facilities § 61.57 Labeling. Each package of waste must be clearly labeled...

  3. 10 CFR 61.57 - Labeling.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 10 Energy 2 2013-01-01 2013-01-01 false Labeling. 61.57 Section 61.57 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) LICENSING REQUIREMENTS FOR LAND DISPOSAL OF RADIOACTIVE WASTE Technical Requirements for Land Disposal Facilities § 61.57 Labeling. Each package of waste must be clearly labeled...

  4. 9 CFR 317.4 - Labeling approval.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... approval to the Food Labeling Division, Regulatory Programs, Food Safety and Inspection Service, and... approval as set forth in § 317.4(a) shall be submitted in duplicate to the Food Labeling Division... deemed deficient in some particular may be granted by the Food Labeling Division. Temporary approvals...

  5. 9 CFR 317.4 - Labeling approval.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... approval to the Food Labeling Division, Regulatory Programs, Food Safety and Inspection Service, and... approval as set forth in § 317.4(a) shall be submitted in duplicate to the Food Labeling Division... deemed deficient in some particular may be granted by the Food Labeling Division. Temporary approvals...

  6. 9 CFR 317.4 - Labeling approval.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... approval to the Food Labeling Division, Regulatory Programs, Food Safety and Inspection Service, and... approval as set forth in § 317.4(a) shall be submitted in duplicate to the Food Labeling Division... deemed deficient in some particular may be granted by the Food Labeling Division. Temporary approvals...

  7. 75 FR 81943 - Appliance Labeling Rule

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-12-29

    ... for its new light bulb labeling requirements (published on July 19, 2010, 75 FR 41696) to January 1... (75 FR 41696), the Commission published amendments to the Appliance Labeling Rule (Rule) creating new... new labeling requirements. \\5\\ 59 FR 25176 (May 13, 1994). \\6\\ Pursuant to the revised rule, after...

  8. 21 CFR 660.35 - Labeling.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... STANDARDS FOR DIAGNOSTIC SUBSTANCES FOR LABORATORY TESTS Reagent Red Blood Cells § 660.35 Labeling. In... or end of the label, oustide of the main panel. (2) If washing the cells is required by the manufacturer, the container label shall include appropriate instructions; if the cells should not be...

  9. Cigarette Warning Labels as Educational Devices.

    ERIC Educational Resources Information Center

    Newman, Ian M.; And Others

    This paper reports an investigation on the educational impact of warning labels on cigarette packages on adolescents. Subjects were asked to identify the locations of warning labels on cigarette packages and advertising and to restate the warning label. Results indicated that official warnings may be well known in general terms but poorly known in…

  10. 21 CFR 701.11 - Identity labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 7 2010-04-01 2010-04-01 false Identity labeling. 701.11 Section 701.11 Food and... COSMETIC LABELING Package Form § 701.11 Identity labeling. (a) The principal display panel of a cosmetic in package form shall bear as one of its principal features a statement of the identity of the commodity....

  11. 40 CFR 205.158 - Labeling requirements.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 26 2012-07-01 2011-07-01 true Labeling requirements. 205.158 Section 205.158 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) NOISE ABATEMENT PROGRAMS... color that contrasts with the background of the label. (5) The label must contain the...

  12. 30 CFR 47.42 - Label contents.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 30 Mineral Resources 1 2011-07-01 2011-07-01 false Label contents. 47.42 Section 47.42 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING HAZARD COMMUNICATION (HazCom) Container Labels and Other Forms of Warning § 47.42 Label contents. When an operator...

  13. 40 CFR 211.104 - Label content.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... Required by U.S. EPA regulation 40 CFR part 211, subpart ___.” EC01FE92.055 ... 40 Protection of Environment 25 2011-07-01 2011-07-01 false Label content. 211.104 Section 211.104... LABELING General Provisions § 211.104 Label content. The following data and information must be on...

  14. 10 CFR 431.31 - Labeling requirements.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 10 Energy 3 2012-01-01 2012-01-01 false Labeling requirements. 431.31 Section 431.31 Energy DEPARTMENT OF ENERGY ENERGY CONSERVATION ENERGY EFFICIENCY PROGRAM FOR CERTAIN COMMERCIAL AND INDUSTRIAL EQUIPMENT Electric Motors Labeling § 431.31 Labeling requirements. (a) Electric motor nameplate—(1)...

  15. 10 CFR 431.31 - Labeling requirements.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 10 Energy 3 2013-01-01 2013-01-01 false Labeling requirements. 431.31 Section 431.31 Energy DEPARTMENT OF ENERGY ENERGY CONSERVATION ENERGY EFFICIENCY PROGRAM FOR CERTAIN COMMERCIAL AND INDUSTRIAL EQUIPMENT Electric Motors Labeling § 431.31 Labeling requirements. (a) Electric motor nameplate—(1)...

  16. 10 CFR 431.31 - Labeling requirements.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 10 Energy 3 2014-01-01 2014-01-01 false Labeling requirements. 431.31 Section 431.31 Energy DEPARTMENT OF ENERGY ENERGY CONSERVATION ENERGY EFFICIENCY PROGRAM FOR CERTAIN COMMERCIAL AND INDUSTRIAL EQUIPMENT Electric Motors Labeling § 431.31 Labeling requirements. (a) Electric motor nameplate—(1)...

  17. 21 CFR 349.80 - Professional labeling.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... labeling. The labeling of any OTC ophthalmic demulcent drug product provided to health professionals (but... 21 Food and Drugs 5 2011-04-01 2011-04-01 false Professional labeling. 349.80 Section 349.80 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS...

  18. 21 CFR 349.80 - Professional labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... labeling. The labeling of any OTC ophthalmic demulcent drug product provided to health professionals (but... 21 Food and Drugs 5 2010-04-01 2010-04-01 false Professional labeling. 349.80 Section 349.80 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS...

  19. 21 CFR 357.280 - Professional labeling.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 5 2013-04-01 2013-04-01 false Professional labeling. 357.280 Section 357.280 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS... Drug Products § 357.280 Professional labeling. The labeling provided to health professionals (but...

  20. Learning Words from Labeling and Directive Speech

    ERIC Educational Resources Information Center

    Callanan, Maureen A.; Akhtar, Nameera; Sussman, Lisa

    2014-01-01

    Despite the common intuition that labeling may be the best way to teach a new word to a child, systematic testing is needed of the prediction that children learn words better from labeling utterances than from directive utterances. Two experiments compared toddlers' label learning in the context of hearing words used in directive versus labeling…

  1. 30 CFR 47.42 - Label contents.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 30 Mineral Resources 1 2014-07-01 2014-07-01 false Label contents. 47.42 Section 47.42 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING HAZARD COMMUNICATION (HazCom) Container Labels and Other Forms of Warning § 47.42 Label contents. When an operator...

  2. 16 CFR 1500.128 - Label comment.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FEDERAL HAZARDOUS SUBSTANCES ACT REGULATIONS HAZARDOUS SUBSTANCES AND ARTICLES; ADMINISTRATION AND ENFORCEMENT REGULATIONS § 1500.128 Label comment. The... hazardous substance if furnished with: (a) Complete labeling or proposed labeling, which may be in...

  3. 16 CFR 1500.128 - Label comment.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FEDERAL HAZARDOUS SUBSTANCES ACT REGULATIONS HAZARDOUS SUBSTANCES AND ARTICLES; ADMINISTRATION AND ENFORCEMENT REGULATIONS § 1500.128 Label comment. The... hazardous substance if furnished with: (a) Complete labeling or proposed labeling, which may be in...

  4. 16 CFR 1500.128 - Label comment.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FEDERAL HAZARDOUS SUBSTANCES ACT REGULATIONS HAZARDOUS SUBSTANCES AND ARTICLES; ADMINISTRATION AND ENFORCEMENT REGULATIONS § 1500.128 Label comment. The... hazardous substance if furnished with: (a) Complete labeling or proposed labeling, which may be in...

  5. 16 CFR 1500.128 - Label comment.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FEDERAL HAZARDOUS SUBSTANCES ACT REGULATIONS HAZARDOUS SUBSTANCES AND ARTICLES; ADMINISTRATION AND ENFORCEMENT REGULATIONS § 1500.128 Label comment. The... hazardous substance if furnished with: (a) Complete labeling or proposed labeling, which may be in...

  6. 16 CFR 1500.128 - Label comment.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FEDERAL HAZARDOUS SUBSTANCES ACT REGULATIONS HAZARDOUS SUBSTANCES AND ARTICLES; ADMINISTRATION AND ENFORCEMENT REGULATIONS § 1500.128 Label comment. The... hazardous substance if furnished with: (a) Complete labeling or proposed labeling, which may be in...

  7. 21 CFR 660.28 - Labeling.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... label—(1) Color coding. The final container label of all Blood Grouping Reagents shall be completely... panel. Blood grouping reagent Color of label paper Anti-A Blue. Anti-B Yellow. Slide and rapid tube test... STANDARDS FOR DIAGNOSTIC SUBSTANCES FOR LABORATORY TESTS Blood Grouping Reagent § 660.28 Labeling....

  8. 21 CFR 660.28 - Labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... label—(1) Color coding. The final container label of all Blood Grouping Reagents shall be completely... panel. Blood grouping reagent Color of label paper Anti-A Blue. Anti-B Yellow. Slide and rapid tube test... STANDARDS FOR DIAGNOSTIC SUBSTANCES FOR LABORATORY TESTS Blood Grouping Reagent § 660.28 Labeling....

  9. 10 CFR 431.31 - Labeling requirements.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... manufacturer or private labeler, pursuant to § 431.36(f), and applicable to that motor. Such CC number must be... EQUIPMENT Electric Motors Labeling § 431.31 Labeling requirements. (a) Electric motor nameplate—(1) Required information. The permanent nameplate of an electric motor for which standards are prescribed in § 431.25...

  10. Sulphur tracer experiments in laboratory animals using 34S-labelled yeast.

    PubMed

    Martínez-Sierra, J Giner; Moreno Sanz, F; Herrero Espílez, P; Marchante Gayón, J M; Rodríguez Fernández, J; García Alonso, J I

    2013-03-01

    We have evaluated the use of (34)S-labelled yeast to perform sulphur metabolic tracer experiments in laboratory animals. The proof of principle work included the selection of the culture conditions for the preparation of sulphur labelled yeast, the study of the suitability of this labelled yeast as sulphur source for tracer studies using in vitro gastrointestinal digestion and the administration of the (34)S-labelled yeast to laboratory animals to follow the fate and distribution of (34)S in the organism. For in vitro gastrointestinal digestion, the combination of sodium dodecyl sulphate-polyacrylamide gel electrophoresis and high-performance liquid chromatography and inductively coupled plasma mass spectrometry (HPLC-ICP-MS) showed that labelled methionine, cysteine and other low molecular weight sulphur-containing biomolecules were the major components in the digested extracts of the labelled yeast. Next, in vivo kinetic experiments were performed in healthy Wistar rats after the oral administration of (34)S-labelled yeast. The isotopic composition of total sulphur in tissues, urine and faeces was measured by double-focusing inductively coupled plasma mass spectrometry after microwave digestion. It was observed that measurable isotopic enrichments were detected in all samples. Finally, initial investigations on sulphur isotopic composition of serum and urine samples by HPLC-ICP-MS have been carried out. For serum samples, no conclusive data were obtained. Interestingly, chromatographic analysis of urine samples showed differential isotope enrichment for several sulphur-containing biomolecules.

  11. Use of In-labeled autologous leukocytes to image an abdominal abscess in a horse

    SciTech Connect

    Koblik, P.D.; Lofstedt, J.; Jakowski, R.M.; Johnson, K.L.

    1985-06-15

    Indium 111-labeled autologous leukocytes were used to image an abdominal abscess in a horse with a palpable abdominal mass and history of Streptococcus equi infection. A focal area of radioactivity was identified in the location corresponding to the abscess. Imaging of this focal uptake was optimal 48 hours after injection. Similar scans obtained in 2 clinically normal horses revealed no evidence of focal radioactivity in this region. The cell labeling procedure gave acceptable labeling efficiency (87.5%) but an excessive number of damaged WBC, resulting in persistent lung radioactivity on all images. No adverse effects were noted. Radiation measured in the horse and its excreta were well within acceptable limits.

  12. Gender, status, and psychiatric labels.

    PubMed

    Kroska, Amy; Harkness, Sarah K; Brown, Ryan P; Thomas, Lauren S

    2015-11-01

    We examine a key modified labeling theory proposition-that a psychiatric label increases vulnerability to competence-based criticism and rejection-within task- and collectively oriented dyads comprised of same-sex individuals with equivalent education. Drawing on empirical work that approximates these conditions, we expect the proposition to hold only among men. We also expect education, operationalized with college class standing, to moderate the effects of gender by reducing men's and increasing women's criticism and rejection. But, we also expect the effect of education to weaken when men work with a psychiatric patient. As predicted, men reject suggestions from teammates with a psychiatric history more frequently than they reject suggestions from other teammates, while women's resistance to influence is unaffected by their teammate's psychiatric status. Men also rate psychiatric patient teammates as less powerful but no lower in status than other teammates, while women's teammate assessments are unaffected by their teammate's psychiatric status. Also as predicted, education reduces men's resistance to influence when their teammate has no psychiatric history. Education also increases men's ratings of their teammate's power, as predicted, but has no effect on women's resistance to influence or teammate ratings. We discuss the implications of these findings for the modified labeling theory of mental illness and status characteristics theory.

  13. 27 CFR 19.642 - Statements required on labels under an exemption from label approval.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... meaning given, and be stated in the manner provided in 27 CFR part 5. (Sec. 201, Pub. L. 85-859, 72 Stat... labels under an exemption from label approval. 19.642 Section 19.642 Alcohol, Tobacco Products and... PLANTS Liquor Bottle and Label Requirements Bottle Label Requirements § 19.642 Statements required...

  14. Abandoning a label doesn’t make it disappear: The perseverance of labeling effects

    PubMed Central

    Foroni, Francesco; Rothbart, Myron

    2012-01-01

    Labels exert strong influence on perception and judgment. The present experiment examines the possibility that such effects may persist even when labels are abandoned. Participants judged the similarity of pairs of silhouette drawings of female body types, ordered on a continuum from very thin to very heavy, under conditions where category labels were, and were not, superimposed on the ordered stimuli. Consistent with earlier research, labels had strong effects on perceived similarity, with silhouettes sharing the same label judged as more similar than those having different labels. Moreover, when the labels were removed and no longer present, the effect of the labels, although diminished, persisted. It did not make any difference whether the labels were simply abandoned or, in addition, had their validity challenged. The results are important for our understanding of categorization and labeling processes. The potential theoretical and practical implications of these results for social processes are discussed. PMID:23105148

  15. Stigma of a label: educational expectations for high school students labeled with learning disabilities.

    PubMed

    Shifrer, Dara

    2013-01-01

    Poorer outcomes for youth labeled with learning disabilities (LDs) are often attributed to the student's own deficiencies or cumulative disadvantage; but the more troubling possibility is that special education placement limits rather than expands these students' opportunities. Labeling theory partially attributes the poorer outcomes of labeled persons to stigma related to labels. This study uses data on approximately 11,740 adolescents and their schools from the Education Longitudinal Survey of 2002 to determine if stigma influences teachers' and parents' educational expectations for students labeled with LDs and labeled adolescents' expectations for themselves. Supporting the predictions of labeling theory, teachers and parents are more likely to perceive disabilities in, and hold lower educational expectations for labeled adolescents than for similarly achieving and behaving adolescents not labeled with disabilities. The negative effect of being labeled with LDs on adolescents' educational expectations is partially mechanized through parents' and particularly teachers' lower expectations.

  16. Stigma of a label: educational expectations for high school students labeled with learning disabilities.

    PubMed

    Shifrer, Dara

    2013-01-01

    Poorer outcomes for youth labeled with learning disabilities (LDs) are often attributed to the student's own deficiencies or cumulative disadvantage; but the more troubling possibility is that special education placement limits rather than expands these students' opportunities. Labeling theory partially attributes the poorer outcomes of labeled persons to stigma related to labels. This study uses data on approximately 11,740 adolescents and their schools from the Education Longitudinal Survey of 2002 to determine if stigma influences teachers' and parents' educational expectations for students labeled with LDs and labeled adolescents' expectations for themselves. Supporting the predictions of labeling theory, teachers and parents are more likely to perceive disabilities in, and hold lower educational expectations for labeled adolescents than for similarly achieving and behaving adolescents not labeled with disabilities. The negative effect of being labeled with LDs on adolescents' educational expectations is partially mechanized through parents' and particularly teachers' lower expectations. PMID:24311756

  17. Mass loss

    NASA Technical Reports Server (NTRS)

    Goldberg, Leo

    1987-01-01

    Observational evidence for mass loss from cool stars is reviewed. Spectra line profiles are used for the derivation of mass-loss rates with the aid of the equation of continuity. This equation implies steady mass loss with spherical symmetry. Data from binary stars, Mira variables, and red giants in globular clusters are examined. Silicate emission is discussed as a useful indicator of mass loss in the middle infrared spectra. The use of thermal millimeter-wave radiation, Very Large Array (VLA) measurement of radio emission, and OH/IR masers are discussed as a tool for mass loss measurement. Evidence for nonsteady mass loss is also reviewed.

  18. Use of Symbols in Labeling. Final rule.

    PubMed

    2016-06-15

    The Food and Drug Administration (FDA or the Agency) is issuing this final rule revising its medical device and certain biological product labeling regulations to explicitly allow for the optional inclusion of graphical representations of information, or symbols, in labeling (including labels) without adjacent explanatory text (referred to in this document as "stand-alone symbols") if certain requirements are met. The final rule also specifies that the use of symbols, accompanied by adjacent explanatory text continues to be permitted. FDA is also revising its prescription device labeling regulations to allow the use of the symbol statement "Rx only" or "[rx] only" in the labeling for prescription devices.

  19. Use of Symbols in Labeling. Final rule.

    PubMed

    2016-06-15

    The Food and Drug Administration (FDA or the Agency) is issuing this final rule revising its medical device and certain biological product labeling regulations to explicitly allow for the optional inclusion of graphical representations of information, or symbols, in labeling (including labels) without adjacent explanatory text (referred to in this document as "stand-alone symbols") if certain requirements are met. The final rule also specifies that the use of symbols, accompanied by adjacent explanatory text continues to be permitted. FDA is also revising its prescription device labeling regulations to allow the use of the symbol statement "Rx only" or "[rx] only" in the labeling for prescription devices. PMID:27311137

  20. Label-Free Identification and Quantification of SUMO Target Proteins.

    PubMed

    Hendriks, Ivo A; Vertegaal, Alfred C O

    2016-01-01

    Mass spectrometry-based approaches are utilized with increasing frequency to facilitate identification of novel SUMO target proteins and to elucidate the dynamics of SUMOylation in response to cellular stresses. Here, we describe a robust method for the identification of SUMO target proteins, and the relative quantification of SUMOylation dynamics, using a label-free approach. The method relies on a decahistidine (His10)-tagged SUMO, which is expressed at a low level in a mammalian cell line or model organism. The His10-tag allows for a single-step, high-yield, and high-purity enrichment of SUMOylated proteins, which are then digested and analyzed by high-resolution mass spectrometry. Matching between runs and label-free quantification integrated in the freely available MaxQuant software allow for a high rate and accuracy of quantification, providing a strong alternative to laborious sample or cell labeling techniques. The method described here allows for identification of >1000 SUMO target proteins, and characterization of their SUMOylation dynamics, without requiring sample fractionation. The purification procedure, starting from total lysate, can be performed in ~4 days. PMID:27631806

  1. In Vitro Metabolic Labeling of Intestinal Microbiota for Quantitative Metaproteomics.

    PubMed

    Zhang, Xu; Ning, Zhibin; Mayne, Janice; Deeke, Shelley A; Li, Jennifer; Starr, Amanda E; Chen, Rui; Singleton, Ruth; Butcher, James; Mack, David R; Stintzi, Alain; Figeys, Daniel

    2016-06-21

    Intestinal microbiota is emerging as one of the key environmental factors influencing or causing the development of numerous human diseases. Metaproteomics can provide invaluable information on the functional activities of intestinal microbiota and on host-microbe interactions as well. However, the application of metaproteomics in human microbiota studies is still largely limited, in part due to the lack of accurate quantitative intestinal metaproteomic methods. Most current metaproteomic microbiota studies are based on label-free quantification, which may suffer from variability during the separate sample processing and mass spectrometry runs. In this study, we describe a quantitative metaproteomic strategy, using in vitro stable isotopically ((15)N) labeled microbiota as a spike-in reference, to study the intestinal metaproteomes. We showed that the human microbiota were efficiently labeled (>95% (15)N enrichment) within 3 days under in vitro conditions, and accurate light-to-heavy protein/peptide ratio measurements were obtained using a high-resolution mass spectrometer and the quantitative proteomic software tool Census. We subsequently employed our approach to study the in vitro modulating effects of fructo-oligosaccharide and five different monosaccharides on the microbiota. Our methodology improves the accuracy of quantitative intestinal metaproteomics, which would promote the application of proteomics for functional studies of intestinal microbiota. PMID:27248155

  2. 46 CFR 160.133-17 - Marking and labeling.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... Marking and labeling. (a) Each hook body of a release mechanism must be marked with a plate or label...) The plate or label must be in English, but may also be in other languages. (c) The plate or label...

  3. 27 CFR 4.32 - Mandatory label information.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... October 6, 1984. (d) (e) Declaration of sulfites. There shall be stated on a front label, back label, strip label or neck label, the statement “Contains sulfites” or “Contains (a) sulfiting agent(s)” or...

  4. Abdominal mass

    MedlinePlus

    Mass in the abdomen ... care provider make a diagnosis. For example, the abdomen can be divided into four areas: Right-upper ... pain or masses include: Epigastric -- center of the abdomen just below the rib cage Periumbilical -- area around ...

  5. Identification of RNA sequence isomer by isotope labeling and LC-MS/MS.

    PubMed

    Li, Siwei; Limbach, Patrick A

    2014-11-01

    Recently, we developed a method for modified ribonucleic acid (RNA) analysis based on the comparative analysis of RNA digests (CARD). Within this CARD approach, sequence or modification differences between two samples are identified through differential isotopic labeling of two samples. Components present in both samples will each be labeled, yielding doublets in the CARD mass spectrum. Components unique to only one sample should be detected as singlets. A limitation of the prior singlet identification strategy occurs when the two samples contain components of unique sequence but identical base composition. At the first stage of mass spectrometry, these sequence isomers cannot be differentiated and would appear as doublets rather than singlets. However, underlying sequence differences should be detectable by collision-induced dissociation tandem mass spectrometry (CID MS/MS), as y-type product ions will retain the original enzymatically incorporated isotope label. Here, we determine appropriate instrumental conditions that enable CID MS/MS of isotopically labeled ribonuclease T1 (RNase T1) digestion products such that the original isotope label is maintained in the product ion mass spectrum. Next, we demonstrate how y-type product ions can be used to differentiate singlets and doublets from isomer sequences. We were then able to extend the utility of this approach by using CID MS/MS for the confirmation of an expected RNase T1 digestion product within the CARD analysis of an Escherichia coli mutant strain even in the presence of interfering and overlapping digestion products from other transfer RNAs.

  6. Accelerator mass spectrometry as a bioanalytical tool for nutritional research

    SciTech Connect

    Vogel, J.S.; Turteltaub, K.W.

    1997-09-01

    Accelerator Mass Spectrometry is a mass spectrometric method of detecting long-lived radioisotopes without regard to their decay products or half-life. The technique is normally applied to geochronology, but recently has been developed for bioanalytical tracing. AMS detects isotope concentrations to parts per quadrillion, quantifying labeled biochemicals to attomole levels in milligram- sized samples. Its advantages over non-isotopeic and stable isotope labeling methods are reviewed and examples of analytical integrity, sensitivity, specificity, and applicability are provided.

  7. Quantifying plant phenotypes with isotopic labeling & metabolic flux analysis.

    PubMed

    Allen, Doug K

    2016-02-01

    Analyses of metabolic flux using stable isotopes in plants have traditionally been restricted to tissues with presumed homogeneous cell populations and long metabolic steady states such as developing seeds, cell suspensions, or cultured roots and root tips. It is now possible to describe these and other metabolically more dynamic tissues such as leaves in greater detail using novel methods in mass spectrometry, isotope labeling strategies, and transient labeling-based flux analyses. Such studies are necessary for a systems level description of plant function that more closely represents biological reality, and provides insights into the genes that will need to be modified as natural resources become ever more limited and environments change. PMID:26613198

  8. Hemoglobin Labeled by Radioactive Lysine

    DOE R&D Accomplishments Database

    Bale, W. F.; Yuile, C. L.; DeLaVergne, L.; Miller, L. L.; Whipple, G. H.

    1949-12-08

    This paper reports on the utilization of tagged epsilon carbon of DL-lysine by a dog both anemic and hypoproteinemic due to repeated bleeding plus a diet low in protein. The experiment extended over period of 234 days, a time sufficient to indicate an erythrocyte life span of at least 115 days based upon the rate of replacement of labeled red cell proteins. The proteins of broken down red cells seem not to be used with any great preference for the synthesis of new hemoglobin.

  9. Label-free cell profiling.

    PubMed

    Schasfoort, Richard B M; Bentlage, Arthur E H; Stojanovic, Ivan; van der Kooi, Alex; van der Schoot, Ellen; Terstappen, Leon W M M; Vidarsson, Gestur

    2013-08-01

    A surface plasmon resonance (SPR) array imaging method is outlined for label-free cell profiling. Red blood cells (RBCs) were injected into a flow chamber on top of a spotted sensor surface. Spots contained antibodies to various RBC membrane antigens. A typical sensorgram showed an initial response corresponding to cell sedimentation (S) followed by a specific upward response (T) corresponding to specific binding of cells during a critical wash step. The full analysis cycle for RBC profiling was less than 6 min. The sensor surface could be regenerated at least 100 times, allowing the determination of a cell surface antigen profile of RBCs.

  10. Particle-based labeling: Fast point-feature labeling without obscuring other visual features.

    PubMed

    Luboschik, Martin; Schumann, Heidrun; Cords, Hilko

    2008-01-01

    In many information visualization techniques, labels are an essential part to communicate the visualized data. To preserve the expressiveness of the visual representation, a placed label should neither occlude other labels nor visual representatives (e.g., icons, lines) that communicate crucial information. Optimal, non-overlapping labeling is an NP-hard problem. Thus, only a few approaches achieve a fast non-overlapping labeling in highly interactive scenarios like information visualization. These approaches generally target the point-feature label placement (PFLP) problem, solving only label-label conflicts. This paper presents a new, fast, solid and flexible 2D labeling approach for the PFLP problem that additionally respects other visual elements and the visual extent of labeled features. The results (number of placed labels, processing time) of our particle-based method compare favorably to those of existing techniques. Although the esthetic quality of non-real-time approaches may not be achieved with our method, it complies with practical demands and thus supports the interactive exploration of information spaces. In contrast to the known adjacent techniques, the flexibility of our technique enables labeling of dense point clouds by the use of non-occluding distant labels. Our approach is independent of the underlying visualization technique, which enables us to demonstrate the application of our labeling method within different information visualization scenarios.

  11. Enzyme-gold affinity labelling of cellulose.

    PubMed

    Berg, R H; Erdos, G W; Gritzali, M; Brown, R D

    1988-04-01

    The enzyme-linked colloidal gold affinity labelling technique was tested as a method to localize cellulose on thin sections of plant cell walls and slime mold spores. Commercially available cellulase from cultures of Trichoderma reesei, the main components being cellobiohydrolase I and II (CBH I, CBH II) and endoglucanase (EG), was linked to colloidal gold by using standard techniques and applied as a dilute, buffered suspension to thin sections. After brief exposure, e.g., 15-30 minutes, cellulose exposed on the surface of sections was labelled with the enzyme-gold complex. Poststaining did not appear to have a deleterious effect on the labelled sections. The specificity of labelling was demonstrated by its complete inhibition when carboxymethylcellulose was incorporated in the labelling mixture, by lack of labelling of 1,4-beta-mannans or 1,3-beta-xylans in noncellulosic walls of marine algae, by lack of labelling of 1,4-beta-glucans in chitin, by much lower labelling density when done at 4 degrees C, and by lack of labelling when sections were predigested with cellulase. Labelling with the crude commercial cellulase was compared to labelling with purified CBH I-, CBH II-, and EG-linked colloidal gold, and the labelling pattern was similar. This method was found useful on conventionally fixed material and required no special preparation other than the use of inert (Ni or Au) grids and 0.5% gelatin to reduce nonspecific binding of the gold complex. Labelling was similar in the several embedding resins tested: LR White, Lowicryl K4M, Epon 812, and Spurr's.(ABSTRACT TRUNCATED AT 250 WORDS)

  12. Label-free photoacoustic nanoscopy

    PubMed Central

    Danielli, Amos; Maslov, Konstantin; Garcia-Uribe, Alejandro; Winkler, Amy M.; Li, Chiye; Wang, Lidai; Chen, Yun; Dorn, Gerald W.; Wang, Lihong V.

    2014-01-01

    Abstract. Super-resolution microscopy techniques—capable of overcoming the diffraction limit of light—have opened new opportunities to explore subcellular structures and dynamics not resolvable in conventional far-field microscopy. However, relying on staining with exogenous fluorescent markers, these techniques can sometimes introduce undesired artifacts to the image, mainly due to large tagging agent sizes and insufficient or variable labeling densities. By contrast, the use of endogenous pigments allows imaging of the intrinsic structures of biological samples with unaltered molecular constituents. Here, we report label-free photoacoustic (PA) nanoscopy, which is exquisitely sensitive to optical absorption, with an 88 nm resolution. At each scanning position, multiple PA signals are successively excited with increasing laser pulse energy. Because of optical saturation or nonlinear thermal expansion, the PA amplitude depends on the nonlinear incident optical fluence. The high-order dependence, quantified by polynomial fitting, provides super-resolution imaging with optical sectioning. PA nanoscopy is capable of super-resolution imaging of either fluorescent or nonfluorescent molecules. PMID:25104412

  13. Chemical kin label in seabirds.

    PubMed

    Célérier, Aurélie; Bon, Cécile; Malapert, Aurore; Palmas, Pauline; Bonadonna, Francesco

    2011-12-23

    Chemical signals yield critical socio-ecological information in many animals, such as species, identity, social status or sex, but have been poorly investigated in birds. Recent results showed that chemical signals are used to recognize their nest and partner by some petrel seabirds whose olfactory anatomy is well developed and which possess a life-history propitious to olfactory-mediated behaviours. Here, we investigate whether blue petrels (Halobaena caerulea) produce some chemical labels potentially involved in kin recognition and inbreeding avoidance. To overcome methodological constraints of chemical analysis and field behavioural experiments, we used an indirect behavioural approach, based on mice olfactory abilities in discriminating odours. We showed that mice (i) can detect odour differences between individual petrels, (ii) perceive a high odour similarity between a chick and its parents, and (iii) perceive this similarity only before fledging but not during the nestling developmental stage. Our results confirm the existence of an individual olfactory signature in blue petrels and show for the first time, to our knowledge, that birds may exhibit an olfactory kin label, which may have strong implications for inbreeding avoidance. PMID:21525047

  14. Inulin determination for food labeling.

    PubMed

    Zuleta, A; Sambucetti, M E

    2001-10-01

    Inulin and oligofructose exhibit valuable nutritional and functional attributes, so they are used as supplements as soluble fiber or as macronutrient substitutes. As classic analytical methods for dietary fiber measurement are not effective, several specific methods have been proposed. These methods measure total fructans and are based on one or more enzymatic sample treatments and determination of released sugars. To determine inulin for labeling purposes, we developed an easy and rapid anion-exchange high-performance liquid chromatography (HPLC) method following water extraction of inulin. HPLC conditions included an Aminex HPX- 87C column (Bio-Rad), deionized water at 85 degrees C as the mobile phase and a refractive index detector. The tested foods included tailor-made food products containing known amounts of inulin and commercial products (cookies, milk, ice creams, cheese, and cereal bars). The average recovery was 97%, and the coefficient of variation ranged from 1.1 to 5% in the food matrixes. The obtained results showed that this method provides an easier, faster and cheaper alternative than previous techniques for determining inulin with enough accuracy and precision for routine labeling purposes by direct determination of inulin by HPLC with refractive index detection. PMID:11599989

  15. Label-free photoacoustic nanoscopy.

    PubMed

    Danielli, Amos; Maslov, Konstantin; Garcia-Uribe, Alejandro; Winkler, Amy M; Li, Chiye; Wang, Lidai; Chen, Yun; Dorn, Gerald W; Wang, Lihong V

    2014-08-01

    Super-resolution microscopy techniques - capable of overcoming the diffraction limit of light - have opened new opportunities to explore subcellular structures and dynamics not resolvable in conventional far-field microscopy. However, relying on staining with exogenous fluorescent markers, these techniques can sometimes introduce undesired artifacts to the image, mainly due to large tagging agent sizes and insufficient or variable labeling densities. By contrast, the use of endogenous pigments allows imaging of the intrinsic structures of biological samples with unaltered molecular constituents. Here, we report label-free photoacoustic (PA) nanoscopy, which is exquisitely sensitive to optical absorption, with an 88 nm resolution. At each scanning position, multiple PA signals are successively excited with increasing laser pulse energy. Because of optical saturation or nonlinear thermal expansion, the PA amplitude depends on the nonlinear incident optical fluence. The high-order dependence, quantified by polynomial fitting, provides super-resolution imaging with optical sectioning. PA nanoscopy is capable of super-resolution imaging of either fluorescent or nonfluorescent molecules.

  16. Novel approach for labeling of biopolymers with DOTA complexes using in situ click chemistry for quantification.

    PubMed

    He, Yide; Esteban-Fernández, Diego; Linscheid, Michael W

    2015-03-01

    In this work, we present a two-step labeling approach for the efficient tagging with lanthanide-containing complexes. For this purpose, derivatization of the cysteine residues with an alkyne group acting as linker was done before the DOTA complex was introduced using in situ click chemistry. The characterization of this new methodology is presented including the optimization of the labeling process, demonstration of the quantitative capabilities using both electrospray ionization mass spectrometry (ESI-MS) and inductively coupled plasma mass spectrometry (ICP-MS) detection, and study of the fragmentation behavior of the labeled peptides by collision-induced dissociation (CID) for identification purposes. The results show that, in terms of labeling efficiency, this new methodology improves previously developed DOTA-based label strategies, such as MeCAT-maleimide (metal-coded affinity tag, MeCAT-Mal) and MeCAT-iodoacetamide (MeCAT-IA) reagents. The goal of reducing the steric hindrance caused by the voluminous DOTA complex was fulfilled allowing both, quantification and identification of labeled biopolymers.

  17. Neutrino mass

    SciTech Connect

    Bowles, T.J.

    1994-04-01

    The existence of a finite neutrino mass would have important consequences in particle physics, astrophysics, and cosmology. Experimental sensitivities have continued to be pushed down without any confirmed evidence for a finite neutrino mass. Yet there are several observations of discrepancies between theoretical predictions and observations which might be possible indications of a finite neutrino mass. Thus, extensive theoretical and experimental work is underway to resolve these issues.

  18. 21 CFR 820.120 - Device labeling.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... accuracy including, where applicable, the correct unique device identifier (UDI) or universal product code... manner that provides proper identification and is designed to prevent mixups. (d) Labeling...

  19. Simultaneous segmentation and statistical label fusion

    NASA Astrophysics Data System (ADS)

    Asman, Andrew J.; Landman, Bennett A.

    2012-02-01

    Labeling or segmentation of structures of interest in medical imaging plays an essential role in both clinical and scientific understanding. Two of the common techniques to obtain these labels are through either fully automated segmentation or through multi-atlas based segmentation and label fusion. Fully automated techniques often result in highly accurate segmentations but lack the robustness to be viable in many cases. On the other hand, label fusion techniques are often extremely robust, but lack the accuracy of automated algorithms for specific classes of problems. Herein, we propose to perform simultaneous automated segmentation and statistical label fusion through the reformulation of a generative model to include a linkage structure that explicitly estimates the complex global relationships between labels and intensities. These relationships are inferred from the atlas labels and intensities and applied to the target using a non-parametric approach. The novelty of this approach lies in the combination of previously exclusive techniques and attempts to combine the accuracy benefits of automated segmentation with the robustness of a multi-atlas based approach. The accuracy benefits of this simultaneous approach are assessed using a multi-label multi-atlas whole-brain segmentation experiment and the segmentation of the highly variable thyroid on computed tomography images. The results demonstrate that this technique has major benefits for certain types of problems and has the potential to provide a paradigm shift in which the lines between statistical label fusion and automated segmentation are dramatically blurred.

  20. MASS SPECTROMETER

    DOEpatents

    White, F.A.

    1960-08-23

    A mass spectrometer is designed with a first adjustable magnetic field for resolving an ion beam into beams of selected masses, a second adjustable magnetic field for further resolving the ion beam from the first field into beams of selected masses, a thin foil disposed in the path of the beam between the first and second magnets to dissociate molecular ions incident thereon, an electrostatic field for further resolving the ion beam from the second field into beams of selected masses, and a detector disposed adjacent to the electrostatic field to receive the ion beam.

  1. 21 CFR 1302.04 - Location and size of symbol on label and labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... AND PACKAGING REQUIREMENTS FOR CONTROLLED SUBSTANCES § 1302.04 Location and size of symbol on label... substance. The symbol on labels shall be clear and large enough to afford easy identification of...

  2. Labeled nucleotide phosphate (NP) probes

    SciTech Connect

    Korlach, Jonas; Webb, Watt W.; Levene, Michael; Turner, Stephen; Craighead, Harold G.; Foquet, Mathieu

    2009-02-03

    The present invention is directed to a method of sequencing a target nucleic acid molecule having a plurality of bases. In its principle, the temporal order of base additions during the polymerization reaction is measured on a molecule of nucleic acid, i.e. the activity of a nucleic acid polymerizing enzyme on the template nucleic acid molecule to be sequenced is followed in real time. The sequence is deduced by identifying which base is being incorporated into the growing complementary strand of the target nucleic acid by the catalytic activity of the nucleic acid polymerizing enzyme at each step in the sequence of base additions. A polymerase on the target nucleic acid molecule complex is provided in a position suitable to move along the target nucleic acid molecule and extend the oligonucleotide primer at an active site. A plurality of labelled types of nucleotide analogs are provided proximate to the active site, with each distinguishable type of nucleotide analog being complementary to a different nucleotide in the target nucleic acid sequence. The growing nucleic acid strand is extended by using the polymerase to add a nucleotide analog to the nucleic acid strand at the active site, where the nucleotide analog being added is complementary to the nucleotide of the target nucleic acid at the active site. The nucleotide analog added to the oligonucleotide primer as a result of the polymerizing step is identified. The steps of providing labelled nucleotide analogs, polymerizing the growing nucleic acid strand, and identifying the added nucleotide analog are repeated so that the nucleic acid strand is further extended and the sequence of the target nucleic acid is determined.

  3. F-18 labeled 3-fluorodiazepam

    SciTech Connect

    Luxen, A.; Barrio, J.R.; Bida, G.T.; Satyamurthy, N.; Phelps, M.E.

    1985-05-01

    3-Fluorodiazepam is a new and potent antianxiety agent with prolonged action. The authors found that molecular fluorine (0.5% in Ne) reacts cleanly with diazepam in freon or chloroform at room temperature to produce 3-fluorodiazepam in good yields. Successful syntheses have employed 2:1 to 5:1 molar ratios diazepam: fluorine to minimize the formation of byproducts. (/sup 18/F) 3-Fluorodiazepam, a potential candidate for PET studies, (specific activity 3-5 Ci/mmol) has been synthesized from /sup 18/F-F/sub 2/ using the same procedure, followed by column chromatographic purification (Silicagel, dichloromethane: ethyl acetate, 5:1) with a radiochemical yield of 12-20% (50% maximum) and a chemical and radiochemical purity >99% as judged by reversed-phase high pressure liquid chromatography analysis (Ultrasyl octyl column, 10 ..mu.. m, 4.6 x 250 mm i.d., 60% MeOH 40% water; flow rate, 1.0 ml/min; retention time for (/sup 18/F) fluorodiazepam, 11.4 min; for diazepam, 13.5 min; radioactivity and ultraviolet detectors). Lower radiochemical yields (5-7%), and significant formation of by-products were observed when (/sup 18/F)acetylhypofluorite, prepared in the gasphase, was used as the reagent. Readily accessible routes to /sup 18/F-labeled benzodiazepines of higher specific activity were also investigated. Approaches to the synthesis of high specific activity (>200 Ci/mmol) (/sup 18/F)3-fluorodiazepam involve nucleophilic displacement at carbon-3 (e.g. from 3-chlorodiazepam) with (/sup 18/F)fluoride ion. The results presented here demonstrate the synthetic accessibility of /sup 18/F-labeled benzodiazepines for application in neurotransmitter ligand studies with PET.

  4. Diagnostic enteral gastrointestinal radiopaque human prescription drugs class labeling guideline for professional labeling. Final report

    SciTech Connect

    Chun, M.Y.

    1982-09-21

    The Food and Drug Administration (FDA) announces the availability of a class labeling guideline for the professional labeling of diagnostic enteral gastrointestinal radiopaque agents. Class labeling is appropriate for these products because they are all closely related in chemical structure, pharmacology, therapeutic activity, and adverse reactions. The guideline is intended to promote the use of identical professional labeling for each member of the diagnostic enteral gastrointestinal radiopaque drug class.

  5. Inertial Mass

    ERIC Educational Resources Information Center

    King, Kenneth P.

    2007-01-01

    The inertial balance is one device that can help students to quantify the quality of inertia--a body's resistance to a change in movement--in more generally understood terms of mass. In this hands-on activity, students use the inertial balance to develop a more quantitative idea of what mass means in an inertial sense. The activity also helps…

  6. The reappropriation of stigmatizing labels: the reciprocal relationship between power and self-labeling.

    PubMed

    Galinsky, Adam D; Wang, Cynthia S; Whitson, Jennifer A; Anicich, Eric M; Hugenberg, Kurt; Bodenhausen, Galen V

    2013-10-01

    We present a theoretical model of reappropriation--taking possession of a slur previously used exclusively by dominant groups to reinforce another group's lesser status. Ten experiments tested this model and established a reciprocal relationship between power and self-labeling with a derogatory group term. We first investigated precursors to self-labeling: Group, but not individual, power increased participants' willingness to label themselves with a derogatory term for their group. We then examined the consequences of such self-labeling for both the self and observers. Self-labelers felt more powerful after self-labeling, and observers perceived them and their group as more powerful. Finally, these labels were evaluated less negatively after self-labeling, and this attenuation of stigma was mediated by perceived power. These effects occurred only for derogatory terms (e.g., queer, bitch), and not for descriptive (e.g., woman) or majority-group (e.g., straight) labels. These results suggest that self-labeling with a derogatory label can weaken the label's stigmatizing force.

  7. 76 FR 46671 - Food Labeling; Gluten-Free Labeling of Foods; Reopening of the Comment Period

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-08-03

    ... could use to establish a gluten threshold level to define the food labeling term ``gluten-free'' (72 FR... HUMAN SERVICES Food and Drug Administration 21 CFR Part 101 RIN 0910-ZA26 Food Labeling; Gluten-Free Labeling of Foods; Reopening of the Comment Period AGENCY: Food and Drug Administration, HHS....

  8. Portion Size Labeling and Intended Soft Drink Consumption: The Impact of Labeling Format and Size Portfolio

    ERIC Educational Resources Information Center

    Vermeer, Willemijn M.; Steenhuis, Ingrid H. M.; Leeuwis, Franca H.; Bos, Arjan E. R.; de Boer, Michiel; Seidell, Jacob C.

    2010-01-01

    Objective: To assess what portion size labeling "format" is most promising in helping consumers selecting appropriate soft drink sizes, and whether labeling impact depends on the size portfolio. Methods: An experimental study was conducted in fast-food restaurants in which 2 labeling formats (ie, reference portion size and small/medium/large…

  9. 21 CFR 660.28 - Labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... antibody or antibodies present as set forth in paragraph (d) of this section. (ii) Name, address (including... antibody designation on the labels of a final container with a capacity of less than 5 milliliters shall be not less than 12 point. The type size for the specificity of the antibody designations on the label...

  10. 40 CFR 1033.135 - Labeling.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR POLLUTION CONTROLS CONTROL OF EMISSIONS FROM LOCOMOTIVES Emission Standards and Related Requirements § 1033.135 Labeling. As described in... information: (A) The label heading: “ORIGINAL LOCOMOTIVE EMISSION CONTROL INFORMATION.”...

  11. 40 CFR 1033.135 - Labeling.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR POLLUTION CONTROLS CONTROL OF EMISSIONS FROM LOCOMOTIVES Emission Standards and Related Requirements § 1033.135 Labeling. As described in... information: (A) The label heading: “ORIGINAL LOCOMOTIVE EMISSION CONTROL INFORMATION.”...

  12. 40 CFR 1033.135 - Labeling.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR POLLUTION CONTROLS CONTROL OF EMISSIONS FROM LOCOMOTIVES Emission Standards and Related Requirements § 1033.135 Labeling. As described in... information: (A) The label heading: “ORIGINAL LOCOMOTIVE EMISSION CONTROL INFORMATION.”...

  13. 21 CFR 201.313 - Estradiol labeling.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... Pharmacopeia under the designation “Alpha Estradiol.” The substance should no longer be referred to in drug labeling as “Alpha Estradiol.” The Food and Drug Administration would not object to label references to the... referred to the presence of “Estradiol (formerly known as Alpha Estradiol).”...

  14. 21 CFR 201.313 - Estradiol labeling.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... Pharmacopeia under the designation “Alpha Estradiol.” The substance should no longer be referred to in drug labeling as “Alpha Estradiol.” The Food and Drug Administration would not object to label references to the... referred to the presence of “Estradiol (formerly known as Alpha Estradiol).”...

  15. 21 CFR 201.313 - Estradiol labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... Pharmacopeia under the designation “Alpha Estradiol.” The substance should no longer be referred to in drug labeling as “Alpha Estradiol.” The Food and Drug Administration would not object to label references to the... referred to the presence of “Estradiol (formerly known as Alpha Estradiol).”...

  16. 21 CFR 201.313 - Estradiol labeling.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... Pharmacopeia under the designation “Alpha Estradiol.” The substance should no longer be referred to in drug labeling as “Alpha Estradiol.” The Food and Drug Administration would not object to label references to the... referred to the presence of “Estradiol (formerly known as Alpha Estradiol).”...

  17. 21 CFR 201.313 - Estradiol labeling.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... Pharmacopeia under the designation “Alpha Estradiol.” The substance should no longer be referred to in drug labeling as “Alpha Estradiol.” The Food and Drug Administration would not object to label references to the... referred to the presence of “Estradiol (formerly known as Alpha Estradiol).”...

  18. 40 CFR 204.55-4 - Labeling.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... label: (i) The label heading: Compressor Noise Emission Control Information; (ii) Full corporate name... following acts or the causing thereof by any person are prohibited by the Noise Control Act of 1972: (A) The... any noise control device or element of design (listed in the owner's manual) incorporated into...

  19. 40 CFR 204.55-4 - Labeling.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... label: (i) The label heading: Compressor Noise Emission Control Information; (ii) Full corporate name... following acts or the causing thereof by any person are prohibited by the Noise Control Act of 1972: (A) The... any noise control device or element of design (listed in the owner's manual) incorporated into...

  20. 40 CFR 204.55-4 - Labeling.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... label: (i) The label heading: Compressor Noise Emission Control Information; (ii) Full corporate name... following acts or the causing thereof by any person are prohibited by the Noise Control Act of 1972: (A) The... any noise control device or element of design (listed in the owner's manual) incorporated into...

  1. 10 CFR 20.1904 - Labeling containers.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 10 Energy 1 2010-01-01 2010-01-01 false Labeling containers. 20.1904 Section 20.1904 Energy....1904 Labeling containers. (a) The licensee shall ensure that each container of licensed material bears... handling or using the containers, or working in the vicinity of the containers, to take precautions...

  2. 21 CFR 640.70 - Labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Source Plasma § 640.70 Labeling. Link to an amendment published... information shall appear on the label affixed to each container of Source Plasma: (1) The proper name of the... shall follow the proper name in the same size and type of print as the proper name. If the Source...

  3. 42 CFR 84.257 - Labeling requirements.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... Labeling requirements. (a) A warning shall be placed on the label of each gas mask, chemical-cartridge... performance of any gas mask, chemical-cartridge respirator, or powered air-purifying respirator approved under... this subpart shall be specified as follows: Chemical-cartridge respirator 1 hour. Gas mask 4...

  4. 42 CFR 84.257 - Labeling requirements.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... Labeling requirements. (a) A warning shall be placed on the label of each gas mask, chemical-cartridge... performance of any gas mask, chemical-cartridge respirator, or powered air-purifying respirator approved under... this subpart shall be specified as follows: Chemical-cartridge respirator 1 hour. Gas mask 4...

  5. 42 CFR 84.257 - Labeling requirements.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... Labeling requirements. (a) A warning shall be placed on the label of each gas mask, chemical-cartridge... performance of any gas mask, chemical-cartridge respirator, or powered air-purifying respirator approved under... this subpart shall be specified as follows: Chemical-cartridge respirator 1 hour. Gas mask 4...

  6. 42 CFR 84.257 - Labeling requirements.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... Labeling requirements. (a) A warning shall be placed on the label of each gas mask, chemical-cartridge... performance of any gas mask, chemical-cartridge respirator, or powered air-purifying respirator approved under... this subpart shall be specified as follows: Chemical-cartridge respirator 1 hour. Gas mask 4...

  7. 42 CFR 84.257 - Labeling requirements.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... Labeling requirements. (a) A warning shall be placed on the label of each gas mask, chemical-cartridge... performance of any gas mask, chemical-cartridge respirator, or powered air-purifying respirator approved under... this subpart shall be specified as follows: Chemical-cartridge respirator 1 hour. Gas mask 4...

  8. 75 FR 67615 - Appliance Labeling Rule

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-11-03

    ... Appliance Labeling Rule (16 CFR Part 305) which appeared in the Federal Register on October 23, 2008 (73 FR... (comparability ranges for instantaneous gas water heaters) on August 29, 2007 (72 FR 49948). The correct capacity... Labeling Rule (16 CFR Part 305). This document republishes the text of Sec. 305.20(f) concerning...

  9. 19 CFR 12.18 - Labels.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 19 Customs Duties 1 2013-04-01 2013-04-01 false Labels. 12.18 Section 12.18 Customs Duties U.S. CUSTOMS AND BORDER PROTECTION, DEPARTMENT OF HOMELAND SECURITY; DEPARTMENT OF THE TREASURY SPECIAL CLASSES OF MERCHANDISE Viruses, Serums, and Toxins for Treatment of Domestic Animals § 12.18 Labels. Each separate container of such virus, serum,...

  10. 21 CFR 331.80 - Professional labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... HUMAN USE ANTACID PRODUCTS FOR OVER-THE-COUNTER (OTC) HUMAN USE Labeling § 331.80 Professional labeling.... (i) Prolonged use of aluminum-containing antacids in patients with renal failure may result in or... of aluminum-containing antacids by normophosphatemic patients may result in hypophosphatemia...

  11. 21 CFR 331.80 - Professional labeling.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... HUMAN USE ANTACID PRODUCTS FOR OVER-THE-COUNTER (OTC) HUMAN USE Labeling § 331.80 Professional labeling.... (i) Prolonged use of aluminum-containing antacids in patients with renal failure may result in or... of aluminum-containing antacids by normophosphatemic patients may result in hypophosphatemia...

  12. 21 CFR 331.80 - Professional labeling.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... HUMAN USE ANTACID PRODUCTS FOR OVER-THE-COUNTER (OTC) HUMAN USE Labeling § 331.80 Professional labeling.... (i) Prolonged use of aluminum-containing antacids in patients with renal failure may result in or... of aluminum-containing antacids by normophosphatemic patients may result in hypophosphatemia...

  13. 21 CFR 331.80 - Professional labeling.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... HUMAN USE ANTACID PRODUCTS FOR OVER-THE-COUNTER (OTC) HUMAN USE Labeling § 331.80 Professional labeling.... (i) Prolonged use of aluminum-containing antacids in patients with renal failure may result in or... of aluminum-containing antacids by normophosphatemic patients may result in hypophosphatemia...

  14. 16 CFR 1209.9 - Labeling requirement.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... SAFETY STANDARD FOR CELLULOSE INSULATION The Standard § 1209.9 Labeling requirement. (a) Manufacturers, importers, and private labelers of cellulose insulation shall place on all containers of cellulose... corrosiveness of cellulose insulation. To meet this requirement manufacturers, importers, and private...

  15. 16 CFR 1209.9 - Labeling requirement.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... SAFETY STANDARD FOR CELLULOSE INSULATION The Standard § 1209.9 Labeling requirement. (a) Manufacturers, importers, and private labelers of cellulose insulation shall place on all containers of cellulose... corrosiveness of cellulose insulation. To meet this requirement manufacturers, importers, and private...

  16. 16 CFR 1209.9 - Labeling requirement.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... SAFETY STANDARD FOR CELLULOSE INSULATION The Standard § 1209.9 Labeling requirement. (a) Manufacturers, importers, and private labelers of cellulose insulation shall place on all containers of cellulose... corrosiveness of cellulose insulation. To meet this requirement manufacturers, importers, and private...

  17. 16 CFR 1209.9 - Labeling requirement.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... SAFETY STANDARD FOR CELLULOSE INSULATION The Standard § 1209.9 Labeling requirement. (a) Manufacturers, importers, and private labelers of cellulose insulation shall place on all containers of cellulose... corrosiveness of cellulose insulation. To meet this requirement manufacturers, importers, and private...

  18. 21 CFR 701.11 - Identity labeling.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) COSMETICS COSMETIC LABELING Package Form § 701.11 Identity labeling. (a) The principal display panel of a cosmetic in...) Such statement of identity shall be in terms of: (1) The common or usual name of the cosmetic; or...

  19. 21 CFR 701.11 - Identity labeling.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) COSMETICS COSMETIC LABELING Package Form § 701.11 Identity labeling. (a) The principal display panel of a cosmetic in...) Such statement of identity shall be in terms of: (1) The common or usual name of the cosmetic; or...

  20. 21 CFR 701.11 - Identity labeling.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) COSMETICS COSMETIC LABELING Package Form § 701.11 Identity labeling. (a) The principal display panel of a cosmetic in...) Such statement of identity shall be in terms of: (1) The common or usual name of the cosmetic; or...

  1. 21 CFR 701.11 - Identity labeling.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) COSMETICS COSMETIC LABELING Package Form § 701.11 Identity labeling. (a) The principal display panel of a cosmetic in...) Such statement of identity shall be in terms of: (1) The common or usual name of the cosmetic; or...

  2. 9 CFR 112.3 - Diluent labels.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Diluent labels. 112.3 Section 112.3 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS PACKAGING AND LABELING § 112.3...

  3. 76 FR 45715 - Appliance Labeling Rule

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-08-01

    ... July 19, 2010 (75 FR 41696), the Commission published new light bulb \\1\\ labeling requirements and... authority in requiring labels for LED bulbs, reflector lamps, and three-way lamps. 75 FR 41696, 41698 (Jul...-watt incandescent bulbs, which represent about \\1/5\\ of the incandescent market. 76 FR 20233 (Apr....

  4. Influence of Food Labels on Adolescent Diet.

    ERIC Educational Resources Information Center

    Misra, Ranjita

    2002-01-01

    Provides information on food nutrition labels and discusses the benefits of adolescents' using them to plan healthy diets. Suggests that teachers and educators should encourage appropriate label reading education for adolescents to promote healthy eating practices. Provides definitions of nutrient content claims. (SG)

  5. 27 CFR 18.55 - Label.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2011-04-01 2011-04-01 false Label. 18.55 Section 18.55 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS PRODUCTION OF VOLATILE FRUIT-FLAVOR CONCENTRATE Operations § 18.55 Label. Each...

  6. 27 CFR 18.55 - Label.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Label. 18.55 Section 18.55 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS PRODUCTION OF VOLATILE FRUIT-FLAVOR CONCENTRATE Operations § 18.55 Label. Each...

  7. 78 FR 18272 - Energy Labeling Rule

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-03-26

    ... INFORMATION: The Commission is reopening the comment period for its January 9, 2013 (78 FR 1779) Notice of... CFR Part 305 Energy Labeling Rule AGENCY: Federal Trade Commission (``FTC'' or ``Commission''). ACTION... in the SUPPLEMENTARY INFORMATION section below. Write ``Energy Label Ranges, Matter No. R611004''...

  8. 16 CFR 1630.5 - Labeling.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 16 Commercial Practices 2 2010-01-01 2010-01-01 false Labeling. 1630.5 Section 1630.5 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE SURFACE FLAMMABILITY OF CARPETS AND RUGS (FF 1-70) The Standard § 1630.5 Labeling. If the carpet or rug has had a...

  9. 16 CFR 1630.5 - Labeling.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 16 Commercial Practices 2 2011-01-01 2011-01-01 false Labeling. 1630.5 Section 1630.5 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE SURFACE FLAMMABILITY OF CARPETS AND RUGS (FF 1-70) The Standard § 1630.5 Labeling. If the carpet or rug has had a...

  10. 16 CFR 1630.5 - Labeling.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 16 Commercial Practices 2 2012-01-01 2012-01-01 false Labeling. 1630.5 Section 1630.5 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE SURFACE FLAMMABILITY OF CARPETS AND RUGS (FF 1-70) The Standard § 1630.5 Labeling. If the carpet or rug has had a...

  11. 16 CFR 1630.5 - Labeling.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 16 Commercial Practices 2 2014-01-01 2014-01-01 false Labeling. 1630.5 Section 1630.5 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE SURFACE FLAMMABILITY OF CARPETS AND RUGS (FF 1-70) The Standard § 1630.5 Labeling. If the carpet or rug has had a...

  12. 16 CFR 1630.5 - Labeling.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 16 Commercial Practices 2 2013-01-01 2013-01-01 false Labeling. 1630.5 Section 1630.5 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE SURFACE FLAMMABILITY OF CARPETS AND RUGS (FF 1-70) The Standard § 1630.5 Labeling. If the carpet or rug has had a...

  13. 47 CFR 15.19 - Labelling requirements.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... specified under paragraph (b)(1) of this section on it, such as for a CPU board or a plug-in circuit board...) and the logo must be displayed on the device. (4) The label shall not be a stick-on, paper label. The... 90, etc., shall bear the following statement in a conspicuous location on the device: This...

  14. 47 CFR 15.19 - Labelling requirements.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... specified under paragraph (b)(1) of this section on it, such as for a CPU board or a plug-in circuit board...) and the logo must be displayed on the device. (4) The label shall not be a stick-on, paper label. The... 90, etc., shall bear the following statement in a conspicuous location on the device: This...

  15. Linguistic Labels: Conceptual Markers or Object Features?

    ERIC Educational Resources Information Center

    Sloutsky, Vladimir M.; Fisher, Anna V.

    2012-01-01

    Linguistic labels affect inductive generalization; however, the mechanism underlying these effects remains unclear. According to one similarity-based model, SINC (similarity, induction, naming, and categorization), early in development labels are features of objects contributing to the overall similarity of compared entities, with early induction…

  16. 40 CFR 92.212 - Labeling.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... remanufacturer in the manner described in this section at the time of manufacture or remanufacture. (b... distinguish this label from the engine label described in paragraph (c) of this section. (B) Full corporate... U.S. EPA regulations applicable to locomotives originally manufactured prior to January 1, 2002;...

  17. 40 CFR 92.212 - Labeling.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... remanufacturer in the manner described in this section at the time of manufacture or remanufacture. (b... distinguish this label from the engine label described in paragraph (c) of this section. (B) Full corporate... U.S. EPA regulations applicable to locomotives originally manufactured prior to January 1, 2002;...

  18. 40 CFR 92.212 - Labeling.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... remanufacturer in the manner described in this section at the time of manufacture or remanufacture. (b... distinguish this label from the engine label described in paragraph (c) of this section. (B) Full corporate... U.S. EPA regulations applicable to locomotives originally manufactured prior to January 1, 2002;...

  19. 40 CFR 92.212 - Labeling.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... remanufacturer in the manner described in this section at the time of manufacture or remanufacture. (b... distinguish this label from the engine label described in paragraph (c) of this section. (B) Full corporate... U.S. EPA regulations applicable to locomotives originally manufactured prior to January 1, 2002;...

  20. 9 CFR 112.3 - Diluent labels.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Diluent labels. 112.3 Section 112.3 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS PACKAGING AND LABELING § 112.3...