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Sample records for assisted laser desorption

  1. Overview literature on matrix assisted laser desorption ionization ...

    Indian Academy of Sciences (India)

    Unknown

    Overview literature on matrix assisted laser desorption ionization mass spectroscopy (MALDI MS): basics and its .... Overview literature on MALDI MS. 517 mined as opposed to obtaining relative molecular ...... accurate representation of the overall molecular mass distribution in each of the fractionated materials. This.

  2. Investigations into ultraviolet matrix-assisted laser desorption

    Energy Technology Data Exchange (ETDEWEB)

    Heise, Theodore W. [Iowa State Univ., Ames, IA (United States)

    1993-07-01

    Matrix-assisted laser desorption (MALD) is a technique for converting large biomolecules into gas phase ions. Some characteristics of the commonly used uv matrices are determined. Solubilities in methanol range from 0.1 to 0.5 M. Solid phase absorption spectra are found to be similar to solution, but slightly red-shifted. Acoustic and quartz crystal microbalance signals are investigated as possible means of uv-MALD quantitation. Evidence for the existence of desorption thresholds is presented. Threshold values are determined to be in the range of 2 to 3 MW/cm2. A transient imaging technique based on laser-excited fluorescence for monitoring MALD plumes is described. Sensitivity is well within the levels required for studying matrix-assisted laser desorption, where analyte concentrations are significantly lower than those in conventional laser desorption. Results showing the effect of film morphology, particularly film thickness, on plume dynamics are presented. In particular, MALD plumes from thicker films tend to exhibit higher axial velocities. Fluorescent labeling of protein and of DNA is used to allow imaging of their uv-MALD generated plumes. Integrated concentrations are available with respect to time, making it possible to assess the rate of fragmentation. The spatial and temporal distributions are important for the design of secondary ionization schemes to enhance ion yields and for the optimization of ion collection in time-of-flight MS instruments to maximize resolution. Such information could also provide insight into whether ionization is closely associated with the desorption step or whether it is a result of subsequent collisions with the matrix gas (e.g., proton transfer). Although the present study involves plumes in a normal atmosphere, adaptation to measurements in vacuum (e.g., inside a mass spectrometer) should be straightforward.

  3. Matrix-assisted laser desorption/ionization imaging mass spectrometry.

    Science.gov (United States)

    Zaima, Nobuhiro; Hayasaka, Takahiro; Goto-Inoue, Naoko; Setou, Mitsutoshi

    2010-01-01

    Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is a powerful tool that enables the simultaneous detection and identification of biomolecules in analytes. MALDI-imaging mass spectrometry (MALDI-IMS) is a two-dimensional MALDI-mass spectrometric technique used to visualize the spatial distribution of biomolecules without extraction, purification, separation, or labeling of biological samples. MALDI-IMS has revealed the characteristic distribution of several biomolecules, including proteins, peptides, amino acids, lipids, carbohydrates, and nucleotides, in various tissues. The versatility of MALDI-IMS has opened a new frontier in several fields such as medicine, agriculture, biology, pharmacology, and pathology. MALDI-IMS has a great potential for discovery of unknown biomarkers. In this review, we describe the methodology and applications of MALDI-IMS for biological samples.

  4. Ionization Mechanism of Matrix-Assisted Laser Desorption/Ionization

    Science.gov (United States)

    Lu, I.-Chung; Lee, Chuping; Lee, Yuan-Tseh; Ni, Chi-Kung

    2015-07-01

    In past studies, mistakes in determining the ionization mechanism in matrix-assisted laser desorption/ionization (MALDI) were made because an inappropriate ion-to-neutral ratio was used. The ion-to-neutral ratio of the analyte differs substantially from that of the matrix in MALDI. However, these ratios were not carefully distinguished in previous studies. We begin by describing the properties of ion-to-neutral ratios and reviews early experimental measurements. A discussion of the errors committed in previous theoretical studies and a comparison of recent experimental measurements follow. We then describe a thermal proton transfer model and demonstrate how the model appropriately describes ion-to-neutral ratios and the total ion intensity. Arguments raised to challenge thermal ionization are then discussed. We demonstrate how none of the arguments are valid before concluding that thermal proton transfer must play a crucial role in the ionization process of MALDI.

  5. Quantitative analysis of biopolymers by matrix-assisted laser desorption

    Energy Technology Data Exchange (ETDEWEB)

    Tang, K.; Allman, S.L.; Jones, R.B.; Chen, C.H. (Oak Ridge National Lab., TN (United States))

    1993-08-01

    During the past few years, major efforts have been made to use mass spectrometry to measure biopolymers because of the great potential benefit to biological and medical research. Although the theoretical details of laser desorption and ionization mechanisms of MALDI are not yet fully understood, several models have been presented to explain the production of large biopolymer ions. In brief, it is very difficult to obtain reliable measurements of the absolute quantity of analytes by MALDI. If MALDI is going to become a routine analytical tool, it is obvious that quantitative measurement capability must be pursued. Oligonucleotides and protein samples used in this work were purchased from commercial sources. Nicotinic acid was used as matrix for both types of biopolymers. From this experiment, it is seen that it is difficult to obtain absolute quantitative measurements of biopolymers using MALDI. However, internal calibration with molecules having similar chemical properties can be used to resolve these difficulties. Chemical reactions between biopolymers must be avoided to prevent the destruction of the analyte materials. 10 refs., 8 figs.

  6. Characterisation of bacteria by matrix-assisted laser desorption/ionisation and electrospray mass spectrometry

    NARCIS (Netherlands)

    Baar, B.L.M. van

    2000-01-01

    Chemical analysis for the characterisation of micro-organisms is rapidly evolving, after the recent advent of new ionisation methods in mass spectrometry (MS): electrospray (ES) and matrix-assisted laser desorption/ionisation (MALDI). These methods allow quick characterisation of micro-organisms,

  7. Electrospray ionization and matrix assisted laser desorption/ionization mass spectrometry: powerful analytical tools in recombinant protein chemistry

    DEFF Research Database (Denmark)

    Andersen, Jens S.; Svensson, B; Roepstorff, P

    1996-01-01

    Electrospray ionization and matrix assisted laser desorption/ionization are effective ionization methods for mass spectrometry of biomolecules. Here we describe the capabilities of these methods for peptide and protein characterization in biotechnology. An integrated analytical strategy is presen......Electrospray ionization and matrix assisted laser desorption/ionization are effective ionization methods for mass spectrometry of biomolecules. Here we describe the capabilities of these methods for peptide and protein characterization in biotechnology. An integrated analytical strategy...

  8. Aerosol matrix-assisted laser desorption ionization for liquid chromatography/time-of-flight mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Murray, K.K.; Lewis, T.M.; Beeson, M.D.; Russell, D.H. (Texas A M Univ., College Station, TX (United States))

    1994-05-15

    We report the application of aerosol matrix-assisted laser desorption ionization (MALDI) to liquid chromatography/mass spectrometry (LC/MS). The aerosol MALDI experiment uses aerosol liquid introduction in conjunction with pulsed UV laser ionization to form ions from large biomolecules in solution. Mass analysis is achieved in a time-of-flight mass spectrometer. In the LC/MALDI-MS experiment, the matrix solution is combined with the column effluent in a mixing tee, LC/MALDI-MS is demonstrated for the separation of bradykinin, gramicidin S, and myoglobin. 32 refs., 8 figs., 1 tab.

  9. Matrix-assisted laser desorption/ionisation mass spectrometry of transfer ribonucleic acids isolated from yeast.

    Science.gov (United States)

    Gruic-Sovulj, I; Lüdemann, H C; Hillenkamp, F; Weygand-Durasevic, I; Kucan, Z; Peter-Katalinic, J

    1997-01-01

    tRNATyr and tRNASer purified from bulk brewer's yeast tRNA were subjected to analysis by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry. Choosing a mixture of 2,4,6- and 2,3,4-trihydroxy-acetophenone and diammonium citrate as matrix a mass resolution of up to 220 (FWHM) was achieved in the linear mode of operation. Cation adduct suppression by addition of cation exchange beads and a chelating agent (CDTA) is shown to substantially improve mass resolution for this class of molecules. PMID:9108172

  10. Cosmetic Analysis Using Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Imaging (MALDI-MSI

    Directory of Open Access Journals (Sweden)

    Rodrigo Ramos Catharino

    2013-03-01

    Full Text Available A new “omic” platform—Cosmetomics—that proves to be extremely simple and effective in terms of sample preparation and readiness for data acquisition/interpretation is presented. This novel approach employing Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Imaging (MALDI-MSI for cosmetic analysis has proven to readily identify and quantify compounds of interest. It also allows full control of all the production phases, as well as of the final product, by integration of both analytical and statistical data. This work has focused on products of daily use, namely nail polish, lipsticks and eyeliners of multiple brands sold in the worldwide market.

  11. Manganese oxide nanoparticle-assisted laser desorption/ionization mass spectrometry for medical applications

    Directory of Open Access Journals (Sweden)

    Shu Taira, Kenji Kitajima, Hikaru Katayanagi, Eiichiro Ichiishi and Yuko Ichiyanagi

    2009-01-01

    Full Text Available We prepared and characterized manganese oxide magnetic nanoparticles (d =5.6 nm and developed nanoparticle-assited laser desorption/ionization (nano-PALDI mass spectrometry. The nanoparticles had MnO2 and Mn2O3 cores conjugated with hydroxyl and amino groups, and showed paramagnetism at room temperature. The nanoparticles worked as an ionization assisting reagent in mass spectroscopy. The mass spectra showed no background in the low m/z. The nanoparticles could ionize samples of peptide, drug and proteins (approx. 5000 Da without using matrix, i.e., 2,5-dihydroxybenzoic acid (DHB, 4-hydroxy-α-cinnamic acid (CHCA and liquid matrix, as conventional ionization assisting reagents. Post source decay spectra by nano-PALDI mass spectrometry will yield information of the chemical structure of analytes.

  12. Drawing a different picture with pencil lead as matrix-assisted laser desorption/ionization matrix for fullerene derivatives.

    Science.gov (United States)

    Nye, Leanne C; Hungerbühler, Hartmut; Drewello, Thomas

    2017-01-01

    Inspired by reports on the use of pencil lead as a matrix-assisted laser desorption/ionization matrix, paving the way towards matrix-free matrix-assisted laser desorption/ionization, the present investigation evaluates its usage with organic fullerene derivatives. Currently, this class of compounds is best analysed using the electron transfer matrix trans-2-[3-(4-tert-butylphenyl)-2-methyl-2-propenylidene] malononitrile (DCTB), which was employed as the standard here. The suitability of pencil lead was additionally compared to direct (i.e. no matrix) laser desorption/ionization-mass spectrometry. The use of (DCTB) was identified as the by far gentler method, producing spectra with abundant molecular ion signals and much reduced fragmentation. Analytically, pencil lead was found to be ineffective as a matrix, however, appears to be an extremely easy and inexpensive method for producing sodium and potassium adducts.

  13. Identification of Molds by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

    Science.gov (United States)

    Posteraro, Brunella

    2016-01-01

    ABSTRACT Although to a lesser extent than diagnostic bacteriology, matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) has recently revolutionized the diagnostic mycology workflow. With regard to filamentous fungi (or molds), the precise recognition of pathogenic species is important for rapid diagnosis and appropriate treatment, especially for invasive diseases. This review summarizes the current experience with MALDI-TOF MS-based identification of common and uncommon mold species of Aspergillus, Fusarium, Mucorales, dimorphic fungi, and dermatophytes. This experience clearly shows that MALDI-TOF MS holds promise as a fast and accurate identification tool, particularly with common species or typical strains of filamentous fungi. PMID:27807151

  14. Identification of Molds by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry.

    Science.gov (United States)

    Sanguinetti, Maurizio; Posteraro, Brunella

    2017-02-01

    Although to a lesser extent than diagnostic bacteriology, matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has recently revolutionized the diagnostic mycology workflow. With regard to filamentous fungi (or molds), the precise recognition of pathogenic species is important for rapid diagnosis and appropriate treatment, especially for invasive diseases. This review summarizes the current experience with MALDI-TOF MS-based identification of common and uncommon mold species of Aspergillus, Fusarium, Mucorales, dimorphic fungi, and dermatophytes. This experience clearly shows that MALDI-TOF MS holds promise as a fast and accurate identification tool, particularly with common species or typical strains of filamentous fungi. Copyright © 2017 American Society for Microbiology.

  15. Use of matrix-assisted laser desorption/ionisation mass spectrometry in cancer research.

    Science.gov (United States)

    Bateson, Hannah; Saleem, Saira; Loadman, Paul M; Sutton, Chris W

    2011-01-01

    Cancer significantly affects millions of people worldwide. It is possible to use proteomic techniques to aid in detection, monitoring of treatment and progression, as well as gaining an increased understanding of cancer. Matrix-assisted laser desorption/ionisation (MALDI) mass spectrometry can be utilised to detect the presence of proteins and peptides within various samples from the body, including blood, biological fluids and tumour tissue. This review aims to introduce MALDI mass spectrometry and discuss a range of applications in the field of cancer research, from quantitative to qualitative methods. Also described is MALDI imaging mass spectrometry which differs from typical sample preparation methods, as analytes are ionised directly from the tissue. Finally, presented is a brief summary of the status of biomarker discovery using blood/serum and biological fluids samples, and the implications in the clinic. Copyright © 2011 Elsevier Inc. All rights reserved.

  16. Matrix-assisted laser desorption/ionization-collision induced dissociation of poly(styrene).

    Science.gov (United States)

    Jackson, A T; Yates, H T; Scrivens, J H; Green, M R; Bateman, R H

    1998-04-01

    Matrix-assisted laser desorption/ionization-collision induced dissociation (MALDI-CID) has been employed for the analysis of poly(styrene) in a tandem hybrid sector-time of flight instrument. Spectra are shown for adducts of poly(styrene) with copper and silver ions. The distributions of fragment ion peaks were found to be consistent from precursor ions containing both metal ions. It is shown how the masses of the end groups of the polymer may be inferred from the mass-to-charge ratios of two of the series of ion peaks that are seen in the MALDI-CID spectra. Mechanisms are proposed for the formation of some of the other series of ion peaks that are observed in the spectra.

  17. Affinity surface-assisted laser desorption/ionization mass spectrometry for peptide enrichment.

    Science.gov (United States)

    Coffinier, Yannick; Nguyen, Nhung; Drobecq, Hervé; Melnyk, Oleg; Thomy, Vincent; Boukherroub, Rabah

    2012-12-07

    In this paper, we report on the functionalization of silicon nanostructured (NanoSi) surface with an organic layer of nitrilotriacetic acid (NTA) and its subsequent use as an affinity surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS) interface for histidine-tagged peptide enrichment and mass spectrometry analysis. The NTA terminal groups are immobilized onto the NanoSi surface via very stable Si-C covalent bonds. The NTA-modified NanoSi (NTA-NanoSi) interface was characterized by contact angle measurements, Fourier transform infrared (FTIR) spectroscopy and X-ray photoelectron spectroscopy (XPS). The NTA-NanoSi interface has shown a good selectivity toward His-tagged peptide and permits its enrichment from an artificial mixture of both tagged and untagged peptides and its subsequent mass spectrometry detection with good signal/noise ratio.

  18. Direct Surface Analysis of Fungal Species by Matrix-assisted Laser Desorption/Ionization Mass Spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Valentine, Nancy B.(BATTELLE (PACIFIC NW LAB)); Wahl, Jon H.(BATTELLE (PACIFIC NW LAB)); Kingsley, Mark T.(BATTELLE (PACIFIC NW LAB)); Wahl, Karen L.(BATTELLE (PACIFIC NW LAB))

    2001-12-01

    Intact spores and/or hyphae of Aspergillus niger, Rhizopus oryzae, Trichoderma reesei and Phanerochaete chrysosporium are analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). This study investigates various methods of sample preparation and matrices to determine optimum collection and analysis criteria for fungal analysis by MALDI-MS. Fungi are applied to the MALDI sample target as untreated, sonicated, acid/heat treated, or blotted directly from the fungal culture with double-stick tape. Ferulic acid or sinapinic acid matrix solution is layered over the dried samples and analyzed by MALDI-MS. Statistical analysis of the data show that simply using double stick tape to collect and transfer to a MALDI sample plate typically worked as well as the other preparation methods, but requires the least sample handling.

  19. Phenotypic identification of Porphyromonas gingivalis validated with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

    NARCIS (Netherlands)

    Rams, Thomas E; Sautter, Jacqueline D; Getreu, Adam; van Winkelhoff, Arie J

    OBJECTIVE: Porphyromonas gingivalis is a major bacterial pathogen in human periodontitis. This study used matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry to assess the accuracy of a rapid phenotypic identification scheme for detection of cultivable P.

  20. Development of matrix-assisted laser desorption ionization-mass spectrometry imaging (MALDI-MSI) for plant metabolite analysis

    Energy Technology Data Exchange (ETDEWEB)

    Korte, Andrew R [Iowa State Univ., Ames, IA (United States)

    2014-12-01

    This thesis presents efforts to improve the methodology of matrix-assisted laser desorption ionization-mass spectrometry imaging (MALDI-MSI) as a method for analysis of metabolites from plant tissue samples. The first chapter consists of a general introduction to the technique of MALDI-MSI, and the sixth and final chapter provides a brief summary and an outlook on future work.

  1. Metal powder substrate-assisted laser desorption/ionization mass spectrometry for polyethylene analysis.

    Science.gov (United States)

    Yalcin, Talat; Wallace, William E; Guttman, Charles M; Li, Liang

    2002-09-15

    Polyethylene is one of the most important industrial polymers and is also one of the most challenging polymers to be characterized by mass spectrometry. We have developed a substrate-assisted laser desorption/ionization (LDI) mass spectrometric method for polyethylene analysis. In this method, cobalt, copper, nickel, or iron metal powders are used as a sample substrate and silver nitrate is used as the cationization reagent. Using a conventional UV LDI time-of-flight mass spectrometer, intact oligomer ions having masses up to 5000 u can be detected. Cobalt is found to produce spectra with the highest signal-to-noise ratio and the lowest level of fragmentation. Cobalt powder size is shown to have some effect on the spectra produced. The best results are obtained with the use of cobalt powders with diameters ranging from 30 to 100 microm. Fragmentation cannot be totally eliminated, but the fragment ion peaks can be readily discerned from the intact polyethylene ions in the substrate-assisted LDI spectrum. Thus, the average molecular masses of low-mass polyethylene samples can be determined by using this method. A rapid heating model is used to account for the effectiveness of using the coarse metal powders to assist the analysis of intact polyethylene molecules by LDI.

  2. On the Primary Ionization Mechanism(s in Matrix-Assisted Laser Desorption Ionization

    Directory of Open Access Journals (Sweden)

    Laura Molin

    2012-01-01

    Full Text Available A mechanism is proposed for the first step of ionization occurring in matrix-assisted laser desorption ionization, leading to protonated and deprotonated matrix (Ma molecules ([Ma+H]+ and [Ma-H]- ions. It is based on observation that in solid state, for carboxyl-containing MALDI matrices, the molecules form strong hydrogen bonds and their carboxylic groups can act as both donors and acceptors. This behavior leads to stable dimeric structures. The laser irradiation leads to the cleavage of these hydrogen bonds, and theoretical calculations show that both [Ma+H]+ and [Ma-H]- ions can be formed through a two-photon absorption process. Alternatively, by the absorption of one photon only, a heterodissociation of one of the O–H bonds can lead to a stable structure containing both cationic and anionic sites. This structure could be considered an intermediate that, through the absorption of a further photon, leads to the formation of matrix ions. Some experiments have been performed to evaluate the role of thermal ionization and indicate that its effect is negligible. Some differences have been observed for different matrices in the formation of analyte molecule (M ion [M+H]+, [M-H]-, M+•, and [M-2H]-•, and they have been explained in terms of ionization energies, pKa values, and thermodynamic stability.

  3. Iron oxide nanomatrix facilitating metal ionization in matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Obena, Rofeamor P; Lin, Po-Chiao; Lu, Ying-Wei; Li, I-Che; del Mundo, Florian; Arco, Susan dR; Nuesca, Guillermo M; Lin, Chung-Chen; Chen, Yu-Ju

    2011-12-15

    The significance and epidemiological effects of metals to life necessitate the development of direct, efficient, and rapid method of analysis. Taking advantage of its simple, fast, and high-throughput features, we present a novel approach to metal ion detection by matrix-functionalized magnetic nanoparticle (matrix@MNP)-assisted MALDI-MS. Utilizing 21 biologically and environmentally relevant metal ion solutions, the performance of core and matrix@MNP against conventional matrixes in MALDI-MS and laser desorption ionization (LDI) MS were systemically tested to evaluate the versatility of matrix@MNP as ionization element. The matrix@MNPs provided 20- to >100-fold enhancement on detection sensitivity of metal ions and unambiguous identification through characteristic isotope patterns and accurate mass (potentials (IP), we formulated a metal ionization mechanism model, alluding to the role of exciton pooling in matrix@MNP-assisted MALDI-MS. Moreover, the detection of Cu in spiked tap water demonstrated the practicability of this new approach as an efficient and direct alternative tool for fast, sensitive, and accurate determination of trace metal ions in real samples.

  4. Bioaerosol detection by aerosol TOF-mass spectrometry: Application of matrix assisted laser desorption/ionisation

    NARCIS (Netherlands)

    Wuijckhuijse, A.L. van; Stowers, M.A.; Kientz, Ch.E.; Marijnissen, J.C.M.; Scarlett, B.

    2000-01-01

    In previous publications the use of an aerosol time of flight mass spectrometer was reported for the on-line measurements of aerosols (Weiss 1997, Kievit 1995). The apparatus is capable of measuring the size as well as the chemical composition, by the use of Laser Desorption/Ionisation (LDI), of an

  5. Matrix assisted laser desorption time of flight mass spectrometry (MALDI-TOF MS) in clinical microbiology.

    Science.gov (United States)

    Angeletti, Silvia

    2017-07-01

    The microbiological management of patients with suspected bacterial infection includes the identification of the pathogen and the determination of the antibiotic susceptibility. These traditional approaches, based on the pure culture of the microorganism, require at least 36-48h. A new method, Matrix Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS), has been recently developed to profile bacterial proteins from whole cell extracts and obtain a bacterial fingerprint able to discriminate microorganisms from different genera and species. By whole cell-mass spectrometry, microbial identification can be achieved within minutes from cultured isolate, rather than traditional phenotypic or genotypic characterizations. From the year 2009 an explosion of applications of this technology has been observed with promising results. Several studies have been performed and showed that MALDI-TOF represents a reliable alternative method for rapid bacteria and fungi identification in clinical setting. A future area of expansion is represented by the application of MALDI-TOF technology to the antibiotic susceptibility test. In conclusion, the revision of the literature available up to date demonstrated that MALDI-TOF MS represents an innovative technology for the rapid and accurate identification of bacterial and fungal isolates in clinical settings. By an earlier microbiological diagnosis, MALDI-TOF MS contributes to a reduced mortality and hospitalization time of the patients and consequently has a significant impact on cost savings and public health. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. Matrix-assisted ultraviolet laser desorption/ionization mass spectrometry applied to multiple forms of lipases.

    Science.gov (United States)

    Hedrich, H C; Isobe, K; Stahl, B; Nokihara, K; Kordel, M; Schmid, R D; Karas, M; Hillenkamp, F; Spener, F

    1993-06-01

    Matrix-assisted ultraviolet laser desorption/ionization mass spectrometry was used to investigate heterogeneous patterns and molecular masses of microbial lipases from Penicillium camembertii, Geotrichum candidum, and Pseudomonas sp. Mass spectral peaks of the native, glycosylated lipases from P. camembertii and G. candidum were broader than those of the corresponding deglycosylated enzymes, indicative of heterogeneous glycosylations. The broader peaks in the mass spectra were caused by an overlapping of unresolved peaks, derived from single glycoprotein species. Molecular masses determined for the deglycosylated proteins were in excellent agreement with those deduced from amino acid composition and sequence data, whereas with conventional biochemical methods (gelfiltration, sodium dodecyl sulfate-polyacrylamide gel electrophoresis) only very rough estimations of molecular masses were possible. By mass spectrometric analysis of the four fractions of chromatographically separated P. camembertii lipase molecular masses of 29,990, 34,030, 31,990, and 32,140 Da were found before and 29,960, 29,980, 29,990 and 30,010 Da, respectively, after deglycosylation. Thus from the four native fractions of P. camembertii lipase three were glycoproteins. G. candidum lipase showed an average molecular mass of 63,500 Da for the heterogeneously deglycosylated native form and a molecular mass of 59,650 Da for the deglycosylated enzyme. For the Pseudomonas lipase, which could only be isolated with lipids firmly attached, a molecular mass of 32,890 Da was determined, in close agreement with that derived from the cDNA sequence.

  7. Properties of matrix-assisted laser desorption. Measurements with a time-to-digital converter.

    Science.gov (United States)

    Ens, W; Mao, Y; Mayer, F; Standing, K G

    1991-03-01

    Some properties of matrix-assisted laser desorption have been studied using single-ion-counting methods and a time-to-digital converter. The methods allow examination of the process for irradiances near the reported threshold for observation with a transient recorder. All measurements were made using bovine insulin as a test compound. We present direct evidence that an irradiance threshold near 10(6) W cm-2 exists for ion production, and that the process is a collective effect, either involving a large number of molecular ions (approximately 10(4) in a successful event or none at all. Above the threshold, the yield is found to scale with a high power (4th to 6th) of the irradiance. Measurements of initial velocity distributions indicate an axial velocity spread corresponding to approximately 50 eV and a radial velocity spread corresponding to approximately 2.4 eV. Thus the ejection or extraction mechanism appears to be strongly asymmetric.

  8. Nanoparticle-assisted laser desorption/ionization mass spectrometry: Novel sample preparation methods and nanoparticle screening for plant metabolite imaging

    Energy Technology Data Exchange (ETDEWEB)

    Yagnik, Gargey B. [Iowa State Univ., Ames, IA (United States)

    2016-02-19

    The main goal of the presented research is development of nanoparticle based matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS). This dissertation includes the application of previously developed data acquisition methods, development of novel sample preparation methods, application and comparison of novel nanoparticle matrices, and comparison of two nanoparticle matrix application methods for MALDI-MS and MALDI-MS imaging.

  9. Rapid Detection of OXA-48-Producing Enterobacteriaceae by Matrix-Assisted Laser Desorption Ionization−Time of Flight Mass Spectrometry

    Science.gov (United States)

    Oviaño, Marina; Barba, Maria José; Fernández, Begoña; Ortega, Adriana; Aracil, Belén; Oteo, Jesús; Campos, José

    2015-01-01

    A rapid and sensitive (100%) matrix-assisted laser desorption ionization−time of flight mass spectrometry (MALDI-TOF MS) assay was developed to detect OXA-48-type producers, using 161 previously characterized clinical isolates. Ertapenem was monitored to detect carbapenem resistance, and temocillin was included in the assay as a marker for OXA-48-producers. Structural analysis of temocillin is described. Data are obtained within 60 min. PMID:26677247

  10. Identification of Nocardia Species by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

    Science.gov (United States)

    Khot, Prasanna D.; Bird, Brian A.; Durrant, Robert J.

    2015-01-01

    Matrix-assisted laser desorption ionization–time of flight mass spectrometry for identification of Nocardia species remains challenging. By identifying 83.1% (64 of 77) and 80% (8 of 10) to the species and complex levels, respectively, and 94.3% (82 of 87) to the genus level, we show that an approach using routine sample preparation, an up-to-date commercial database minimally augmented with custom spectra, and testing at an early stage of growth is promising. PMID:26269617

  11. The value of total protein in guiding management of infectious parapneumonic effusion by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

    National Research Council Canada - National Science Library

    Chiu, Chih-Yung; Hsieh, Sen-Yung; Wong, Kin-Sun; Lai, Shen-Hao; Chen, Jen-Kun; Huang, Jing-Long

    2015-01-01

    .... The purpose of this study was to investigate the value of total protein analysis in guiding management of infectious PE by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry...

  12. GUMBOS matrices of variable hydrophobicity for matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Al Ghafly, Hashim; Siraj, Noureen; Das, Susmita; Regmi, Bishnu P; Magut, Paul K S; Galpothdeniya, Waduge Indika S; Murray, Kermit K; Warner, Isiah M

    2014-11-15

    Detection of hydrophobic peptides remains a major obstacle for matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). This stems from the fact that most matrices for MALDI are hydrophilic and therefore have low affinities for hydrophobic peptides. Herein, 1-aminopyrene (AP) and AP-derived group of uniform materials based on organic salts (GUMBOS) as novel matrices for MALDI-MS analyses of peptides were investigated for hydrophobic and hydrophilic peptides. A number of solid-phase AP-based GUMBOS are synthesized with variable hydrophobicity simply by changing the counterions. Structures were confirmed by use of (1)H NMR and electrospray ionization mass spectrometry (ESI-MS). 1-Octanol/water partition coefficients (Ko/w) were used to measure the hydrophobicity of the matrices. A dried-droplet method was used for sample preparation. All spectra were obtained using a MALDI-TOF mass spectrometer in positive ion reflectron mode. A series of AP-based GUMBOS was synthesized including [AP][chloride] ([AP][Cl]), [AP][ascorbate] ([AP][Asc]) and [AP][bis(trifluoromethane)sulfonimide] ([AP][NTf2]). The relative hydrophobicities of these compounds and α-cyano-4-hydroxycinnamic acid (CHCA, a common MALDI matrix) indicated that AP-based GUMBOS can be tuned to be much more hydrophobic than CHCA. A clear trend is observed between the signal intensities of hydrophobic peptides and hydrophobicity of the matrix. MALDI matrices of GUMBOS with tunable hydrophobicities are easily obtained simply by varying the counterion. We have found that hydrophobic matrix materials are very effective for MALDI determination of hydrophobic peptides and, similarly, the more hydrophilic peptides displayed greater intensity in the more hydrophilic matrix. Copyright © 2014 John Wiley & Sons, Ltd.

  13. Detection and mapping of Cannabinoids in single hair samples through rapid derivatization- Matrix-Assisted Laser Desorption Ionization Mass Spectrometry

    OpenAIRE

    Beasley, Emma; Francese, Simona; Bassindale, Thomas

    2016-01-01

    The sample preparation method reported in this work has permitted for the first time the application of Matrix Assisted Laser Desorption Ionization Mass Spectrometry Profiling and Imaging (MALDI-MSP and MALDI-MSI) for the detection and mapping of cannabinoids in a single hair sample. MALDI-MSI analysis of hair samples has recently been suggested as an alternative technique to traditional methods of GC-MS and LC-MS due to simpler sample preparation, the ability to detect a narrower time frame ...

  14. Species identification of clinical Prevotella isolates by matrix-assisted laser desorption ionization-time of flight mass spectrometry.

    Science.gov (United States)

    Wybo, Ingrid; Soetens, Oriane; De Bel, Annelies; Echahidi, Fedoua; Vancutsem, Ellen; Vandoorslaer, Kristof; Piérard, Denis

    2012-04-01

    The performance of matrix-assisted laser desorption-ionization time of flight mass spectrometry (MALDI-TOF MS) for species identification of Prevotella was evaluated and compared with 16S rRNA gene sequencing. Using a Bruker database, 62.7% of the 102 clinical isolates were identified to the species level and 73.5% to the genus level. Extension of the commercial database improved these figures to, respectively, 83.3% and 89.2%. MALDI-TOF MS identification of Prevotella is reliable but needs a more extensive database.

  15. Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry applied to virus identification

    OpenAIRE

    Calderaro, Adriana; Arcangeletti, Maria-Cristina; Rodighiero, Isabella; Buttrini, Mirko; Gorrini, Chiara; Motta, Federica; Germini, Diego; Medici, Maria-Cristina; Chezzi, Carlo; De Conto, Flora

    2014-01-01

    Virus detection and/or identification traditionally rely on methods based on cell culture, electron microscopy and antigen or nucleic acid detection. These techniques are good, but often expensive and/or time-consuming; furthermore, they not always lead to virus identification at the species and/or type level. In this study, Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) was tested as an innovative tool to identify human polioviruses and to identif...

  16. Nanomaterials as Assisted Matrix of Laser Desorption/Ionization Time-of-Flight Mass Spectrometry for the Analysis of Small Molecules

    Science.gov (United States)

    Lu, Minghua; Yang, Xueqing; Yang, Yixin; Qin, Peige; Wu, Xiuru; Cai, Zongwei

    2017-01-01

    Matrix-assisted laser desorption/ionization (MALDI), a soft ionization method, coupling with time-of-flight mass spectrometry (TOF MS) has become an indispensible tool for analyzing macromolecules, such as peptides, proteins, nucleic acids and polymers. However, the application of MALDI for the analysis of small molecules (mass region. To overcome this drawback, more attention has been paid to explore interference-free methods in the past decade. The technique of applying nanomaterials as matrix of laser desorption/ionization (LDI), also called nanomaterial-assisted laser desorption/ionization (nanomaterial-assisted LDI), has attracted considerable attention in the analysis of low-molecular weight compounds in TOF MS. This review mainly summarized the applications of different types of nanomaterials including carbon-based, metal-based and metal-organic frameworks as assisted matrices for LDI in the analysis of small biological molecules, environmental pollutants and other low-molecular weight compounds. PMID:28430138

  17. Benefits of 2.94 μm infrared matrix-assisted laser desorption/ionization for analysis of labile molecules by Fourier transform mass spectrometry

    DEFF Research Database (Denmark)

    Budnik, Bogdan A.; Jensen, Kenneth Bendix; Jørgensen, Thomas J. D.

    2000-01-01

    A 2.94 microm Er:YAG laser was used together with a commercial Fourier transform mass spectrometer to study labile biomolecules. The combination has shown superior performance over conventional 337 nm ultraviolet matrix-assisted laser desorption/ionization (UV-MALDI) Fourier transform mass...

  18. Spatially resolved protein hydrogen exchange measured by matrix-assisted laser desorption ionization in-source decay

    DEFF Research Database (Denmark)

    Rand, Kasper D; Bache, Nicolai; Nedertoft, Morten M

    2011-01-01

    Mass spectrometry has become a powerful tool for measuring protein hydrogen exchange and thereby reveal the structural dynamics of proteins in solution. Here we describe the successful application of a matrix-assisted laser desorption ionization (MALDI) mass spectrometry approach based on in......-source decay (ISD) to measure spatially resolved amide backbone hydrogen exchange. By irradiating deuterated protein molecules in a crystalline matrix with a high laser fluence, they undergo prompt fragmentation. Spatially resolved deuteration levels are readily obtained by mass analysis of consecutive...... fragment ions. MALDI ISD analysis of deuterated cytochrome c yielded an extensive series of c-fragment ions which originate from cleavage of nearly all N-C(α) bonds (Cys17 to Glu104) allowing for a detailed analysis of the deuterium content of the backbone amides. While hydrogen scrambling can be major...

  19. Matrix-Assisted Laser Desorption/Ionization Mass Spectrometric Analysis of Uncomplexed Highly Sulfated Oligosaccharides Using Ionic Liquid Matrices

    Science.gov (United States)

    Laremore, Tatiana N.; Murugesan, Saravanababu; Park, Tae-Joon; Avci, Fikri Y.; Zagorevski, Dmitri V.; Linhardt, Robert J.

    2014-01-01

    Direct UV matrix-assisted laser desorption/ionization (MALDI) mass spectrometric analysis of uncomplexed, underivatized, highly sulfated oligosaccharides has been carried out using ionic liquids as matrices. Under conventionally used MALDI time-of-flight experimental conditions, uncomplexed polysulfated oligosaccharides do not produce any signal. We report that 1-methylimidazolium α-cyano-4-hydroxycinnamate and butylammonium 2,5-dihydroxybenzoate ionic liquid matrices allow the detection of picomole amounts of the sodium salts of a disaccharide, sucrose octasulfate, and an octasulfated pentasaccharide, Arixtra. The experimental results indicate that both analytes undergo some degree of thermal fragmentation with a mass loss corresponding to cleavage of O–SO3Na bonds in the matrix upon laser irradiation, reflecting lability of sulfo groups. PMID:16536411

  20. Matrix-assisted laser desorption/ionisation mass spectrometry imaging and its development for plant protein imaging

    Science.gov (United States)

    2011-01-01

    Matrix-Assisted Laser Desorption/Ionisation (MALDI) mass spectrometry imaging (MSI) uses the power of high mass resolution time of flight (ToF) mass spectrometry coupled to the raster of lasers shots across the cut surface of tissues to provide new insights into the spatial distribution of biomolecules within biological tissues. The history of this technique in animals and plants is considered and the potential for analysis of proteins by this technique in plants is discussed. Protein biomarker identification from MALDI-MSI is a challenge and a number of different approaches to address this bottleneck are discussed. The technical considerations needed for MALDI-MSI are reviewed and these are presented alongside examples from our own work and a protocol for MALDI-MSI of proteins in plant samples. PMID:21726462

  1. Rapid detection of undesired cosmetic ingredients by matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Ouyang, Jie; An, Dongli; Chen, Tengteng; Lin, Zhiwei

    2017-10-01

    In recent years, cosmetic industry profits soared due to the widespread use of cosmetics, which resulted in illicit manufacturers and products of poor quality. Therefore, the rapid and accurate detection of the composition of cosmetics has become crucial. At present, numerous methods, such as gas chromatography and liquid chromatography-mass spectrometry, were available for the analysis of cosmetic ingredients. However, these methods present several limitations, such as failure to perform comprehensive and rapid analysis of the samples. Compared with other techniques, matrix-assisted laser desorption ionization time-of-flight mass spectrometry offered the advantages of wide detection range, fast speed and high accuracy. In this article, we briefly summarized how to select a suitable matrix and adjust the appropriate laser energy. We also discussed the rapid identification of undesired ingredients, focusing on antibiotics and hormones in cosmetics.

  2. Solvent separating secondary metabolites directly from biosynthetic tissue for surface-assisted laser desorption ionisation mass spectrometry.

    Science.gov (United States)

    Rudd, David; Benkendorff, Kirsten; Voelcker, Nicolas H

    2015-03-16

    Marine bioactive metabolites are often heterogeneously expressed in tissues both spatially and over time. Therefore, traditional solvent extraction methods benefit from an understanding of the in situ sites of biosynthesis and storage to deal with heterogeneity and maximize yield. Recently, surface-assisted mass spectrometry (MS) methods namely nanostructure-assisted laser desorption ionisation (NALDI) and desorption ionisation on porous silicon (DIOS) surfaces have been developed to enable the direct detection of low molecular weight metabolites. Since direct tissue NALDI-MS or DIOS-MS produce complex spectra due to the wide variety of other metabolites and fragments present in the low mass range, we report here the use of "on surface" solvent separation directly from mollusc tissue onto nanostructured surfaces for MS analysis, as a mechanism for simplifying data annotation and detecting possible artefacts from compound delocalization during the preparative steps. Water, ethanol, chloroform and hexane selectively extracted a range of choline esters, brominated indoles and lipids from Dicathais orbita hypobranchial tissue imprints. These compounds could be quantified on the nanostructured surfaces by comparison to standard curves generated from the pure compounds. Surface-assisted MS could have broad utility for detecting a broad range of secondary metabolites in complex marine tissue samples.

  3. Quantitation of peptides and proteins by matrix-assisted laser desorption/ionization mass spectrometry using (18)O-labeled internal standards

    DEFF Research Database (Denmark)

    Mirgorodskaya, O A; Kozmin, Y P; Titov, M I

    2000-01-01

    A method for quantitating proteins and peptides in the low picomole and sub-picomole range has been developed using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) with internal (18)O-labeled standards. A simple procedure is proposed to produce such internal standards for...... inhibitor, were quantified by MALDI-time-of-flight (TOF) mass spectrometry.......A method for quantitating proteins and peptides in the low picomole and sub-picomole range has been developed using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) with internal (18)O-labeled standards. A simple procedure is proposed to produce such internal standards...

  4. Furoic and mefenamic acids as new matrices for matrix assisted laser desorption/ionization-(MALDI)-mass spectrometry.

    Science.gov (United States)

    Abdelhamid, Hani Nasser; Wu, Hui-Fen

    2013-10-15

    The present study introduces two novel organic matrices for matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS) for the analysis of small molecules. The first matrix is "2-amino-4,5-diphenylfuran-3-carboxylic acid" (also called furoic acid, FA) which was synthesized and then characterized by ultraviolet (UV), infrared (FTIR), nuclear magnetic resonance NMR ((1)H and (13)C) and mass spectrometry. The compound has organic semiconductor properties and exhibits intense UV-absorption which is suitable for the UV-MALDI laser (N2 laser, 337 nm). The second matrix is mefenamic acid (MA). The two matrices can be successfully applied for various classes of compounds including adenosine-5'-triphosphate (ATP, 0.5 µL(10.0 nmol)), spectinomycin (spect, 0.5 µL(14.0 nmol)), glutathione (GSH, 0.5 µL(9.0 nmol)), sulfamethazole (SMT, 0.5 µL(2.0 nmol)) and mixture of peptides gramicidin D (GD, 0.5µL (9.0 nmol)). The two matrices can effectively absorb the laser energy, resulting in excellent desorption/ionization of small molecules. The new matrices offer a significant enhancement of ionization, less fragmentation, few interferences, nice reproducibility, and excellent stability under vacuum. Theoretical calculations of the physical parameters demonstrated increase in polarizability, molar volume and refractivity than the conventional organic matrices which can effectively enhance the proton transfer reactions between the matrices with the analyte molecules. While the reduction in density, surface tension and index of refraction can enhance homogeneity between the two new matrices with the analytes. Due to the sublimation energy of mefenamic acid is (1.2 times) higher than that of the DHB, it is more stable to be used in the vacuum. Copyright © 2013 Elsevier B.V. All rights reserved.

  5. Enhanced capabilities for imaging gangliosides in murine brain with matrix-assisted laser desorption/ionization and desorption electrospray ionization mass spectrometry coupled to ion mobility separation.

    Science.gov (United States)

    Škrášková, Karolina; Claude, Emmanuelle; Jones, Emrys A; Towers, Mark; Ellis, Shane R; Heeren, Ron M A

    2016-07-15

    The increased interest in lipidomics calls for improved yet simplified methods of lipid analysis. Over the past two decades, mass spectrometry imaging (MSI) has been established as a powerful technique for the analysis of molecular distribution of a variety of compounds across tissue surfaces. Matrix-assisted laser desorption/ionization (MALDI) MSI is widely used to study the spatial distribution of common lipids. However, a thorough sample preparation and necessity of vacuum for efficient ionization might hamper its use for high-throughput lipid analysis. Desorption electrospray ionization (DESI) is a relatively young MS technique. In DESI, ionization of molecules occurs under ambient conditions, which alleviates sample preparation. Moreover, DESI does not require the application of an external matrix, making the detection of low mass species more feasible due to the lack of chemical matrix background. However, irrespective of the ionization method, the final information obtained during an MSI experiment is very complex and its analysis becomes challenging. It was shown that coupling MSI to ion mobility separation (IMS) simplifies imaging data interpretation. Here we employed DESI and MALDI MSI for a lipidomic analysis of the murine brain using the same IMS-enabled instrument. We report for the first time on the DESI IMS-MSI of multiply sialylated ganglioside species, as well as their acetylated versions, which we detected directly from the murine brain tissue. We show that poly-sialylated gangliosides can be imaged as multiply charged ions using DESI, while they are clearly separated from the rest of the lipid classes based on their charge state using ion mobility. This represents a major improvement in MSI of intact fragile lipid species. We additionally show that complementary lipid information is reached under particular conditions when DESI is compared to MALDI MSI. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. Enhancing carbohydrate ion yield by controlling crystalline structures in matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Lee, Hsun; Lai, Yin-Hung; Ou, Yu-Meng; Tsao, Chien-Wei; Jheng, Ya-Jin; Kuo, Shu-Yun; Chang, Huan-Tsung; Wang, Yi-Sheng

    2017-11-22

    Carbohydrate analysis is challenging due to lack of sensitive detection and efficient separation methods. Although matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) is a sensitive tool, the low ionization efficiency of carbohydrates makes mass analyses inefficient. This work systematically examines the correlation between MALDI-MS sensitivity and carbohydrate sample morphology. Depending on the properties of the matrix used, the morphology changes through sample recrystallization after drying or imposition of hydrodynamic flows during droplet drying. Observation shows that amorphous solids and finer crystals offer higher carbohydrate sensitivity and spatial homogeneity than larger crystals. Clear evidences of an inverse correlation between sensitivity and crystal size are obtained when various kinds of carbohydrates are mixed with different matrixes. Similar experiments on proteins and peptides showed a negative or negligible effect. The result serves as a general guideline for improving efficiency in routine carbohydrate analysis. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Phosphopeptide detection and sequencing by matrix-assisted laser desorption/ionization quadrupole time-of-flight tandem mass spectrometry

    DEFF Research Database (Denmark)

    Bennett, Keiryn L; Stensballe, Allan; Podtelejnikov, Alexandre V

    2002-01-01

    A prototype matrix-assisted laser desorption/ionization quadrupole time-of-flight (MALDI-TOF) tandem mass spectrometer was used to sequence a series of phosphotyrosine-, phosphothreonine- and phosphoserine-containing peptides. The high mass resolution and mass accuracy of the instrument allowed...... ion at m/z 216.04 was observed in these MALDI low-energy CID tandem mass spectra. Elimination of phosphate groups was evident from the triphosphorylated peptide but not from the monophosphorylated species. The main fragmentation pathway for the synthetic phosphothreonine-containing peptide...... the localization of one, three or four phosphorylated amino acid residues in phosphopeptides up to 3.1 kDa. Tandem mass spectra of two different phosphotyrosine peptides permitted amino acid sequence determination and localization of one and three phosphorylation sites, respectively. The phosphotyrosine immonium...

  8. Pigments and proteins in green bacterial chlorosomes studied by matrix-assisted laser desorption ionization mass spectrometry

    DEFF Research Database (Denmark)

    Persson, S; Sönksen, C P; Frigaard, N-U

    2000-01-01

    We have used matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) for mass determination of pigments and proteins in chlorosomes, the light-harvesting organelles from the photosynthetic green sulfur bacterium Chlorobium tepidum. By applying a small volume (1...... microL) of a concentrated suspension of isolated chlorosomes directly to the target of the mass spectrometer we have been able to detect bacteriochlorophyll a and all the major homologs of bacteriochlorophyll c. The peak heights of the different bacteriochlorophyll c homologs in the MALDI spectra were...... homologs in a small amount of green bacterial cells. In addition to information on pigments, the MALDI spectra also contained peaks from chlorosome proteins. Thus we have been able with high precision to confirm the molecular masses of the chlorosome proteins CsmA and CsmE which have been previously...

  9. Electrowetting-based microfluidics for analysis of peptides and proteins by matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Wheeler, Aaron R; Moon, Hyejin; Kim, Chang-Jin; Loo, Joseph A; Garrell, Robin L

    2004-08-15

    A new technique for preparing samples for matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is reported. The technique relies on electrowetting-on-dielectric (EWOD) to move droplets containing proteins or peptides and matrix to specific locations on an array of electrodes for analysis. Standard MALDI-MS reagents, analytes, concentrations, and recipes are demonstrated to be compatible with the technique. Mass spectra are comparable to those collected by conventional methods. Nonspecific adsorption of analytes to device surfaces is demonstrated to be negligible. The results suggest that EWOD may be a useful tool for automating sample preparation for high-throughput proteomics and other applications of MALDI-MS.

  10. Performance of matrix-assisted laser desorption-time of flight mass spectrometry for identification of clinical yeast isolates

    DEFF Research Database (Denmark)

    Rosenvinge, Flemming S; Dzajic, Esad; Knudsen, Elisa

    2013-01-01

    Accurate and fast yeast identification is important when treating patients with invasive fungal disease as susceptibility to antifungal agents is highly species related. Matrix-assisted laser desorption-time of flight mass spectrometry (MALDI-TOF-MS) provides a powerful tool with a clear potential...... to improve current diagnostic practice. Two MALDI-TOF-MS-systems (BioTyper/Bruker and Saramis/AXIMA) were evaluated using: (i) A collection of 102 archived, well characterised yeast isolates representing 14 different species and (ii) Prospectively collected isolates obtained from clinical samples at two...... identification, respectively, whereas the other laboratory identified 83/98 (85%) to species level by both BioTyper/Bruker and conventional identification. Both MALDI-TOF-MS systems are fast, have built-in databases that cover the majority of clinically relevant Candida species, and have an accuracy...

  11. Characterization of unknown compounds from stainless steel plates in matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Yang, Hyo-Jik; Park, Kyu Hwan; Kim, Hyun Sik; Kim, Jeongkwon

    2010-12-01

    Peaks originating from unknown compounds on stainless steel plates used in matrix-assisted laser desorption/ionization (MALDI) mass spectrometers are observed around m/z 304.3, 332.3, 360.4, and 388.4 regardless of the matrix and/or solvent, and are even observed with bare plates. These peaks were characterized using three different types of MALDI-MS instrumentation: MALDI-TOF MS, MALDI-TOF/TOF MS, and MALDI-FTMS. The fragmentation data from MALDI-TOF/TOF MS and accurate mass determination by MALDI-FTMS enabled identification of the chemical formulae and structures. The unknown compounds are, in fact, likely benzylalkylmethylammonium salts, as confirmed by closely matching fragmentation patterns with a commercially available benzalkonium chloride. Copyright © 2010 American Society for Mass Spectrometry. Published by Elsevier Inc. All rights reserved.

  12. Rapid discrimination of environmental Vibrio by matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Eddabra, Rkia; Prévost, Gilles; Scheftel, Jean-Michel

    2012-04-20

    The aim of this study was to discriminate 30 Vibrio strains isolated from two wastewater treatment plants from Agadir, Morocco by two molecular typing methods, pulsed-field gel electrophoresis (PFGE) and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Out of the 30 strains of Vibrio examined in this study, 5 isolates could not be typed by PFGE and consistently appeared as a smear on the gel. In general, high genetic biodiversity among the Vibrio strains was found regardless to the isolation source. The results of MALDI TOF analysis show a high congruence of strain grouping demonstrating the accuracy and reliability of MALDI-TOF MS. Copyright © 2011 Elsevier GmbH. All rights reserved.

  13. 5-Methoxysalicylic Acid Matrix for Ganglioside Analysis with Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry

    Science.gov (United States)

    Lee, Dongkun; Cha, Sangwon

    2015-03-01

    In this note, we report that high quality ganglioside profiles with minimal loss of sialic acid residues can be obtained in the positive ion mode by using a 5-methoxysalicylic acid (MSA) matrix for matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS). Our results showed that MSA produced much less sialic acid losses from gangliosides than DHB, although MSA and DHB are differ only by their functional groups at their 5-positions (-OH for DHB and -OCH3 for MSA). Furthermore, our data also demonstrated that addition of an alkali metal additive was effective for simplifying ganglioside profiles, but not necessary for stabilizing glycosidic bonds of gangliosides if MSA was used as a matrix. This suggests that MALDI MS with MSA has a potential to gain additional benefits from the positive-ion mode analyses without losing performance in ganglioside profiling.

  14. Generation of CsI cluster ions for mass calibration in matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Lou, Xianwen; van Dongen, Joost L J; Meijer, E W

    2010-07-01

    A simple method was developed for the generation of cesium iodide (CsI) cluster ions up to m/z over 20,000 in matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). Calibration ions in both positive and negative ion modes can readily be generated from a single MALDI spot of CsI(3) with 2-[(2E)-3-(4-tert-butylphenyl)-2-methylprop-2-enylidene] malononitrile (DCTB) matrix. The major cluster ion series observed in the positive ion mode is [(CsI)(n)Cs](+), and in the negative ion mode is [(CsI)(n)I](-). In both cluster series, ions spread evenly every 259.81 units. The easy method described here for the production of CsI cluster ions should be useful for MALDI MS calibrations. Copyright 2010 American Society for Mass Spectrometry. Published by Elsevier Inc. All rights reserved.

  15. On plate graphite supported sample processing for simultaneous lipid and protein identification by matrix assisted laser desorption ionization mass spectrometry.

    Science.gov (United States)

    Calvano, Cosima Damiana; van der Werf, Inez Dorothé; Sabbatini, Luigia; Palmisano, Francesco

    2015-05-01

    The simultaneous identification of lipids and proteins by matrix assisted laser desorption ionization-mass spectrometry (MALDI-MS) after direct on-plate processing of micro-samples supported on colloidal graphite is demonstrated. Taking advantages of large surface area and thermal conductivity, graphite provided an ideal substrate for on-plate proteolysis and lipid extraction. Indeed proteins could be efficiently digested on-plate within 15 min, providing sequence coverages comparable to those obtained by conventional in-solution overnight digestion. Interestingly, detection of hydrophilic phosphorylated peptides could be easily achieved without any further enrichment step. Furthermore, lipids could be simultaneously extracted/identified without any additional treatment/processing step as demonstrated for model complex samples such as milk and egg. The present approach is simple, efficient, of large applicability and offers great promise for protein and lipid identification in very small samples. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. Alkaloid profiling of the Chinese herbal medicine Fuzi by combination of matrix-assisted laser desorption ionization mass spectrometry with liquid chromatography-mass spectrometry

    NARCIS (Netherlands)

    Wang, J.; Heijden, R. van der; Spijksma, G.; Reijmers, T.; Wang, M.; Xu, G.; Hankemeier, T.; Greef, J. van der

    2009-01-01

    A matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS) method was developed for the high throughput and robust qualitative profiling of alkaloids in Fuzi-the processed lateral roots of the Chinese herbal medicine Aconitum carmichaeli Debx (A. carmichaeli). After optimization,

  17. Analysis of Phospholipid Mixtures from Biological Tissues by Matrix-Assisted Laser Desorption and Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS): A Laboratory Experiment

    Science.gov (United States)

    Eibisch, Mandy; Fuchs, Beate; Schiller, Jurgen; Sub, Rosmarie; Teuber, Kristin

    2011-01-01

    Matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF MS) is increasingly used to investigate the phospholipid (PL) compositions of tissues and body fluids, often without previous separation of the total mixture into the individual PL classes. Therefore, the questions of whether all PL classes are detectable…

  18. Gene analysis using mass spectrometric cleaved amplified polymorphic sequence (MS-CAPS) with matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF).

    Science.gov (United States)

    Kajiwara, Hideyuki

    2015-01-01

    Mass spectrometric cleaved amplified polymorphic sequence (MS-CAPS) is a method for detecting genes using a combination of short PCR and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). MS-CAPS can identify a single nucleotide polymorphism (SNP) in less than one hour and is suitable for plants, animals, bacteria, and food.

  19. Differentiation of Clinically Relevant mucorales Rhizopus microsporus and R. arrhizus by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS)

    NARCIS (Netherlands)

    Dolatabadi, S.; Kolecka, A.; Versteeg, Matthijs; de Hoog, Sybren G; Boekhout, Teun

    This study addresses the usefulness of Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) for reliable identification of the two most frequently occuring clinical species of Rhizopus, namely R. arrhizus with its two varieties arrhizus and delemar and R.

  20. Identification of barley and rye varieties using matrix- assisted laser desorption/ionisation time-of-flight mass spectrometry with neural networks

    DEFF Research Database (Denmark)

    Bloch, H.A.; Petersen, Marianne Kjerstine; Sperotto, Maria Maddalena

    2001-01-01

    developed, which combines analysis of alcohol-soluble wheat proteins (gliadins) using matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry with neural networks. Here we have applied the same method for the identification of both barley (Hordeum vulgare L.) and rye (Secale cereale L...

  1. Differentiation of Raoultella ornithinolytica/planticola and Klebsiella oxytoca clinical isolates by matrix-assisted laser desorption/ionization-time of flight mass spectrometry.

    NARCIS (Netherlands)

    Jong, Eefje de; Jong, A.S. de; Smidts-van den Berg, N.; Rentenaar, R.J.

    2013-01-01

    Ninety-nine clinical isolates previously identified as Klebsiella oxytoca were evaluated using matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS). Eight isolates were identified as Raoultella spp., being 5 Raoultella spp. and 3 K. oxytoca, by 16S rRNA

  2. Structural analysis of (methyl-esterified) oligogalacturonides using post-source decay matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

    NARCIS (Netherlands)

    Alebeek, van G.J.W.M.; Zabotina, O.; Beldman, G.; Schols, H.A.; Voragen, A.G.J.

    2000-01-01

    The use of post-source decay matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for the structural analysis of ((partly) methyl-esterified) oligogalacturonides (oligoGalA) is described. The fragmentation behavior of purified (un)saturated oligoGalA (degree of polymerization

  3. Direct analysis of triterpenes from high-salt fermented cucumbers using infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI)

    Science.gov (United States)

    High-salt samples present a challenge to mass spectrometry (MS) analysis, particularly when electrospray ionization (ESI) is used, requiring extensive sample preparation steps such as desalting, extraction, and purification. In this study, infrared matrix-assisted laser desorption electrospray ioniz...

  4. In situ liquid-liquid extraction as a sample preparation method for matrix-assisted laser desorption/ionization MS analysis of polypeptide mixtures

    DEFF Research Database (Denmark)

    Kjellström, Sven; Jensen, Ole Nørregaard

    2003-01-01

    A novel liquid-liquid extraction (LLE) procedure was investigated for preparation of peptide and protein samples for matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). LLE using ethyl acetate as the water-immiscible organic solvent enabled segregation of hydrophobic...

  5. Identification of multiple target sites for a glutathione conjugate on glutathione-S-transferase by matrix-assisted laser desorption/ionization mass spectrometry

    NARCIS (Netherlands)

    Jespersen, S.; Ploemen, J.H.T.M.; Bladeren, P.J.; Niessen, W.M.A.; Tjaden, U.R.; Greef, J. van der

    1996-01-01

    A mass spectrometric method providing qualitative site-specific information regarding covalent modification of proteins is described. The method involves comparison of unmodified and modified proteins by matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) peptide mapping in

  6. Distinct features of matrix-assisted 6 microm infrared laser desorption/ionization mass spectrometry in biomolecular analysis.

    Science.gov (United States)

    Tajiri, Michiko; Takeuchi, Takae; Wada, Yoshinao

    2009-08-15

    Midinfrared-matrix-assisted laser desorption/ionization mass spectrometry (mid-IR-MALDI MS) with a laser emission in the 6 microm wavelength range, which utilizes energy absorption at the C=O double-bond stretch region, was applied to biomolecular analysis. The softness of IR-MALDI MS was evident in the negative ion mode yielding clean mass spectra of [M - H](-) ions for acidic biomolecules with sulfate, phosphate, or carboxylate groups, resulting in better sensitivity than ultraviolet (UV)-MALDI MS. There was no substantial loss of sialic acid due to the prompt fragmentation occurring in IR-MALDI of sialylated glycoconjugates such as gangliosides. Furthermore, the advantage of the low photon energy of IR is that, for the first time, intact protonated molecules of S-nitrosylated peptides can be detected by MALDI MS. In the analysis of redox-sensitive molecules including methylene blue and riboflavin, reductive hydrogenation was minimal, suggesting few hydrogen radicals to have formed in the plume, in contrast to UV-MALDI. In conjunction with a potent new matrix, oxamide, requiring smaller laser fluence, distinct features of the 6 microm IR wavelength range are anticipated to remove one of the limitations of MALDI MS for biomolecular analysis.

  7. Identification and localization of trauma-related biomarkers using matrix assisted laser desorption/ionization imaging mass spectrometry

    Science.gov (United States)

    Jones, Kirstin; Reilly, Matthew A.; Glickman, Randolph D.

    2017-02-01

    Current treatments for ocular and optic nerve trauma are largely ineffective and may have adverse side effects; therefore, new approaches are needed to understand trauma mechanisms. Identification of trauma-related biomarkers may yield insights into the molecular aspects of tissue trauma that can contribute to the development of better diagnostics and treatments. The conventional approach for protein biomarker measurement largely relies on immunoaffinity methods that typically can only be applied to analytes for which antibodies or other targeting means are available. Matrix assisted laser-assisted desorption/ionization imaging mass spectrometry (MALDI-IMS) is a specialized application of mass spectrometry that not only is well suited to the discovery of novel or unanticipated biomarkers, but also provides information about the spatial localization of biomarkers in tissue. We have been using MALDI-IMS to find traumarelated protein biomarkers in retina and optic nerve tissue from animal models subjected to ocular injury produced by either blast overpressure or mechanical torsion. Work to date by our group, using MALDI-IMS, found that the pattern of protein expression is modified in the injured ocular tissue as soon as 24 hr post-injury, compared to controls. Specific proteins may be up- or down-regulated by trauma, suggesting different tissue responses to a given injury. Ongoing work is directed at identifying the proteins affected and mapping their expression in the ocular tissue, anticipating that systematic analysis can be used to identify targets for prospective therapies for ocular trauma.

  8. Basic poly(propylene glycols) as reference compounds for internal mass calibration in positive-ion matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Gross, Jürgen H

    2017-12-01

    Basic poly(propylene glycols), commercially available under the trade name Jeffamine, are evaluated for their potential use as internal mass calibrants in matrix-assisted laser desorption/ionization-time-of-flight-mass spectrometry. Due to their basic amino endgroups Jeffamines are expected to deliver [M+H] + ions in higher yields than neutral poly(propylene glycols) or poly(ethylene glycols). Aiming at accurate mass measurements and molecular formula determinations by matrix-assisted laser desorption/ionization-time-of-flight-mass spectrometry, four Jeffamines (M-600, M-2005, D-400, D-230) were thus compared. As a result, Jeffamine M-2005 is introduced as a new mass calibrant for positive-ion matrix-assisted laser desorption/ionization-time-of-flight-mass spectrometry in the range of m/z 200-1200 and the reference mass list is provided. While Jeffamine M-2005 is compatible with α-cyano-4-hydroxycinnamic acid, 2,5-dihydroxybenzoic acid, and 2-[(2 E)-3-(4- tert-butylphenyl)-2-methylprop-2-enylidene]malonitrile matrix, its use in combination with 2-[(2 E)-3-(4- tert-butylphenyl)-2-methylprop-2-enylidene]malonitrile provides best results due to low laser fluence requirements. Applications to PEG 300, PEG 600, the ionic liquid trihexyl(tetradecyl)-phosphonium tris(pentafluoroethyl)-trifluorophosphate, and [60]fullerene demonstrate mass accuracies of 2-5 ppm.

  9. Measurement of laser activated electron tunneling from semiconductor zinc oxide to adsorbed organic molecules by a matrix assisted laser desorption ionization mass spectrometer.

    Science.gov (United States)

    Zhong, Hongying; Fu, Jieying; Wang, Xiaoli; Zheng, Shi

    2012-06-04

    Measurement of light induced heterogeneous electron transfer is important for understanding of fundamental processes involved in chemistry, physics and biology, which is still challenging by current techniques. Laser activated electron tunneling (LAET) from semiconductor metal oxides was observed and characterized by a MALDI (matrix assisted laser desorption ionization) mass spectrometer in this work. Nanoparticles of ZnO were placed on a MALDI sample plate. Free fatty acids and derivatives were used as models of organic compounds and directly deposited on the surface of ZnO nanoparticles. Irradiation of UV laser (λ=355 nm) with energy more than the band gap of ZnO produces ions that can be detected in negative mode. When TiO(2) nanoparticles with similar band gap but much lower electron mobility were used, these ions were not observed unless the voltage on the sample plate was increased. The experimental results indicate that laser induced electron tunneling is dependent on the electron mobility and the strength of the electric field. Capture of low energy electrons by charge-deficient atoms of adsorbed organic molecules causes unpaired electron-directed cleavages of chemical bonds in a nonergodic pathway. In positive detection mode, electron tunneling cannot be observed due to the reverse moving direction of electrons. It should be able to expect that laser desorption ionization mass spectrometry is a new technique capable of probing the dynamics of electron tunneling. LAET offers advantages as a new ionization dissociation method for mass spectrometry. Copyright © 2012 Elsevier B.V. All rights reserved.

  10. Matrix-assisted laser desorption/ionization mass spectrometric analysis of aliphatic biodegradable photoluminescent polymers using new ionic liquid matrices.

    Science.gov (United States)

    Serrano, Carlos A; Zhang, Yi; Yang, Jian; Schug, Kevin A

    2011-05-15

    In this study, two novel ionic liquid matrices (ILMs), N,N-diisopropylethylammonium 3-oxocoumarate and N,N-diisopropylethylammonium dihydroxymonooxoacetophenoate, were tested for the structural elucidation of recently developed aliphatic biodegradable polymers by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). The polymers, formed by a condensation reaction of three components, citric acid, octane diol, and an amino acid, are fluorescent, but the exact mechanism behind their luminescent properties has not been fully elucidated. In the original studies, which introduced the polymer class (J. Yang et al., Proc. Natl. Acad. Sci. USA 2009, 106, 10086-10091), a hyper-conjugated cyclic structure was proposed as the source for the photoluminescent behavior. With the use of the two new ILMs, we present evidence that supports the presence of the proposed cyclization product. In addition, the new ILMs, when compared with a previously established ILM, N,N-diisopropylethylammonium α-cyano-3-hydroxycinnimate, provided similar signal intensities and maintained similar spectral profiles. This research also established that the new ILMs provided good spot-to-spot reproducibility and high ionization efficiency compared with corresponding crystalline matrix preparations. Many polymer features revealed through the use of the ILMs could not be observed with crystalline matrices. Ultimately, the new ILMs highlighted the composition of the synthetic polymers, as well as the loss of water that was expected for the formation of the proposed cyclic structure on the polymer backbone. Copyright © 2011 John Wiley & Sons, Ltd.

  11. Matrix Assisted Laser Desorption Ionization Mass Fingerprinting for Identification of Acacia Gum in Microsamples from Works of Art.

    Science.gov (United States)

    Granzotto, Clara; Sutherland, Ken

    2017-03-07

    This paper reports an improved method for the identification of Acacia gum in cultural heritage samples using matrix assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) after enzymatic digestion of the polysaccharide component. The analytical strategy was optimized using a reference Acacia gum (gum arabic, sp. A. senegal) and provided an unambiguous MS profile of the gum, characterized by specific and recognized oligosaccharides, from as little as 0.1 μg of material. The enhanced experimental approach with reduced detection limit was successfully applied to the analysis of naturally aged (∼80 year) gum arabic samples, pure and mixed with lead white pigment, and allowed the detection of gum arabic in samples from a late painting (1949/1954) by Georges Braque in the collection of the Art Institute of Chicago. This first application of the technique to characterize microsamples from a painting, in conjunction with analyses by gas chromatography/mass spectrometry (GC/MS), provided important insights into Braque's unusual mixed paint media that are also helpful to inform appropriate conservation treatments for his works. The robustness of the analytical strategy due to the reproducibility of the gum MS profile, even in the presence of other organic and inorganic components, together with the minimal sample size required, demonstrate the value of this new MALDI-TOF MS method as an analytical tool for the identification of gum arabic in microsamples from museum artifacts.

  12. Newborn screening by matrix-assisted laser desorption/ionization mass spectrometry based on parylene-matrix chip.

    Science.gov (United States)

    Kim, Jo-Il; Noh, Joo-Yoon; Kim, Mira; Park, Jong-Min; Song, Hyun-Woo; Kang, Min-Jung; Pyun, Jae-Chul

    2017-08-01

    Newborn screening for diagnosis of phenylketonuria, homocystinuria, and maple syrup urine disease have been conducted by analyzing the concentration of target amino acids using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-ToF MS) based on parylene-matrix chip. Parylene-matrix chip was applied to MALDI-ToF MS analysis reducing the matrix peaks significantly at low mass-to-charge ratio range (m/z mass spectrometric analysis showing 13.3 to 45% of extraction efficiency. Calibration curves for diagnosis of neonatal metabolic disorders were obtained by analyzing methanol-extracted serum spiked with target amino acids using MALDI-ToF MS. They showed good linearity (R2 > 0.98) and the LODs were ranging from 9.0 to 22.9 μg/mL. Effect of proteins in serum was estimated by comparing MALDI-ToF mass spectra of amino acids-spiked serum before and after the methanol extraction. Interference of other amino acids on analysis of target analyte was determined to be insignificant. From these results, MALDI-ToF MS based on parylene-matrix chip could be applicable to medical diagnosis of neonatal metabolic disorders. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. Differentiation of virulence of Helicobacter pylori by matrix-assisted laser desorption/ionization mass spectrometry and multivariate analyses.

    Science.gov (United States)

    Cho, Yi-Tzu; Kuo, Chao-Hung; Wang, Sophie S W; Chen, Yu-Syuan; Weng, Bi-Chuang; Lee, Yi-Chern; Cheng, Chu-Nian; Shiea, Jentaie; Wu, Deng-Chyang

    2013-09-23

    The ability to determine the virulence of a Helicobacter pylori strain would be helpful for predicting the development of gastrointestinal disease and suggesting medical treatment. A protocol based on matrix-assisted laser desorption ionization/time-of-flight mass spectrometry (MALDI-TOF MS) was established for the efficient detection of peptides and proteins in extracts of H. pylori cells. Two multivariate statistical methods-principal component analysis (PCA) and hierarchical clustering analysis-were used to analyze the resulting MALDI mass spectra of reference strains and clinical isolated/inoculated strains. Based on differences in their peptide and protein profiles, H. pylori strains having similar virulence genotypes were grouped together on the PCA score plot. Hierarchical cluster analysis revealed high conformity between the protein profiles and the respective virulence genotypes. The inoculated H. pylori strain, which was clustered in the same group with the high-virulence reference strains, also resulted in severe histopathological lesions in gerbils. MALDI-TOF MS combined with multivariate analyses shows the ability to rapidly differentiate H. pylori strains in terms of their virulence. Copyright © 2013 Elsevier B.V. All rights reserved.

  14. Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry applied to virus identification.

    Science.gov (United States)

    Calderaro, Adriana; Arcangeletti, Maria-Cristina; Rodighiero, Isabella; Buttrini, Mirko; Gorrini, Chiara; Motta, Federica; Germini, Diego; Medici, Maria-Cristina; Chezzi, Carlo; De Conto, Flora

    2014-10-30

    Virus detection and/or identification traditionally rely on methods based on cell culture, electron microscopy and antigen or nucleic acid detection. These techniques are good, but often expensive and/or time-consuming; furthermore, they not always lead to virus identification at the species and/or type level. In this study, Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) was tested as an innovative tool to identify human polioviruses and to identify specific viral protein biomarkers in infected cells. The results revealed MALDI-TOF MS to be an effective and inexpensive tool for the identification of the three poliovirus serotypes. The method was firstly applied to Sabin reference strains, and then to isolates from different clinical samples, highlighting its value as a time-saving, sensitive and specific technique when compared to the gold standard neutralization assay and casting new light on its possible application to virus detection and/or identification.

  15. Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry in Clinical Microbiology: What Are the Current Issues?

    Science.gov (United States)

    Welker, Martin; Pincus, David; Charrier, Jean-Philippe; Girard, Victoria

    2017-01-01

    Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has revolutionized the identification of microbial species in clinical microbiology laboratories. MALDI-TOF-MS has swiftly become the new gold-standard method owing to its key advantages of simplicity and robustness. However, as with all new methods, adoption of the MALDI-TOF MS approach is still not widespread. Optimal sample preparation has not yet been achieved for several applications, and there are continuing discussions on the need for improved database quality and the inclusion of additional microbial species. New applications such as in the field of antimicrobial susceptibility testing have been proposed but not yet translated to the level of ease and reproducibility that one should expect in routine diagnostic systems. Finally, during routine identification testing, unexpected results are regularly obtained, and the best methods for transmitting these results into clinical care are still evolving. We here discuss the success of MALDI-TOF MS in clinical microbiology and highlight fields of application that are still amenable to improvement. PMID:28840984

  16. Rapid assignment of malting barley varieties by matrix-assisted laser desorption-ionisation - Time-of-flight mass spectrometry.

    Science.gov (United States)

    Šedo, Ondrej; Kořán, Michal; Jakešová, Michaela; Mikulíková, Renata; Boháč, Michal; Zdráhal, Zbyněk

    2016-09-01

    A method for discriminating malting barley varieties based on direct matrix-assisted laser desorption-ionisation - time-of-flight mass spectrometry (MALDI-TOF MS) fingerprinting of proteins was developed. Signals corresponding to hordeins were obtained by simple mixing of powdered barley grain with a MALDI matrix solution containing 12.5mgmL(-1) of ferulic acid in an acetonitrile:water:formic acid 50:33:17 v/v/v mixture. Compared to previous attempts at MALDI-TOF mass spectrometric analysis of barley proteins, the extraction and fractionation steps were practically omitted, resulting in a significant reduction in analytical time and costs. The discriminatory power was examined on twenty malting barley varieties and the practicability of the method was tested on sixty barley samples acquired from Pilsner Urquell Brewery. The method is proposed as a rapid tool for variety assignment and purity determination of malting barley that may replace gel electrophoresis currently used for this purpose. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. Identification of Leishmania at the species level with matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Cassagne, C; Pratlong, F; Jeddi, F; Benikhlef, R; Aoun, K; Normand, A-C; Faraut, F; Bastien, P; Piarroux, R

    2014-06-01

    Matrix-assisted laser desorption ionization time-of-flightMALDI-TOF mass spectrometry (MS) is now widely recognized as a powerful tool with which to identify bacteria and fungi at the species level, and sometimes in a rapid and accurate manner. We report herein an approach to identify, at the species level, Leishmania promastigotes from in vitro culture. We first constructed a reference database of spectra including the main Leishmania species known to cause human leishmaniasis. Then, the performance of the reference database in identifying Leishmania promastigotes was tested on a panel of 69 isolates obtained from patients. Our approach correctly identified 66 of the 69 isolates tested at the species level with log (score) values superior to 2. Two Leishmania isolates yielded non-interpretable MALDI-TOF MS patterns, owing to low log (score) values. Only one Leishmania isolate of Leishmania peruviana was misidentified as the closely related species Leishmania braziliensis, with a log (score) of 2.399. MALDI-TOF MS is a promising approach, providing rapid and accurate identification of Leishmania from in vitro culture at the species level. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  18. Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry in Clinical Microbiology: What Are the Current Issues?

    Science.gov (United States)

    van Belkum, Alex; Welker, Martin; Pincus, David; Charrier, Jean Philippe; Girard, Victoria

    2017-11-01

    Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has revolutionized the identification of microbial species in clinical microbiology laboratories. MALDI-TOF-MS has swiftly become the new gold-standard method owing to its key advantages of simplicity and robustness. However, as with all new methods, adoption of the MALDI-TOF MS approach is still not widespread. Optimal sample preparation has not yet been achieved for several applications, and there are continuing discussions on the need for improved database quality and the inclusion of additional microbial species. New applications such as in the field of antimicrobial susceptibility testing have been proposed but not yet translated to the level of ease and reproducibility that one should expect in routine diagnostic systems. Finally, during routine identification testing, unexpected results are regularly obtained, and the best methods for transmitting these results into clinical care are still evolving. We here discuss the success of MALDI-TOF MS in clinical microbiology and highlight fields of application that are still amenable to improvement. © The Korean Society for Laboratory Medicine.

  19. Improved analysis of membrane protein by PVDF-aided, matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Chang, Chih-Yang; Liao, Hsin-Kai; Juo, Chiun-Gung; Chen, Shu-Hua; Chen, Yu-Ju

    2006-01-18

    Characterization of membrane proteins remains an analytical challenge because of difficulties associated with tedious isolation and purification. This study presents the utility of the polyvinylidene difluoride (PVDF) membrane for direct sub-proteome profiling and membrane protein characterization by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). The hydrophobic adsorption of protein, particularly membrane proteins, on the PVDF surface enables efficient on-PVDF washing to remove high concentrations of detergents and salts, such as up to 5% sodium dodecyl sulfate (SDS). The enhanced spectrum quality for MALDI detection is particularly notable for high molecular weight proteins. By using on-PVDF washing prior to MALDI detection, we obtained protein profiles of the detergent-containing and detergent-insoluble membrane fractions from Methylococcus capsulatus (Bath). Similar improvements of signal-to-noise ratios were shown on the MALDI spectra for proteins electroblotted from SDS-polyacrylamide gel electrophoresis (SDS-PAGE) onto the PVDF membrane. We have applied this strategy to obtain intact molecular weights of the particulate methane monooxygenase (pMMO) composed of three intrinsic membrane-bound proteins, PmoA, PmoB, and PmoC. Together with peptide sequencing by tandem mass spectrometry, post-translational modifications including N-terminal acetylation of PmoA and PmoC and alternative C-terminal truncation of PmoB were identified. The above results show that PVDF-aided MALDI-MS can be an effective approach for profiling and characterization of membrane proteins.

  20. Principles of hydrogen radical mediated peptide/protein fragmentation during matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Asakawa, Daiki

    2016-07-01

    Matrix-assisted laser desorption/ionization in-source decay (MALDI-ISD) is a very easy way to obtain large sequence tags and, thereby, reliable identification of peptides and proteins. Recently discovered new matrices have enhanced the MALDI-ISD yield and opened new research avenues. The use of reducing and oxidizing matrices for MALDI-ISD of peptides and proteins favors the production of fragmentation pathways involving "hydrogen-abundant" and "hydrogen-deficient" radical precursors, respectively. Since an oxidizing matrix provides information on peptide/protein sequences complementary to that obtained with a reducing matrix, MALDI-ISD employing both reducing and oxidizing matrices is a potentially useful strategy for de novo peptide sequencing. Moreover, a pseudo-MS(3) method provides sequence information about N- and C-terminus extremities in proteins and allows N- and C-terminal side fragments to be discriminated within the complex MALDI-ISD mass spectrum. The combination of high mass resolution of a Fourier transform-ion cyclotron resonance (FTICR) analyzer and the software suitable for MALDI-ISD facilitates the interpretation of MALDI-ISD mass spectra. A deeper understanding of the MALDI-ISD process is necessary to fully exploit this method. Thus, this review focuses first on the mechanisms underlying MALDI-ISD processes, followed by a discussion of MALDI-ISD applications in the field of proteomics. © 2014 Wiley Periodicals, Inc., Mass Spec Rev 35:535-556, 2016. © 2014 Wiley Periodicals, Inc.

  1. Silver cluster interferences in matrix-assisted laser desorption/ionization (MALDI) mass spectrometry of nonpolar polymers.

    Science.gov (United States)

    Macha, S F; Limbach, P A; Hanton, S D; Owens, K G

    2001-06-01

    Potential difficulties associated with background silver salt clusters during matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) of nonpolar polymers are reported. Silver salt cluster ions were observed from m/z 1500 to 7000 when acidic, polar matrices, such as 2,5-dihydroxybenzoic acid (DHB), all-trans-retinoic acid (RTA) or 2-(4-hydroxyphenylazo)benzoic acid (HABA), were used for the analysis of nonpolar polymers. These background signals could be greatly reduced or eliminated by the use of nonpolar matrices such as anthracene or pyrene. Representative examples of these background interferences are demonstrated during the analysis of low molecular weight nonpolar polymers including polybutadiene and polystyrene. Nonpolar polymers analyzed with acidic, polar matrices (e.g., RTA) and silver cationization reagents can yield lower quality mass spectral results when interferences due to silver clusters are present. Replacing the polar matrices with nonpolar matrices or the silver salts with copper salts substantially improved the quality of the analytical results. In addition, it was found that silver contamination cannot be completely removed from standard stainless steel sample plates, although the presence of silver contamination was greatly reduced after thorough cleaning of the sample plate with aluminum oxide grit. Carry-over silver may cationize polymer samples and complicate the interpretation of data obtained using nonpolar matrices in the absence of added cationization reagents.

  2. Adduct simplification in the analysis of cyanobacterial toxins by matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Howard, Karen L; Boyer, Gregory L

    2007-01-01

    A novel method for simplifying adduct patterns to improve the detection and identification of peptide toxins using matrix-assisted laser desorption/ionization (MALDI) time-of-flight (TOF) mass spectrometry is presented. Addition of 200 microM zinc sulfate heptahydrate (ZnSO(4) . 7H(2)O) to samples prior to spotting on the target enhances detection of the protonated molecule while suppressing competing adducts. This produces a highly simplified spectrum with the potential to enhance quantitative analysis, particularly for complex samples. The resulting improvement in total signal strength and reduction in the coefficient of variation (from 31.1% to 5.2% for microcystin-LR) further enhance the potential for sensitive and accurate quantitation. Other potential additives tested, including 18-crown-6 ether, alkali metal salts (lithium chloride, sodium chloride, potassium chloride), and other transition metal salts (silver chloride, silver nitrate, copper(II) nitrate, copper(II) sulfate, zinc acetate), were unable to achieve comparable results. Application of this technique to the analysis of several microcystins, potent peptide hepatotoxins from cyanobacteria, is illustrated. Copyright (c) 2007 John Wiley & Sons, Ltd.

  3. Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry applied to virus identification

    Science.gov (United States)

    Calderaro, Adriana; Arcangeletti, Maria-Cristina; Rodighiero, Isabella; Buttrini, Mirko; Gorrini, Chiara; Motta, Federica; Germini, Diego; Medici, Maria-Cristina; Chezzi, Carlo; De Conto, Flora

    2014-01-01

    Virus detection and/or identification traditionally rely on methods based on cell culture, electron microscopy and antigen or nucleic acid detection. These techniques are good, but often expensive and/or time-consuming; furthermore, they not always lead to virus identification at the species and/or type level. In this study, Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) was tested as an innovative tool to identify human polioviruses and to identify specific viral protein biomarkers in infected cells. The results revealed MALDI-TOF MS to be an effective and inexpensive tool for the identification of the three poliovirus serotypes. The method was firstly applied to Sabin reference strains, and then to isolates from different clinical samples, highlighting its value as a time-saving, sensitive and specific technique when compared to the gold standard neutralization assay and casting new light on its possible application to virus detection and/or identification. PMID:25354905

  4. Peptide Profiling Using Matrix-Assisted Laser Desorption/Ionization-Time-of-Flight Mass Spectrometry for Identification of Animal Fibers.

    Science.gov (United States)

    Izuchi, Yukari; Tokuhara, Mutsumi; Takashima, Tsuneo; Kuramoto, Kanya

    2013-01-01

    Identification of fibers for verification of their specific animal origin is necessary for maintaining quality and value in the clothing industry. In order to examine adulteration in animal fibers, there is a commercially accepted method of microscopy analysis. However, this method is subjective and time-consuming due to its reliance on an operator identifying magnified fibers from their scale image and other features. Therefore, alternative reliable identification methods are required. In this study, peptide analysis using matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOFMS) is presented and used to distinguish between cashmere, wool, mohair, yak, camel, angora, and alpaca in untreated and treated fibers (dyed, chlorinated wool). Typical m/z values for each specific type of animal fiber were identified. Predictive models that could identify seven types of animal fibers as well as 50% blended samples were successfully constructed using multivariate analyses such as PCA and PLS regression. This technique is therefore extremely useful for complementing the conventional tests for detecting adulteration in animal fiber fabrics and clothing.

  5. Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry Identification of Mycobacteria in Routine Clinical Practice

    Science.gov (United States)

    El Khéchine, Amel; Couderc, Carine; Flaudrops, Christophe; Raoult, Didier; Drancourt, Michel

    2011-01-01

    Background Non-tuberculous mycobacteria recovered from respiratory tract specimens are emerging confounder organisms for the laboratory diagnosis of tuberculosis worldwide. There is an urgent need for new techniques to rapidly identify mycobacteria isolated in clinical practice. Matrix-assisted laser desorption time-of-flight mass spectrometry (MALDI-TOF MS) has previously been proven to effectively identify mycobacteria grown in high-concentration inocula from collections. However, a thorough evaluation of its use in routine laboratory practice has not been performed. Methodology We set up an original protocol for the MALDI-TOF MS identification of heat-inactivated mycobacteria after dissociation in Tween-20, mechanical breaking of the cell wall and protein extraction with formic acid and acetonitrile. By applying this protocol to as few as 105 colony-forming units of reference isolates of Mycobacterium tuberculosis, Mycobacterium avium, and 20 other Mycobacterium species, we obtained species-specific mass spectra for the creation of a local database. Using this database, our protocol enabled the identification by MALDI-TOF MS of 87 M. tuberculosis, 25 M. avium and 12 non-tuberculosis clinical isolates with identification scores ≥2 within 2.5 hours. Conclusions Our data indicate that MALDI-TOF MS can be used as a first-line method for the routine identification of heat-inactivated mycobacteria. MALDI-TOF MS is an attractive method for implementation in clinical microbiology laboratories in both developed and developing countries. PMID:21935444

  6. The protein profile of Theobroma cacao L. seeds as obtained by matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Bertazzo, Antonella; Agnolin, Fabio; Comai, Stefano; Zancato, Mirella; Costa, Carlo V L; Seraglia, Roberta; Traldi, Pietro

    2011-07-30

    The water-soluble protein profile of the seeds of green, red, and yellow Theobroma cacao L. fruits has been determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-ToF-MS). The seeds were powdered under liquid nitrogen and defatted. The residues were dialyzed and lyophilized. The obtained samples were suspended in the matrix solution of sinapinic acid. The obtained MALDI mass spectra showed the presence of a wide number of proteins with molecular weight ranging from 8000 to 13,000 Da and a cluster of peaks centered at 21,000 Da that were attributed to albumin. The abundance of this peak was found to depend on the different portion of the seed (husk, apical and cortical parts); however, the MALDI mass spectra obtained from the different varieties of cocoa were practically superimposable. Changes in the protein profiles were also observed after the cocoa seeds were treated by fermentation and roasting, which are processes usually employed for the commercial production of cocoa. Copyright © 2011 John Wiley & Sons, Ltd.

  7. Analysis of Microbial Mixtures by Matrix-assisted Laser Desorption/Ionization time-of-flight Mass Spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Wahl, Karen L.; Wunschel, Sharon C.; Jarman, Kristin H.; Valentine, Nancy B.; Petersen, Catherine E.; Kingsley, Mark T.; Zartolas, Kimberly A.; Saenz, Adam J.

    2002-12-15

    Many different laboratories are currently developing mass-spectrometric techniques to analyze and identify microorganisms. However, minimal work has been done with mixtures of bacteria. To demonstrate that microbial mixtures could be analyzed by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), mixed bacterial cultures were analyzed in a double-blind fashion. Nine different bacterial species currently in our MALDI-MS fingerprint library were used to generate 50 different simulated mixed bacterial cultures similar to that done for an initial blind study previously reported.(1) The samples were analyzed by MALDI-MS with automated data extraction and analysis algorithms developed in our laboratory. The components present in the sample were identified correctly to the species level in all but one of the samples. However, correctly eliminating closely related organisms was challenging for the current algorithms, especially in differentiating Serratia marcescens, Escherichia coli, and Yersinia enterocolitica, which have some similarities in their MALDI-MS fingerprints. Efforts to improve the specificity of the algorithms are in progress.

  8. Determination of paclitaxel distribution in solid tumors by nano-particle assisted laser desorption ionization mass spectrometry imaging.

    Science.gov (United States)

    Morosi, Lavinia; Spinelli, Pietro; Zucchetti, Massimo; Pretto, Francesca; Carrà, Andrea; D'Incalci, Maurizio; Giavazzi, Raffaella; Davoli, Enrico

    2013-01-01

    A sensitive, simple and reproducible protocol for nanoparticle-assisted laser desorption/ionization mass spectrometry imaging technique is described. The use of commercially available TiO2 nanoparticles abolishes heterogeneous crystallization, matrix background interferences and enhances signal detection, especially in the low mass range. Molecular image normalization was based on internal standard deposition on tissues, allowing direct comparison of drug penetration and distribution between different organs and tissues. The method was applied to analyze the distribution of the anticancer drug paclitaxel, inside normal and neoplastic mouse tissue sections. Spatial resolution was good, with a linear response between different in vivo treatments and molecular imaging intensity using therapeutic drug doses. This technique distinguishes the different intensity of paclitaxel distribution in control organs of mice, such as liver and kidney, in relation to the dose. Animals treated with 30 mg/kg of paclitaxel had half of the concentration of those treated with 60 mg/kg. We investigated the spatial distribution of paclitaxel in human melanoma mouse xenografts, following different dosage schedules and found a more homogeneous drug distribution in tumors of mice given repeated doses (5×8 mg/kg) plus a 60 mg/kg dose than in those assigned only a single 60 mg/kg dose. The protocol can be readily applied to investigate anticancer drug distribution in neoplastic lesions and to develop strategies to optimize and enhance drug penetration through different tumor tissues.

  9. Determination of paclitaxel distribution in solid tumors by nano-particle assisted laser desorption ionization mass spectrometry imaging.

    Directory of Open Access Journals (Sweden)

    Lavinia Morosi

    Full Text Available A sensitive, simple and reproducible protocol for nanoparticle-assisted laser desorption/ionization mass spectrometry imaging technique is described. The use of commercially available TiO2 nanoparticles abolishes heterogeneous crystallization, matrix background interferences and enhances signal detection, especially in the low mass range. Molecular image normalization was based on internal standard deposition on tissues, allowing direct comparison of drug penetration and distribution between different organs and tissues. The method was applied to analyze the distribution of the anticancer drug paclitaxel, inside normal and neoplastic mouse tissue sections. Spatial resolution was good, with a linear response between different in vivo treatments and molecular imaging intensity using therapeutic drug doses. This technique distinguishes the different intensity of paclitaxel distribution in control organs of mice, such as liver and kidney, in relation to the dose. Animals treated with 30 mg/kg of paclitaxel had half of the concentration of those treated with 60 mg/kg. We investigated the spatial distribution of paclitaxel in human melanoma mouse xenografts, following different dosage schedules and found a more homogeneous drug distribution in tumors of mice given repeated doses (5×8 mg/kg plus a 60 mg/kg dose than in those assigned only a single 60 mg/kg dose. The protocol can be readily applied to investigate anticancer drug distribution in neoplastic lesions and to develop strategies to optimize and enhance drug penetration through different tumor tissues.

  10. Usefulness of matrix-assisted laser desorption ionisation-time-of-flight mass spectrometry for identifying clinical Trichosporon isolates.

    Science.gov (United States)

    de Almeida Júnior, J N; Figueiredo, D S Y; Toubas, D; Del Negro, G M B; Motta, A L; Rossi, F; Guitard, J; Morio, F; Bailly, E; Angoulvant, A; Mazier, D; Benard, G; Hennequin, C

    2014-08-01

    Trichosporon spp. have recently emerged as significant human pathogens. Identification of these species is important, both for epidemiological purposes and for therapeutic management, but conventional identification based on biochemical traits is hindered by the lack of updates to the species databases provided by the different commercial systems. In this study, 93 strains, or isolates, belonging to 16 Trichosporon species were subjected to both molecular identification using IGS1 gene sequencing and matrix-assisted laser desorption ionisation-time-of-flight (MALDI-TOF) analysis. Our results confirmed the limits of biochemical systems for identifying Trichosporon species, because only 27 (36%) of the isolates were correctly identified using them. Different protein extraction procedures were evaluated, revealing that incubation for 30 min with 70% formic acid yields the spectra with the highest scores. Among the six different reference spectra databases that were tested, a specific one composed of 18 reference strains plus seven clinical isolates allowed the correct identification of 67 of the 68 clinical isolates (98.5%). Although until recently it has been less widely applied to the basidiomycetous fungi, MALDI-TOF appears to be a valuable tool for identifying clinical Trichosporon isolates at the species level. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  11. Enhanced sensitivity for high spatial resolution lipid analysis by negative ion mode matrix assisted laser desorption ionization imaging mass spectrometry.

    Science.gov (United States)

    Angel, Peggi M; Spraggins, Jeffrey M; Baldwin, H Scott; Caprioli, Richard

    2012-02-07

    We have achieved enhanced lipid imaging to a ~10 μm spatial resolution using negative ion mode matrix assisted laser desorption ionization (MALDI) imaging mass spectrometry, sublimation of 2,5-dihydroxybenzoic acid as the MALDI matrix, and a sample preparation protocol that uses aqueous washes. We report on the effect of treating tissue sections by washing with volatile buffers at different pHs prior to negative ion mode lipid imaging. The results show that washing with ammonium formate, pH 6.4, or ammonium acetate, pH 6.7, significantly increases signal intensity and number of analytes recorded from adult mouse brain tissue sections. Major lipid species measured were glycerophosphoinositols, glycerophosphates, glycerolphosphoglycerols, glycerophosphoethanolamines, glycerophospho-serines, sulfatides, and gangliosides. Ion images from adult mouse brain sections that compare washed and unwashed sections are presented and show up to 5-fold increases in ion intensity for washed tissue. The sample preparation protocol has been found to be applicable across numerous organ types and significantly expands the number of lipid species detectable by imaging mass spectrometry at high spatial resolution. © 2012 American Chemical Society

  12. Imaging of Phospholipids in Formalin Fixed Rat Brain Sections by Matrix Assisted Laser Desorption/Ionization Mass Spectrometry

    Science.gov (United States)

    Carter, Claire L.; McLeod, Cameron W.; Bunch, Josephine

    2011-11-01

    Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is a valuable tool for the analysis of molecules directly from tissue. Imaging of phospholipids is gaining widespread interest, particularly as these lipids have been implicated in a variety of pathologic processes. Formalin fixation (FF) is the standard protocol used in histology laboratories worldwide to preserve tissue for analysis, in order to aid in the diagnosis and prognosis of diseases. This study assesses MALDI imaging of phospholipids directly in formalin fixed tissue, with a view to future analysis of archival tissue. This investigation proves the viability of MALDI-MSI for studying the distribution of lipids directly in formalin fixed tissue, without any pretreatment protocols. High quality molecular images for several phosphatidylcholine (PC) and sphingomyelin (SM) species are presented. Images correspond well with previously published data for the analysis of lipids directly from freshly prepared tissue. Different ionization pathways are observed when analyzing fixed tissue compared with fresh, and this change was found to be associated with formalin buffers employed in fixation protocols. The ability to analyze lipids directly from formalin fixed tissue opens up new doors in the investigation of disease profiles. Pathologic specimens taken for histologic investigation can be analyzed by MALDI-MS to provide greater information on the involvement of lipids in diseased tissue.

  13. Novel ionic liquid matrices for qualitative and quantitative detection of carbohydrates by matrix assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Zhao, Xiaoyong; Shen, Shanshan; Wu, Datong; Cai, Pengfei; Pan, Yuanjiang

    2017-09-08

    Analysis of carbohydrates based on matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is still challenging and researchers have been devoting themselves to efficient matrices discovery. In the present study, the design, synthesis, qualitative and quantitative performance of non-derivative ionic liquid matrices (ILMs) were reported. DHB/N-methylaniline (N-MA) and DHB/N-ethylaniline (N-EA), performing best for carbohydrate detection, have been screened out. The limit of detection for oligosaccharide provided by DHB/N-MA and DHB/N-EA were as low as 10 fmol. DHB/N-MA and DHB/N-EA showed significantly higher ion generation efficiency than DHB. The comparison of capacity to probe polysaccharide between these two ILMs and DHB also revealed their powerful potential. Their outstanding performance were probably due to lower proton affinities and stronger UV absorption at λ = 355 nm. What is more, taking DHB/N-MA as an example, quantitative analysis of fructo-oligosaccharide mixtures extracted and identified from rice noodles has been accomplished sensitively using an internal standard method. Overall, DHB/N-MA and DHB/N-EA exhibited excellent performance and might be significant sources as the carbohydrate matrices. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Filamentous fungal characterizations by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Santos, C; Paterson, R R M; Venâncio, A; Lima, N

    2010-02-01

    Matrix-assisted laser desorption/ionization time-of-flight intact cell mass spectrometry (MALDI-TOF ICMS) is coming of age for the identification and characterization of fungi. The procedure has been used extensively with bacteria. UV-absorbing matrices function as energy mediators that transfer the absorbed photoenergy from an irradiation source to the surrounding sample molecules, resulting in minimum fragmentation. A surprisingly high number of fungal groups have been studied: (i) the terverticillate penicillia, (ii) aflatoxigenic, black and other aspergilli, (iii) Fusarium, (iv) Trichoderma, (iv) wood rotting fungi (e.g. Serpula lacrymans) and (v) dermatophytes. The technique has been suggested for optimizing quality control of fungal Chinese medicines (e.g. Cordyceps). MALDI-TOF ICMS offers advantages over PCR. The method is now used in taxonomic assessments (e.g. Trichoderma) as distinct from only strain characterization. Low and high molecular mass natural products (e.g. peptaibols) can be analysed. The procedure is rapid and requires minimal pretreatment. However, issues of reproducibility need to be addressed further in terms of strains of species tested and between run variability. More studies into the capabilities of MALDI-TOF ICMS to identify fungi are required.

  15. Characterization of some synthetic Ru and Ir complexes by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

    NARCIS (Netherlands)

    Lou, X.; Buijtenen, J. van; Bastiaansen, J.J.A.M.; Waal, B.F.M. de; Langeveld, B.M.W.; Dongen, J.L.J. van

    2005-01-01

    Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) was applied to the analysis of Ru(OCOCF 3)2(CO)(PPh3)2, Ru(OCOC 3F7)2(CO)(PPh3)2, Ir(tBuppy)3 and Ir(ppy)2(acac) complexes. A troublesome problem in the MALDI-TOFMS characterization of these metal complexes is

  16. Influence of Culture Media on Detection of Carbapenem Hydrolysis by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry.

    Science.gov (United States)

    Ramos, Ana Carolina; Carvalhaes, Cecília Godoy; Cordeiro-Moura, Jhonatha Rodrigo; Rockstroh, Anna Carolina; Machado, Antonia Maria Oliveira; Gales, Ana Cristina

    2016-07-01

    In this study, we evaluated the influence of distinct bacterial growth media on detection of carbapenemase hydrolysis by matrix-assisted laser desorption ionization-time of flight mass spectrometry. False-negative results were observed for OXA-25-, OXA-26-, and OXA-72-producing Acinetobacter baumannii isolates grown on MacConkey agar medium. The other culture media showed 100% sensitivity and 100% specificity for detecting carbapenemase. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  17. Identification of different respiratory viruses, after a cell culture step, by matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS)

    OpenAIRE

    Adriana Calderaro; Maria Cristina Arcangeletti; Isabella Rodighiero; Mirko Buttrini; Sara Montecchini; Rosita Vasile Simone; Maria Cristina Medici; Carlo Chezzi; Flora De Conto

    2016-01-01

    In this study matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS), a reliable identification method for the diagnosis of bacterial and fungal infections, is presented as an innovative tool to investigate the protein profile of cell cultures infected by the most common viruses causing respiratory tract infections in humans. MALDI-TOF MS was applied to the identification of influenza A and B viruses, adenovirus C species, parainfluenza virus types 1, 2 an...

  18. MIL-101(Cr) as matrix for sensitive detection of quercetin by matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Han, Guobin; Zeng, Qiaoling; Jiang, Zhongwei; Xing, Tiantian; Huang, Chengzhi; Li, Yuanfang

    2017-03-01

    Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) has been proven as a useful and advanced technique in the identification of polymers and proteins. However, MALDI-TOF-MS still has the unavoidable drawback of self-signal interference with traditional organic matrices, which could suppress and overlap with the analyte signals in the low-mass region. In this work, MIL-101(Cr), a kind of metal-organic frameworks which possess high molecular weight, π-conjugated 3-D structure, coordinately unsaturated chromium sites (CUS) and strong absorption in the UV range, was employed to replace traditional organic matrices, and it was found that MIL-101(Cr) can dramatically eliminate the background peaks, showing high signal-to-noise level in the analysis of small molecules. As proof-of-concept, quercetin, daidzein, genistein and naringenin, members of flavonol family which widely exists in food and natural products, were successfully determined by utilizing MIL-101(Cr) as the surface-assisted matrix, and the detection of quercetin was sensitive with good salt tolerance and reproducibility. Under optimal conditions, the mass peak intensity exhibited good linear relationships in the range from 0.25µg/mL-7.00µg/mL for quercetin (R2=0.996) with detection limit 2.11ng/mL (3σ/k), making the identification of quercetin in sophora japonica successfully. With this strategy we have demonstrated the potentiality of MIL-101(Cr) nanomaterials as MALDI-MS matrix for the detection of small molecules. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. Rapid Identification of the Foodborne Pathogen Trichinella spp. by Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry.

    Science.gov (United States)

    Mayer-Scholl, Anne; Murugaiyan, Jayaseelan; Neumann, Jennifer; Bahn, Peter; Reckinger, Sabine; Nöckler, Karsten

    2016-01-01

    Human trichinellosis occurs through consumption of raw or inadequately processed meat or meat products containing larvae of the parasitic nematodes of the genus Trichinella. Currently, nine species and three genotypes are recognized, of which T. spiralis, T. britovi and T. pseudospiralis have the highest public health relevance. To date, the differentiation of the larvae to the species and genotype level is based primarily on molecular methods, which can be relatively time consuming and labor intensive. Due to its rapidness and ease of use a matrix assisted laser desorption / ionization time of flight mass spectrometry (MALDI-TOF MS) reference spectra database using Trichinella strains of all known species and genotypes was created. A formicacid/acetonitrile protein extraction was carried out after pooling 10 larvae of each Trichinella species and genotype. Each sample was spotted 9 times using α-cyano 4-hydoxy cinnamic acid matrix and a MicroFlex LT mass spectrometer was used to acquire 3 spectra (m/z 2000 to 20000 Da) from each spot resulting in 27 spectra/species or genotype. Following the spectra quality assessment, Biotyper software was used to create a main spectra library (MSP) representing nine species and three genotypes of Trichinella. The evaluation of the spectra generated by MALDI-TOF MS revealed a classification which was comparable to the results obtained by molecular methods. Also, each Trichinella species utilized in this study was distinct and distinguishable with a high confidence level. Further, different conservation methods such as freezing and conservation in alcohol and the host species origin of the isolated larvae did not have a significant influence on the generated spectra. Therefore, the described MALDI-TOF MS can successfully be implemented for both genus and species level identification and represents a major step forward in the use of this technique in foodborne parasitology.

  20. Evaporative derivatization of phenols with 2-sulfobenzoic anhydride for detection by matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Yao, Yuanyuan; Wang, Poguang; Giese, Roger

    2014-03-30

    Phenols are an important class of analytes, for example as bioactive environmental contaminants. Towards a goal of improving their detection by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) or MALDI-tandem time-of-flight (TOF/TOF)-MS, we studied their derivatization with 2-sulfobenzoic anhydride (SBA). We chose SBA for this purpose since it is commercially available, inexpensive, and forms an anionic derivative. Under the selected conditions developed here for phenols, a reaction mixture of one or more of such compounds in acetonitrile containing SBA and 4-dimethylaminopyridine (DMAP) is evaporated to a solid, heated at 60 °C for 1 h, redissolved in 50% acetonitrile containing matrix, spotted onto a MALDI target, and subjected to negative ion MALDI-TOF/TOF-MS. While conventional (solution-phase) reaction of 4-phenylphenol (model analyte) with SBA and DMAP only gave a 47% yield of SBA-tagged 4-phenylphenol, evaporative derivatization as above gave a 96% yield, and 25 pmol (4.3 ng) of 4-phenylphenol could be detected in this way by MALDI-TOF/TOF-MS at signal-to-noise ratio (S/N) = 260, whereas even 1 nmol of the nonderivatized phenol was not detected in the absence of derivatization. A wide range of responses was observed when a mixture of 15 phenols was derivatized, with the higher responses coming from phenols with a pKa value above 9. Without derivatization, phenols with pKa values below 5 were the most readily detected. Evaporative derivatization with SBA (a convenient reagent) can improve the detection of phenols with relatively high pKa values (above 9) by negative ion MALDI-TOF-MS, and accomplish this in the absence of post-derivatization reaction cleanup. Copyright © 2014 John Wiley & Sons, Ltd.

  1. Identification of Tsetse (Glossina spp.) Using Matrix-Assisted Laser Desorption/Ionisation Time of Flight Mass Spectrometry

    Science.gov (United States)

    Hoppenheit, Antje; Murugaiyan, Jayaseelan; Bauer, Burkhard; Steuber, Stephan; Clausen, Peter-Henning; Roesler, Uwe

    2013-01-01

    Glossina (G.) spp. (Diptera: Glossinidae), known as tsetse flies, are vectors of African trypanosomes that cause sleeping sickness in humans and nagana in domestic livestock. Knowledge on tsetse distribution and accurate species identification help identify potential vector intervention sites. Morphological species identification of tsetse is challenging and sometimes not accurate. The matrix-assisted laser desorption/ionisation time of flight mass spectrometry (MALDI TOF MS) technique, already standardised for microbial identification, could become a standard method for tsetse fly diagnostics. Therefore, a unique spectra reference database was created for five lab-reared species of riverine-, savannah- and forest- type tsetse flies and incorporated with the commercial Biotyper 3.0 database. The standard formic acid/acetonitrile extraction of male and female whole insects and their body parts (head, thorax, abdomen, wings and legs) was used to obtain the flies' proteins. The computed composite correlation index and cluster analysis revealed the suitability of any tsetse body part for a rapid taxonomical identification. Phyloproteomic analysis revealed that the peak patterns of G. brevipalpis differed greatly from the other tsetse. This outcome was comparable to previous theories that they might be considered as a sister group to other tsetse spp. Freshly extracted samples were found to be matched at the species level. However, sex differentiation proved to be less reliable. Similarly processed samples of the common house fly Musca domestica (Diptera: Muscidae; strain: Lei) did not yield any match with the tsetse reference database. The inclusion of additional strains of morphologically defined wild caught flies of known origin and the availability of large-scale mass spectrometry data could facilitate rapid tsetse species identification in the future. PMID:23875040

  2. Direct matrix-assisted laser desorption ionisation time-of-flight mass spectrometry identification of mycobacteria from colonies.

    Science.gov (United States)

    Zingue, D; Flaudrops, C; Drancourt, M

    2016-12-01

    Matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF-MS) identification of mycobacteria requires a standard acetonitrile/formic acid pre-MALDI-TOF-MS. We prospectively compared this standard protocol with direct deposit with matrix for the identification of mycobacteria cultured on solid media. We first verified that Mycobacterium tuberculosis was killed after it was mixed with matrix. Then, 111 Mycobacterium isolates previously identified by partial rpoB gene sequencing were tested in parallel by the two protocols. An identification score >1.7 was obtained in 86/111 (77.5 %) isolates after protein extraction versus 97/111 (87.4 %) isolates after direct deposit (p = 0.039, Chi-squared test). In a third step, we determined that direct deposit achieved identification for as few as 2.104 M. tuberculosis organisms. In a fourth step, we evaluated direct deposit of one colony for 116 solid medium-cultured clinical isolates finally identified as representative of 12 species (63.8 % M. tuberculosis). For 114/116 (98.3 %) isolates with an identification score >1.2, the MALDI-TOF-MS identification was in complete agreement with the reference rpoB gene sequencing identification. One isolate with a MALDI-TOF-MS identification score of 1.22 for M. fortuitum was identified as M. avium by partial rpoB gene sequencing. One other isolate with a MALDI-TOF-MS identification score of 1.22 for M. tuberculosis was identified as M. tuberculosis by genotyping. All the original MALDI-TOF-MS spectra reported here have been deposited in a public database. Direct deposit of one colony on a MALDI-TOF-MS plate allows for an accurate identification of mycobacteria for an identification score >1.3.

  3. Identification of Tsetse (Glossina spp. using matrix-assisted laser desorption/ionisation time of flight mass spectrometry.

    Directory of Open Access Journals (Sweden)

    Antje Hoppenheit

    Full Text Available Glossina (G. spp. (Diptera: Glossinidae, known as tsetse flies, are vectors of African trypanosomes that cause sleeping sickness in humans and nagana in domestic livestock. Knowledge on tsetse distribution and accurate species identification help identify potential vector intervention sites. Morphological species identification of tsetse is challenging and sometimes not accurate. The matrix-assisted laser desorption/ionisation time of flight mass spectrometry (MALDI TOF MS technique, already standardised for microbial identification, could become a standard method for tsetse fly diagnostics. Therefore, a unique spectra reference database was created for five lab-reared species of riverine-, savannah- and forest- type tsetse flies and incorporated with the commercial Biotyper 3.0 database. The standard formic acid/acetonitrile extraction of male and female whole insects and their body parts (head, thorax, abdomen, wings and legs was used to obtain the flies' proteins. The computed composite correlation index and cluster analysis revealed the suitability of any tsetse body part for a rapid taxonomical identification. Phyloproteomic analysis revealed that the peak patterns of G. brevipalpis differed greatly from the other tsetse. This outcome was comparable to previous theories that they might be considered as a sister group to other tsetse spp. Freshly extracted samples were found to be matched at the species level. However, sex differentiation proved to be less reliable. Similarly processed samples of the common house fly Musca domestica (Diptera: Muscidae; strain: Lei did not yield any match with the tsetse reference database. The inclusion of additional strains of morphologically defined wild caught flies of known origin and the availability of large-scale mass spectrometry data could facilitate rapid tsetse species identification in the future.

  4. Discovery of a solvent effect preventing quantitative profiling by matrix-assisted laser desorption/ionization and its treatment.

    Science.gov (United States)

    Park, Kyung Man; Moon, Jeong Hee; Kim, Jae Hyung; Song, Un Tak; Lee, Seong Hoon; Kim, Myung Soo

    2016-02-15

    In analyte profiling by matrix-assisted laser desorption/ionization (MALDI), drawing a quantitative profile map is an outstanding problem. Recently, we developed a method to quantify an analyte by MALDI, which is needed to solve the problem. Another requirement for quantitative profiling is the quantitative sample-to-matrix analyte transfer, which is investigated in this work. MALDI-time-of-flight (TOF) spectra were acquired for samples produced by two methods. In one, a sample solution containing a matrix and an analyte was loaded with a pipet and dried. In the other, a sample was prepared by a consecutive process, i.e., loading-drying of an analyte solution followed by that of a matrix solution. Two different micro-spotters were used in the second method. Various mixtures of organic solvents with water were used to prepare matrix solutions. The organic solvent, matrix, and analyte used in the study did not affect the analyte transfer efficiency, whereas it improved as the water content in the solvent increased. It also improved as the liquid droplet emitted by a micro-spotter got larger. Use of a more polar solvent or a larger droplet increases the contact time between a solution droplet and the sample surface, which seems to be responsible for the improvement in the transfer efficiency. Sample-to-matrix analyte transfer occurred efficiently when polar solvents and/or large liquid droplets were used to produce solid samples for MALDI profiling with a micro-spotter. A long contact time between the sample surface and a matrix solution droplet is one of the requirements for quantitative profiling. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

  5. Identification of Acinetobacter Species Using Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry

    Science.gov (United States)

    Jeong, Seri; Hong, Jun Sung; Kim, Jung Ok; Kim, Keon-Han; Lee, Woonhyoung; Bae, Il Kwon; Lee, Kyungwon

    2016-01-01

    Background Acinetobacter baumannii has a greater clinical impact and exhibits higher antimicrobial resistance rates than the non-baumannii Acinetobacter species. Therefore, the correct identification of Acinetobacter species is clinically important. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) has recently become the method of choice for identifying bacterial species. The purpose of this study was to evaluate the ability of MALDI-TOF MS (Bruker Daltonics GmbH, Germany) in combination with an improved database to identify various Acinetobacter species. Methods A total of 729 Acinetobacter clinical isolates were investigated, including 447 A. baumannii, 146 A. nosocomialis, 78 A. pittii, 18 A. ursingii, 9 A. bereziniae, 9 A. soli, 4 A. johnsonii, 4 A. radioresistens, 3 A. gyllenbergii, 3 A. haemolyticus, 2 A. lwoffii, 2 A. junii, 2 A. venetianus, and 2 A. genomospecies 14TU. After 212 isolates were tested with the default Bruker database, the profiles of 63 additional Acinetobacter strains were added to the default database, and 517 isolates from 32 hospitals were assayed for validation. All strains in this study were confirmed by rpoB sequencing. Results The addition of the 63 Acinetobacter strains' profiles to the default Bruker database increased the overall concordance rate between MALDI-TOF MS and rpoB sequencing from 69.8% (148/212) to 100.0% (517/517). Moreover, after library modification, all previously mismatched 64 Acinetobacter strains were correctly identified. Conclusions MALDI-TOF MS enables the prompt and accurate identification of clinically significant Acinetobacter species when used with the improved database. PMID:27139605

  6. Matrix-assisted laser desorption-ionization time-of-flight mass spectrometry for the identification of Neisseria gonorrhoeae.

    Science.gov (United States)

    Buchanan, R; Ball, D; Dolphin, H; Dave, J

    2016-09-01

    Matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF MS) was compared with the API NH biochemical method for the identification of Neisseria gonorrhoeae in routine clinical samples. A retrospective review of laboratory records for 1090 isolates for which both biochemical and MALDI-TOF MS identifications were available was performed. Cases of discrepant results were examined in detail for evidence supportive of a particular organism identification. Of 1090 isolates, 1082 were identified as N. gonorrhoeae by API NH. MALDI-TOF MS successfully identified 984 (91%) of these after one analysis, rising to 1081 (99.9%) after two analyses, with a positive predictive value of 99.3%. For those isolates requiring a repeat analysis, failure to generate an identifiable proteomic signature was the reason in 76% of cases, with alternative initial identifications accounting for the remaining 24%. MALDI-TOF MS identified eight isolates as N. gonorrhoeae that were not identified as such by API NH-examination of these discrepant results suggested that the MALDI-TOF MS identification may be the more reliable. MALDI-TOF MS is at least as accurate and reliable a method of identifying N. gonorrhoeae as API NH. We propose that MALDI-TOF MS could potentially be used as a single method for N. gonorrhoeae identification in routine cases by laboratories with access to this technology. Copyright © 2016 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  7. Microorganisms direct identification from blood culture by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Ferreira, L; Sánchez-Juanes, F; Porras-Guerra, I; García-García, M I; García-Sánchez, J E; González-Buitrago, J M; Muñoz-Bellido, J L

    2011-04-01

    Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) allows a fast and reliable bacterial identification from culture plates. Direct analysis of clinical samples may increase its usefulness in samples in which a fast identification of microorganisms can guide empirical treatment, such as blood cultures (BC). Three hundred and thirty BC, reported as positive by the automated BC incubation device, were processed by conventional methods for BC processing, and by a fast method based on direct MALDI-TOF MS. Three hundred and eighteen of them yield growth on culture plates, and 12 were false positive. The MALDI-TOF MS-based method reported that no peaks were found, or the absence of a reliable identification profile, in all these false positive BC. No mixed cultures were found. Among these 318 BC, we isolated 61 Gram-negatives (GN), 239 Gram-positives (GP) and 18 fungi. Microorganism identifications in GN were coincident with conventional identification, at the species level, in 83.3% of BC and, at the genus level, in 96.6%. In GP, identifications were coincident with conventional identification in 31.8% of BC at the species level, and in 64.8% at the genus level. Fungaemia was not reliably detected by MALDI-TOF. In 18 BC positive for Candida species (eight C. albicans, nine C. parapsilosis and one C. tropicalis), no microorganisms were identified at the species level, and only one (5.6%) was detected at the genus level. The results of the present study show that this fast, MALDI-TOF MS-based method allows bacterial identification directly from presumptively positive BC in a short time (<30 min), with a high accuracy, especially when GN bacteria are involved. © 2010 The Authors. Clinical Microbiology and Infection © 2010 European Society of Clinical Microbiology and Infectious Diseases.

  8. Rapid identification of siderophores by combined thin-layer chromatography/matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Hayen, Heiko; Volmer, Dietrich A

    2005-01-01

    The investigation of a combined thin-layer chromatography/matrix-assisted laser desorption/ionization mass spectrometry (TLC/MALDI-MS) method for the analysis of siderophores from microbial samples is described. The investigated siderophores were enterobactin, ferrioxamine B, ferrichrome, ferrirhodin, rhodotorulic acid and coprogen. Solid-phase extraction was employed to recover the siderophores from the microbial samples. After visualization of the spots via spraying with ferric chloride or chrome azurol sulfonate assay solution, the MALDI matrix was applied to the gel surface. Several TLC/MALDI experimental parameters were optimized, such as type and concentration of MALDI matrix, as well as the type and composition of solvent to facilitate analyte transport from the inside of the TLC gel to the surface. The impact of these parameters on sensitivity, precision and ion formation of the various siderophores was studied. The detection limits for the investigated siderophores were in the range 1-4 pmol. These values were about 4-24 times higher than the detection limits obtained directly from stainless steel MALDI targets. The differences were most likely due to incomplete transport of the 'trapped' analyte molecules from the deeper layers of the TLC gel to the surface and into the matrix layer. In addition, chromatographic band broadening spread the analyte further in TLC as compared with the steel plates, resulting in less analyte per surface area. The identification of the siderophores was aided by concurrently applying a Ga(III) nitrate solution to the TLC plate during the visualization step. The resulting formation of Ga(III) complexes lead to distinctive (69)Ga/(71)Ga isotope patterns in the mass spectra. The versatility of the TLC/MALDI-MS assay was demonstrated by using it to analyze siderophores in a Pseudomonas aeruginosa sample. An iron-binding compound was identified in the sample, namely pyochelin (2-(2-o-hydroxyphenyl-2-thiazolin-4-yl)-3

  9. Differentiation of Streptococcus pneumoniae conjunctivitis outbreak isolates by matrix-assisted laser desorption ionization-time of flight mass spectrometry.

    Science.gov (United States)

    Williamson, Yulanda M; Moura, Hercules; Woolfitt, Adrian R; Pirkle, James L; Barr, John R; Carvalho, Maria Da Gloria; Ades, Edwin P; Carlone, George M; Sampson, Jacquelyn S

    2008-10-01

    Streptococcus pneumoniae (pneumococcus [Pnc]) is a causative agent of many infectious diseases, including pneumonia, septicemia, otitis media, and conjunctivitis. There have been documented conjunctivitis outbreaks in which nontypeable (NT), nonencapsulated Pnc has been identified as the etiological agent. The use of mass spectrometry to comparatively and differentially analyze protein and peptide profiles of whole-cell microorganisms remains somewhat uncharted. In this report, we discuss a comparative proteomic analysis between NT S. pneumoniae conjunctivitis outbreak strains (cPnc) and other known typeable or NT pneumococcal and streptococcal isolates (including Pnc TIGR4 and R6, Streptococcus oralis, Streptococcus mitis, Streptococcus pseudopneumoniae, and Streptococcus pyogenes) and nonstreptococcal isolates (including Escherichia coli, Enterococcus faecalis, and Staphylococcus aureus) as controls. cPnc cells and controls were grown to mid-log phase, harvested, and subsequently treated with a 10% trifluoroacetic acid-sinapinic acid matrix mixture. Protein and peptide fragments of the whole-cell bacterial isolate-matrix combinations ranging in size from 2 to 14 kDa were evaluated by matrix-assisted laser desorption ionization-time of flight mass spectrometry. Additionally Random Forest analytical tools and dendrogramic representations (Genesis) suggested similarities and clustered the isolates into distinct clonal groups, respectively. Also, a peak list of protein and peptide masses was obtained and compared to a known Pnc protein mass library, in which a peptide common and unique to cPnc isolates was tentatively identified. Information gained from this study will lead to the identification and validation of proteins that are commonly and exclusively expressed in cPnc strains which could potentially be used as a biomarker in the rapid diagnosis of pneumococcal conjunctivitis.

  10. Evaluation of Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry for Identification of Nocardia Species▿

    Science.gov (United States)

    Verroken, A.; Janssens, M.; Berhin, C.; Bogaerts, P.; Huang, T.-D.; Wauters, G.; Glupczynski, Y.

    2010-01-01

    The identification of Nocardia species, usually based on biochemical tests together with phenotypic in vitro susceptibility and resistance patterns, is a difficult and lengthy process owing to the slow growth and limited reactivity of these bacteria. In this study, a panel of 153 clinical and reference strains of Nocardia spp., altogether representing 19 different species, were characterized by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). As reference methods for species identification, full-length 16S rRNA gene sequencing and phenotypical biochemical and enzymatic tests were used. In a first step, a complementary homemade reference database was established by the analysis of 110 Nocardia isolates (pretreated with 30 min of boiling and extraction) in the MALDI BioTyper software according to the manufacturer's recommendations for microflex measurement (Bruker Daltonik GmbH, Leipzig, Germany), generating a dendrogram with species-specific cluster patterns. In a second step, the MALDI BioTyper database and the generated database were challenged with 43 blind-coded clinical isolates of Nocardia spp. Following addition of the homemade database in the BioTyper software, MALDI-TOF MS provided reliable identification to the species level for five species of which more than a single isolate was analyzed. Correct identification was achieved for 38 of the 43 isolates (88%), including 34 strains identified to the species level and 4 strains identified to the genus level according to the manufacturer's log score specifications. These data suggest that MALDI-TOF MS has potential for use as a rapid (Nocardia species without any substantial costs for consumables. PMID:20861335

  11. Rapid Identification of the Foodborne Pathogen Trichinella spp. by Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry.

    Directory of Open Access Journals (Sweden)

    Anne Mayer-Scholl

    Full Text Available Human trichinellosis occurs through consumption of raw or inadequately processed meat or meat products containing larvae of the parasitic nematodes of the genus Trichinella. Currently, nine species and three genotypes are recognized, of which T. spiralis, T. britovi and T. pseudospiralis have the highest public health relevance. To date, the differentiation of the larvae to the species and genotype level is based primarily on molecular methods, which can be relatively time consuming and labor intensive. Due to its rapidness and ease of use a matrix assisted laser desorption / ionization time of flight mass spectrometry (MALDI-TOF MS reference spectra database using Trichinella strains of all known species and genotypes was created. A formicacid/acetonitrile protein extraction was carried out after pooling 10 larvae of each Trichinella species and genotype. Each sample was spotted 9 times using α-cyano 4-hydoxy cinnamic acid matrix and a MicroFlex LT mass spectrometer was used to acquire 3 spectra (m/z 2000 to 20000 Da from each spot resulting in 27 spectra/species or genotype. Following the spectra quality assessment, Biotyper software was used to create a main spectra library (MSP representing nine species and three genotypes of Trichinella. The evaluation of the spectra generated by MALDI-TOF MS revealed a classification which was comparable to the results obtained by molecular methods. Also, each Trichinella species utilized in this study was distinct and distinguishable with a high confidence level. Further, different conservation methods such as freezing and conservation in alcohol and the host species origin of the isolated larvae did not have a significant influence on the generated spectra. Therefore, the described MALDI-TOF MS can successfully be implemented for both genus and species level identification and represents a major step forward in the use of this technique in foodborne parasitology.

  12. Matrix-assisted laser desorption ionization time-of-flight mass spectrometric analysis of glycosphingolipids including gangliosides.

    Science.gov (United States)

    Taketomi, T; Hara, A; Uemura, K; Sugiyama, E

    1998-01-01

    Long chain base compositions of gangliosides containing mainly stearic acid could be determined without any chemical modification by matrix-assisted laser desorption ionization time-of-flight mass spectrometry with delayed ion extraction (DE MALDI-TOF MS). The analytical results for the long chain base compositions of various samples of GM1 from the brain tissues of patients with different diseases at different ages confirmed that the proportion of d20:1 (icosasphingosine) and d20 (icosa-sphinganine) of the total sphingosine bases increased quickly until adolescent or adult age and then remained constant slightly exceeding 50%; this value was evidently higher than the proportion of d20:1 and d20 of GM1 in various adult mammalian brains. A long chain base composition of GM1 from the brain tissue of a patient with infantile type of GM1-gangliosidosis at 4y2m was abnormal and so was in two sibling patients with Spielmeyer-Vogt type of juvenile amaurotic idiocy at 19y and 21y in spite of that in the latter there was no accumulation of GM1 in the brain tissue. On the other hand, a patient with adult type of GM1 gangliosidosis at 66y showed a local accumulation of GM1 in the putamen and caudate nucleus, but its long chain base composition was found to be normal. It was of interest that the white matter of Eker rat with hereditary renal carcinoma contained a large amount of plasmalocerebroside as compared with the amount of cerebroside and sphingomyelin. The individual molecular species of plasmalocerebroside were identified by DE MALDI-TOF MS.

  13. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for rapid identification of fungal rhinosinusitis pathogens.

    Science.gov (United States)

    Huang, Yanfei; Wang, Jinglin; Zhang, Mingxin; Zhu, Min; Wang, Mei; Sun, Yufeng; Gu, Haitong; Cao, Jingjing; Li, Xue; Zhang, Shaoya; Lu, Xinxin

    2017-03-01

    Filamentous fungi are among the most important pathogens, causing fungal rhinosinusitis (FRS). Current laboratory diagnosis of FRS pathogens mainly relies on phenotypic identification by culture and microscopic examination, which is time consuming and expertise dependent. Although matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS has been employed to identify various fungi, its efficacy in the identification of FRS fungi is less clear. A total of 153 FRS isolates obtained from patients were analysed at the Clinical Laboratory at the Beijing Tongren Hospital affiliated to the Capital Medical University, between January 2014 and December 2015. They were identified by traditional phenotypic methods and Bruker MALDI-TOF MS (Bruker, Biotyper version 3.1), respectively. Discrepancies between the two methods were further validated by sequencing. Among the 153 isolates, 151 had correct species identification using MALDI-TOF MS (Bruker, Biot 3.1, score ≥2.0 or 2.3). MALDI-TOF MS enabled identification of some very closely related species that were indistinguishable by conventional phenotypic methods, including 1/10 Aspergillus versicolor, 3/20 Aspergillus flavus, 2/30 Aspergillus fumigatus and 1/20 Aspergillus terreus, which were misidentified by conventional phenotypic methods as Aspergillus nidulans, Aspergillus oryzae, Aspergillus japonicus and Aspergillus nidulans, respectively. In addition, 2/2 Rhizopus oryzae and 1/1 Rhizopus stolonifer that were identified only to the genus level by the phenotypic method were correctly identified by MALDI-TOF MS. MALDI-TOF MS is a rapid and accurate technique, and could replace the conventional phenotypic method for routine identification of FRS fungi in clinical microbiology laboratories.

  14. Phenotypic identification of Porphyromonas gingivalis validated with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Rams, Thomas E; Sautter, Jacqueline D; Getreu, Adam; van Winkelhoff, Arie J

    2016-05-01

    Porphyromonas gingivalis is a major bacterial pathogen in human periodontitis. This study used matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry to assess the accuracy of a rapid phenotypic identification scheme for detection of cultivable P. gingivalis in human subgingival plaque biofilms. A total of 314 fresh cultivable subgingival isolates from 38 adults with chronic periodontitis were presumptively identified on anaerobically-incubated enriched Brucella blood agar primary isolation plates as P. gingivalis based on dark-pigmented colony morphology, lack of a brick-red autofluorescence reaction under long-wave ultraviolet light, and a positive CAAM fluorescence test for trypsin-like enzyme activity. Each presumptive P. gingivalis isolate, and a panel of other human subgingival bacterial species, were subjected to MALDI-TOF mass spectrometry analysis using a benchtop mass spectrometer equipped with software containing mass spectra for P. gingivalis in its reference library of bacterial protein profiles. A MALDI-TOF mass spectrometry log score of ≥1.7 was required for species identification of the subgingival isolates. All 314 (100%) presumptive P. gingivalis subgingival isolates were confirmed as P. gingivalis with MALDI-TOF mass spectrometry analysis (Cohen's kappa coefficient = 1.0). MALDI-TOF mass spectrometry log scores between 1.7 and 1.9, and ≥2.0, were found for 92 (29.3%) and 222 (70.7%), respectively, of the presumptive P. gingivalis clinical isolates. No other tested bacterial species was identified as P. gingivalis by MALDI-TOF mass spectrometry. Rapid phenotypic identification of cultivable P. gingivalis in human subgingival biofilm specimens was found to be 100% accurate with MALDI-TOF mass spectrometry. These findings provide validation for the continued use of P. gingivalis research data based on this species identification methodology. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. Ceria nanocubic-ultrasonication assisted dispersive liquid-liquid microextraction coupled with matrix assisted laser desorption/ionization mass spectrometry for pathogenic bacteria analysis.

    Science.gov (United States)

    Abdelhamid, Hani Nasser; Bhaisare, Mukesh L; Wu, Hui-Fen

    2014-03-01

    A new ceria (CeO2) nanocubic modified surfactant is used as the basis of a novel nano-based microextraction technique for highly sensitive detection of pathogenic bacteria (Pseudomonas aeruginosa and Staphylococcus aureus). The technique uses ultrasound enhanced surfactant-assisted dispersive liquid-liquid microextraction (UESA-DLLME) with and without ceria (CeO2) followed by matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS). In order to achieve high separation efficiency, we investigated the influential parameters, including extraction time of ultrasonication, type and volume of the extraction solvent and surfactant. Among various surfactants, the cationic surfactants can selectively offer better extraction efficiency on bacteria analysis than that of the anionic surfactants due to the negative charges of bacteria cell membranes. Extractions of the bacteria lysate from aqueous samples via UESA-DLLME-MALDI-MS were successfully achieved by using cetyltrimethyl ammonium bromide (CTAB, 10.0 µL, 1.0×10(-3) M) as surfactants in chlorobenzene (10.0 µL) and chloroform (10.0 µL) as the optimal extracting solvent for P. aeruginosa and S. aureus, respectively. Ceria nanocubic was synthesized, and functionalized with CTAB (CeO2@CTAB) and then characterized using transmission electron microscopy (TEM) and optical spectroscopy (UV and FTIR). CeO2@CTAB demonstrates high extraction efficiency, improve peaks ionization, and enhance resolution. The prime reasons for these improvements are due to the large surface area of nanoparticles, and its absorption that coincides with the wavelength of MALDI laser (337 nm, N2 laser). CeO2@CTAB-based microextraction offers lowest detectable concentrations tenfold lower than that of without nanoceria. The present approach has been successfully applied to detect pathogenic bacteria at low concentrations of 10(4)-10(5) cfu/mL (without ceria) and at 10(3)-10(4) cfu/mL (with ceria) from bacteria suspensions. Finally, the

  16. Composite glycerol/graphite/aromatic acid matrices for thin-layer chromatography/matrix-assisted laser desorption/ionization mass spectrometry of heterocyclic compounds.

    Science.gov (United States)

    Esparza, Cesar; Borisov, R S; Varlamov, A V; Zaikin, V G

    2016-10-28

    New composite matrices have been suggested for the analysis of mixtures of different synthetic organic compounds (N-containing heterocycles and erectile dysfunction drugs) by thin layer chromatography/matrix-assisted laser desorption ionization time-of-flight mass spectrometry (TLC/MALDI-TOF). Different mixtures of classical MALDI matrices and graphite particles dispersed in glycerol were used for the registration of MALDI mass spectra directly from TLC plates after analytes separation. In most of cases, the mass spectra possessed [M+H](+) ions; however, for some analytes only [M+Na](+) and [M+K](+) ions were observed. These ions have been used to generate visualized TLC chromatograms. The described approach increases the desorption/ionization efficiencies of analytes separated by TLC, prevent spot blurring, simplifies and decrease time for sample preparation. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Coupling thin-layer chromatography with vibrational cooling matrix-assisted laser desorption/ionization Fourier transform mass spectrometry for the analysis of ganglioside mixtures.

    Science.gov (United States)

    Ivleva, Vera B; Elkin, Yuri N; Budnik, Bogdan A; Moyer, Susanne C; O'Connor, Peter B; Costello, Catherine E

    2004-11-01

    Thin-layer chromatography (TLC), which is widely used for separation of glycolipids, oligosaccharides, lipids, and compounds of environmental and pharmaceutical interest, can be readily coupled to matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometers, but this arrangement usually compromises mass spectral resolution due to the irregularity of the TLC surface. However, TLC can be coupled to an external ion source MALDI-Fourier transform (FT) MS instrument without compromising mass accuracy and resolution of the spectra. Furthermore, when the FTMS has a vibrationally cooled MALDI ion source, fragile glycolipids can be desorbed from TLC plates without fragmentation, even to the point that desorption of intact molecules from "hot"matrixes such as alpha-cyano-4-hydroxycinnamic acid is possible. In this work, whole brain gangliosides are separated using TLC; the TLC plates are attached directly to the MALDI target, where the gangliosides are desorbed, ionized, and detected in the FTMS with >70 000 resolving power.

  18. Identification of Microorganisms by FilmArray and Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry Prior to Positivity in the Blood Culture System

    OpenAIRE

    Almuhayawi, Mohammed; Altun, Osman; Strålin, Kristoffer; Özenci, Volkan

    2014-01-01

    In this study, we investigated the performance of the FilmArray and matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) in identifying microorganisms from blood culture (BC) bottles prior to positivity. First, we used simulated BacT/Alert FA Plus BC bottles with five each for Escherichia coli and Staphylococcus aureus isolates. The FilmArray identified all 10 isolates before BC positivity with 9/10 at 5 h and 1 at 7.5 h after incubation in the BC system...

  19. Species identification of clinical isolates of anaerobic bacteria: a comparison of two matrix-assisted laser desorption ionization-time of flight mass spectrometry systems

    DEFF Research Database (Denmark)

    Justesen, Ulrik Stenz; Holm, Anette; Knudsen, Elisa

    2011-01-01

    We compared two matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) systems (Shimadzu/SARAMIS and Bruker) on a collection of consecutive clinically important anaerobic bacteria (n = 290). The Bruker system had more correct identifications to the species level...... (67.2% versus 49.0%), but also more incorrect identifications (7.9% versus 1.4%). The system databases need to be optimized to increase identification levels. However, MALDI-TOF MS in its present version seems to be a fast and inexpensive method for identification of most clinically important...

  20. Detection and quantification of neurotensin in human brain tissue by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

    DEFF Research Database (Denmark)

    Gobom, J; Kraeuter, K O; Persson, R

    2000-01-01

    A method was developed for mass spectrometric detection of neurotensin (NT)-like immunoreactivity and quantification of NT in human brain tissue. The method is based on immunoprecipitation followed by analysis using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI...... of less than 5%. The method allowed detection of low-femtomole amounts of NT, staring from low-milligram amounts of lyophilized brain tissue. In addition to NT, several other peptides were detected in the purified samples, most of which, according to their molecular masses, corresponded to fragments of NT...

  1. Rapid identification of positive blood cultures by matrix-assisted laser desorption ionization-time of flight mass spectrometry using prewarmed agar plates.

    Science.gov (United States)

    Bhatti, M M; Boonlayangoor, S; Beavis, K G; Tesic, V

    2014-12-01

    This study describes an inexpensive and straightforward method for identifying bacteria by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) directly from positive blood cultures using prewarmed agar plates. Different inoculation methods and incubation times were evaluated to determine the optimal conditions. The two methods using pelleted material from positive culture bottles performed best. In particular, the pellet streak method correctly identified 94% of the Gram negatives following 4 h of incubation and 98% of the Gram positives following 6 h of incubation. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  2. A novel type of matrix for surface-assisted laser desorption-ionization mass spectrometric detection of biomolecules using metal-organic frameworks.

    Science.gov (United States)

    Fu, Chien-Ping; Lirio, Stephen; Liu, Wan-Ling; Lin, Chia-Her; Huang, Hsi-Ya

    2015-08-12

    A 3D metal-organic framework (MOF) nanomaterial as matrix for surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS) and tandem mass spectrometry (MS/MS) was developed for the analysis of complex biomolecules. Unlike other nanoparticle matrices, this MOF nanomaterial does not need chemical modification prior to use. An exceptional signal reproducibility as well as very low background interferences in analyzing mono-/di-saccharides, peptides and complex starch digests demonstrate its high potential for biomolecule assays, especially for small molecules. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. Semiconductor Nanomaterials-Based Fluorescence Spectroscopic and Matrix-Assisted Laser Desorption/Ionization (MALDI Mass Spectrometric Approaches to Proteome Analysis

    Directory of Open Access Journals (Sweden)

    Suresh Kumar Kailasa

    2013-12-01

    Full Text Available Semiconductor quantum dots (QDs or nanoparticles (NPs exhibit very unusual physico-chemcial and optical properties. This review article introduces the applications of semiconductor nanomaterials (NMs in fluorescence spectroscopy and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS for biomolecule analysis. Due to their unique physico-chemical and optical properties, semiconductors NMs have created many new platforms for investigating biomolecular structures and information in modern biology. These semiconductor NMs served as effective fluorescent probes for sensing proteins and cells and acted as affinity or concentrating probes for enriching peptides, proteins and bacteria proteins prior to MALDI-MS analysis.

  4. Structure Determination of β-Glucans from Ganoderma lucidum with Matrix-assisted Laser Desorption/ionization (MALDI Mass Spectrometry

    Directory of Open Access Journals (Sweden)

    Wen-Bin Yang

    2008-08-01

    Full Text Available A novel method that uses matrix-assisted laser desorption/ionization (MALDI mass spectrometry to analyze molecular weight and sequencing of glucan in Ganoderma lucidum is presented. Thus, β-glucan, which was isolated from fruiting bodies of G. lucidum, was measured in a direct and fast way using MALDI mass spectrometry. In addition, tandem mass spectrometry of permethylated glucans of G. lucidum, dextran, curdlan and maltohexaose were also pursued and different fragment patterns were obtained. The G. lucidum glucan structure was determined and this method for linkage analysis of permethylated glucan has been proven feasible.

  5. Rapid identification of pathogens directly from blood culture bottles by Bruker matrix-assisted laser desorption laser ionization-time of flight mass spectrometry versus routine methods.

    Science.gov (United States)

    Jamal, Wafaa; Saleem, Rola; Rotimi, Vincent O

    2013-08-01

    The use of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) for identification of microorganisms directly from blood culture is an exciting dimension to the microbiologists. We evaluated the performance of Bruker SepsiTyper kit™ (STK) for direct identification of bacteria from positive blood culture. This was done in parallel with conventional methods. Nonrepetitive positive blood cultures from 160 consecutive patients were prospectively evaluated by both methods. Of 160 positive blood cultures, the STK identified 114 (75.6%) isolates and routine conventional method 150 (93%). Thirty-six isolates were misidentified or not identified by the kit. Of these, 5 had score of >2.000 and 31 had an unreliable low score of time using the STK was 35 min, including extraction steps and 30:12 to 36:12 h with routine method. The STK holds promise for timely management of bacteremic patients. Copyright © 2013 Elsevier Inc. All rights reserved.

  6. Direct Imaging Mass Spectrometry of Plant Leaves Using Surface-assisted Laser Desorption/Ionization with Sputter-deposited Platinum Film.

    Science.gov (United States)

    Ozawa, Tomoyuki; Osaka, Issey; Hamada, Satoshi; Murakami, Tatsuya; Miyazato, Akio; Kawasaki, Hideya; Arakawa, Ryuichi

    2016-01-01

    Plant leaves administered with systemic insecticides as agricultural chemicals were analyzed using imaging mass spectrometry (IMS). Matrix-assisted laser desorption/ionization (MALDI) is inadequate for the detection of insecticides on leaves because of the charge-up effect that occurs on the non-conductive surface of the leaves. In this study, surface-assisted laser desorption/ionization with a sputter-deposited platinum film (Pt-SALDI) was used for direct analysis of chemicals in plant leaves. Sputter-deposited platinum (Pt) films were prepared on leaves administered with the insecticides. A sputter-deposited Pt film with porous structure was used as the matrix for Pt-SALDI. Acephate and acetamiprid contained in the insecticides on the leaves could be detected using Pt-SALDI-MS, but these chemical components could not be adequately detected using MALDI-MS because of the charge-up effect. Enhancement of ion yields for the insecticides was achieved using Pt-SALDI, accompanied by prevention of the charge-up effect by the conductive Pt film. The movement of systemic insecticides in plants could be observed clearly using Pt-SALDI-IMS. The distribution and movement of components of systemic insecticides on leaves could be analyzed directly using Pt-SALDI-IMS. Additionally, changes in the properties of the chemicals with time, as an indicator of the permeability of the insecticides, could be evaluated.

  7. A comparative study of matrix- and nano-assisted laser desorption/ionisation time-of-flight mass spectrometry of isolated and synthetic lignin.

    Science.gov (United States)

    Yoshioka, Koichi; Ando, Daisuke; Watanabe, Takashi

    2012-01-01

    Lignin is the second most abundant biopolymer next to cellulose. However, because of the complexity of the heterogeneous macromolecules, it is difficult to elucidate the polymeric structures of lignin by conventional analytical methods. To obtain the detailed structures of lignin, we comparatively applied nano-assisted laser desorption/ionisation time-of-flight mass spectrometry (NALDI-TOF MS) and matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS). Synthetic lignin from coniferyl alcohol and an isolated lignin from Pinus densiflora were subjected to NALDI- and MALDI-TOF MS. We first obtained NALDI-TOF MS of synthetic and isolated lignin. Mass increments of 178 and 196 Da were observed in NALDI- and MALDI-TOF mass spectra of the synthetic and isolated lignin. The mass intervals indicated that radical coupling forming β-O-4 bonds is the major pathway. Peaks in the low molecular mass region between m/z 500 and 800 were observed more extensively using NALDI-TOF MS than MALDI-TOF MS, which enabled detailed analysis of the interunit linkages in lignin. Owing to the ionisation profile differentiation from MALDI-TOF MS, NALDI-TOF MS is useful for the structural analysis of lignin. Copyright © 2011 John Wiley & Sons, Ltd.

  8. The value of total protein in guiding management of infectious parapneumonic effusion by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Chiu, Chih-Yung; Hsieh, Sen-Yung; Wong, Kin-Sun; Lai, Shen-Hao; Chen, Jen-Kun; Huang, Jing-Long

    2015-10-01

    Infectious parapneumonic effusion (PE) contains proteins originating from circulation as well as proteins locally released by inflammatory pulmonary cells. The purpose of this study was to investigate the value of total protein analysis in guiding management of infectious PE by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Fifty-seven children with pneumonia followed by PE were consecutively enrolled into our study. Protein profiles generated by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry after fractionating samples with functionalized magnetic beads (C8) were used for differentiating complicated PE (CPE) from non-CPE. A training set was used to generate classification models and the clinical efficacy of these models in detecting CPE and the need for intervention was then evaluated in an independent set. The MS spectra derived from PE were analyzed, and classification models were constructed in the training set. A total of 123 mass/charge (m/z) values were identified and 23 m/z values which were significant with p management of infectious PE. Copyright © 2013. Published by Elsevier B.V.

  9. Short communication: Identification of subclinical cow mastitis pathogens in milk by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Barreiro, J R; Ferreira, C R; Sanvido, G B; Kostrzewa, M; Maier, T; Wegemann, B; Böttcher, V; Eberlin, M N; dos Santos, M V

    2010-12-01

    Subclinical mastitis is a common and easily disseminated disease in dairy herds. Its routine diagnosis via bacterial culture and biochemical identification is a difficult and time-consuming process. In this work, we show that matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) allows bacterial identification with high confidence and speed (1 d for bacterial growth and analysis). With the use of MALDI-TOF MS, 33 bacterial culture isolates from milk of different dairy cows from several farms were analyzed, and the results were compared with those obtained by classical biochemical methods. This proof-of-concept case demonstrates the reliability of MALDI-TOF MS bacterial identification, and its increased selectivity as illustrated by the additional identification of coagulase-negative Staphylococcus species and mixed bacterial cultures. Matrix-assisted laser desorption-ionization mass spectrometry considerably accelerates the diagnosis of mastitis pathogens, especially in cases of subclinical mastitis. More immediate and efficient animal management strategies for mastitis and milk quality control in the dairy industry can therefore be applied. Copyright © 2010 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  10. Nanomaterial based affinity matrix-assisted laser desorption/ionization mass spectrometry for biomolecules and pathogenic bacteria.

    Science.gov (United States)

    Chiu, Tai-Chia; Huang, Li-Shing; Lin, Po-Chiao; Chen, Yu-Chie; Chen, Yu-Ju; Lin, Chun-Cheng; Chang, Huan-Tsung

    2007-01-01

    This paper describes mass spectrometry (MS) using nanoparticles (NPs) for the analysis of biomolecules such as aminothiol compounds, drugs, peptides, proteins, and bacteria. Papers and patents dealing with preparation and use of several NPs in MS have been briefly reviewed, including carbon nanotubes, gold NPs, and magnetic NPs. The NPs or bioconjugated NPs were used for selective concentration and/or assisted matrices for desorption and ionization of analytes of interest. When compared to conventional organic matrixes, the NPs provide low MS background in low-mass region and low shot-to-shot variation. MS techniques using NPs and bioconjugated NPs for the analysis of disease-associated biomarkers and bacteria in real samples such as blood and urine are highlighted, showing the advantages of high sensitivity, reproducibility, and simplicity.

  11. Soft-landing ion mobility of silver clusters for small-molecule matrix-assisted laser desorption ionization mass spectrometry and imaging of latent fingerprints.

    Science.gov (United States)

    Walton, Barbara L; Verbeck, Guido F

    2014-08-19

    Matrix-assisted laser desorption ionization (MALDI) imaging is gaining popularity, but matrix effects such as mass spectral interference and damage to the sample limit its applications. Replacing traditional matrices with silver particles capable of equivalent or increased photon energy absorption from the incoming laser has proven to be beneficial for low mass analysis. Not only can silver clusters be advantageous for low mass compound detection, but they can be used for imaging as well. Conventional matrix application methods can obstruct samples, such as fingerprints, rendering them useless after mass analysis. The ability to image latent fingerprints without causing damage to the ridge pattern is important as it allows for further characterization of the print. The application of silver clusters by soft-landing ion mobility allows for enhanced MALDI and preservation of fingerprint integrity.

  12. Analysis of noncovalent chitinase-chito-oligosaccharide complexes by infrared-matrix assisted laser desorption ionization and nanoelectrospray ionization mass spectrometry.

    Science.gov (United States)

    Dybvik, Anette I; Norberg, Anne Line; Schute, Veronika; Soltwisch, Jens; Peter-Katalinić, Jasna; Vårum, Kjell M; Eijsink, Vincent G H; Dreisewerd, Klaus; Mormann, Michael; Sørlie, Morten

    2011-06-01

    Transferring noncovalently bound complexes from the condensed phase into the gas phase represents a challenging task due to weak intermolecular bonds that have to be maintained during the phase transition. Currently, electrospray ionization (ESI) is the standard mass spectrometric (MS) technique to analyze noncovalent complexes. Although infrared matrix-assisted laser desorption ionization (IR-MALDI)-MS also provides particular soft desorption/ionization conditions, this method has so far hardly been applied for the analysis of noncovalent complexes. In this study, we employed IR-MALDI orthogonal time-of-flight (o-TOF)-MS in combination with the liquid matrix glycerol to characterize the specific complex formation of chito-oligosaccharide (CHOS) ligands with two variants of Chitinase A (ChiA) from Serratia marcescens, the inactive E315Q mutant and the active W167A mutant, respectively. The IR-MALDI-o-TOF-MS results were compared to those obtained using nano-ESI-quadrupole (q)-TOF-MS and ultraviolet (UV)-MALDI-o-TOF-MS. Using IR-MALDI-o-TOF-MS, specific noncovalent complexes between ChiA and CHOS were detected with distributions between enzymes with bound oligosaccharides vs free enzymes that were essentially identical to those obtained by nano-ESI-q-TOF-MS. Chitinase-CHOS complexes were not detected when UV-MALDI was employed for desorption/ionization. The results show that IR-MALDI-MS can be a valuable tool for fast and simple screening of noncovalent enzyme-ligand interactions.

  13. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) coupled to XAD fractionation: Method to algal organic matter characterization.

    Science.gov (United States)

    Nicolau, Rudy; Leloup, Maud; Lachassagne, Delphine; Pinault, Emilie; Feuillade-Cathalifaud, Geneviève

    2015-05-01

    This work is focused on the development of an analytical procedure for the improvement of the Organic Matter structure characterization, particularly the algal matter. Two fractions of algal organic matter from laboratory cultures of algae (Euglena gracilis) and cyanobacteria (Microcystis aeruginosa) were extracted with XAD resins. The fractions were studied using laser desorption ionization (LDI) and Matrix-Assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF). A comparison with the natural organic matter characteristics from commercial humic acids and fulvic acids extracted from Suwannee River was performed. Results show that algal and natural organic matters have unique quasi-polymeric structures. Significant repeating patterns were identified. Different fractions extracted from organic matter with common origin had common structures. Thus, 44, 114 and 169Da peaks separation for fractions from E. gracilis organic matter and 28, 58 and 100Da for M. aeruginosa ones were clearly observed. Using the developed protocol, a structural scheme and organic matter composition were obtained. The range 600-2000Da contained more architectural composition differences than the range 100-600Da, suggesting that organic matter is composed of an assembly of common small molecules. Associated to specific monomers, particular patterns were common to all samples but assembly and resulting structure were unique for each organic matter. Thus, XAD fractionation coupled to mass spectroscopy allowed determining a specific fingerprint for each organic matter. Copyright © 2015 Elsevier B.V. All rights reserved.

  14. Surface-assisted laser desorption/ionization mass spectrometric detection of biomolecules by using functional single-walled carbon nanohorns as the matrix.

    Science.gov (United States)

    Ma, Rongna; Lu, Minghua; Ding, Lin; Ju, Huangxian; Cai, Zongwei

    2013-01-02

    A surface-assisted laser desorption/ionization time-of-flight mass spectrometric (SALDI-TOF MS) method was developed for the analysis of small biomolecules by using functional single-walled carbon nanohorns (SWNHs) as matrix. The functional SWNHs could transfer energy to the analyte under laser irradiation for accelerating its desorption and ionization, which led to low matrix effect, avoided fragmentation of the analyte, and provided high salt tolerance. Biomolecules including amino acids, peptides, and fatty acids could successfully be analyzed with about 3- and 5-fold higher signals than those obtained using conventional matrix. By integrating the advantages of SWNHs and the recognition ability of aptamers, a selective approach was proposed for simultaneous capture, enrichment, ionization, and MS detection of adenosine triphosphate (ATP). This method showed a greatly improved detection limit (1.0 μM) for the analysis of ATP in complex biological samples. This newly designed protocol not only opened a new application of SWNHs, but also offered a new technique for selective MS analysis of biomolecules based on aptamer recognition systems. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Species Identification of Clinical Prevotella Isolates by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

    Science.gov (United States)

    Soetens, Oriane; De Bel, Annelies; Echahidi, Fedoua; Vancutsem, Ellen; Vandoorslaer, Kristof; Piérard, Denis

    2012-01-01

    The performance of matrix-assisted laser desorption–ionization time of flight mass spectrometry (MALDI-TOF MS) for species identification of Prevotella was evaluated and compared with 16S rRNA gene sequencing. Using a Bruker database, 62.7% of the 102 clinical isolates were identified to the species level and 73.5% to the genus level. Extension of the commercial database improved these figures to, respectively, 83.3% and 89.2%. MALDI-TOF MS identification of Prevotella is reliable but needs a more extensive database. PMID:22301022

  16. Detection of carbapenemases using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) meropenem hydrolysis assay.

    Science.gov (United States)

    Hrabák, Jaroslav

    2015-01-01

    Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been recently introduced to many diagnostic microbiological laboratories. Besides the identification of bacteria and fungi, that technique provides a potentially useful tool for the detection of antimicrobial resistance, especially of that conferred by β-lactamases. Here, we describe an assay allowing a detection of meropenem hydrolysis in clinical isolates of Enterobacteriaceae, Pseudomonas spp., and Acinetobacter baumannii using MALDI-TOF MS. This method is able to confirm carbapenemases within 3 h. The results are important for proper and fast intervention to limit the spread of carbapenemase-producing bacteria and provide information for appropriate initial therapy of the infections caused by these microbes.

  17. Direct Analysis of Triterpenes from High-Salt Fermented Cucumbers Using Infrared Matrix-Assisted Laser Desorption Electrospray Ionization (IR-MALDESI)

    Science.gov (United States)

    Ekelöf, Måns; McMurtrie, Erin K.; Nazari, Milad; Johanningsmeier, Suzanne D.; Muddiman, David C.

    2017-02-01

    High-salt samples present a challenge to mass spectrometry (MS) analysis, particularly when electrospray ionization (ESI) is used, requiring extensive sample preparation steps such as desalting, extraction, and purification. In this study, infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) coupled to a Q Exactive Plus mass spectrometer was used to directly analyze 50-μm thick slices of cucumber fermented and stored in 1 M sodium chloride brine. From the several hundred unique substances observed, three triterpenoid lipids produced by cucumbers, β-sitosterol, stigmasterol, and lupeol, were putatively identified based on exact mass and selected for structural analysis. The spatial distribution of the lipids were imaged, and the putative assignments were confirmed by tandem mass spectrometry performed directly on the same cucumber, demonstrating the capacity of the technique to deliver confident identifications from highly complex samples in molar concentrations of salt without the need for sample preparation.

  18. Detection and Quantification of 4-Methylimidazole in Cola by Matrix-assisted Laser Desorption Ionization Mass Spectrometry with Fe2O3Nanoparticles on Zeolite.

    Science.gov (United States)

    Fujii, Yosuke; Ding, Yuqi; Umezawa, Taichi; Akimoto, Takafumi; Xu, Jiawei; Uchida, Takashi; Fujino, Tatsuya

    2018-01-01

    Food additives generally used in carbonated drinks, such as 4-methylimidazole (4MI), caffeine (Caf?), citric acid (CA), and aspartame (Apm), were measured by matrix-assisted laser desorption ionization mass spectrometry (MALDI MS) using nanometer-sized particles of iron oxide (Fe 2 O 3 NPs). The quantification of 4MI in Coca Cola (C-cola) was carried out. In order to improve the reproducibility of the peak intensities, Fe 2 O 3 NPs loaded on ZSM5 zeolite were used as the matrix for quantification. By using 2-ethylimidazole (2EI) as the internal standard, the amount of 4MI in C-cola was determined to range from 88 to 65 μg/355 mL. The results agree with the published value (approx. 72 μg/355 mL). It was found that MALDI using Fe 2 O 3 was applicable to the quantification of 4MI in C-cola.

  19. Copper(I) chloride: a simple salt for enhancement of polystyrene cationization in matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Kéki, S; Deák, G; Zsuga, M

    2001-01-01

    The possibility of using copper(I) chloride as a doping salt to enhance the cationization of polystyrene in matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) was investigated. It was shown that copper(I) chloride possesses sufficient solubility in tetrahydrofuran. The parameters of the MALDI mass spectra of different polystyrene samples, such as the number-average (M(n)) and mass-average (M(w)) molecular mass values, obtained by copper(I) cationization were compared with those obtained by means of silver(I) cationization, and good agreement was found. It was also shown that application of copper(I) chloride as a doping salt, and dithranol as a matrix, ensured good MALDI mass spectra of the sample spots even after storage for 1 month. Copyright 2001 John Wiley & Sons, Ltd.

  20. Analysis of acidic carbohydrates as their quaternary ammonium or phosphonium salts by matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Ueki, Masaaki; Yamaguchi, Miyuki

    2005-07-25

    New two-component systems using quaternary ammonium or phosphonium salts as a co-matrix have been developed for the analysis of acidic carbohydrates by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). In the analysis of the sodium salt of heparin disaccharide I-S, the combination of 2-amino-5-nitropyridine with tetraphenylphosphonium bromide gave the best result. In the analysis of gangliosides containing the sialic acid moiety, the combination of 2,4,6-trihydroxyacetophenone with dimethyldipalmitylammonium bromide was determined to be the system of choice. Under optimum conditions all acidic carbohydrates gave molecular ions in the form of [M(Q(n))-Q]-, where M(Q(n)) is the molecular mass of a molecule containing n molecules of quaternary ions as salt.

  1. Analysis of low molecular weight acids by negative mode matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Shroff, Rohit; Muck, Alexander; Svatos, Ales

    2007-01-01

    Free 9-aminoacridine base is demonstrated to be a suitable matrix for negative mode matrix-assisted laser desorption/ionization time-of-flight mass spectrometric (MALDI-TOFMS) analysis of a wide range of low molecular weight organic acids including aliphatic (from acetic to palmitic acid), aromatic acids, phytohormones (e.g. jasmonic and salicylic acids), and amino acids. Low limits of quantitation in the femtomolar range (jasmonic - 250 fmol; caffeic - 160 fmol and salicylic - 12.5 fmol) and linear detector response over two concentration orders in the pico- and femtomolar range are extremely encouraging for the direct study of such acids in complex biological matrices. Copyright (c) 2007 John Wiley & Sons, Ltd.

  2. [Separation and identification of bovine lactoferricin by high performance liquid chromatography-matrix-assisted laser desorption/ionization time of flight/ time of flight mass spectrometry].

    Science.gov (United States)

    An, Meichen; Liu, Ning

    2010-02-01

    A high performance liquid chromatography-matrix-assisted laser desorption/ionization time of flight/time of flight mass spectrometry (HPLC-MALDI-TOF/TOF MS) method was developed for the separation and identification of bovine lactoferricin (LfcinB). Bovine lactoferrin was hydrolyzed by pepsin and then separated by ion exchange chromatography and reversed-phase liquid chromatography (RP-LC). The antibacterial activities of the fractions from RP-LC separation were determined and the protein concentration of the fraction with the highest activity was measured, whose sequence was indentified by MALDI-TOF/TOF MS. The relative molecular mass of LfcinB was 3 124.89 and the protein concentration was 18.20 microg/mL. The method of producing LfcinB proposed in this study has fast speed, high accuracy and high resolution.

  3. Proteome analysis of Arabidopsis thaliana by two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionisation-time of flight mass spectrometry.

    Science.gov (United States)

    Giavalisco, Patrick; Nordhoff, Eckhard; Kreitler, Thomas; Klöppel, Klaus-Dieter; Lehrach, Hans; Klose, Joachim; Gobom, Johan

    2005-05-01

    In the present study we show results of a large-scale proteome analysis of the recently sequenced plant Arabidopsis thaliana. On the basis of a previously published sequential protein extraction protocol, we prepared protein extracts from eight different A. thaliana tissues (primary leaf, leaf, stem, silique, seedling, seed, root, and inflorescence) and analysed these by two-dimensional gel electrophoresis. A total of 6000 protein spots, from three of these tissues, namely primary leaf, silique and seedling, were excised and the contained proteins were analysed by matrix assisted laser desorption/ionisation time of flight mass spectrometry peptide mass fingerprinting. This resulted in the identification of the proteins contained in 2943 spots, which were found to be products of 663 different genes. In this report we present and discuss the methodological and biological results of our plant proteome analysis.

  4. The effect of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry instrumentation parameters on the matrix-assisted laser desorption/ionization simulated size exclusion chromatography number-mass, average-weight and polydispersity values of dextran against corresponding values obtained by size exclusion chromatography.

    Science.gov (United States)

    Bashir, S; Giannakopulos, A E; Liu, J

    2017-12-01

    The matrix-assisted laser desorption/ionization simulated size exclusion chromatography (SECPC) average-number mass, weight average and polydispersity of dextran 1000 were determined by matrix-assisted laser desorption/ionization (MALDI) time-of-flight (TOF) mass spectrometry. The instrument parameters were varied and the SECPC value determined via the Bruker XMASS software was compared to the value obtained from aqueous-phase size exclusion chromatography. The aqueous-phase size exclusion chromatography values for average-number mass, weight average and polydispersity were 1223 Da, 1500 Da and 1.23 (1010 Da, 1270 Da and 1.26 from manufacturer), whereas the SECPC value varied on the instrumental parameters. The factors that had the greatest effect on the average-number mass, weight average and polydispersity were: (most effect on SECPC value) laser attenuation > matrix-analyte molar concentration > matrix-analyte molar ratios > delay extraction time > solvent-system composition > detector delay (least effect on SECPC value). The oligosaccharide signal distribution as a function of laser attenuation indicate that two distinct regions exist in dextran 1000, where one corresponds to the higher mass oligosaccharides (hexasaccharide or greater), while another region corresponds to lower oligosaccharides (tetra-saccharide). This distribution depends upon the crystallization of the biopolymer and the efficiency of desorption/ionization, which yields the SECPC value. There was broad agreement between the SECPC values and size exclusion chromatography values for dextran, although the polydispersity indicated by SECPC was less than size exclusion chromatography (1.10 vs. 1.26). It can be shown that for narrow polydisperse biopolymers the instrumental conditions are less critical in the determination of average-number mass, weight average and polydispersity, although the SECPC Mn, and weight average values are often higher than the corresponding values

  5. Validation of a modified algorithm for the identification of yeast isolates using matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS)

    NARCIS (Netherlands)

    van Herendael, B.H.; Bruynseels, P.; Bensaid, M.; Boekhout, T.; de Baere, T.; Surmont, I.; Mertens, A.H.

    2011-01-01

    Optimising antifungal treatment requires the fast and species-specific identification of yeast isolates. We evaluated a modified protocol for the rapid identification of clinical yeast isolates using matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) technology. First, we

  6. Structure determination of two conotoxins from Conus textile by a combination of matrix-assisted laser desorption/ionization time-of-flight and electrospray ionization mass spectrometry and biochemical methods

    DEFF Research Database (Denmark)

    Kalume, D E; Stenflo, J; Czerwiec, E

    2000-01-01

    Two highly modified conotoxins from the mollusc Conus textile, epsilon-TxIX and Gla(1)-TxVI, were characterized by matrix-assisted laser desorption/ionization and electrospray mass spectrometry and also by electrospray ionization tandem and triple mass spectrometry in combination with enzymatic c...

  7. Classification of wheat varieties: Use of two-dimensional gel electrophoresis for varieties that can not be classified by matrix assisted laser desorption/ionization-time of flight-mass spectrometry and an artificial neural network

    DEFF Research Database (Denmark)

    Jacobsen, Susanne; Nesic, Ljiljana; Petersen, Marianne Kjerstine

    2001-01-01

    Analyzing a gliadin extract by matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI- TOF-MS) combined with an artificial neural network (ANN) is a suitable method for identification of wheat varieties. However, the ANN can not distinguish between all different wheat...

  8. The importance of matrix-assisted laser desorption ionization–time of flight mass spectrometry for correct identification of Clostridium difficile isolated from chromID C. difficile chromogenic agar

    Directory of Open Access Journals (Sweden)

    Jonathan H.K. Chen

    2017-10-01

    Full Text Available The clinical workflow of using chromogenic agar and matrix-assisted laser desorption ionization time-of-fight mass spectrometry (MALDI-TOF MS for Clostridium difficile identification was evaluated. The addition of MALDI-TOF MS identification after the chromID C. difficile chromogenic agar culture could significantly improve the diagnostic accuracy of C. difficile.

  9. Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry Analysis of Gram-Positive, Catalase-Negative Cocci Not Belonging to the Streptococcus or Enterococcus Genus and Benefits of Database Extension

    DEFF Research Database (Denmark)

    Christensen, Jens Jørgen; Dargis, Rimtas; Hammer, Monja

    2012-01-01

    Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry with a Bruker Daltonics microflex LT system was applied to 90 well-characterized catalase-negative, Gram-positive cocci not belonging to the streptococci or enterococci. Biotyper version 2.0.43.1 software...

  10. An Approach to Glycobiology from Glycolipidomics: Ganglioside Molecular Scanning in the Brains of Patients with Alzheimer's Disease by TLC-Blot/Matrix Assisted Laser Desorption/Ionization-Time of Flight MS

    National Research Council Canada - National Science Library

    Taki, Takao

    2012-01-01

    ...)-Blot/matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MS) system. This new approach consists of a combination of a method for transferring lipids separated on a TLC-plate to a poly-vinylidene difluoride...

  11. Quantum theory of laser-stimulated desorption

    Science.gov (United States)

    Slutsky, M. S.; George, T. F.

    1978-01-01

    A quantum theory of laser-stimulated desorption (LSDE) is presented and critically analyzed. It is shown how LSDE depends on laser-pulse characteristics and surface-lattice dynamics. Predictions of the theory for a Debye model of the lattice dynamics are compared to recent experimental results.

  12. Analysis of complex phthalic acid based polyesters by the combination of size exclusion chromatography and matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Pretorius, Nadine O; Rode, Karsten; Simpson, Jaylin M; Pasch, Harald

    2014-01-15

    Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used in conjunction with size exclusion chromatography (SEC) to investigate a model polyester system based on phthalic anhydride-1,2-propylene glycol. The polyesters were synthesized with a 30% molar excess of glycol, with kinetic samples being removed during different intervals of the polyesterification reaction. SEC was used to track the course of the reaction by determining the molecular weight and molecular weight distributions before subsequent off-line coupling with MALDI-TOF MS as a selective detection method to determine the chemical composition, identify the functionality type distributions as well as assist in assigning structural conformations. Mass spectrometry analysis proved to be a highly effective tool to facilitate the identification of the narrowly dispersed fractions obtained from the chromatographic separations as well as serve as a core method to investigate the heterogeneous nature of the bulk kinetic samples. Through the hyphenation of these sophisticated polymer characterization techniques, information on the molecular heterogeneity of the model polyesters, showing a complex variety of possible distributions, was obtained. Copyright © 2013 Elsevier B.V. All rights reserved.

  13. Decision peptide-driven: a free software tool for accurate protein quantification using gel electrophoresis and matrix assisted laser desorption ionization time of flight mass spectrometry.

    Science.gov (United States)

    Santos, Hugo M; Reboiro-Jato, Miguel; Glez-Peña, Daniel; Nunes-Miranda, J D; Fdez-Riverola, Florentino; Carvallo, R; Capelo, J L

    2010-09-15

    The decision peptide-driven tool implements a software application for assisting the user in a protocol for accurate protein quantification based on the following steps: (1) protein separation through gel electrophoresis; (2) in-gel protein digestion; (3) direct and inverse (18)O-labeling and (4) matrix assisted laser desorption ionization time of flight mass spectrometry, MALDI analysis. The DPD software compares the MALDI results of the direct and inverse (18)O-labeling experiments and quickly identifies those peptides with paralleled loses in different sets of a typical proteomic workflow. Those peptides are used for subsequent accurate protein quantification. The interpretation of the MALDI data from direct and inverse labeling experiments is time-consuming requiring a significant amount of time to do all comparisons manually. The DPD software shortens and simplifies the searching of the peptides that must be used for quantification from a week to just some minutes. To do so, it takes as input several MALDI spectra and aids the researcher in an automatic mode (i) to compare data from direct and inverse (18)O-labeling experiments, calculating the corresponding ratios to determine those peptides with paralleled losses throughout different sets of experiments; and (ii) allow to use those peptides as internal standards for subsequent accurate protein quantification using (18)O-labeling. In this work the DPD software is presented and explained with the quantification of protein carbonic anhydrase. Copyright (c) 2010 Elsevier B.V. All rights reserved.

  14. Direct analysis of hCGβcf glycosylation in normal and aberrant pregnancy by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Iles, Ray K; Cole, Laurence A; Butler, Stephen A

    2014-06-05

    The analysis of human chorionic gonadotropin (hCG) in clinical chemistry laboratories by specific immunoassay is well established. However, changes in glycosylation are not as easily assayed and yet alterations in hCG glycosylation is associated with abnormal pregnancy. hCGβ-core fragment (hCGβcf) was isolated from the urine of women, pregnant with normal, molar and hyperemesis gravidarum pregnancies. Each sample was subjected to matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI TOF MS) analysis following dithiothreitol (DTT) reduction and fingerprint spectra of peptide hCGβ 6-40 were analyzed. Samples were variably glycosylated, where most structures were small, core and largely mono-antennary. Larger single bi-antennary and mixtures of larger mono-antennary and bi-antennary moieties were also observed in some samples. Larger glycoforms were more abundant in the abnormal pregnancies and tri-antennary carbohydrate moieties were only observed in the samples from molar and hyperemesis gravidarum pregnancies. Given that such spectral profiling differences may be characteristic, development of small sample preparation for mass spectral analysis of hCG may lead to a simpler and faster approach to glycostructural analysis and potentially a novel clinical diagnostic test.

  15. Flexible xxx-asp/asn and gly-xxx residues of equine cytochrome C in matrix-assisted laser desorption/ionization in-source decay mass spectrometry.

    Science.gov (United States)

    Takayama, Mitsuo

    2012-01-01

    The backbone flexibility of a protein has been studied from the standpoint of the susceptibility of amino acid residues to in-source decay (ISD) in matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). Residues more susceptible to MALDI-ISD, namely Xxx-Asp/Asn and Gly-Xxx, were identified from the discontinuous intense peak of c'-ions originating from specific cleavage at N-Cα bonds of the backbone of equine cytochrome c. The identity of the residues susceptible to ISD was consistent with the known flexible backbone amides as estimated by hydrogen/deuterium exchange (HDX) experiments. The identity of these flexible amino acid residues (Asp, Asn, and Gly) is consistent with the fact that these residues are preferred in flexible secondary structure free from intramolecular hydrogen-bonded structures such as α-helix and β-sheet. The MALDI-ISD spectrum of equine cytochrome c gave not only intense N-terminal side c'-ions originating from N-Cα bond cleavage at Xxx-Asp/Asn and Gly-Xxx residues, but also C-terminal side complement z'-ions originating from the same cleavage sites. The present study implies that MALDI-ISD can give information about backbone flexibility of proteins, comparable with the protection factors estimated by HDX.

  16. Flexible Xxx–Asp/Asn and Gly–Xxx Residues of Equine Cytochrome c in Matrix-Assisted Laser Desorption/Ionization In-Source Decay Mass Spectrometry

    Science.gov (United States)

    Takayama, Mitsuo

    2012-01-01

    The backbone flexibility of a protein has been studied from the standpoint of the susceptibility of amino acid residues to in-source decay (ISD) in matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). Residues more susceptible to MALDI-ISD, namely Xxx–Asp/Asn and Gly–Xxx, were identified from the discontinuous intense peak of c′-ions originating from specific cleavage at N–Cα bonds of the backbone of equine cytochrome c. The identity of the residues susceptible to ISD was consistent with the known flexible backbone amides as estimated by hydrogen/deuterium exchange (HDX) experiments. The identity of these flexible amino acid residues (Asp, Asn, and Gly) is consistent with the fact that these residues are preferred in flexible secondary structure free from intramolecular hydrogen-bonded structures such as α-helix and β-sheet. The MALDI-ISD spectrum of equine cytochrome c gave not only intense N-terminal side c′-ions originating from N–Cα bond cleavage at Xxx–Asp/Asn and Gly–Xxx residues, but also C-terminal side complement z′-ions originating from the same cleavage sites. The present study implies that MALDI-ISD can give information about backbone flexibility of proteins, comparable with the protection factors estimated by HDX. PMID:24349908

  17. Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass-Spectrometry (MALDI-TOF MS) Based Microbial Identifications: Challenges and Scopes for Microbial Ecologists.

    Science.gov (United States)

    Rahi, Praveen; Prakash, Om; Shouche, Yogesh S

    2016-01-01

    Matrix-assisted laser desorption/ionization time-of-flight mass-spectrometry (MALDI-TOF MS) based biotyping is an emerging technique for high-throughput and rapid microbial identification. Due to its relatively higher accuracy, comprehensive database of clinically important microorganisms and low-cost compared to other microbial identification methods, MALDI-TOF MS has started replacing existing practices prevalent in clinical diagnosis. However, applicability of MALDI-TOF MS in the area of microbial ecology research is still limited mainly due to the lack of data on non-clinical microorganisms. Intense research activities on cultivation of microbial diversity by conventional as well as by innovative and high-throughput methods has substantially increased the number of microbial species known today. This important area of research is in urgent need of rapid and reliable method(s) for characterization and de-replication of microorganisms from various ecosystems. MALDI-TOF MS based characterization, in our opinion, appears to be the most suitable technique for such studies. Reliability of MALDI-TOF MS based identification method depends mainly on accuracy and width of reference databases, which need continuous expansion and improvement. In this review, we propose a common strategy to generate MALDI-TOF MS spectral database and advocated its sharing, and also discuss the role of MALDI-TOF MS based high-throughput microbial identification in microbial ecology studies.

  18. Multi-imaging of Cytokinin and Abscisic Acid on the Roots of Rice (Oryza sativa) Using Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry.

    Science.gov (United States)

    Shiono, Katsuhiro; Hashizaki, Riho; Nakanishi, Toyofumi; Sakai, Tatsuko; Yamamoto, Takushi; Ogata, Koretsugu; Harada, Ken-Ichi; Ohtani, Hajime; Katano, Hajime; Taira, Shu

    2017-09-06

    Plant hormones act as important signaling molecules that regulate responses to abiotic stress as well as plant growth and development. Because their concentrations of hormones control the physiological responses in the target tissue, it is important to know the distributions and concentrations in the tissues. However, it is difficult to determine the hormone concentration on the plant tissue as a result of the limitations of conventional methods. Here, we report the first multi-imaging of two plant hormones, one of cytokinin [i.e., trans-zeatin (tZ)] and abscisic acid (ABA) using a new technology, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) imaging. Protonated signals of tZ (m/z 220.1) and ABA (m/z 265.3) were chosen on longitudinal sections of rice roots for MS imaging. tZ was broadly distributed about 40 mm behind the root apex but was barely detectable at the apex, whereas ABA was mainly detected at the root apex. Multi-imaging using MALDI-TOF-MS enabled the visualization of the localization and quantification of plant hormones. Thus, this tool is applicable to a wide range of plant species growing under various environmental conditions.

  19. Rapid Characterization of Microalgae and Microalgae Mixtures Using Matrix-Assisted Laser Desorption Ionization Time-Of-Flight Mass Spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Barbano, Duane; Diaz, Regina; Zhang, Lin; Sandrin, Todd; Gerken, Henri; Dempster, Thomas

    2015-01-01

    Current molecular methods to characterize microalgae are time-intensive and expensive. Matrix Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) may represent a rapid and economical alternative approach. The objectives of this study were to determine whether MALDI-TOF MS can be used to: 1) differentiate microalgae at the species and strain levels and 2) characterize simple microalgal mixtures. A common protein extraction sample preparation method was used to facilitate rapid mass spectrometry-based analysis of 31 microalgae. Each yielded spectra containing between 6 and 56 peaks in the m/z 2,000 to 20,000 range. The taxonomic resolution of this approach appeared higher than that of 18S rDNA sequence analysis. For example, two strains of Scenedesmus acutus differed only by two 18S rDNA nucleotides, but yielded distinct MALDI-TOF mass spectra. Mixtures of two and three microalgae yielded relatively complex spectra that contained peaks associated with members of each mixture. Interestingly, though, mixture-specific peaks were observed at m/z 11,048 and 11,230. Our results suggest that MALDI-TOF MS affords rapid characterization of individual microalgae and simple microalgal mixtures.

  20. Discrimination of Bacillus anthracis Spores by Direct in-situ Analysis of Matrix-Assisted Laser Desorption/Ionization Time-Of-Flight Mass Spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Jeong, Youngsu; Lee, Jonghee; Kim, Seongsoo [Agency for Defense Development, Daejeon (Korea, Republic of)

    2013-09-15

    The rapid and accurate identification of biological agents is a critical step in the case of bio-terror and biological warfare attacks. Recently, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry has been widely used for the identification of microorganisms. In this study, we describe a method for the rapid and accurate discrimination of Bacillus anthracis spores using MALDI-TOF MS. Our direct in-situ analysis of MALDI-TOF MS does not involve subsequent high-resolution mass analyses and sample preparation steps. This method allowed the detection of species-specific biomarkers from each Bacillus spores. Especially, B. anthracis spores had specific biomarker peaks at 2503, 3089, 3376, 6684, 6698, 6753, and 6840 m/z. Cluster and PCA analyses of the mass spectra of Bacillus spores revealed distinctively separated clusters and within-groups similarity. Therefore, we believe that this method is effective in the real-time identification of biological warfare agents such as B. anthracis as well as other microorganisms in the field.

  1. [Performance of two matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS) models for identification of bacteria isolated from blood culture].

    Science.gov (United States)

    Itoh, Eisuke; Watari, Tomohisa; Azuma, Yuka; Watanabe, Naoki; Tomoda, Yutaka; Akasaka, Kazumi; Kino, Shuichi

    2013-05-01

    We compared the results of two bacterial identification methods: 1) a traditional method based on phenotypic identification of the causative organism using gram-staining, culture and biochemical markers and 2) matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). A total of 111 isolates, including 107 strains of common bacteria species and 4 strains of 3 yeast species, were tested by the traditional method and MALDI-TOF MS method(VITEK MS and Micro flex LT). Data obtained using MALDI-TOF MS were classified as Level 1 and Level 2 according to the confidence level of identification results from the VITEK MS ver. 1.0 database (VITEK MS) and MALDI Biotyper ver. 2.0 database (Microflex LT). The proportions of measured samples identified as Level 1 were 98.2% with the VITEK MS database and 87.4% with the MALDI Biotyper database. The concordance rates of the traditional method were 93.7% with the VITEK MS database and 82.0% with the MALDI Biotyper database. Identification results of five strains were mismatched between the traditional method and MALDI-TOF MS. Their ribosomal RNA sequences were identical to the results obtained from MALDI-TOF MS. We concluded that the performance of VITEK MS is superior to that of the traditional method and Microflex LT.

  2. Detection of melamine in a human renal uric acid stone by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Liu, Chia-Chu; Wu, Chia-Fang; Shiea, Jentaie; Cho, Yi-Tzu; Hsieh, Tusty-Jiuan; Chou, Yii-Her; Chen, Bai-Hsiun; Huang, Shu-Pin; Wu, Wen-Jeng; Shen, Jung-Tsung; Chang, Mei-Yu; Huang, Chun-Hsiung; Chang, Ai-Wen; Wu, Ming-Tsang

    2012-10-09

    The link between melamine-contaminated daily foodstuffs and urolithiasis formation has drawn an international concern. However, detection of melamine levels in urine may not completely represent external melamine exposure. Thus, finding an additional analytical method for the study of environmental melamine exposure and its adverse effect in humans is crucial. Eleven adult patients diagnosed with uric acid urolithiasis were retrospectively analyzed. Melamine levels in their overnight one-spot urine samples were measured by a triple quadrupole liquid chromatography tandem mass spectrometry (LC-MS/MS). The compositions of stone samples were analyzed by the Fourier transform infrared (FTIR) spectrophotometer and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Seven (63.6%) out of 11 patients had detectable melamine levels in their urine specimens (method of detection limit: 0.8 ng/ml). Three patients (27.3%) were highly suspected of having melamine-containing urolithiasis in FTIR spectra. In one of those three cases who still had available stored stone specimens, MALDI-TOF MS further confirmed melamine components in this male patient's stone specimens. In contrast, his urinary melamine level was below the detection limit by LC-MS/MS. Direct analysis of melamine in the composition of urolithiasis by MALDI-TOF MS can be an additional analytical method to evaluate for external melamine exposure. Copyright © 2012 Elsevier B.V. All rights reserved.

  3. Matrix-Assisted Laser Desorption Ionization–Time of Flight (MALDI-TOF) Mass Spectrometry for Detection of Antibiotic Resistance Mechanisms: from Research to Routine Diagnosis

    Science.gov (United States)

    Chudáčková, Eva; Walková, Radka

    2013-01-01

    Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) has been successfully applied as an identification procedure in clinical microbiology and has been widely used in routine laboratory practice because of its economical and diagnostic benefits. The range of applications of MALDI-TOF MS has been growing constantly, from rapid species identification to labor-intensive proteomic studies of bacterial physiology. The purpose of this review is to summarize the contribution of the studies already performed with MALDI-TOF MS concerning antibiotic resistance and to analyze future perspectives in this field. We believe that current research should continue in four main directions, including the detection of antibiotic modifications by degrading enzymes, the detection of resistance mechanism determinants through proteomic studies of multiresistant bacteria, and the analysis of modifications of target sites, such as ribosomal methylation. The quantification of antibiotics is suggested as a new approach to study influx and efflux in bacterial cells. The results of the presented studies demonstrate that MALDI-TOF MS is a relevant tool for the detection of antibiotic resistance and opens new avenues for both clinical and experimental microbiology. PMID:23297261

  4. Principle component analysis combined with matrix-assisted laser desorption ionization mass spectrometry for rapid diagnosing the sera of patients with major depression.

    Science.gov (United States)

    Huang, Tiao-Lai; Cho, Yi-Tzu; Su, Hong; Shiea, Jentaie

    2013-09-23

    Previously, we demonstrated that 6M HCl hydrolysis followed by matrix-assisted laser desorption/ionization (MALDI-TOF) mass spectrometry is a useful technique to detect potential protein biomarkers in the sera collected from major depression (MD) patients and from healthy controls. In this study, the effects of various organic acids in hydrolyzing proteins in serum were first examined. The organic matrixes commonly used in MALDI analysis were also examined for characterizing the hydrolyzed peptides. Finally, principle component analysis (PCA) was used to analyze the MALDI mass spectra of acid-hydrolyzed serum samples. It was found that 20% TFA and sinapinic acid were the optimal reagents for hydrolysis and MALDI matrix. Samples collected from MD patients and healthy controls were readily classified through PCA analysis. A receiver operating characteristic (ROC) curve based on the ratio of the intensities of the two fragment ions (m/z 8606 and 9287) indicated by PCA plot was also constructed. The area under the curve was 0.845; the sensitivity and specificity were both 80%. An analytical platform employing trifluoroacetic acid to hydrolyze serum proteins followed by MALDI-TOF/MS and PCA analysis was developed to rapidly differentiate the sera between MD patients and healthy controls. Copyright © 2013 Elsevier B.V. All rights reserved.

  5. Proteogenomic biomarkers for identification of Francisella species and subspecies by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry.

    Science.gov (United States)

    Durighello, Emie; Bellanger, Laurent; Ezan, Eric; Armengaud, Jean

    2014-10-07

    Francisella tularensis is the causative agent of tularemia. Because some Francisella strains are very virulent, this species is considered by the Centers for Disease Control and Prevention to be a potential category A bioweapon. A mass spectrometry method to quickly and robustly distinguish between virulent and nonvirulent Francisella strains is desirable. A combination of shotgun proteomics and whole-cell matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF) mass spectrometry on the Francisella tularensis subsp. holarctica LVS defined three protein biomarkers that allow such discrimination: the histone-like protein HU form B, the 10 kDa chaperonin Cpn10, and the 50S ribosomal protein L24. We established that their combined detection by whole-cell MALDI-TOF spectrum could enable (i) the identification of Francisella species, and (ii) the prediction of their virulence level, i.e., gain of a taxonomical level with the identification of Francisella tularensis subspecies. The detection of these biomarkers by MALDI-TOF mass spectrometry is straightforward because of their abundance and the absence of other abundant protein species closely related in terms of m/z. The predicted molecular weights for the three biomarkers and their presence as intense peaks were confirmed with MALDI-TOF/MS spectra acquired on Francisella philomiragia ATCC 25015 and on Francisella tularensis subsp. tularensis CCUG 2112, the most virulent Francisella subspecies.

  6. Barcode DNA-Mediated Signal Amplifying Strategy for Ultrasensitive Biomolecular Detection on Matrix-Assisted Laser Desorption Ionization Time of Flight (MALDI-TOF) Mass Spectrometry.

    Science.gov (United States)

    Ahmad, Raheel; Jang, Hyowon; Batule, Bhagwan S; Park, Hyun Gyu

    2017-09-05

    We have devised a barcode DNA-mediated signal amplifying strategy for ultrasensitive biomolecular detection by utilizing matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). As a model target, thrombin was first captured by specific aptamer15 functionalized on magnetic beads (MBs-apt15) and sandwiched through the simultaneous interaction with gold nanoparticles modified with another specific aptamer29 and barcode DNA molecules (apt29-AuNPs-bcDNAs). The sandwiched complex was collected by convenient magnetic separation and then treated with potassium cyanide (KCN) to dissolve the gold nanoparticles (AuNPs) and consequently release the barcode DNA molecules (bcDNAs), which were then again magnetically separated and analyzed by using MALDI-TOF MS. Under optimized conditions, this strategy revealed an excellent sensitivity with a limit of detection of 0.89 aM in a wide linear detection range from 0 aM to 0.1 nM and exhibited an acceptable recovery for thrombin detection in complex biological matrices. This signal amplifying strategy based on MALDI-TOF MS could greatly enable the ultrasensitive detection of various low abundant biomolecules.

  7. In vitro detection of bacterial contamination in platelet concentrates by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry: a preliminary study.

    Science.gov (United States)

    Chetouane, Yasmine; Dubourg, Gregory; Gallian, Pierre; Delerce, Jeremy; Levasseur, Anthony; Flaudrops, Christophe; Chabrière, Eric; Chiaroni, Jacques; Raoult, Didier; Camoin-Jau, Laurence

    2017-11-01

    Platelet concentrates are at risk of transfusion-related sepsis. The microbial detection methods currently available have reached their limits, as they do not completely prevent transfusion-related bacterial contamination.The aim of this study was to develop a new strategy to detect the risk of platelet transfusion-related bacterial contamination using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). In vitro, platelet concentrates were seeded with known concentrations of bacterial strains. Protein mass profiles were acquired by using a Microflex MALDI-TOF instrument. Dedicated 'Platelet' software was used as a spectrum subtraction tool to reveal specific peaks caused by the presence of pathogens in samples. The MALDI-TOF spectra of platelets were characterized and the reproducibility over time, regardless of the blood donor, was demonstrated with a positive predictive value of 100 %. In addition, the negative predictive value of the total number of specific peaks to predict contamination was 100 %. Detecting bacteria in platelet concentrates using the MALDI-TOF approach and analysing spectra with the Platelet software present the advantage of combining the precocity of results and sufficient sensitivity (10 c.f.u. ml-1). Further research will be conducted to compare this novel method with the current conventional method in order to validate our results, the objective being to reduce the risk of platelet transfusion-related bacterial contamination.

  8. Application of Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Imaging Mass Spectrometry (MALDI-TOF IMS) for Premalignant Gastrointestinal Lesions.

    Science.gov (United States)

    Ko, Kwang Hyun; Kwon, Chang Il; Park, So Hye; Han, Na Young; Lee, Hoo Keun; Kim, Eun Hee; Hahm, Ki Baik

    2013-11-01

    Imaging mass spectrometry (IMS) is currently receiving large attention from the mass spectrometric community, although its use is not yet well known in the clinic. As matrix-assisted laser desorption/ionization time-of-flight (MALDI)-IMS can show the biomolecular changes in cells as well as tissues, it can be an ideal tool for biomedical diagnostics as well as the molecular diagnosis of clinical specimens, especially aimed at the prompt detection of premalignant lesions much earlier before overt mass formation, or for obtaining histologic clues from endoscopic biopsy. Besides its use for pathologic diagnosis, MALDI-IMS is also a powerful tool for the detection and localization of drugs, proteins, and lipids in tissue. Measurement of parameters that define and control the implications, challenges, and opportunities associated with the application of IMS to biomedical tissue studies might be feasible through a deep understanding of mass spectrometry. In this focused review series, new insights into the molecular processes relevant to IMS as well as other field applications are introduced.

  9. Matrix-Assisted Laser Desorption-Ionization Time-of-Flight Mass Spectrometry (MALDI- TOF MS) for Early Identification of Septic Patients.

    Science.gov (United States)

    Sekercioglu, Ali O; Cekin, Yesim; Ogunc, Dilara; Ongut, Gozde; Baysan, Betil O; Colak, Dilek; Gunseren, Filiz; Donmez, Levent

    2017-04-01

    Matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a powerful technique for the rapid identification of bacteria from growing colonies in routine cultures. In this study, we evaluated the feasibility of a 5-hour incubation on solid medium after sub-cultivation of positive blood culture broth without any preparation steps in order to speed up the identification of bacteria. In addition to standard laboratory protocols, a Columbia agar plate with 5% sheep blood was inoculated with 1 drop from the blood culture broth. After a 5-hour incubation period, a colony from the culture plate was submitted to MALDI-TOF MS. A total of 1351 positive blood cultures (1299 monomicrobial and 51 polymicrobial) were analyzed. When compared to routine identification procedure results for positive blood cultures, 79.3% of isolates were correctly identified to the species level. When manufacturer-recommended score values were taken into account, MALDITOF MS correctly identified 98.4% of the isolates to the species level with a score of > 2.0, 89.1% with a score between 1.7 and 2.0, and 75.4% with a score of MALDI-TOF MS identification after short-term culture is directly related to the sufficient growth of microorganisms at 5 hours.

  10. Quantitation of lysergic acid diethylamide in urine using atmospheric pressure matrix-assisted laser desorption/ionization ion trap mass spectrometry.

    Science.gov (United States)

    Cui, Meng; McCooeye, Margaret A; Fraser, Catharine; Mester, Zoltán

    2004-12-01

    A quantitative method was developed for analysis of lysergic acid diethylamide (LSD) in urine using atmospheric pressure matrix-assisted laser desorption/ionization ion trap mass spectrometry (AP MALDI-ITMS). Following solid-phase extraction of LSD from urine samples, extracts were analyzed by AP MALDI-ITMS. The identity of LSD was confirmed by fragmentation of the [M + H](+) ion using tandem mass spectrometry. The quantification of LSD was achieved using stable-isotope-labeled LSD (LSD-d(3)) as the internal standard. The [M + H](+) ion fragmented to produce a dominant fragment ion, which was used for a selected reaction monitoring (SRM) method for quantitative analysis of LSD. SRM was compared with selected ion monitoring and produced a wider linear range and lower limit of quantification. For SRM analysis of samples of LSD spiked in urine, the calibration curve was linear in the range of 1-100 ng/mL with a coefficient of determination, r(2), of 0.9917. This assay was used to determine LSD in urine samples and the AP MALDI-MS results were comparable to the HPLC/ ESI-MS results.

  11. Matrix-assisted laser desorption ionisation time-of-flight mass spectrometry rapid detection of carbapenamase activity in Acinetobacter baumannii isolates.

    Science.gov (United States)

    Abouseada, Noha; Raouf, May; El-Attar, Eman; Moez, Pacinte

    2017-01-01

    Carbapenamase-producing Acinetobacter baumannii are an increasing threat in hospitals and Intensive Care Units. Accurate and rapid detection of carbapenamase producers has a great impact on patient improvement and aids in implementation of infection control measures. In this study, we describe the use of matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI TOF MS) to identify carbapenamase-producing A. baumannii isolates in up to 3 h. Isolates and Methods: A total of 50 A. baumannii isolates (of which 39 were carabapenamase producers) were tested using MALDI TOF MS. Isolates were incubated for 3 h with 0.25 mg/ml up to 2 mg/ml of imipenem (IMP) at 37°C. Supernatants were analysed by MALDI TOF to analyse peaks corresponding to IMP (300 Da) and an IMP metabolite (254 Da) using UltrafleXtreme (Bruker Daltonics, Bremen, Germany). All carbapenamase-producing isolates were evidenced by the disappearance or reduction in intensity of the 300 Da peak of IPM and the appearance of a 254 Da peak of the IPM metabolite. In isolates that did not produce carbapenamase, the IPM 300 Da peak remained intact. MALDI TOF is a promising tool in the field of diagnostic microbiology that has the ability to transfer identification and antimicrobial susceptibility testing time from days to hours.

  12. Identification of N-glycans from Ebola virus glycoproteins by matrix-assisted laser desorption/ionisation time-of-flight and negative ion electrospray tandem mass spectrometry

    Science.gov (United States)

    Ritchie, Gayle; Harvey, David J.; Stroeher, Ute; Feldmann, Friederike; Feldmann, Heinz; Wahl-Jensen, Victoria; Royle, Louise; Dwek, Raymond A.; Rudd, Pauline M.

    2012-01-01

    The larger fragment of the transmembrane glycoprotein (GP1) and the soluble glycoprotein (sGP) of Ebola virus were expressed in human embryonic kidney cells and the secreted products were purified from the supernatant for carbohydrate analysis. The N-glycans were released with PNGase F from within sodium dodecyl sulphate/polyacrylamide gel electrophoresis (SDS-PAGE) gels. Identification of the glycans was made with normal-phase high-performance liquid chromatography (HPLC), matrix-assisted laser desorption/ionisation mass spectrometry, negative ion electrospray ionisation fragmentation mass spectrometry and exoglycosidase digestion. Most glycans were complex bi-, tri-and tetra-antennary compounds with reduced amounts of galactose. No bisected compounds were detected. Triantennary glycans were branched on the 6-antenna; fucose was attached to the core GlcNAc residue. Sialylated glycans were present on sGP but were largely absent from GP1, the larger fragment of the transmembrane glycoprotein. Consistent with this was the generally higher level of processing of carbohydrates found on sGP as evidenced by a higher percentage of galactose and lower levels of high-mannose glycans than were found on GP1. These results confirm and expand previous findings on partial characterisation of the Ebola virus transmembrane glycoprotein. They represent the first detailed data on carbohydrate structures of the Ebola virus sGP. PMID:20131323

  13. Investigation of colloidal graphite as a matrix for matrix-assisted laser desorption/ionisation mass spectrometry of low molecular weight analytes.

    Science.gov (United States)

    Warren, Alexander D; Conway, Ulric; Arthur, Christopher J; Gates, Paul J

    2016-07-01

    The analysis of low molecular weight compounds by matrix-assisted laser desorption/ionisation mass spectrometry is problematic due to the interference and suppression of analyte ionisation by the matrices typically employed - which are themselves low molecular weight compounds. The application of colloidal graphite is demonstrated here as an easy to use matrix that can promote the ionisation of a wide range of analytes including low molecular weight organic compounds, complex natural products and inorganic complexes. Analyte ionisation with colloidal graphite is compared with traditional organic matrices along with various other sources of graphite (e.g. graphite rods and charcoal pencils). Factors such as ease of application, spectra reproducibility, spot longevity, spot-to-spot reproducibility and spot homogeneity (through single spot imaging) are explored. For some analytes, considerable matrix suppression effects are observed resulting in spectra completely devoid of matrix ions. We also report the observation of radical molecular ions [M(-●) ] in the negative ion mode, particularly with some aromatic analytes. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  14. A simple method for rapid microbial identification from positive monomicrobial blood culture bottles through matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Lin, Jung-Fu; Ge, Mao-Cheng; Liu, Tsui-Ping; Chang, Shih-Cheng; Lu, Jang-Jih

    2017-06-30

    Rapid identification of microbes in the bloodstream is crucial in managing septicemia because of its high disease severity, and direct identification from positive blood culture bottles through matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) can shorten the turnaround time. Therefore, we developed a simple method for rapid microbiological identification from positive blood cultures by using MALDI-TOF MS. We modified previously developed methods to propose a faster, simpler and more economical method, which includes centrifugation and hemolysis. Specifically, our method comprises two-stage centrifugation with gravitational acceleration (g) at 600g and 3000g, followed by the addition of a lysis buffer and another 3000g centrifugation. In total, 324 monomicrobial bacterial cultures were identified. The success rate of species identification was 81.8%, which is comparable with other complex methods. The identification success rate was the highest for Gram-negative aerobes (85%), followed by Gram-positive aerobes (78.2%) and anaerobes (67%). The proposed method requires less than 10 min, costs less than US$0.2 per usage, and facilitates batch processing. We conclude that this method is feasible for clinical use in microbiology laboratories, and can serve as a reference for treatments or further complementary diagnostic testing. Copyright © 2017. Published by Elsevier B.V.

  15. Identification of Cronobacter species by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry with an optimized analysis method.

    Science.gov (United States)

    Wang, Qi; Zhao, Xiao-Juan; Wang, Zi-Wei; Liu, Li; Wei, Yong-Xin; Han, Xiao; Zeng, Jing; Liao, Wan-Jin

    2017-08-01

    Rapid and precise identification of Cronobacter species is important for foodborne pathogen detection, however, commercial biochemical methods can only identify Cronobacter strains to genus level in most cases. To evaluate the power of mass spectrometry based on matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF MS) for Cronobacter species identification, 51 Cronobacter strains (eight reference and 43 wild strains) were identified by both MALDI-TOF MS and 16S rRNA gene sequencing. Biotyper RTC provided by Bruker identified all eight reference and 43 wild strains as Cronobacter species, which demonstrated the power of MALDI-TOF MS to identify Cronobacter strains to genus level. However, using the Bruker's database (6903 main spectra products) and Biotyper software, the MALDI-TOF MS analysis could not identify the investigated strains to species level. When MALDI-TOF MS analysis was performed using the combined in-house Cronobacter database and Bruker's database, bin setting, and unweighted pair group method with arithmetic mean (UPGMA) clustering, all the 51 strains were clearly identified into six Cronobacter species and the identification accuracy increased from 60% to 100%. We demonstrated that MALDI-TOF MS was reliable and easy-to-use for Cronobacter species identification and highlighted the importance of establishing a reliable database and improving the current data analysis methods by integrating the bin setting and UPGMA clustering. Copyright © 2017. Published by Elsevier B.V.

  16. A novel matrix-assisted laser desorption/ionisation mass spectrometry imaging based methodology for the identification of sexual assault suspects.

    Science.gov (United States)

    Bradshaw, Robert; Wolstenholme, Rosalind; Blackledge, Robert D; Clench, Malcolm R; Ferguson, Leesa S; Francese, Simona

    2011-02-15

    An increase in the use of condoms by sexual offenders has been observed. This is likely to be due both to the risk of sexually transmitted diseases and to prevent the transfer of DNA evidence. In this scenario the detection of condom lubricants at a crime scene could aid in proving corpus delicti. Here we show a novel application of Matrix-Assisted Laser Desorption/Ionisation Mass Spectrometry Imaging (MALDI MSI) for mapping the fingermark ridge pattern simultaneously to the detection of the condom lubricant within the fingermark itself. Two condom brands have been investigated to prove the concept. Condoms were handled producing lubricant-contaminated fingermarks. Images of the ridge pattern were obtained simultaneously to the detection of two lubricants, even several weeks after the fingermark deposition. The results therefore show the potential of MALDI MSI to link the suspect (identification through fingermark ridge pattern) to the crime (detection of condom lubricant) in one analysis. This would enable forensic scientists to provide evidence with stronger support in alleged cases of sexual assault. Copyright © 2011 John Wiley & Sons, Ltd.

  17. Matrix-assisted laser desorption/ionization mass spectrometry for quantitative determination of β-blocker drugs in one-drop of human serum sample.

    Science.gov (United States)

    Shrivas, Kamlesh; Patel, Devesh Kumar

    2011-01-01

    Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) has been applied for the quantitative determination of β-blocker drugs in one-drop of human serum samples using drop-to-drop solvent microextraction (DDSME) as a preconcentrating probe. The optimum experimental conditions for β-blocker drugs were investigated and 1.8 μL volume of toluene for 10 min extraction time with the 5% addition of NaCl under pH 11.0 was found to be the best conditions for the separation and preconcentration of drugs from 30 μL of serum sample from a patient with high blood pressure. The optimized methodologies for DDSME/MALDI-MS analyses exhibited a good linearity with intra- and inter day precision value of 8.5-10.5% and 9.4-12.6%, respectively. The proposed DDSME/MALDI-MS offers a very simple, rapid and low-cost technique for the determination of β-blocker drugs in one drop of serum sample. The reported method has been successfully applied for the determination of propranolol and nadolol in small volume of serum sample from patient suffering from high blood pressure. In future, this technique could be applied for pharmacokinetic and clinical studies. Copyright © 2010 Elsevier B.V. All rights reserved.

  18. Determination of immunosuppressive drugs in human urine and serum by surface-assisted laser desorption/ionization mass spectrometry with dispersive liquid-liquid microextraction.

    Science.gov (United States)

    Chen, Pin-Shiuan; Cheng, Yu-Han; Lin, Sheng-Yu; Chang, Sarah Y

    2016-01-01

    A rapid and sensitive method for the determination of immunosuppressive drugs through surface-assisted laser desorption/ionization mass spectrometric detection (SALDI/MS) was developed. Colloidal Pd and α-cyano-4-hydroxycinnamic acid (CHCA) were used as the SALDI co-matrix. To eliminate interference and enhance the sensitivity, dispersive liquid-liquid microextraction (DLLME) was employed to extract the immunosuppressive drugs from the aqueous solutions. Under optimal extraction and detection conditions, calibration curves for cyclosporine and everolimus in aqueous solutions were linear over a concentration range from 0.01 to 1.20 μM. For sirolimus, the linear concentration range of the calibration curve was from 0.05 to 2.00 μM. The limits of detection (LODs) were calculated to be 3, 3, and 14 nM for cyclosporine, everolimus, and sirolimus, respectively. The enrichment factors of DLLME were calculated to be 108, 122, and 101 for cyclosporine, everolimus, and sirolimus, respectively. This novel method was successfully applied for the determination of immunosuppressive drugs in human urine and serum samples.

  19. Rapid determination of rivaroxaban in human urine and serum using colloidal palladium surface-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Cheng, Yu-Han; Chen, Wen-Chi; Chang, Sarah Y

    2015-11-15

    Rivaroxaban is a new anticoagulant drug that has recently been introduced for clinical applications. To ensure optimum efficacy while minimizing the risk of toxicity and other adverse effects, a simple and sensitive analytical procedure for monitoring the concentration of rivaroxaban in biological fluids is required. Rivaroxaban was extracted from aqueous solutions by dispersive liquid-liquid microextraction (DLLME). Detection of rivaroxaban was achieved through surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS) using colloidal palladium as the SALDI matrix. The calibration curve for rivaroxaban in aqueous solutions was linear over the concentration range from 5 to 500 nM. The limit of detection (LOD) for rivaroxaban at a signal-to-noise ratio of 3 was 2 nM. With a sample-to-extract volume ratio of 200, the enrichment factors were calculated to be 141. This method was successfully applied for the determination of rivaroxaban in human urine and serum samples. The LODs for rivaroxaban in urine and serum were calculated to be 6 nM and 60 nM, respectively. The analysis speed, together with the ease of operation and high sensitivity, allows SALDI-MS method to be particularly suitable for the high-throughput screening of rivaroxaban levels in human urine and serum samples. Copyright © 2015 John Wiley & Sons, Ltd.

  20. Analysis of cancer cell lipids using matrix-assisted laser desorption/ionization 15-T Fourier transform ion cyclotron resonance mass spectrometry.

    Science.gov (United States)

    Yang, Hyo-Jik; Park, Kyu Hwan; Lim, Dong Wan; Kim, Hyun Sik; Kim, Jeongkwon

    2012-03-30

    A combination of methodologies using the extremely high mass accuracy and resolution of 15-T Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry (MS) was introduced for the identification of intact cancer cell phospholipids. Lipids from a malignant glioma cell line were initially analyzed at a resolution of >200,000 and identified by setting the mass tolerance to ±1 mDa using matrix-assisted laser desorption/ionization (MALDI) 15-T FT-ICR MS in positive ion mode. In most cases, a database search of potential lipid candidates using the exact masses of the lipids yielded only one possible chemical composition. Extremely high mass accuracy (800,000), yielded well-resolved isotopic fine structures allowing for the identification of lipids by MALDI 15-T FT-ICR MS without using tandem mass spectrometric (MS/MS) analysis. Using this method, a total of 38 unique lipids were successfully identified. Copyright © 2012 John Wiley & Sons, Ltd.

  1. Hydrophilic interaction chromatography coupled matrix assisted laser desorption/ionization mass spectrometry for molecular analysis of organic compounds in medicines, tea, and coffee

    KAUST Repository

    Wang, Renqi

    2013-01-01

    Natural occurring organic compounds from food, natural organic matter, as well as metabolic products have received intense attention in current chemical and biological studies. Examination of unknown compounds in complex sample matrices is hampered by the limited choices for data readout and molecular elucidation. Herein, we report a generic method of hydrophilic interaction chromatography (HILIC) coupled with matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS) for the rapid characterization of ingredients in pharmaceutical compounds, tea, and coffee. The analytes were first fractionated using a cationic HILIC column prior to MALDI-MS analyses. It was found that the retention times of a compound arising from different samples were consistent under the same conditions. Accordingly, molecules can be readily characterized by both the mass and chromatographic retention time. The retention behaviors of acidic and basic compounds on the cationic HILIC column were found to be significantly influenced by the pH of mobile phases, whereas neutral compounds depicted a constant retention time at different pH. The general HILIC-MALDI-MS method is feasible for fast screening of naturally occurring organic compounds. A series of homologs can be determined if they have the same retention behavior. Their structural features can be elucidated by considering their mass differences and hydrophilic properties as determined by HILIC chromatogram. © 2013 The Royal Society of Chemistry.

  2. Effect of matrix and solvent on the analysis of novel poly(phenylenevinylene) derivatives by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Zhang, Zhenying; Deng, Huimin; Deng, Qinying; Zhao, Shankai

    2004-01-01

    Six novel poly(phenylenevinylene) (PPV) derivatives carrying butoxy or myrtanyl groups, including poly(2-butoxy-m-phenylenevinylene) (Bu-MPV), poly(2,5-dibutoxy-p-phenylenevinylene-alter-p-phenylenevinylene) (Bu-PPPV), poly(2,5-dibutoxy-p-phenylenevinylene-alter-m-phenylenevinylene) (Bu-PMPV), poly(2-myrtanyl-m-phenylenevinylene) (Myr-MPV), poly(2,5-dimyrtanyl-p-phenylenevinylene-alter-p-phenylenevinylene) (Myr-PPPV), and poly(2,5-dimyrtanyl-p-phenylenevinylene-alter-m-phenylenevinylene) (Myr-PMPV), were characterized by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). The repeat unit mass and the end-group structures of each sample were obtained. Distinctly different spectra with different ion series and/or different ion signal intensities were observed for the analytes Bu-MPV, Myr-MPV, Bu-PPPV, Myr-PPPV and Myr-PMPV when different matrices were used, and different ion series were acquired when different solvents were used for Myr-PPPV and Myr-PMPV. The results show that the PPV oligomers with different shapes and/or with different end groups can be selectively desorbed and ionized in MALDI by using different matrices. Copyright 2004 John Wiley & Sons, Ltd.

  3. Direct Analysis of hCGβcf Glycosylation in Normal and Aberrant Pregnancy by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry

    Directory of Open Access Journals (Sweden)

    Ray K. Iles

    2014-06-01

    Full Text Available The analysis of human chorionic gonadotropin (hCG in clinical chemistry laboratories by specific immunoassay is well established. However, changes in glycosylation are not as easily assayed and yet alterations in hCG glycosylation is associated with abnormal pregnancy. hCGβ-core fragment (hCGβcf was isolated from the urine of women, pregnant with normal, molar and hyperemesis gravidarum pregnancies. Each sample was subjected to matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI TOF MS analysis following dithiothreitol (DTT reduction and fingerprint spectra of peptide hCGβ 6–40 were analyzed. Samples were variably glycosylated, where most structures were small, core and largely mono-antennary. Larger single bi-antennary and mixtures of larger mono-antennary and bi-antennary moieties were also observed in some samples. Larger glycoforms were more abundant in the abnormal pregnancies and tri-antennary carbohydrate moieties were only observed in the samples from molar and hyperemesis gravidarum pregnancies. Given that such spectral profiling differences may be characteristic, development of small sample preparation for mass spectral analysis of hCG may lead to a simpler and faster approach to glycostructural analysis and potentially a novel clinical diagnostic test.

  4. Examination of the translocation of sulfonylurea herbicides in sunflower plants by matrix-assisted laser desorption/ionisation mass spectrometry imaging.

    Science.gov (United States)

    Anderson, David M G; Carolan, Vikki A; Crosland, Susan; Sharples, Kate R; Clench, Malcolm R

    2010-11-30

    Pesticides are widely used in agriculture to control weeds, pests and diseases. Successful control is dependent on the compound reaching the target site within the organism after spray or soil application. Conventional methods for determining uptake and movement of herbicides and pesticides include autoradiography, liquid scintillation and chromatographic techniques such as high-performance liquid chromatography (HPLC). Autoradiography using radiolabelled compounds provides the best indication of a compound's movement within the plant system. Autoradiography is an established technique but it relies on the synthesis of radiolabelled compounds. The distribution of four sulfonylurea herbicides in sunflower plants has been studied 24  h after foliar application. The use of matrix-assisted laser desorption/ionisation mass spectrometry imaging (MALDI-MSI) images of protonated molecules and fragment ions (resulting from fragmentation at the urea bond within the sulfonylurea herbicides) has provided evidence for translocation above and below the application point. The translocation of nicosulfuron and azoxystrobin within the same plant system has also been demonstrated following their application to the plant stem. This study provides evidence that MALDI-MSI has great potential as an analytical technique to detect and assess the foliar, root and stem uptake of agrochemicals, and to reveal their distribution through the plant once absorbed and translocated. 2010 John Wiley & Sons, Ltd.

  5. Successful Identification of Clinical Dermatophyte and Neoscytalidium Species by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

    Science.gov (United States)

    Alshawa, Kinda; Beretti, Jean-Luc; Lacroix, Claire; Feuilhade, Martine; Dauphin, Brunhilde; Quesne, Gilles; Hassouni, Noura; Nassif, Xavier

    2012-01-01

    Dermatophytes are keratinolytic fungi responsible for a wide variety of diseases of glabrous skin, nails, and hair. Their identification, currently based on morphological criteria, is hindered by intraspecies morphological variability and the atypical morphology of some clinical isolates. The aim of this study was to evaluate matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) as a routine tool for identifying dermatophyte and Neoscytalidium species, both of which cause dermatomycoses. We first developed a spectral database of 12 different species of common and unusual dermatophytes and two molds responsible for dermatomycoses (Neoscytalidium dimidiatum and N. dimidiatum var. hyalinum). We then prospectively tested the performance of the database on 381 clinical dermatophyte and Neoscytalidium isolates. Correct identification of the species was obtained for 331/360 dermatophytes (91.9%) and 18/21 Neoscytalidium isolates (85.7%). The results of MALDI-TOF MS and standard identification disagreed for only 2 isolates. These results suggest that MALDI-TOF MS could be a useful tool for routine and fast identification of dermatophytes and Neoscytalidium spp. in clinical mycology laboratories. PMID:22535981

  6. Matrix-assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS Can Precisely Discriminate the Lineages of Listeria monocytogenes and Species of Listeria.

    Directory of Open Access Journals (Sweden)

    Teruyo Ojima-Kato

    Full Text Available The genetic lineages of Listeria monocytogenes and other species of the genus Listeria are correlated with pathogenesis in humans. Although matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS has become a prevailing tool for rapid and reliable microbial identification, the precise discrimination of Listeria species and lineages remains a crucial issue in clinical settings and for food safety. In this study, we constructed an accurate and reliable MS database to discriminate the lineages of L. monocytogenes and the species of Listeria (L. monocytogenes, L. innocua, L. welshimeri, L. seeligeri, L. ivanovii, L. grayi, and L. rocourtiae based on the S10-spc-alpha operon gene encoded ribosomal protein mass spectrum (S10-GERMS proteotyping method, which relies on both genetic information (genomics and observed MS peaks in MALDI-TOF MS (proteomics. The specific set of eight biomarkers (ribosomal proteins L24, L6, L18, L15, S11, S9, L31 type B, and S16 yielded characteristic MS patterns for the lineages of L. monocytogenes and the different species of Listeria, and led to the construction of a MS database that was successful in discriminating between these organisms in MALDI-TOF MS fingerprinting analysis followed by advanced proteotyping software Strain Solution analysis. We also confirmed the constructed database on the proteotyping software Strain Solution by using 23 Listeria strains collected from natural sources.

  7. Hydrophilic interaction chromatography coupled matrix assisted laser desorption/ionization mass spectrometry for molecular analysis of organic compounds in medicines, tea, and coffee.

    Science.gov (United States)

    Wang, Ren-Qi; Bao, Kai; Croué, Jean-Philippe; Ng, Siu Choon

    2013-11-21

    Natural occurring organic compounds from food, natural organic matter, as well as metabolic products have received intense attention in current chemical and biological studies. Examination of unknown compounds in complex sample matrices is hampered by the limited choices for data readout and molecular elucidation. Herein, we report a generic method of hydrophilic interaction chromatography (HILIC) coupled with matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS) for the rapid characterization of ingredients in pharmaceutical compounds, tea, and coffee. The analytes were first fractionated using a cationic HILIC column prior to MALDI-MS analyses. It was found that the retention times of a compound arising from different samples were consistent under the same conditions. Accordingly, molecules can be readily characterized by both the mass and chromatographic retention time. The retention behaviors of acidic and basic compounds on the cationic HILIC column were found to be significantly influenced by the pH of mobile phases, whereas neutral compounds depicted a constant retention time at different pH. The general HILIC-MALDI-MS method is feasible for fast screening of naturally occurring organic compounds. A series of homologs can be determined if they have the same retention behavior. Their structural features can be elucidated by considering their mass differences and hydrophilic properties as determined by HILIC chromatogram.

  8. Cultivable Methylobacterium species diversity in rice seeds identified with whole-cell matrix-assisted laser desorption/ionization time-of-flight mass spectrometric analysis.

    Science.gov (United States)

    Okumura, Marie; Fujitani, Yoshiko; Maekawa, Masahiko; Charoenpanich, Jittima; Murage, Hunja; Kimbara, Kazuhide; Sahin, Nurettin; Tani, Akio

    2017-02-01

    Methylobacterium species are methylotrophic bacteria that widely inhabit plant surfaces. In addition to studies on methylotrophs as model organisms, research has also been conducted on their mechanism of plant growth promotion as well as the species-species specificity of plant-microbe interaction. We employed whole-cell matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (WC-MS) analysis, which enables the rapid and accurate identification of bacteria at the species level, to identify Methylobacterium isolates collected from the rice seeds of different cultivars harvested in Japan, Thailand, and Kenya. Rice seeds obtained from diverse geographical locations showed different communities of Methylobacterium species. We found that M. fujisawaense, M. aquaticum, M. platani, and M. radiotolerans are the most frequently isolated species, but none were isolated as common species from 18 seed samples due to the highly biased communities in some samples. These findings will contribute to the development of formulations containing selected species that promote rice growth, though it may be necessary to customize the formulations depending on the cultivars and farm conditions. Copyright © 2016 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  9. Matrix-assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) Can Precisely Discriminate the Lineages of Listeria monocytogenes and Species of Listeria.

    Science.gov (United States)

    Ojima-Kato, Teruyo; Yamamoto, Naomi; Takahashi, Hajime; Tamura, Hiroto

    2016-01-01

    The genetic lineages of Listeria monocytogenes and other species of the genus Listeria are correlated with pathogenesis in humans. Although matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has become a prevailing tool for rapid and reliable microbial identification, the precise discrimination of Listeria species and lineages remains a crucial issue in clinical settings and for food safety. In this study, we constructed an accurate and reliable MS database to discriminate the lineages of L. monocytogenes and the species of Listeria (L. monocytogenes, L. innocua, L. welshimeri, L. seeligeri, L. ivanovii, L. grayi, and L. rocourtiae) based on the S10-spc-alpha operon gene encoded ribosomal protein mass spectrum (S10-GERMS) proteotyping method, which relies on both genetic information (genomics) and observed MS peaks in MALDI-TOF MS (proteomics). The specific set of eight biomarkers (ribosomal proteins L24, L6, L18, L15, S11, S9, L31 type B, and S16) yielded characteristic MS patterns for the lineages of L. monocytogenes and the different species of Listeria, and led to the construction of a MS database that was successful in discriminating between these organisms in MALDI-TOF MS fingerprinting analysis followed by advanced proteotyping software Strain Solution analysis. We also confirmed the constructed database on the proteotyping software Strain Solution by using 23 Listeria strains collected from natural sources.

  10. Matrix-assisted laser desorption ionization-time of flight mass spectrometry: a fundamental shift in the routine practice of clinical microbiology.

    Science.gov (United States)

    Clark, Andrew E; Kaleta, Erin J; Arora, Amit; Wolk, Donna M

    2013-07-01

    Within the past decade, clinical microbiology laboratories experienced revolutionary changes in the way in which microorganisms are identified, moving away from slow, traditional microbial identification algorithms toward rapid molecular methods and mass spectrometry (MS). Historically, MS was clinically utilized as a high-complexity method adapted for protein-centered analysis of samples in chemistry and hematology laboratories. Today, matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) MS is adapted for use in microbiology laboratories, where it serves as a paradigm-shifting, rapid, and robust method for accurate microbial identification. Multiple instrument platforms, marketed by well-established manufacturers, are beginning to displace automated phenotypic identification instruments and in some cases genetic sequence-based identification practices. This review summarizes the current position of MALDI-TOF MS in clinical research and in diagnostic clinical microbiology laboratories and serves as a primer to examine the "nuts and bolts" of MALDI-TOF MS, highlighting research associated with sample preparation, spectral analysis, and accuracy. Currently available MALDI-TOF MS hardware and software platforms that support the use of MALDI-TOF with direct and precultured specimens and integration of the technology into the laboratory workflow are also discussed. Finally, this review closes with a prospective view of the future of MALDI-TOF MS in the clinical microbiology laboratory to accelerate diagnosis and microbial identification to improve patient care.

  11. Identification of Lactobacillus from the saliva of adult patients with caries using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Directory of Open Access Journals (Sweden)

    Yifei Zhang

    Full Text Available Matrix-assisted laser desorption/ionization (MALDI time-of-flight (TOF mass spectrometry (MS has been presented as a superior method for the detection of microorganisms in body fluid samples (e.g., blood, saliva, pus, etc. However, the performance of MALDI-TOF MS in routine identification of caries-related Lactobacillus isolates from saliva of adult patients with caries has not been determined. In the present study, we introduced a new MALDI-TOF MS system for identification of lactobacilli. Saliva samples were collected from 120 subjects with caries. Bacteria were isolated and cultured, and each isolate was identified by both 16S rRNA sequencing and MALDI-TOF MS. The identification results obtained by MALDI-TOF MS were concordant at the genus level with those of conventional 16S rRNA-based sequencing for 88.6% of lactobacilli (62/70 and 95.5% of non-lactobacilli (21/22. Up to 96 results could be obtained in parallel on a single MALDI target, suggesting that this is a reliable high-throughput approach for routine identification of lactobacilli. However, additional reference strains are necessary to increase the sensitivity and specificity of species-level identification.

  12. Discrimination of Streptococcus equi subsp. equi and Streptococcus equi subsp. zooepidemicus using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Mani, Rinosh J; Thachil, Anil J; Ramachandran, Akhilesh

    2017-09-01

    Accurate and timely identification of infectious etiologies is of great significance in veterinary microbiology, especially for critical diseases such as strangles, a highly contagious disease of horses caused by Streptococcus equi subsp. equi. We evaluated a matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) platform for use in species- and subspecies-level identification of S. equi isolates from horses and compared it with an automated biochemical system. We used 25 clinical isolates each of S. equi subsp. equi and S. equi subsp. zooepidemicus. Using the MALDI-TOF MS platform, it was possible to correctly identify all 50 isolates to the species level. Unique mass peaks were identified in the bacterial peptide mass spectra generated by MALDI-TOF MS, which can be used for accurate subspecies-level identification of S. equi. Mass peaks (mass/charge, m/ z) 6,751.9 ± 1.4 (mean ± standard deviation) and 5,958.1 ± 1.3 were found to be unique to S. equi subsp. equi and S. equi subsp. zooepidemicus, respectively. The automated biochemical system correctly identified 47 of 50 of the isolates to the species level as S. equi, whereas at the subspecies level, 24 of 25 S. equi subsp. equi isolates and 22 of 25 S. equi subsp. zooepidemicus isolates were correctly identified. Our results indicate that MALDI-TOF MS can be used for accurate species- and subspecies-level identification of S. equi.

  13. Improved detection of phosphopeptides by negative ion matrix-assisted laser desorption/ionization mass spectrometry using a proton sponge co-matrix

    Energy Technology Data Exchange (ETDEWEB)

    Zhang Shu [State Key Laboratory for Chirosciences and Department of Applied Biology and Chemical Technology, Hong Kong Polytechnic University, Hung Hom, Kowloon (Hong Kong); Yao Zhongping, E-mail: bczpyao@inet.polyu.edu.hk [State Key Laboratory for Chirosciences and Department of Applied Biology and Chemical Technology, Hong Kong Polytechnic University, Hung Hom, Kowloon (Hong Kong)

    2012-01-20

    Highlights: Black-Right-Pointing-Pointer Use of ATT/DMAN/DHC matrix for detection of phosphopeptides by negative MALDI-MS. Black-Right-Pointing-Pointer Lower limit of detection. Black-Right-Pointing-Pointer Reduced signal suppression effects. Black-Right-Pointing-Pointer Improved position-to-position reproducibility. - Abstract: Analysis of phosphopeptides is an important task in proteomic studies. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is a technique very commonly used for such a purpose. Analysis of phosphopeptides by MALDI-MS is, however, still a challenging task due to the low ionization efficiency of phosphopeptides. In this study, we reported that by using a proton sponge 1,8-bis(dimethyl-amino)naphthalene (DMAN) as a co-matrix, detection of phosphopeptides by negative ion MALDI-MS could be greatly improved. Combination of DMAN with another matrix 6-aza-2-thiothymine (ATT) and additive diammonium hydrogen citrate (DHC) allowed much lower limit of detection, significantly reduced signal suppression effects and improved position-to-position reproducibility for detection of phosphopeptides by negative ion MALDI-MS. Potential applications of the matrix system in qualification of phosphopeptides and analysis of proteolytic digests of phosphorylated proteins were also demonstrated in this study.

  14. Calibration laws based on multiple linear regression applied to matrix-assisted laser desorption/ionization Fourier transform ion cyclotron resonance mass spectrometry.

    Science.gov (United States)

    Williams, D Keith; Chadwick, M Ashley; Williams, Taufika Islam; Muddiman, David C

    2008-12-01

    Operation of any mass spectrometer requires implementation of mass calibration laws to translate experimentally measured physical quantities into a m/z range. While internal calibration in Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS) offers several attractive features, including exposure of calibrant and analyte ions to identical experimental conditions (e.g. space charge), external calibration affords simpler pulse sequences and higher throughput. The automatic gain control method used in hybrid linear trap quadrupole (LTQ) FT-ICR-MS to consistently obtain the same ion population is not readily amenable to matrix-assisted laser desorption/ionization (MALDI) FT-ICR-MS, due to the heterogeneous nature and poor spot-to-spot reproducibility of MALDI. This can be compensated for by taking external calibration laws into account that consider magnetic and electric fields, as well as relative and total ion abundances. Herein, an evaluation of external mass calibration laws applied to MALDI-FT-ICR-MS is performed to achieve higher mass measurement accuracy (MMA). Copyright (c) 2008 John Wiley & Sons, Ltd.

  15. Solvent selection for matrix-assisted laser desorption/ionization time-of-flight mass spectrometric analysis of synthetic polymers employing solubility parameters.

    Science.gov (United States)

    Brandt, Heike; Ehmann, Thomas; Otto, Matthias

    2010-08-30

    The principle relating to the selection of a proper matrix, cationization reagent, and solvent for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) of synthetic polymers is still a topic of research. In this work we focused on the selection of a suitable MALDI solvent. Polystyrene PS7600 and poly(ethylene glycol) PEG4820 were analyzed by MALDI-TOF MS using various solvents which were selected based on the Hansen solubility parameter system. For polystyrene (PS), dithranol was used as the matrix and silver trifluoroacetate as the cationization reagent whereas, for poly(ethylene glycol) (PEG), the combination of 2,5-dihydroxybenzoic acid and sodium trifluoroacetate was used for all experiments. When employing solvents which dissolve PS and PEG, reliable MALDI mass spectra were obtained while samples in non-solvents (solvents which are not able to dissolve the polymer) failed to provide spectra. It seems that the solubility of the matrix and the cationization reagent are less important than the polymer solubility. 2010 John Wiley & Sons, Ltd.

  16. Selective identification by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry of different types of gluten in foods made with cereal mixtures.

    Science.gov (United States)

    Camafeita, E; Solís, J; Alfonso, P; López, J A; Sorell, L; Méndez, E

    1998-10-09

    The gluten toxic fractions responsible for the mucosal damage in coeliac disease (CD), so-called gliadins, hordeins, secalins and avenins from a large number (30-40) of wheat, barley, rye and oats cultivars respectively, have been mass analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Gliadin, secalin and avenin characteristic mass profiles are nearly identical amongst distinct cultivars from the corresponding cereal, while hordeins profiles show more variability depending on the particular barley cultivar. On the basis of these four distinguishable characteristic mass patterns spreading within the 20,000-40,000 Da range, MALDI-TOF-MS has permitted the direct and simultaneous visualization of gliadins, hordeins, secalins and avenins in foods elaborated with cereal mixtures of wheat, barley, rye and oats. This capacity has been demonstrated by mass analyzing foods made with these four cereals in varying ratios. Thus MALDI-TOF-MS can be preliminarily established as a unique system with the ability to discriminate the specific type of gluten toxic fractions present in food samples.

  17. Differentiation of Lactobacillus brevis strains using Matrix-Assisted-Laser-Desorption-Ionization-Time-of-Flight Mass Spectrometry with respect to their beer spoilage potential.

    Science.gov (United States)

    Kern, Carola C; Vogel, Rudi F; Behr, Jürgen

    2014-06-01

    Lactobacillus (L.) brevis is one of the most frequently encountered bacteria in beer-spoilage incidents. As the species Lactobacillus brevis comprises strains showing varying ability to grow in beer, ranging from growth in low hopped wheat to highly hopped pilsner beer, differentiation and classification of L. brevis with regard to their beer-spoiling ability is of vital interest for the brewing industry. Matrix-Assisted-Laser-Desorption-Ionization-Time-Of-Flight Mass Spectrometry (MALDI-TOF MS) has been shown as a powerful tool for species and sub-species differentiation of bacterial isolates and is increasingly used for strain-level differentiation. Seventeen L. brevis strains, representative of different spoilage types, were characterized according to their tolerance to iso-alpha-acids and their growth in wheat-, lager- and pilsner beer. MALDI-TOF MS spectra were acquired to perform strain-level identification, cluster analysis and biomarker detection. Strain-level identification was achieved in 90% out of 204 spectra. Misidentification occurred nearly exclusively among strains belonging to the same spoilage type. Though spectra of strongly beer-spoiling strains showed remarkable similarity, no decisive single markers were detected to be present in all strains of one group. However, MALDI-TOF MS spectra can be reliably assigned to the corresponding strain and thus allow to track single strains and connect them to their physiological properties. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. Sample-first preparation: a method for surface-assisted laser desorption/ionization time-of-flight mass spectrometry analysis of cyclic oligosaccharides.

    Science.gov (United States)

    Wu, Hsin-Pin; Su, Chih-Lin; Chang, Hui-Chiu; Tseng, Wei-Lung

    2007-08-15

    A new sample preparation method for the analysis of cyclic oligosaccharides in surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS) is presented. We call this new technique "sample first method", in which a sample is deposited first and then bare gold nanoparticles (AuNPs), which serve as the SALDI matrixes, are added to the top of the sample layer. The use of the sample first method offers significant advantages for improving shot-to-shot reproducibility, enhancing the ionization efficiency of the analyte, and reducing sample preparation time as compared to the dried-droplet method, wherein samples and bare AuNPs are mixed and dried together. The relative standard deviation (RSD) values of the signal intensity as calculated from 65 sample spots was 25% when the sample first methods were applied to the analysis of beta-cyclodextrin. The results were more homogeneous as compared to the outcome using dried-droplet preparation of AuNPs (RSD=66%) and 2,5-dihydroxybenzoic acid (RSD=209%). We also found out that the optimal concentration of AuNP for ionization efficiency is 7.4 nM (4.52x10(12) particles/mL) while the lowest detectable concentration of cyclic oligosaccharides through this approach is 0.25 microM. Except for the cyclic oligosaccharide, the proposed method was also applied to the analyses of other biological samples, including neutral carbohydrate and steroid, aminothiols, and peptides as well as proteins.

  19. Application of nonpolar matrices for the analysis of low molecular weight nonpolar synthetic polymers by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

    Science.gov (United States)

    Macha; Limbach; Savickas

    2000-08-01

    The application of nonpolar matrices for the analysis of low molecular weight nonpolar synthetic polymers using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is demonstrated. Anthracene, pyrene, and acenaphthene were utilized as nonpolar matrices for the analysis of polybutadiene, polyisoprene, and polystyrene samples of various average molecular weights ranging from about 700 to 5,000. The standard MALDI-MS approach for the analysis of these types of polymers involves the use of conventional acidic matrices, such as all-trans-retinoic acid, with an additional cationization reagent. The nonpolar matrices used in this study are shown to be as equally effective as the conventional matrices. The uniform mixing of the nonpolar matrices and the nonpolar analytes enhances the MALDI-MS spectral reproducibility. Silver salts were found to be the best cationization reagents for all of the cases studied. Copper salts worked well for polystyrene, poorly for polyisoprene, and not at all for polybutadiene samples. These matrices should be useful for the characterization of hydrocarbon polymers and other analytes, such as modified polymers, which may potentially be sensitive to acidic matrices.

  20. Quantification of Saccharides in Honey Samples Through Surface-Assisted Laser Desorption/Ionization Mass Spectrometry Using HgTe Nanostructures

    Science.gov (United States)

    Wang, Chia-Wei; Chen, Wen-Tsen; Chang, Huan-Tsung

    2014-07-01

    Quantification of monosaccharides and disaccharides in five honey samples through surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS) using HgTe nanostructures as the matrix and sucralose as an internal standard has been demonstrated. Under optimal conditions (1× HgTe nanostructure, 0.2 mM ammonium citrate at pH 9.0), the SALDI-MS approach allows detection of fructose and maltose at the concentrations down to 15 and 10 μM, respectively. Without conducting tedious sample pretreatment and separation, the SALDI-MS approach allows determination of the contents of monosaccharides and disaccharides in honey samples within 30 min, with reproducibility (relative standard deviation <15%). Unlike only sodium adducts of standard saccharides detected, sodium adducts and potassium adducts with differential amounts have been found among various samples, showing different amounts of sodium and potassium ions in the honey samples. The SALDI-MS data reveal that the contents of monosaccharides and disaccharides in various honey samples are dependent on their nectar sources. In addition to the abundant amounts of monosaccharides and disaccharides, oligosaccharides in m/z range of 650 - 2700 are only detected in pomelo honey. Having advantages of simplicity, rapidity, and reproducibility, this SALDI-MS holds great potential for the analysis of honey samples.

  1. Novel approach for differentiating Shigella species and Escherichia coli by matrix-assisted laser desorption ionization-time of flight mass spectrometry.

    Science.gov (United States)

    Khot, Prasanna D; Fisher, Mark A

    2013-11-01

    Shigella species are so closely related to Escherichia coli that routine matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) cannot reliably differentiate them. Biochemical and serological methods are typically used to distinguish these species; however, "inactive" isolates of E. coli are biochemically very similar to Shigella species and thus pose a greater diagnostic challenge. We used ClinProTools (Bruker Daltonics) software to discover MALDI-TOF MS biomarker peaks and to generate classification models based on the genetic algorithm to differentiate between Shigella species and E. coli. Sixty-six Shigella spp. and 72 E. coli isolates were used to generate and test classification models, and the optimal models contained 15 biomarker peaks for genus-level classification and 12 peaks for species-level classification. We were able to identify 90% of E. coli and Shigella clinical isolates correctly to the species level. Only 3% of tested isolates were misidentified. This novel MALDI-TOF MS approach allows laboratories to streamline the identification of E. coli and Shigella species.

  2. Identification of different respiratory viruses, after a cell culture step, by matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Calderaro, Adriana; Arcangeletti, Maria Cristina; Rodighiero, Isabella; Buttrini, Mirko; Montecchini, Sara; Vasile Simone, Rosita; Medici, Maria Cristina; Chezzi, Carlo; De Conto, Flora

    2016-10-27

    In this study matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS), a reliable identification method for the diagnosis of bacterial and fungal infections, is presented as an innovative tool to investigate the protein profile of cell cultures infected by the most common viruses causing respiratory tract infections in humans. MALDI-TOF MS was applied to the identification of influenza A and B viruses, adenovirus C species, parainfluenza virus types 1, 2 and 3, respiratory syncytial virus, echovirus, cytomegalovirus and metapneumovirus. In this study MALDI-TOF MS was proposed as a model to be applied to the identification of cultivable respiratory viruses using cell culture as a viral proteins enrichment method to the proteome profiling of virus infected and uninfected cell cultures. The reference virus strains and 58 viruses identified from respiratory samples of subjects with respiratory diseases positive for one of the above mentioned viral agents by cell culture were used for the in vitro infection of suitable cell cultures. The isolated viral particles, concentrated by ultracentrifugation, were used for subsequent protein extraction and their spectra profiles were generated by MALDI-TOF MS analysis. The newly created library allowed us to discriminate between uninfected and respiratory virus infected cell cultures.

  3. Identification of different respiratory viruses, after a cell culture step, by matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS)

    Science.gov (United States)

    Calderaro, Adriana; Arcangeletti, Maria Cristina; Rodighiero, Isabella; Buttrini, Mirko; Montecchini, Sara; Vasile Simone, Rosita; Medici, Maria Cristina; Chezzi, Carlo; De Conto, Flora

    2016-01-01

    In this study matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS), a reliable identification method for the diagnosis of bacterial and fungal infections, is presented as an innovative tool to investigate the protein profile of cell cultures infected by the most common viruses causing respiratory tract infections in humans. MALDI-TOF MS was applied to the identification of influenza A and B viruses, adenovirus C species, parainfluenza virus types 1, 2 and 3, respiratory syncytial virus, echovirus, cytomegalovirus and metapneumovirus. In this study MALDI-TOF MS was proposed as a model to be applied to the identification of cultivable respiratory viruses using cell culture as a viral proteins enrichment method to the proteome profiling of virus infected and uninfected cell cultures. The reference virus strains and 58 viruses identified from respiratory samples of subjects with respiratory diseases positive for one of the above mentioned viral agents by cell culture were used for the in vitro infection of suitable cell cultures. The isolated viral particles, concentrated by ultracentrifugation, were used for subsequent protein extraction and their spectra profiles were generated by MALDI-TOF MS analysis. The newly created library allowed us to discriminate between uninfected and respiratory virus infected cell cultures. PMID:27786297

  4. Identification of medically relevant species of arthroconidial yeasts by use of matrix-assisted laser desorption ionization-time of flight mass spectrometry.

    Science.gov (United States)

    Kolecka, Anna; Khayhan, Kantarawee; Groenewald, Marizeth; Theelen, Bart; Arabatzis, Michael; Velegraki, Aristea; Kostrzewa, Markus; Mares, Mihai; Taj-Aldeen, Saad J; Boekhout, Teun

    2013-08-01

    Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) was used for an extensive identification study of arthroconidial yeasts, using 85 reference strains from the CBS-KNAW yeast collection and 134 clinical isolates collected from medical centers in Qatar, Greece, and Romania. The test set included 72 strains of ascomycetous yeasts (Galactomyces, Geotrichum, Saprochaete, and Magnusiomyces spp.) and 147 strains of basidiomycetous yeasts (Trichosporon and Guehomyces spp.). With minimal preparation time, MALDI-TOF MS proved to be an excellent diagnostic tool that provided reliable identification of most (98%) of the tested strains to the species level, with good discriminatory power. The majority of strains were correctly identified at the species level with good scores (>2.0) and seven of the tested strains with log score values between 1.7 and 2.0. The MALDI-TOF MS results obtained were consistent with validated internal transcribed spacer (ITS) and/or large subunit (LSU) ribosomal DNA sequencing results. Expanding the mass spectrum database by increasing the number of reference strains for closely related species, including those of nonclinical origin, should enhance the usefulness of MALDI-TOF MS-based diagnostic analysis of these arthroconidial fungi in medical and other laboratories.

  5. Identification of Medically Relevant Species of Arthroconidial Yeasts by Use of Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

    Science.gov (United States)

    Kolecka, Anna; Khayhan, Kantarawee; Groenewald, Marizeth; Theelen, Bart; Arabatzis, Michael; Velegraki, Aristea; Kostrzewa, Markus; Mares, Mihai; Taj-Aldeen, Saad J.

    2013-01-01

    Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) was used for an extensive identification study of arthroconidial yeasts, using 85 reference strains from the CBS-KNAW yeast collection and 134 clinical isolates collected from medical centers in Qatar, Greece, and Romania. The test set included 72 strains of ascomycetous yeasts (Galactomyces, Geotrichum, Saprochaete, and Magnusiomyces spp.) and 147 strains of basidiomycetous yeasts (Trichosporon and Guehomyces spp.). With minimal preparation time, MALDI-TOF MS proved to be an excellent diagnostic tool that provided reliable identification of most (98%) of the tested strains to the species level, with good discriminatory power. The majority of strains were correctly identified at the species level with good scores (>2.0) and seven of the tested strains with log score values between 1.7 and 2.0. The MALDI-TOF MS results obtained were consistent with validated internal transcribed spacer (ITS) and/or large subunit (LSU) ribosomal DNA sequencing results. Expanding the mass spectrum database by increasing the number of reference strains for closely related species, including those of nonclinical origin, should enhance the usefulness of MALDI-TOF MS-based diagnostic analysis of these arthroconidial fungi in medical and other laboratories. PMID:23678074

  6. A novel cluster of Mycobacterium abscessus complex revealed by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Suzuki, Hiromichi; Yoshida, Shiomi; Yoshida, Atsushi; Okuzumi, Katsuko; Fukusima, Atsuhito; Hishinuma, Akira

    2015-12-01

    Mycobacterium abscessus complex is a rapidly growing mycobacterium consisting of 3 subspecies, M. abscessus, Mycobacterium massiliense, and Mycobacterium bolletii. However, rapid and accurate species identification is difficult. We first evaluated a suitable protocol of matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) for distinguishing these subspecies. Then, we studied spectral signals by MALDI-TOF MS in 59 M. abscessus, 42 M. massiliense, and 2 M. bolletii. Among several specific spectral signals, 4 signals clearly differentiate M. massiliense from the other 2 subspecies, M. abscessus and M. bolletii. Moreover, 6 of the 42 M. massiliense isolates showed a spectral pattern similar to M. abscessus. These isolates correspond to the distinctive class of M. massiliense (cluster D) which is closer to M. abscessus by the previous variable number tandem repeat analysis. These results indicate that MALDI-TOF MS is not only useful for the identification of 3 subspecies of M. abscessus complex but also capable of distinguishing clusters of M. massiliense. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. Matrix-assisted laser desorption ionization mass spectrometry: a new tool for probing interactions between proteins and metal surfaces. Use in dental implantology.

    Science.gov (United States)

    Leize, E M; Leize, E J; Leize, M C; Voegel, J C; Van Dorsselaer, A

    1999-07-15

    The fixation in the bone of an artificial titanium tooth root is believed to be initiated by the rapid adsorption of the proteins present in the surgical cavity on the titanium surface. The study of this adsorption should make it possible to predict the osseointegration capacities of new implant surface treatments. We describe here a new method, based on matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS), for quantifying proteins adsorbed on titanium surfaces fully identical to these designed for implantology. The key step of this method is a new MALDI-MS sample preparation allowing the adsorbed proteins to be removed from the surface and to be homogeneously dispersed in the matrix crystals. The adsorption of a model protein (lysozyme) on two titanium surfaces (polished and sandblasted) was studied in order to evaluate the method. The absolute MALDI-MS intensity was shown to vary linearly with the amount of adsorbed lysozyme. After dipping the titanium surfaces for different times in lysozyme solutions at different concentrations, the maximum amount of adsorbed lysozyme was measured by MALDI-MS and was shown to correspond to a lysozyme monolayer, which is consistent with results described in the literature. Copyright 1999 Academic Press.

  8. Effects of solid-medium type on routine identification of bacterial isolates by use of matrix-assisted laser desorption ionization-time of flight mass spectrometry.

    Science.gov (United States)

    Anderson, Neil W; Buchan, Blake W; Riebe, Katherine M; Parsons, Lauren N; Gnacinski, Stacy; Ledeboer, Nathan A

    2012-03-01

    Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is a rapid method for the identification of bacteria. Factors that may alter protein profiles, including growth conditions and presence of exogenous substances, could hinder identification. Bacterial isolates identified by conventional methods were grown on various media and identified using the MALDI Biotyper (Bruker Daltonics, Billerica, MA) using a direct smear method and an acid extraction method. Specimens included 23 Pseudomonas isolates grown on blood agar, Pseudocel (CET), and MacConkey agar (MAC); 20 Staphylococcus isolates grown on blood agar, colistin-nalidixic acid agar (CNA), and mannitol salt agar (MSA); and 25 enteric isolates grown on blood agar, xylose lysine deoxycholate agar (XLD), Hektoen enteric agar (HE), salmonella-shigella agar (SS), and MAC. For Pseudomonas spp., the identification rate to genus using the direct method was 83% from blood, 78% from MAC, and 94% from CET. For Staphylococcus isolates, the identification rate to genus using the direct method was 95% from blood, 75% from CNA, and 95% from MSA. For enteric isolates, the identification rate to genus using the direct method was 100% from blood, 100% from MAC, 100% from XLD, 92% from HE, and 87% from SS. Extraction enhanced identification rates. The direct method of MALDI-TOF analysis of bacteria from selective and differential media yields identifications of varied confidence. Notably, Staphylococci spp. from CNA exhibit low identification rates. Extraction enhances identification rates and is recommended for colonies from this medium.

  9. Rapid Characterization of Microalgae and Microalgae Mixtures Using Matrix-Assisted Laser Desorption Ionization Time-Of-Flight Mass Spectrometry (MALDI-TOF MS)

    Science.gov (United States)

    Barbano, Duane; Diaz, Regina; Zhang, Lin; Sandrin, Todd; Gerken, Henri; Dempster, Thomas

    2015-01-01

    Current molecular methods to characterize microalgae are time-intensive and expensive. Matrix Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) may represent a rapid and economical alternative approach. The objectives of this study were to determine whether MALDI-TOF MS can be used to: 1) differentiate microalgae at the species and strain levels and 2) characterize simple microalgal mixtures. A common protein extraction sample preparation method was used to facilitate rapid mass spectrometry-based analysis of 31 microalgae. Each yielded spectra containing between 6 and 56 peaks in the m/z 2,000 to 20,000 range. The taxonomic resolution of this approach appeared higher than that of 18S rDNA sequence analysis. For example, two strains of Scenedesmus acutus differed only by two 18S rDNA nucleotides, but yielded distinct MALDI-TOF mass spectra. Mixtures of two and three microalgae yielded relatively complex spectra that contained peaks associated with members of each mixture. Interestingly, though, mixture-specific peaks were observed at m/z 11,048 and 11,230. Our results suggest that MALDI-TOF MS affords rapid characterization of individual microalgae and simple microalgal mixtures. PMID:26271045

  10. Sample deposition device for off-line combination of supercritical fluid chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Planeta, Josef; Rehulka, Pavel; Chmelík, Josef

    2002-08-01

    A new sample deposition device for off-line SFC-MALDI combination of supercritical fluid chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry was assembled. This device was successfully applied to the detailed characterization of synthetic silicone oils. SFC was used to separate samples of silicone oils on micropacked capillary columns and to determine their molecular mass distribution. The separated fractions for the identification studies were obtained from SFC runs at defined time intervals. Using the constructed deposition device, these fractions were sprayed directly from the restrictor on the target probe covered with a proper matrix. MALDI-TOF MS was used for the identification of individual oligomers in the separated fractions and also in the unfractionated sample. The determined molecular mass distributions based on supercritical fluid chromatography with flame ionization detector, MALDI-TOF MS, and combined SFC-MALDI measurements were compared and the results were in a good agreement. The sample deposition device is based on a common plotter unit, complemented by a microcontroller PIC16C84. The unit is connected by an RS-232 interface to a PC with the main control software running under MS Windows. The new sample deposition device made the off-line combination SFC-MALDI simpler, faster, and more sensitive.

  11. Evaluation of Matrix-Assisted Laser Desorption Ionization−Time of Flight Mass Spectrometry for Identification of Mycobacterium species, Nocardia species, and Other Aerobic Actinomycetes

    Science.gov (United States)

    Buckwalter, S. P.; Olson, S. L.; Connelly, B. J.; Lucas, B. C.; Rodning, A. A.; Walchak, R. C.; Deml, S. M.; Wohlfiel, S. L.

    2015-01-01

    The value of matrix-assisted laser desorption ionization−time of flight mass spectrometry (MALDI-TOF MS) for the identification of bacteria and yeasts is well documented in the literature. Its utility for the identification of mycobacteria and Nocardia spp. has also been reported in a limited scope. In this work, we report the specificity of MALDI-TOF MS for the identification of 162 Mycobacterium species and subspecies, 53 Nocardia species, and 13 genera (totaling 43 species) of other aerobic actinomycetes using both the MALDI-TOF MS manufacturer's supplied database(s) and a custom database generated in our laboratory. The performance of a simplified processing and extraction procedure was also evaluated, and, similar to the results in an earlier literature report, our viability studies confirmed the ability of this process to inactivate Mycobacterium tuberculosis prior to analysis. Following library construction and the specificity study, the performance of MALDI-TOF MS was directly compared with that of 16S rRNA gene sequencing for the evaluation of 297 mycobacteria isolates, 148 Nocardia species isolates, and 61 other aerobic actinomycetes isolates under routine clinical laboratory working conditions over a 6-month period. MALDI-TOF MS is a valuable tool for the identification of these groups of organisms. Limitations in the databases and in the ability of MALDI-TOF MS to rapidly identify slowly growing mycobacteria are discussed. PMID:26637381

  12. Evaluation of Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry for Identification of Mycobacterium species, Nocardia species, and Other Aerobic Actinomycetes.

    Science.gov (United States)

    Buckwalter, S P; Olson, S L; Connelly, B J; Lucas, B C; Rodning, A A; Walchak, R C; Deml, S M; Wohlfiel, S L; Wengenack, N L

    2016-02-01

    The value of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for the identification of bacteria and yeasts is well documented in the literature. Its utility for the identification of mycobacteria and Nocardia spp. has also been reported in a limited scope. In this work, we report the specificity of MALDI-TOF MS for the identification of 162 Mycobacterium species and subspecies, 53 Nocardia species, and 13 genera (totaling 43 species) of other aerobic actinomycetes using both the MALDI-TOF MS manufacturer's supplied database(s) and a custom database generated in our laboratory. The performance of a simplified processing and extraction procedure was also evaluated, and, similar to the results in an earlier literature report, our viability studies confirmed the ability of this process to inactivate Mycobacterium tuberculosis prior to analysis. Following library construction and the specificity study, the performance of MALDI-TOF MS was directly compared with that of 16S rRNA gene sequencing for the evaluation of 297 mycobacteria isolates, 148 Nocardia species isolates, and 61 other aerobic actinomycetes isolates under routine clinical laboratory working conditions over a 6-month period. MALDI-TOF MS is a valuable tool for the identification of these groups of organisms. Limitations in the databases and in the ability of MALDI-TOF MS to rapidly identify slowly growing mycobacteria are discussed. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  13. Multicenter Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry Study for Identification of Clinically Relevant Nocardia spp.

    Science.gov (United States)

    Blosser, Sara J; Drake, Steven K; Andrasko, Jennifer L; Henderson, Christina M; Kamboj, Kamal; Antonara, Stella; Mijares, Lilia; Conville, Patricia; Frank, Karen M; Harrington, Susan M; Balada-Llasat, Joan-Miquel; Zelazny, Adrian M

    2016-05-01

    This multicenter study analyzed Nocardia spp., including extraction, spectral acquisition, Bruker matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) identification, and score interpretation, using three Nocardia libraries, the Bruker, National Institutes of Health (NIH), and The Ohio State University (OSU) libraries, and compared the results obtained by each center. A standardized study protocol, 150 Nocardia isolates, and NIH and OSU Nocardia MALDI-TOF MS libraries were distributed to three centers. Following standardized culture, extraction, and MALDI-TOF MS analysis, isolates were identified using score cutoffs of ≥2.0 for species/species complex-level identification and ≥1.8 for genus-level identification. Isolates yielding a score of Nocardia MALDI-TOF MS library with both the OSU and NIH libraries increased the genus-level and species-level identification by 18.2% and 36.9%, respectively. Overall, this study demonstrates the ability of diverse clinical microbiology laboratories to utilize MALDI-TOF MS for the rapid identification of clinically relevant Nocardia spp. and to implement MALDI-TOF MS libraries developed by single laboratories across institutions. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  14. Multicenter Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry Study for Identification of Clinically Relevant Nocardia spp.

    Science.gov (United States)

    Blosser, Sara J.; Drake, Steven K.; Andrasko, Jennifer L.; Henderson, Christina M.; Kamboj, Kamal; Antonara, Stella; Mijares, Lilia; Conville, Patricia; Frank, Karen M.; Harrington, Susan M.; Balada-Llasat, Joan-Miquel

    2016-01-01

    This multicenter study analyzed Nocardia spp., including extraction, spectral acquisition, Bruker matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) identification, and score interpretation, using three Nocardia libraries, the Bruker, National Institutes of Health (NIH), and The Ohio State University (OSU) libraries, and compared the results obtained by each center. A standardized study protocol, 150 Nocardia isolates, and NIH and OSU Nocardia MALDI-TOF MS libraries were distributed to three centers. Following standardized culture, extraction, and MALDI-TOF MS analysis, isolates were identified using score cutoffs of ≥2.0 for species/species complex-level identification and ≥1.8 for genus-level identification. Isolates yielding a score of Nocardia MALDI-TOF MS library with both the OSU and NIH libraries increased the genus-level and species-level identification by 18.2% and 36.9%, respectively. Overall, this study demonstrates the ability of diverse clinical microbiology laboratories to utilize MALDI-TOF MS for the rapid identification of clinically relevant Nocardia spp. and to implement MALDI-TOF MS libraries developed by single laboratories across institutions. PMID:26912758

  15. Application of porous metal enrichment probe sampling to single cell analysis using matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS).

    Science.gov (United States)

    Fu, Qiang; Tang, Jun; Cui, Meng; Xing, Junpeng; Liu, Zhiqiang; Liu, Shuying

    2016-01-01

    There is an increasing need for analyzing metabolism in a single cell, which is important to understand the nature of cellular heterogeneity, disease, growth and specialization, etc. However, single cell analysis is often challenging for the traces of samples. In the present study, porous metal enrichment probe sampling combined with matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS) has been applied for in situ analysis of live onion epidemic cell. Porous probe, treated by corroding copper wire with HCl, was directly inserted into a single cell to get cell solution. A self-made linear actuator was enough to control the penetration of probe into the target cell accurately. Then samples on the tip of probe were eluted and detected by a commercial MALDI-TOF-MS directly. The formation of porous microstructure on the probe surface increased the adsorptive capacity of cell solution. The sensitivity of porous probe sampling was 6 times higher than uncorroded probes generally. This method provides a sensitive and convenient way for the sampling and detection of single cell solution. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

  16. Differentiation of clinically relevant Mucorales Rhizopus microsporus and R. arrhizus by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Dolatabadi, Somayeh; Kolecka, Anna; Versteeg, Matthijs; de Hoog, Sybren G; Boekhout, Teun

    2015-07-01

    This study addresses the usefulness of matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) MS for reliable identification of the two most frequently occurring clinical species of Rhizopus, namely Rhizopus arrhizus with its two varieties, arrhizus and delemar, and Rhizopus microsporus. The test-set comprised 38 isolates of clinical and environmental origin previously identified by internal transcribed spacer (ITS) sequencing of rDNA. Multi-locus sequence data targeting three gene markers (ITS, ACT, TEF ) showed two monophylic clades for Rhizopus arrhizus and Rhizopus microsporus (bootstrap values of 99 %). Cluster analysis confirmed the presence of two distinct clades within Rhizopus arrhizus representing its varieties arrhizus and delemar. The MALDI Biotyper 3.0 Microflex LT platform (Bruker Daltonics) was used to confirm the distinction between Rhizopus arrhizus and Rhizopus microsporus and the presence of two varieties within the species Rhizopus arrhizus. An in-house database of 30 reference main spectra (MSPs) was initially tested for correctness using commercially available databases of Bruker Daltonics. By challenging the database with the same strains of which an in-house database was created, automatic identification runs confirmed that MALDI-TOF MS is able to recognize the strains at the variety level. Based on principal component analysis, two MSP dendrograms were created and showed concordance with the multi-locus tree; thus, MALDI-TOF MS is a useful tool for diagnostics of mucoralean species.

  17. Rapid identification of Mycobacterium avium clinical isolates by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Lin, Chuan-Sheng; Su, Chih-Cheng; Hsieh, Shang-Chen; Lu, Chia-Chen; Wu, Tsu-Lan; Jia, Ju-Hsin; Wu, Ting-Shu; Han, Chau-Chung; Tsai, Wen-Cherng; Lu, Jang-Jih; Lai, Hsin-Chih

    2015-04-01

    Rapid and accurate discrimination of Mycobacterium avium from other mycobacteria is essential for appropriate therapeutic management and timely intervention for infection control. However, routine clinical identification methods for M. avium are both time consuming and labor intensive. In the present study, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used to identify specific cellular protein pattern for rapid identification of M. avium isolates. A total of 40 clinically relevant Mycobacterium strains comprising 13 distinct species were enrolled for the MALDI-TOF MS identification. A 10-minute extraction-free examination procedure was set up to obtain mass spectral fingerprints from whole bacterial cells. The characteristic mass spectral peak patterns in the m/z (mass/charge ratio) range of 5-20 kDa can be obtained within 10 minutes. The species-specific mass spectra for M. avium is identified and can be differentiated from as Mycobacterium strains. This technique shortens and simplifies the identification procedure of MALDI-TOF MS and may further extend the mycobacterial MALDI-TOF MS database. Simplicity and rapidity of identification procedures make MALDI-TOF MS an attractive platform in routine identification of mycobacteria. MALDI-TOF MS is applicable for rapid discrimination of M. avium from other Mycobacterium species, and shows its potential for clinical application. Copyright © 2013. Published by Elsevier B.V.

  18. Rapid identification of clinical mycobacterial isolates by protein profiling using matrix assisted laser desorption ionization-time of flight mass spectrometry.

    Science.gov (United States)

    Panda, A; Kurapati, S; Samantaray, J C; Myneedu, V P; Verma, A; Srinivasan, A; Ahmad, H; Behera, D; Singh, U B

    2013-01-01

    The purpose of this study was to evaluate the identification of Mycobacterium tuberculosis which is often plagued with ambiguity. It is a time consuming process requiring 4-8 weeks after culture positivity, thereby delaying therapeutic intervention. For a successful treatment and disease management, timely diagnosis is imperative. We evaluated a rapid, proteomic based technique for identification of clinical mycobacterial isolates by protein profiling using matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Freshly grown mycobacterial isolates were used. Acetonitrile/trifluoroacetic acid extraction procedure was carried out, following which cinnamic acid charged plates were subjected to identification by MALDI-TOF MS. A comparative analysis of 42 clinical mycobacterial isolates using the MALDI-TOF MS and conventional techniques was carried out. Among these, 97.61% were found to corroborate with the standard methods at genus level and 85.36% were accurate till the species level. One out of 42 was not in accord with the conventional assays because MALDI-TOF MS established it as Mycobacterium tuberculosis (log (score)>2.0) and conventional methods established it to be non-tuberculous Mycobacterium. MALDI-TOF MS was found to be an accurate, rapid, cost effective and robust system for identification of mycobacterial species. This innovative approach holds promise for early therapeutic intervention leading to better patient care.

  19. Application of delayed extraction matrix-assisted laser desorption ionization time-of-flight mass spectrometry for analysis of sphingolipids in cultured skin fibroblasts from sphingolipidosis patients.

    Science.gov (United States)

    Fujiwaki, Takehisa; Yamaguchi, Seiji; Sukegawa, Kazuko; Taketomi, Tamotsu

    2002-04-01

    Sphingolipidoses are caused by defects of enzymes involved in the hydrolysis of sphingolipids. Using delayed extraction matrix-assisted laser desorption ionization time-of-flight mass spectrometry (DE MALDI-TOF-MS), we analyzed sphingolipids in cultured skin fibroblasts from patients with sphingolipidoses, including: (a) Farber disease (FD, acid ceramidase deficiency); (b) Gaucher disease (GD); (c) Niemann-Pick disease type C (NPDC); and (d) GM1-gangliosidosis (GM1G). Crude lipids were extracted from about 50 mg wet weight of cultured skin fibroblasts. After mild alkaline treatment, a sphingolipid fraction was prepared from the crude lipids and analyzed by DE MALDI-TOF-MS. The results were as follows: (a) in fibroblasts from the FD patient, the ceramide/sphingomyelin and ceramide/monohexosylceramide ratios were both significantly high; (b) in the GD patient, the glucosylceramide/sphingomyelin ratio was increased; on the other hand; (c) in the NPDC patient, the monohexosylceramide/sphingomyelin ratio was within normal range; and (d) in the GM1G patient, no specific data were obtained. Sphingolipids in cultured fibroblasts can be evaluated by DE MALDI-TOF-MS, whereas GM1-ganglioside or its asialo derivatives are not detectable. With this DE MALDI-TOF-MS method, ceramide or monohexosylceramide accumulating in cultured fibroblasts from cases of sphingolipidoses, such as FD and GD, respectively, can be easily detected.

  20. Rapid detection of GM1 ganglioside in cerebrospinal fluid in dogs with GM1 gangliosidosis using matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Satoh, Hiroyuki; Yamauchi, Toyofumi; Yamasaki, Masahiro; Maede, Yoshimitsu; Yabuki, Akira; Chang, Hye-Sook; Asanuma, Taketoshi; Yamato, Osamu

    2011-11-01

    The concentration of GM1 (monosialotetrahexosyl ganglioside) in cerebrospinal fluid (CSF) is markedly increased in dogs with GM1 gangliosidosis due to GM1 accumulation in the central nervous system and leakage to the CSF. The present study established a rapid and simple method for detection of accumulated GM1 in the CSF in dogs with GM1 gangliosidosis using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI TOF MS) and discusses the usefulness of this method for the rapid diagnosis and/or high-risk screening of this disease in domestic animals. Cerebrospinal fluid was collected from normal dogs and 4- to 11-month-old Shiba dogs with GM1 gangliosidosis. The MALDI TOF MS analysis was carried out in combination with a special sample plate and a simple desalting step on the plate. Specific signs of GM1 could be detected in the standard GM1 solutions at concentrations of 50 nmol/l or more. The signs were also clearly detected in CSF (131-618 nmol/l) in affected dogs, but not in normal canine CSF (12 ± 5 nmol/l, mean ± standard deviation). The results demonstrated that MALDI TOF MS can detect GM1 accumulated in canine CSF even in the early stage of the disease. In conclusion, the rapid detection of increased CSF GM1 using MALDI TOF MS is a useful method for diagnosis and/or screening for canine GM1 gangliosidosis.

  1. Analysis of brain lipids by directly coupled matrix-assisted laser desorption ionization time-of-flight mass spectrometry and high-performance thin-layer chromatography.

    Science.gov (United States)

    Fuchs, Beate; Nimptsch, Ariane; Suss, Rosmarie; Schiller, Jürgen

    2008-01-01

    Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) is a soft ionization MS technique providing only minor fragmentation of the analyte. Therefore, the method is basically suitable for mixture analysis, although the ion yields strongly depend on the basicity/acidity of the analyte in relation to the applied matrix. Accordingly, less sensitively detectable compounds may be suppressed by more sensitively detectable compounds. Thus, separation of the mixture into the individual compounds is normally indispensable. This paper demonstrates the capabilities and limitations of a direct, simple, and inexpensive MALDI-high-performance thin-layer chromatography (HPTLC) coupling for the analysis of a crude lipid extract from porcine brain. Brain lipids were chosen because they represent a rather complex mixture and are of currently significant research interest. It was found that normal-phase HPTLC-separated lipids can be easily characterized by direct MALDI-TOF-MS analysis with sufficient resolution to allow the assignment of virtually all lipid classes, even rather minor species such as phosphorylated phosphoinositides or complex glycolipids as gangliosides. Advantages and disadvantages of this approach are discussed.

  2. Application of delayed extraction matrix-assisted laser desorption ionization time-of-flight mass spectrometry for analysis of sphingolipids in tissues from sphingolipidosis patients.

    Science.gov (United States)

    Fujiwaki, T; Yamaguchi, S; Sukegawa, K; Taketomi, T

    1999-08-06

    Sphingolipidosis is due to defects in enzymes involved in hydrolysis of sphingolipids. We analyzed sphingolipids in tissues from patients with sphingolipidosis, including Farber disease (FD, acid ceramidase deficiency), Gaucher disease (GD), Niemann-Pick disease type C (NPDC), and GM1-gangliosidosis (GM1G), using delayed extraction matrix-assisted laser desorption ionization time-of-flight mass spectrometry (DE MALDI-TOF-MS). Crude lipids were extracted from about 100 mg wet weight of autopsied tissues, including liver, spleen, cerebrum or cerebellum. After mild alkaline treatment, a sphingolipid fraction was prepared from the crude lipids and analyzed by DE MALDI-TOF-MS. The results were as follows: (a) In FD liver both the ceramide/sphingomyelin and ceramide/monohexosylceramide ratios were significantly high; (b) in both liver and spleen from a GD patient, the glucosylceramide/sphingomyelin ratio was raised; (c) in liver from a NPDC patient, the monohexosylceramide/sphingomyelin ratio was markedly low, suggesting an increase of sphingomyelin; and (d) in all tissues examined in the GM1G patient, GM1-gangliosides or asialo-GM1-gangliosides, that are undetectable in a normal control, were increased. In conclusion, sphingolipids in human tissues could be directly determined by DE MALDI-TOF-MS, with only a small amount of specimens. This method will be useful for the diagnosis and biochemical evaluation of sphingolipidosis patients.

  3. Mass fingerprint analysis of spider mites (Acari) by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for rapid discrimination.

    Science.gov (United States)

    Kajiwara, Hideyuki; Hinomoto, Norihide; Gotoh, Tetsuo

    2016-04-30

    Discrimination of spider mite species is still performed using morphological information, although DNA and other biological approaches have been attempted for identification purposes. These techniques need much time, are expensive, and require specialist staff. As an alternative, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) analysis is applied for rapid discrimination of spider mite species. Spider mites were analyzed using MALDI-TOFMS after extraction with 70% formic acid and acetonitrile. A single spider mite was also analyzed directly on double-sided carbon tape. A dendrogram was compiled from the MS data. Evolutionarily close and morphologically similar spider mites, the Kanzawa (Tetranychus kanzawai) and the two-spotted (T. urticae) spider mites, as well as three other related species of spider mites, could be discriminated by mass fingerprints. Although female adults were mainly used in this report, male adults and nymphs showed almost the same mass fingerprints and were not considered to affect discrimination capability. A single spider mite on double-sided carbon tape was analyzed directly by MALDI-TOFMS. Spider mites could be analyzed directly by MALDI-TOFMS, with evolutionarily and morphologically closely related spider mites showing different mass fingerprints, allowing for their identification. Copyright © 2016 John Wiley & Sons, Ltd.

  4. An investigation on the role of 3-hydroxykynurenine in pigment formation by matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Vogliardi, Susanna; Bertazzo, Antonella; Comai, Stefano; Costa, Carlo V L; Allegri, Graziella; Seraglia, Roberta; Traldi, Pietro

    2004-01-01

    In order to investigate the role of tryptophan and its metabolites in biogenesis of melanins, a study on the enzymatic reaction of 3-hydroxykynurenine with tyrosinase and peroxidase was performed. The reaction at different pH values was monitored by sampling at different times, with ultrafiltration used before analysis by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). The data obtained in this way showed that oligomerization processes take place with both enzymes, but with different behaviour, also depending on pH. 3-Hydroxykynurenine in the presence of tyrosinase at pH 6.0 leads to formation of xanthommatin, and at pH 8.0 hydroxanthommatin is formed in the first step of the reaction followed by formation of black-brown pigments. In contrast, the formation of oligomerization products by peroxidase action is observed in high yields under both acidic and basic conditions; however, at pH 6.0, a more extensive oligomerization process is observed. Thus peroxidase is able to activate oligomerization analogous to that observed in the case of tyrosinase without depending on the variation of pH. Due to the early formation of decarboxylated hydroxykynurenine, hydroxanthommatin and decarboxylated hydroxanthommatin, the enzymatic reaction leads to mixed oligomers, which can be considered as precursors of new pathways in pigment production. Copyright 2004 John Wiley & Sons, Ltd.

  5. Enzyme-coupled nanoparticles-assisted laser desorption ionization mass spectrometry for searching for low-mass inhibitors of enzymes in complex mixtures.

    Science.gov (United States)

    Salwiński, Aleksander; Da Silva, David; Delépée, Raphaël; Maunit, Benoît

    2014-04-01

    In this report, enzyme-coupled magnetic nanoparticles (EMPs) were shown to be an effective affinity-based tool for finding specific interactions between enzymatic targets and the low-mass molecules in complex mixtures using classic MALDI-TOF apparatus. EMPs used in this work act as nonorganic matrix enabling ionization of small molecules without any interference in the low-mass range (enzyme-coupled nanoparticles-assisted laser desorption ionization MS, ENALDI MS) and simultaneously carry the superficial specific binding sites to capture inhibitors present in a studied mixture. We evaluated ENALDI approach in two complementary variations: 'ion fading' (IF-ENALDI), based on superficial adsorption of inhibitors and 'ion hunting' (IH-ENALDI), based on selective pre-concentration of inhibitors. IF-ENALDI was applied for two sets of enzyme-inhibitor pairs: tyrosinase-glabridin and trypsin-leupeptin and for the real plant sample: Sparrmannia discolor leaf and stem methanol extract. The efficacy of IH-ENALDI was shown for the pair of trypsin-leupeptin. Both ENALDI approaches pose an alternative for bioassay-guided fractionation, the common method for finding inhibitors in the complex mixtures.

  6. Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry: a Fundamental Shift in the Routine Practice of Clinical Microbiology

    Science.gov (United States)

    Clark, Andrew E.; Kaleta, Erin J.; Arora, Amit

    2013-01-01

    SUMMARY Within the past decade, clinical microbiology laboratories experienced revolutionary changes in the way in which microorganisms are identified, moving away from slow, traditional microbial identification algorithms toward rapid molecular methods and mass spectrometry (MS). Historically, MS was clinically utilized as a high-complexity method adapted for protein-centered analysis of samples in chemistry and hematology laboratories. Today, matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) MS is adapted for use in microbiology laboratories, where it serves as a paradigm-shifting, rapid, and robust method for accurate microbial identification. Multiple instrument platforms, marketed by well-established manufacturers, are beginning to displace automated phenotypic identification instruments and in some cases genetic sequence-based identification practices. This review summarizes the current position of MALDI-TOF MS in clinical research and in diagnostic clinical microbiology laboratories and serves as a primer to examine the “nuts and bolts” of MALDI-TOF MS, highlighting research associated with sample preparation, spectral analysis, and accuracy. Currently available MALDI-TOF MS hardware and software platforms that support the use of MALDI-TOF with direct and precultured specimens and integration of the technology into the laboratory workflow are also discussed. Finally, this review closes with a prospective view of the future of MALDI-TOF MS in the clinical microbiology laboratory to accelerate diagnosis and microbial identification to improve patient care. PMID:23824373

  7. Emerging and Future Applications of Matrix-Assisted Laser Desorption Ionization Time-of-Flight (MALDI-TOF) Mass Spectrometry in the Clinical Microbiology Laboratory: A Report of the Association for Molecular Pathology.

    Science.gov (United States)

    Doern, Christopher D; Butler-Wu, Susan M

    2016-11-01

    The performance of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MS) for routine bacterial and yeast identification as well as direct-from-blood culture bottle identification has been thoroughly evaluated in the peer-reviewed literature. Microbiologists are now moving beyond these methods to apply MS to other areas of the diagnostic process. This review discusses the emergence of advanced matrix-assisted laser desorption ionization time-of-flight MS applications, including the identification of filamentous fungi and mycobacteria and the current and future state of antimicrobial resistance testing. Copyright © 2016 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

  8. A SIMPLE AND RAPID MATRIX-ASSISTED LASER DESORPTION/IONIZATION TIME OF FLIGHT MASS SPECTROMETRY METHOD TO SCREEN FISH PLASMA SAMPLES FOR ESTROGEN-RESPONSIVE BIOMARKERS

    Science.gov (United States)

    In this study, we describe and evaluate the performance of a simple and rapid mass spectral method for screening fish plasma for estrogen-responsive biomarkers using matrix assisted laster desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) couopled with a short...

  9. Matrix-assisted laser desorption/ionization time of flight mass-spectrometry (MALDI-TOF MS) based typing of extended-spectrum β-lactamase producing E. coli--a novel tool for real-time outbreak investigation.

    Science.gov (United States)

    Egli, Adrian; Tschudin-Sutter, Sarah; Oberle, Michael; Goldenberger, Daniel; Frei, Reno; Widmer, Andreas F

    2015-01-01

    Epidemiologically linked clusters are confirmed by typing strains with molecular typing such as pulsed-field gel electrophoresis (PFGE). We compared six extended-spectrum β-lactamase producing E. coli of a PFGE-related cluster with Matrix-assisted laser desorption/ionization-time of flight mass-spectrometry based typing that confirmed relatedness faster and more cost-effective, but as reliable as PFGE.

  10. An in-house assay is superior to Sepsityper for direct matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry identification of yeast species in blood cultures.

    Science.gov (United States)

    Bidart, Marie; Bonnet, Isabelle; Hennebique, Aurélie; Kherraf, Zine Eddine; Pelloux, Hervé; Berger, François; Cornet, Muriel; Bailly, Sébastien; Maubon, Danièle

    2015-05-01

    We developed an in-house assay for the direct identification, by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry, of yeasts in blood culture. Sixty-one representative strains from 12 species were analyzed in spiked blood cultures. Our assay accurately identified 95 of 107 (88.8%) positive blood cultures and outperformed the commercial Sepsityper kit (81.7% identification). Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  11. Identification of Blood Culture Isolates Directly from Positive Blood Cultures by Use of Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry and a Commercial Extraction System: Analysis of Performance, Cost, and Turnaround Time

    OpenAIRE

    Lagacé-Wiens, Philippe R. S.; Adam, Heather J.; Karlowsky, James A.; Nichol, Kimberly A.; Pang, Paulette F.; Guenther, Jodi; Webb, Amanda A.; Miller, Crystal; Alfa, Michelle J.

    2012-01-01

    Matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) mass spectrometry represents a revolution in the rapid identification of bacterial and fungal pathogens in the clinical microbiology laboratory. Recently, MALDI-TOF has been applied directly to positive blood culture bottles for the rapid identification of pathogens, leading to reductions in turnaround time and potentially beneficial patient impacts. The development of a commercially available extraction kit (Bruker Sepsit...

  12. Misidentification of Saprochaete clavata as Magnusiomyces capitatus in Clinical Isolates: Utility of Internal Transcribed Spacer Sequencing and Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry and Importance of Reliable Databases

    OpenAIRE

    Desnos-Ollivier, Marie; Blanc, Catherine; Garcia-Hermoso, Dea; Hoinard, Damien; Alanio, Alexandre; Dromer, Françoise

    2014-01-01

    Saprochaete clavata and Magnusiomyces capitatus are human pathogens that are frequently mistaken for each other due to their similar phenotypes and erroneous or limited databases. Based on internal transcribed spacer (ITS) sequences, we propose species-specific carbon assimilation patterns and matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) fingerprints that enable the identification of S. clavata, M. capitatus, and Galactomyces candidus to the ...

  13. Misidentification of Saprochaete clavata as Magnusiomyces capitatus in Clinical Isolates: Utility of Internal Transcribed Spacer Sequencing and Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry and Importance of Reliable Databases

    Science.gov (United States)

    Desnos-Ollivier, Marie; Blanc, Catherine; Garcia-Hermoso, Dea; Hoinard, Damien; Alanio, Alexandre

    2014-01-01

    Saprochaete clavata and Magnusiomyces capitatus are human pathogens that are frequently mistaken for each other due to their similar phenotypes and erroneous or limited databases. Based on internal transcribed spacer (ITS) sequences, we propose species-specific carbon assimilation patterns and matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) fingerprints that enable the identification of S. clavata, M. capitatus, and Galactomyces candidus to the species level. PMID:24696028

  14. 1,8-Bis(dimethylamino)naphthalene/9-aminoacridine: A new binary matrix for lipid fingerprinting of intact bacteria by matrix assisted laser desorption ionization mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Calvano, C.D., E-mail: cosimadamiana.calvano@uniba.it [Dipartimento di Chimica, Università degli Studi di Bari Aldo Moro, Via Orabona, 4, 70126 Bari (Italy); Monopoli, A.; Ditaranto, N. [Dipartimento di Chimica, Università degli Studi di Bari Aldo Moro, Via Orabona, 4, 70126 Bari (Italy); Palmisano, F. [Dipartimento di Chimica, Università degli Studi di Bari Aldo Moro, Via Orabona, 4, 70126 Bari (Italy); Centro Interdipartimentale di Ricerca S.M.A.R.T., Università degli Studi di Bari Aldo Moro, Via Orabona, 4, 70126 Bari (Italy)

    2013-10-10

    Graphical abstract: -- Highlights: •New binary matrix for less ionizable lipid analysis with no interfering peaks. •Combined MALDI and X-ray photoelectron spectroscopy (XPS) analyses. •Fast lipid fingerprint on Gram positive and Gram negative bacteria by MALDI MS. •Mapping of phospholipids by XPS imaging. •Very fast membrane lipid extraction procedure. -- Abstract: The effectiveness of a novel binary matrix composed of 1,8-bis(dimethylamino)naphthalene (DMAN; proton sponge) and 9-aminoacridine (9AA) for the direct lipid analysis of whole bacterial cells by matrix assisted laser desorption ionization mass spectrometry (MALDI MS) is demonstrated. Deprotonated analyte signals nearly free of matrix-related ions were observed in negative ion mode. The effect of the most important factors (laser energy, pulse voltage, DMAN/9AA ratio, analyte/matrix ratio) was investigated using a Box–Behnken response surface design followed by multi-response optimization in order to simultaneously maximize signal-to-noise (S/N) ratio and resolution. The chemical surface composition of single or mixed matrices was explored by X-ray photoelectron spectroscopy (XPS). Moreover, XPS imaging was used to map the spatial distribution of a model phospholipid in single or binary matrices. The DMAN/9AA binary matrix was then successfully applied to the analysis of intact Gram positive (Lactobacillus sanfranciscensis) or Gram negative (Escherichia coli) microorganisms. About fifty major membrane components (free fatty acids, mono-, di- and tri-glycerides, phospholipids, glycolipids and cardiolipins) were quickly and easily detected over a mass range spanning from ca. 200 to ca. 1600 m/z. Moreover, mass spectra with improved S/N ratio (compared to single matrices), reduced chemical noise and no formation of matrix-clusters were invariably obtained demonstrating the potential of this binary matrix to improve sensitivity.

  15. Humic acids as both matrix for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and adsorbent for magnetic solid phase extraction.

    Science.gov (United States)

    Zhao, Qin; Xu, Jing; Yin, Jia; Feng, Yu-Qi

    2015-08-19

    In the present study, humic acids (HAs) were applied as both a matrix for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and an adsorbent of magnetic solid phase extraction (MSPE) for the first time. As natural macromolecule compounds, HAs are inherently highly functionalized and contain laser energy absorbing-transferring aromatic structures. This special molecular structure made HAs a good candidate for use as a MALDI matrix in small molecule analysis. At the same time, due to its good adsorption ability, HAs was prepared as MSPE adsorbent via a simple co-mixing method, in which the commercially available HAs were directly mixed with Fe3O4 magnetic nanoparticles (MNPs) in a mortar and grinded evenly and completely. In this process, MNPs were physically wrapped and adhered to tiny HAs leading to the formation of magnetic HAs (MHAs). To verify the bi-function of the MHAs, Rhodamine B (RdB) was chosen as model compound. Our results show that the combination of MHAs-based MSPE and MALDI-TOF-MS can provide a rapid and sensitive method for the determination of RdB in chili oil. The whole analytical procedure could be completed within 30 min for simultaneous determination of more than 20 samples, and the limit of quantitation for RdB was found to be 0.02 μg/g. The recoveries in chili oil were in the range 73.8-81.5% with the RSDs less than 21.3% (intraday) and 20.3% (interday). The proposed strategy has potential applications for high-throughput analysis of small molecules in complex samples. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. Quantification of plant surface metabolites by matrix-assisted laser desorption-ionization mass spectrometry imaging: glucosinolates on Arabidopsis thaliana leaves.

    Science.gov (United States)

    Shroff, Rohit; Schramm, Katharina; Jeschke, Verena; Nemes, Peter; Vertes, Akos; Gershenzon, Jonathan; Svatoš, Aleš

    2015-03-01

    The localization of metabolites on plant surfaces has been problematic because of the limitations of current methodologies. Attempts to localize glucosinolates, the sulfur-rich defense compounds of the order Brassicales, on leaf surfaces have given many contradictory results depending on the method employed. Here we developed a matrix-assisted laser desorption-ionization (MALDI) mass spectrometry protocol to detect surface glucosinolates on Arabidopsis thaliana leaves by applying the MALDI matrix through sublimation. Quantification was accomplished by spotting glucosinolate standards directly on the leaf surface. The A. thaliana leaf surface was found to contain approximately 15 nmol of total glucosinolate per leaf with about 50 pmol mm(-2) on abaxial (bottom) surfaces and 15-30 times less on adaxial (top) surfaces. Of the major compounds detected, 4-methylsulfinylbutylglucosinolate, indol-3-ylmethylglucosinolate, and 8-methylsulfinyloctylglucosinolate were also major components of the leaf interior, but the second most abundant glucosinolate on the surface, 4-methylthiobutylglucosinolate, was only a trace component of the interior. Distribution on the surface was relatively uniform in contrast to the interior, where glucosinolates were distributed more abundantly in the midrib and periphery than the rest of the leaf. These results were confirmed by two other mass spectrometry-based techniques, laser ablation electrospray ionization and liquid extraction surface analysis. The concentrations of glucosinolates on A. thaliana leaf surfaces were found to be sufficient to attract the specialist feeding lepidopterans Plutella xylostella and Pieris rapae for oviposition. The methods employed here should be easily applied to other plant species and metabolites. © 2015 The Authors The Plant Journal © 2015 John Wiley & Sons Ltd.

  17. Matrix-assisted laser desorption/ionization mass spectrometric imaging for the rapid segmental analysis of methamphetamine in a single hair using umbelliferone as a matrix.

    Science.gov (United States)

    Wang, Hang; Wang, Ying

    2017-07-04

    Segmental hair analysis offers a longer period for retrospective drug detection than blood or urine. Hair is a keratinous fiber and is strongly hydrophobic. The embedding of drugs in hydrophobic hair at low concentrations makes it difficult for extraction and detection with matrix-assisted laser desorption/ionization (MALDI) coupled with mass spectrometric imaging (MSI). In this study, a single scalp hair was longitudinally cut with a cryostat section to a length of 4 mm and fixed onto a stainless steel MALDI plate. Umbelliferone was used as a new hydrophobic matrix to enrich and assist the ionization efficiency of methamphetamine in the hair sample. MALDI-Fourier transform ion cyclotron resonance (FTICR)-MS profiling and imaging were performed for direct detection and mapping of methamphetamine on the longitudinal sections of the single hair sample in positive ion mode. Using MALDI-MSI, the distribution of methamphetamine was observed throughout five longitudinally sectioned hair samples from a drug abuser. The changes of methamphetamine were also semi-quantified by comparing the ratios of methamphetamine/internal standard (I.S). This method improves the detection sensitivity of target drugs embedded in a hair matrix for imaging with mass spectrometry. The method could provide a detection level of methamphetamine down to a nanogram per milligram incorporated into hair. The results were also compared with the conventional high performance liquid chromatography -tandem mass spectrometry (HPLC-MS/MS) method. Changes in the imaging results over time by the MSI method showed good semi-quantitative correlation to the results from the HPLC-MS/MS method. This study provides a powerful tool for drug abuse control and forensic medicine analysis in a narrow time frame, and a reduction in the sample amount required. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Deuterium desorption from tungsten using laser heating

    Directory of Open Access Journals (Sweden)

    J.H. Yu

    2017-08-01

    Full Text Available Retention and desorption of hydrogenic species need to be accurately modeled to predict the tritium inventory of next generation fusion devices, which is needed both for tritium fuel recovery and for tritium safety concerns. In this paper, experiments on thermal desorption of deuterium from intrinsic polycrystalline tungsten defects using laser heating are compared to TMAP-7 modeling. The samples during deuterium plasma exposure were at a temperature of 373K for this benchmark study with ion fluence of 0.7–1.0 ×1024Dm−2. Following plasma exposure, a fiber laser (λ= 1100nm heated the samples to peak surface temperatures ranging from ∼500 to 1400K with pulse widths from 10ms to 1s, and 1 to 10 pulses applied to each sample. The remaining deuterium retention was measured using temperature programmed desorption (TPD. Results show that > 95% of deuterium is desorbed when the peak surface temperature reached ∼950K for > 1s. TMAP-7 is used to predict deuterium desorption from tungsten for a range of surface temperatures and heating durations, and is compared to previous work on desorption from beryllium codeposits.

  19. Infrared matrix-assisted laser desorption/ionization orthogonal-time-of-flight mass spectrometry employing a cooling stage and water ice as a matrix.

    Science.gov (United States)

    Pirkl, Alexander; Soltwisch, Jens; Draude, Felix; Dreisewerd, Klaus

    2012-07-03

    Although water ice has been utilized in the past as a matrix for infrared matrix-assisted laser desorption/ionization mass spectrometry (IR-MALDI-MS), it has not found a wider use due to limitations in the analytical performance and technical demands on the employment of the necessary cooling stage. Here, we developed a temperature-controlled sample stage for use with an orthogonal time-of-flight mass spectrometer (MALDI-o-TOF-MS). The stage utilizes a combination of liquid nitrogen cooling and counterheating with a Peltier element. It allows adjustment of the sample temperature between ~-120 °C and room temperature. To identify optimal irradiation conditions for IR-MALDI with the water ice matrix, we first investigated the influence of excitation wavelength, varied between 2.7 and 3.1 μm, and laser fluence on the signal intensities of molecular substance P ions. These data suggest the involvement of transient melting of the ice during the laser pulse and primary energy deposition into liquid water. As a consequence, the best analytical performance is obtained at a wavelength corresponding to the absorption maximum of liquid water of about 2.94 μm. The current data significantly surpass the previously reported analytical features. The particular softness of the method is, for example, exemplified by the analysis of noncovalently bound holo-myoglobin and of ribonuclease B. This is also the first report demonstrating the analysis of an IgG monoclonal antibody (MW ~ 150 kDa) from a water ice matrix. Untypical for MALDI-MS, high charge states of multiply protonated species were moreover observed for some of the investigated peptides and even for lacto-N-fucopentaose II oligosaccharides. Using water ice as matrix is of particular interest for MALDI MS profiling and imaging applications since matrix-free spectra are produced. The MS and tandem MS analysis of metabolites directly from frozen food samples is demonstrated with the example of a strawberry fruit.

  20. Matrix Assisted and/or Laser Desorption Ionization Quadrupole Ion Trap Time-of-Flight Mass Spectrometry of WO3 Clusters Formation in Gas Phase. Nanodiamonds, Fullerene, and Graphene Oxide Matrices.

    Science.gov (United States)

    Ausekar, Mayuri Vilas; Mawale, Ravi Madhukar; Pazdera, Pavel; Havel, Josef

    2018-01-16

    The formation of W x O y+●/-● clusters in the gas phase was studied by laser desorption ionization (LDI) and matrix assisted laser desorption ionization (MALDI) of solid WO3. LDI produced (WO3) n+ ●/- ● (n = 1-7) clusters. In MALDI, when using nano-diamonds (NDs), graphene oxide (GO), or fullerene (C60) matrices, higher mass clusters were generated. In addition to (WO3) n-● clusters, oxygen-rich or -deficient species were found in both LDI and MALDI (with the total number of clusters exceeding one hundred ≈ 137). This is the first time that such matrices have been used for the generation of(WO3) n+●/-● clusters in the gas phase, while new high mass clusters (WO3) n-● (n = 12-19) were also detected. Graphical Abstract.

  1. Identification of Microorganisms by FilmArray and Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry Prior to Positivity in the Blood Culture System

    Science.gov (United States)

    Almuhayawi, Mohammed; Altun, Osman; Strålin, Kristoffer

    2014-01-01

    In this study, we investigated the performance of the FilmArray and matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) in identifying microorganisms from blood culture (BC) bottles prior to positivity. First, we used simulated BacT/Alert FA Plus BC bottles with five each for Escherichia coli and Staphylococcus aureus isolates. The FilmArray identified all 10 isolates before BC positivity with 9/10 at 5 h and 1 at 7.5 h after incubation in the BC system. MALDI-TOF MS failed to identify the isolates prior to positivity. When the bottles were incubated for 2.5 h at room temperature (RT) before we put them into the BC system, the FilmArray identified 6/10 at 2.5 h and the remaining 4 at 5 h. Finally, we tested simulated BC bottles after incubation at RT. Interestingly, 9/10 isolates were identified with the FilmArray after 8 h of incubation at RT. Second, we studied clinical BC bottles in quadruplicate. When three-fourths of the parallel bottles signaled positive, the FilmArray was run on the fourth nonsignaled bottle and was found to be positive in 14/15 such cases. Third, we analyzed the performance of the FilmArray in the identification of microorganisms from clinical BC bottles before incubation in the system. Two milliliters of broth from 400 BC bottles was collected after arrival at the laboratory and stored at −70°C. Sixteen bottles later signaled positive in the system. When the frozen broth from these bottles was analyzed, the FilmArray identified all the microorganisms in 8/16 bottles prior to incubation in the BC system. This study shows that the FilmArray can identify microorganisms from BC bottles prior to positivity and in some cases even prior to incubation in the BC system. PMID:24951811

  2. The use of Gram stain and matrix-assisted laser desorption ionization time-of-flight mass spectrometry on positive blood culture: synergy between new and old technology.

    Science.gov (United States)

    Fuglsang-Damgaard, David; Nielsen, Camilla Houlberg; Mandrup, Elisabeth; Fuursted, Kurt

    2011-10-01

    Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) is promising as an alternative to more costly and cumbersome methods for direct identifications in blood cultures. We wanted to evaluate a simplified pre-treatment method for using MALDI-TOF-MS directly on positive blood cultures using BacT/Alert blood culture system, and to test an algorithm combining the result of the initial microscopy with the result suggested by MALDI-TOF-MS. Using the recommended cut-off score of 1.7 the best results were obtained among Gram-negative rods with correct identifications in 91% of Enterobacteriaceae, 83% in aerobic/non-fermentative Gram-negative rods, whereas results were more modest among Gram-positive cocci with correct identifications in 52% of Staphylococci, 54% in Enterococci and only 20% in Streptococci. Combining the results of Gram stain with the top reports by MALDI-TOF-MS, increased the sensitivity from 91% to 93% in the score range from 1.5 to 1.7 and from 48% to 85% in the score range from 1.3 to 1.5. Thus, using this strategy and accepting a cut-off at 1.3 instead of the suggested 1.7, overall sensitivity could be increased from 88.1% to 96.3%. MALDI-TOF-MS is an efficient method for direct routine identification of bacterial isolates in blood culture, especially when combined with the result of the Gram stain. © 2011 The Authors. APMIS © 2011 APMIS.

  3. Efficient identification of Malassezia yeasts by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Kolecka, A; Khayhan, K; Arabatzis, M; Velegraki, A; Kostrzewa, M; Andersson, A; Scheynius, A; Cafarchia, C; Iatta, R; Montagna, M T; Youngchim, S; Cabañes, F J; Hoopman, P; Kraak, B; Groenewald, M; Boekhout, T

    2014-02-01

    Infections caused by Malassezia yeasts are most likely underdiagnosed, because fatty acid supplementation is needed for growth. Rapid identification of Malassezia species is essential for appropriate treatment of Malassezia-related skin infections, fungaemia and nosocomial outbreaks in neonates, children and adults and can be life-saving for those patients. Ma-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has been reported to be a rapid and reliable diagnostic tool to identify clinically important yeasts, but so far no data have been reported on identification of Malassezia isolates with this technique. To create an extensive database of main mass spectra (MSPs) that will allow quick identification of Malassezia species by MALDI-TOF MS. An in-house library of 113 MSPs was created from 48 reference strains from the CBS-KNAW yeast collection. The in-house library was challenged with two test sets of Malassezia strains, namely 165 reference strains from the CBS collection and 338 isolates collected in Greece, Italy, Sweden and Thailand. MALDI-TOF MS allowed correct identification of all 14 Malassezia spp. MALDI-TOF MS results were concordant with those of sequence analyses of the internal transcribed spacers (ITS1/ITS2) and the D1/D2 domains of the large subunit of the ribosomal DNA. Implementation of the MALDI-TOF MS system as a routine identification tool will contribute to correct identification of Malassezia yeasts with minimal effort and in a short turnaround time, which is especially important for the rapid identification of Malassezia in skin diseases and nosocomial outbreaks. © 2013 British Association of Dermatologists.

  4. Identification and characterization of Clostridium botulinum group III field strains by matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Bano, Luca; Drigo, Ilenia; Tonon, Elena; Pascoletti, Simone; Puiatti, Cinzia; Anniballi, Fabrizio; Auricchio, Bruna; Lista, Florigio; Montecucco, Cesare; Agnoletti, Fabrizio

    2017-12-01

    Animal botulism is primarily due to botulinum neurotoxin (BoNT) types C, D or their chimeric variants C/D or D/C, produced by Clostridium botulinum group III, which appears to include the genetically indistinguishable Clostridium haemolyticum and Clostridium novyi. In the present study, we used matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI TOF MS) to identify and characterize 81 BoNT-producing Clostridia isolated in 47 episodes of animal botulism. The instrument's default database, containing no entries for Clostridium botulinum, permitted reliable identification of 26 strains at the genus level. Although supplementation of the database with reference strains enhanced the instrument's ability to identify the neurotoxic strains at the genus level, resolution was not sufficient to recognize field strains at species level. Characterization by MALDI TOF confirmed the well-documented phenotypic and genetic differences between Clostridium botulinum strains of serotypes normally implicated in human botulism (A, B, E, F) and other Clostridium species able to produce BoNTs type C and D. The chimeric and non-chimeric field strains grouped separately. In particular, very low similarity was found between two non-chimeric type C field strains isolated in the same outbreak and the other field strains. This difference was comparable with the differences among the various Clostridia species included in the study. Characterization by MALDI TOF confirmed that BoNT-producing Clostridia isolated from animals are closely related and indistinguishable at the species level from Clostridium haemolyticum and Clostridium novyi reference strains. On the contrary, there seem to be substantial differences among chimeric and some non-chimeric type C strains. Copyright © 2017 Elsevier Ltd. All rights reserved.

  5. Evaluation of two matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) systems for the identification of Candida species.

    Science.gov (United States)

    Lacroix, C; Gicquel, A; Sendid, B; Meyer, J; Accoceberry, I; François, N; Morio, F; Desoubeaux, G; Chandenier, J; Kauffmann-Lacroix, C; Hennequin, C; Guitard, J; Nassif, X; Bougnoux, M-E

    2014-02-01

    Candida spp. are responsible for severe infections in immunocompromised patients and those undergoing invasive procedures. The accurate identification of Candida species is important because emerging species can be associated with various antifungal susceptibility spectra. Conventional methods have been developed to identify the most common pathogens, but have often failed to identify uncommon species. Several studies have reported the efficiency of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for the identification of clinically relevant Candida species. In this study, we evaluated two commercially available MALDI-TOF systems, Andromas™ and Bruker Biotyper™, for Candida identification in routine diagnosis. For this purpose, we investigated 1383 Candida isolates prospectively collected in eight hospital laboratories during routine practice. MALDI-TOF MS results were compared with those obtained using conventional phenotypic methods. Analysis of rDNA gene sequences with internal transcribed regions or D1-D2 regions is considered the reference standard for identification. Both MALDI-TOF MS systems could accurately identify 98.3% of the isolates at the species level (1359/1383 for Andromas™; 1360/1383 for Bruker Biotyper™) vs. 96.5% for conventional techniques. Furthermore, whereas conventional methods failed to identify rare or emerging species, these were correctly identified by MALDI-TOF MS. Both MALDI-TOF MS systems are accurate and cost-effective alternatives to conventional methods for mycological identification of clinically relevant Candida species and should improve the diagnosis of fungal infections as well as patient management. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  6. Matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) for rapid identification of micro-organisms in the routine clinical microbiology laboratory.

    Science.gov (United States)

    Wattal, C; Oberoi, J K; Goel, N; Raveendran, R; Khanna, S

    2017-05-01

    The study evaluates the utility of matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) Vitek MS for identification of microorganisms in the routine clinical microbiology laboratory. From May 2013 to April 2014, microbial isolates recovered from various clinical samples were identified by Vitek MS. In case of failure to identify by Vitek MS, the isolate was identified using the Vitek 2 system (bioMerieux, France) and serotyping wherever applicable or otherwise by nucleic acid-mediated methods. All the moulds were identified by Lactophenol blue mounts, and mycobacterial isolates were identified by molecular identification systems including AccuProbe (bioMerieux, France) or GenoType Mycobacterium CM (Hain Lifescience, Germany). Out of the 12,003 isolates, the Vitek MS gave a good overall ID at the genus and or species level up to 97.7% for bacterial isolates, 92.8% for yeasts and 80% for filamentous fungi. Of the 26 mycobacteria tested, only 42.3% could be identified using the Saramis RUO (Research Use Only) database. VITEK MS could not identify 34 of the 35 yeast isolates identified as C. haemulonii by Vitek 2. Subsequently, 17 of these isolates were identified as Candida auris (not present in the Vitek MS database) by 18S rRNA sequencing. Using these strains, an in-house superspectrum of C. auris was created in the VITEK MS database. Use of MALDI-TOF MS allows a rapid identification of aerobic bacteria and yeasts in clinical practice. However, improved sample extraction protocols and database upgrades with inclusion of locally representative strains is required, especially for moulds.

  7. Characterization of on-target generated tryptic peptides from Giberella zeae conidia spore proteins by means of matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Dong, Hongjuan; Marchetti-Deschmann, Martina; Allmaier, Günter

    2014-01-01

    Traditionally characterization of microbial proteins is performed by a complex sequence of steps with the final step to be either Edman sequencing or mass spectrometry, which generally takes several weeks or months to be complete. In this work, we proposed a strategy for the characterization of tryptic peptides derived from Giberella zeae (anamorph: Fusarium graminearum) proteins in parallel to intact cell mass spectrometry (ICMS) in which no complicated and time-consuming steps were needed. Experimentally, after a simple washing treatment of the spores, the aliquots of the intact G. zeae macro conidia spores solution, were deposited two times onto one MALDI (matrix-assisted laser desorption ionization) mass spectrometry (MS) target (two spots). One spot was used for ICMS and the second spot was subject to a brief on-target digestion with bead-immobilized or non-immobilized trypsin. Subsequently, one spot was analyzed immediately by MALDI MS in the linear mode (ICMS) whereas the second spot containing the digested material was investigated by MALDI MS in the reflectron mode ("peptide mass fingerprint") followed by protonated peptide selection for MS/MS (post source decay (PSD) fragment ion) analysis. Based on the formed fragment ions of selected tryptic peptides a complete or partial amino acid sequence was generated by manual de novo sequencing. These sequence data were used for homology search for protein identification. Finally four different peptides of varying abundances have been identified successfully allowing the verification that our desorbed/ionized surface compounds were indeed derived from proteins. The presence of three different proteins could be found unambiguously. Interestingly, one of these proteins is belonging to the ribosomal superfamily which indicates that not only surface-associated proteins were digested. This strategy minimized the amount of time and labor required for obtaining deeper information on spore preparations within the

  8. Characterizing changes in snow crab (Chionoecetes opilio) cryptocyanin protein during molting using matrix-assisted laser desorption/ionization mass spectrometry and tandem mass spectrometry.

    Science.gov (United States)

    Demian, Wael L L; Jahouh, Farid M; Stansbury, Don; Randell, Edward; Brown, Robert J; Banoub, Joseph H

    2014-02-28

    We report the matrix-assisted laser desorption/ionization mass spectrometric (MALDI-MS) characterization of the cryptocyanin proteins of the juvenile Chionoecetes opilio crabs during their molting and non-molting phases. In order to assess the structural cryptocyanin protein differences between the molting and non-molting phases, the obtained peptides were sequenced by MALDI low-energy collision-induced dissociation tandem mass spectrometry (CID-MS/MS). The cryptocyanin protein was isolated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and analyzed by MALDI-TOF/TOF-MS. The purified cryptocyanin protein was sequenced, using the 'bottom-up' approach. After tryptic digestion, the peptide mixture was analyzed by MALDI-QqTOF-MS/MS and the data obtained were used for the peptide mass fingerprinting (PMF) identification by means of the Mascot database. It was demonstrated using MALDI-TOF/TOF-MS that the actual molecular weights of the non-molting and molting cryptocyanin proteins were different; these were, respectively, 67.6 kDa and 68.1 kDa. Using low-energy CID-MS/MS we have sequenced the trytic peptides to monitor the differences and similarities between the cryptocyanin molecular structures during the molting and non-molting stages. We have demonstrated for the first time that the actual molecular masses of the cryptocyanin protein during the molting and non-molting phases were different. The MALDI-CID-MS/MS analyses allowed the sequencing of the cryptocyanins after tryptic digestion, during the molting and non-molting stages, and showed some similarities and staggering differences between the identified cryptocyanin peptides. Copyright © 2013 John Wiley & Sons, Ltd.

  9. Rapid and simple analysis of disease-associated biomarkers of Taiwanese patients with schizophrenia using matrix-assisted laser desorption ionization mass spectrometry.

    Science.gov (United States)

    Huang, Tiao-Lai; Lo, Li-Hua; Shiea, Jentaie; Su, Hung

    2017-10-01

    Matrix-assisted laser desorption ionization/time-of-flight mass spectrometry (MALDI-TOF MS) is an extremely sensitive analytical tool for characterizing biological compounds in bio samples. In this study, we applied MALDI-TOF MS to assess potential protein biomarkers in the peripheral blood mononuclear cells (PBMCs) of patients with schizophrenia in the acute phase, recovery phase and healthy controls in Taiwan. We recruited 40 participants, including 20 pairs of patients diagnosed with schizophrenia in the acute phase, after four-week treatment with drug in the recovery phase, and 20 healthy controls. The schizophrenic patients were diagnosed using Structured Clinical Interview for DSM-IV Axis I Disorders (SCID), and severity was assessed by a positive and negative symptom scale at baseline and at endpoint following four-week treatment with drug. The patients' PBMCs biomarkers were rapidly measured using a technique that combines MALDI-TOF MS and principle component analysis. A receiver operating characteristic curve was created for the evaluated biomarker. Significant differences in α-defensins 1-3 were found between the patients in acute phase with schizophrenia and the healthy controls, but not between the schizophrenic patients in recovery phase and healthy controls or between the schizophrenic patients in acute phase and in recovery phase. α-Defensins can be biomarkers of Taiwanese patients with schizophrenia, thus supporting the hypothesis that the inflammatory response and immunity system is correlated with the pathophysiology of schizophrenia. Moreover, the result also implies that α-defensins may be related in schizophrenia-associated disease not in efficacy of drug-treatment. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. Evaluation of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for identification and clustering of Neisseria gonorrhoeae.

    Science.gov (United States)

    Carannante, Anna; De Carolis, Elena; Vacca, Paola; Vella, Antonietta; Vocale, Caterina; De Francesco, Maria Antonia; Cusini, Marco; Del Re, Simonetta; Dal Conte, Ivano; Cristaudo, Antonio; Ober, Patrizia; Sanguinetti, Maurizio; Stefanelli, Paola

    2015-07-24

    The sexually transmitted infection gonorrhea remains a public health concern for becoming resistant to drug treatments available. The purpose of this study was to evaluate the usefulness of the matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) to identify and cluster Neisseria gonorrhoeae. From a current monitoring in Italy, as part of the European Gonococcal Antimicrobial Surveillance Programme (EURO-GASP), 93 gonococci collected from 2007 to 2012 susceptible (44 isolates) and resistant (49 isolates) to cefixime were selected. Minimum Inhibitory Concentration (MIC) values for cefixime was assessed by Etest carried out in agreement with the manufacturer's instructions and interpreted referring to European Committee on Antimicrobial Susceptibility testing (EUCAST) clinical breakpoints criteria. Data obtained by N. gonorrhoeae multiantigen sequence typing (NG-MAST) and the dendrogram based on the concatenation of porB and tbpB genes were evaluated. MALDI-TOF MS, to reconfirm gonorrhea identification, analyzed single colonies from freshly grown isolates and applied directly on a ground-steel MALDI target plate. For the MALDI-TOF dendrogram cluster analysis, MSPs (Main Spectrum Profile) from each isolate were created acquiring 5000 shots from 10 technical replicates obtained from bacteria extraction. Molecular typing by NG-MAST showed 28 sequence types (STs); G1407 was the predominant accounting for 75 gonococci. All the 93 gonococci, except one, were correctly identified at species level by MALDI-TOF MS and G1407 isolates were divided into two clusters. MALDI-TOF MS for a real-time detection and cluster analysis of gonorrhea is a promising tool for surveillance purposes. Moreover, additional studies are required to collect more data on the performance of MALDI-TOF MS for gonococci.

  11. Analysis of methicillin-resistant Staphylococcus aureus major clonal lineages by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Zhang, Tingting; Ding, Jinya; Rao, Xiancai; Yu, Jingbo; Chu, Meiling; Ren, Wei; Wang, Lu; Xue, Wencheng

    2015-10-01

    Methicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen associated with nosocomial infections in many countries. Multilocus sequence typing (MLST) is one of the genetic typing methods used to type MRSA with a high discriminatory power, however, it is labor-intensive, timely, and costly. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) coupled with ClinProTools is a potential tool to discover biomarker peaks and to generate a classification model based on highly sophisticated mathematical algorithms to discriminate clonal lineages. We investigated the performance of MALDI-TOF MS for discriminating 154 MRSA-ST239, 72 MRSA-ST5, 30 MRSA-ST59, 14 MRSA-ST45, and 20 MRSA-OST (other clonal lineages). Our results indicate that the model construction and validation have good potency to discriminate ST45 from other lineages with a sensitivity and a specificity of both 100%, and a sensitivity of 95.80% and a specificity of 94.62% to identify ST239. For Biotyper classification, the sensitivity and specificity were more than of 90% for ST239, ST59 and ST45, whereas only 81.94% sensitivity for ST5. By single-peak analysis, the peaks m/z 4808 and 9614 can correctly discriminate ST45 a sensitivity and a specificity of both 100%; the peak m/z 6554 can also discriminate ST239 with a sensitivity of 91.9% and a specificity of 85.4%. In conclusion, MALDI-TOF MS coupled with ClinProTools has a high detection performance for MRSA typing with obvious advantages of being rapid, highly accurate, and being a low cost in comparison with MLST. Copyright © 2015 Elsevier B.V. All rights reserved.

  12. [Identification of Mycobacterium spp. isolates using matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS)].

    Science.gov (United States)

    Amlerová, J; Studentová, V; Hrabák, J

    2014-09-01

    Matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS) has recently been widely used in diagnostic microbiological laboratories. It is a cheap and rapid method for the identification of bacteria and micromycetes. Apart from this purpose, it is also used for the detection of antibiotic resistance mechanisms. It has the potential to be extended for other purposes in microbiology. The aim of this study was to validate MALDI-TOF MS for the identification of mycobacteria. Thirty isolates of Mycobacterium spp. isolated in the Laboratory of Mycobacteriology of the Plzeň University Hospital were included in the study. The isolates were identified to the species level using biochemical tests, gene probes, and sequencing of the gene encoding 16S rRNA. The identification by MALDI-TOF MS was performed with the use of silica beads. Strain identification by sequencing the gene encoding 16S rRNA was considered as the reference method. MALDI-TOF MS correctly identified all isolates of Mycobacterium spp. (score range 1.461 - 2.168). The species identified were Mycobacterium tuberculosis (n= 5), Mycobacterium kansasii (n=5), Mycobacterium avium (n=6), Mycobacterium intracelullare (n=3), Mycobacterium xenopi (n=3), Mycobacterium gordonae (n=1), Mycobacterium abscessus (n=1), Mycobacterium kumamotonense (n=2), Mycobacterium mantenii (n=1), Mycobacterium lentiflavum (n=1), Mycobacterium fortuitum (n=1), and Mycobacterium scrofulaceum (n=1). MALDI-TOF MS is a suitable tool for the routine identification of Mycobacterium spp. in laboratories using this method for the conventional identification of microbes.

  13. Profiling of Microbial Colonies for High-Throughput Engineering of Multistep Enzymatic Reactions via Optically Guided Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry.

    Science.gov (United States)

    Si, Tong; Li, Bin; Comi, Troy J; Wu, Yuwei; Hu, Pingfan; Wu, Yuying; Min, Yuhao; Mitchell, Douglas A; Zhao, Huimin; Sweedler, Jonathan V

    2017-09-13

    Matrix-assisted laser desorption/ionization time-of-flight (MALDI-ToF) mass spectrometry (MS) imaging has been used for rapid phenotyping of enzymatic activities, but is mainly limited to single-step conversions. Herein we report a label-free method for high-throughput engineering of multistep biochemical reactions based on optically guided MALDI-ToF MS analysis of bacterial colonies. The bacterial cells provide containment of multiple enzymes and access to substrates and cofactors via metabolism. Automated MALDI-ToF MS acquisition from randomly distributed colonies simplifies procedures to prepare strain libraries without liquid handling. MALDI-ToF MS profiling was utilized to screen both substrate and enzyme libraries for natural product biosynthesis. Computational algorithms were developed to process and visualize the resulting mass spectral data sets. For analogues of the peptidic antibiotic plantazolicin, multivariate analyses by t-distributed stochastic neighbor embedding were used to group similar spectra for rapid identification of nonisobaric variants. After MALDI-ToF MS screening, follow-up analyses using high-resolution MS and tandem MS were readily performed on the same sample target. Separately, relative ion intensities of rhamnolipid congeners with various lipid moieties were evaluated to engineer enzymatic specificity. The glycolipid profiles of each colony were overlaid with optical images to facilitate the recovery of desirable mutants. For both the antibiotic and rhamnolipid cases, large populations of colonies were rapidly surveyed at the molecular level, providing information-rich insights not easily obtained with traditional screening assays. Utilizing standard microbiological techniques with routine microscopy and MALDI-ToF MS instruments, this simple yet effective workflow is applicable for a wide range of screening campaigns targeting multistep enzymatic reactions.

  14. Rapid and sensitive serum glucose determination using chemical labeling coupled with black phosphorus-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Wang, Qing; Yu, Lei; Qi, Chu-Bo; Ding, Jun; He, Xiao-Mei; Wang, Ren-Qi; Feng, Yu-Qi

    2018-01-01

    Monitoring the concentration of blood glucose in patients is a key component of good medical diagnoses. Therefore, developing an accurate, rapid and sensitive strategy for monitoring blood glucose is of vital importance. We proposed a strategy for serum glucose determination combining 2-(4-boronobenzyl) isoquinolin-2-ium bromide chemical labeling with black phosphorus assisted laser desorption ionization-time of flight mass spectrometry (CL-BP/ALDI-TOF MS). The entire analytical process consisted of 1min of protein precipitation and 3min of chemical labeling in a microwave oven prior to the BP/ALDI-TOF MS analysis. The analysis can be completed in 5min with high throughput and extremely low sample consumption. Good linearity for glucose was obtained with a correlation coefficient (R) of 0.9986. The limit of detection (LOD) and limit of quantification (LOQ) were 11.5 fmol and 37.5 fmol, respectively. Satisfied reproducibility and reliability were gained by evaluation of the intra- and inter-day precisions with relative standard deviations (RSDs) less than 7.2% and relative recoveries ranging from 87.1% to 108.1%, respectively. The proposed strategy was also applied for the analysis of endogenous glucose in various serum samples and the results were consistent with those obtained using the hexokinase method in a clinical laboratory. Considering the results, the proposed CL-BP/ALDI-TOF MS strategy has proven to be reliable, fast, and sensitive for quantitative analysis of serum glucose. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Diagnostic performance of matrix-assisted laser desorption ionisation time-of-flight mass spectrometry in blood bacterial infections: a systematic review and meta-analysis.

    Science.gov (United States)

    Scott, Jamie S; Sterling, Sarah A; To, Harrison; Seals, Samantha R; Jones, Alan E

    2016-07-01

    Matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF MS) has shown promise in decreasing time to identification of causative organisms compared to traditional methods; however, the utility of MALDI-TOF MS in a heterogeneous clinical setting is uncertain. To perform a systematic review on the operational performance of the Bruker MALDI-TOF MS system and evaluate published cut-off values compared to traditional blood cultures. A comprehensive literature search was performed. Studies were included if they performed direct MALDI-TOF MS analysis of blood culture specimens in human patients with suspected bacterial infections using the Bruker Biotyper software. Sensitivities and specificities of the combined studies were estimated using a hierarchical random effects linear model (REML) incorporating cut-off scores of ≥1.7 and ≥2.0. Fifty publications were identified, with 11 studies included after final review. The estimated sensitivity utilising a cut-off of ≥2.0 from the combined studies was 74.6% (95% CI = 67.9-89.3%), with an estimated specificity of 88.0% (95% CI = 74.8-94.7%). When assessing a cut-off of ≥1.7, the combined sensitivity increases to 92.8% (95% CI = 87.4-96.0%), but the estimated specificity decreased to 81.2% (95% CI = 61.9-96.6%). In this analysis, MALDI-TOF MS showed acceptable sensitivity and specificity in bacterial speciation with the current recommended cut-off point compared to blood cultures; however, lowering the cut-off point from ≥2.0 to ≥1.7 would increase the sensitivity of the test without significant detrimental effect on the specificity, which could improve clinician confidence in their results.

  16. [The research and application of pretreatment method for matrix-assisted laser desorption ionization-time of flight mass spectrometry identification of filamentous fungi].

    Science.gov (United States)

    Huang, Y F; Chang, Z; Bai, J; Zhu, M; Zhang, M X; Wang, M; Zhang, G; Li, X Y; Tong, Y G; Wang, J L; Lu, X X

    2017-08-08

    Objective: To establish and evaluate the feasibility of a pretreatment method for matrix-assisted laser desorption ionization-time of flight mass spectrometry identification of filamentous fungi developed by the laboratory. Methods: Three hundred and eighty strains of filamentous fungi from January 2014 to December 2016 were recovered and cultured on sabouraud dextrose agar (SDA) plate at 28 ℃ to mature state. Meanwhile, the fungi were cultured in liquid sabouraud medium with a vertical rotation method recommended by Bruker and a horizontal vibration method developed by the laboratory until adequate amount of colonies were observed. For the strains cultured with the three methods, protein was extracted with modified magnetic bead-based extraction method for mass spectrum identification. Results: For 380 fungi strains, it took 3-10 d to culture with SDA culture method, and the ratio of identification of the species and genus was 47% and 81%, respectively; it took 5-7 d to culture with vertical rotation method, and the ratio of identification of the species and genus was 76% and 94%, respectively; it took 1-2 d to culture with horizontal vibration method, and the ratio of identification of the species and genus was 96% and 99%, respectively. For the comparison between horizontal vibration method and SDA culture method comparison, the difference was statistically significant (χ(2)=39.026, P method and vertical rotation method recommended by Bruker, the difference was statistically significant(χ(2)=11.310, P method and modified magnetic bead-based extraction method developed by the laboratory is superior to the method recommended by Bruker and SDA culture method in terms of the identification capacity for filamentous fungi, which can be applied in clinic.

  17. A versatile cost-effective method for the analysis of fresh frozen tissue sections via matrix-assisted laser desorption/ionisation imaging mass spectrometry.

    Science.gov (United States)

    O'Rourke, Matthew B; Raymond, Benjamin B A; Djordjevic, Steven P; Padula, Matthew P

    2015-04-15

    There are currently multiple methods available for the preparation of fresh frozen tissue samples for analysis via matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) imaging mass spectrometry (IMS). Although these methods report excellent results, many are expensive automated approaches. With no published attempt to standardise less expensive manual processes, our work aims to provide a robust and repeatable method of sample preparation for MALDI-TOF-IMS that is applicable to a variety of tissue types, well explained, simple and cost effective. Fresh frozen tissue was sectioned at 12 µm and mounted onto liquid nitrocellulose coated slides, washed in a graded alcohol series and then mounted into a modified sublimation apparatus. Matrix is deposited onto the slide to achieve a desired coating of 0.2 mg/cm(2). Once coated, the slide is mounted into a custom-built vapor chamber and recrystallised with 50% acetonitrile (ACN), 0.1% trifluoroacetic acid (TFA) for 1 h at 37°C. The slide is then analysed using MALDI-IMS. We have successfully implemented this method for a host of tissue samples, including brain, liver, kidney and heart, with no variation in relative spectra or processing method required. When the protocol is followed correctly, sublimations and recrystallisations are highly predictable with limited variation between samples and a very low failure rate. Additional apparatuses can be easily constructed by following the included instructions, that perform as per specifications with no variation. We believe that we have described a complete protocol for MALDI-IMS that is easy to use and highly reproducible. The lack of expensive commercially available equipment makes this process very cheap with a relatively low initial outlay and our hope is that more laboratories will begin IMS-based avenues of research based on the work we have performed. Copyright © 2015 John Wiley & Sons, Ltd.

  18. Use of Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry for Identification of Molds of the Fusarium Genus

    Science.gov (United States)

    Stubbe, Dirk; De Cremer, Koen; Piérard, Denis; Normand, Anne-Cécile; Piarroux, Renaud; Detandt, Monique; Hendrickx, Marijke

    2014-01-01

    The rates of infection with Fusarium molds are increasing, and a diverse number of Fusarium spp. belonging to different species complexes can cause infection. Conventional species identification in the clinical laboratory is time-consuming and prone to errors. We therefore evaluated whether matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) is a useful alternative. The 289 Fusarium strains from the Belgian Coordinated Collections of Microorganisms (BCCM)/Institute of Hygiene and Epidemiology Mycology (IHEM) culture collection with validated sequence-based identities and comprising 40 species were used in this study. An identification strategy was developed, applying a standardized MALDI-TOF MS assay and an in-house reference spectrum database. In vitro antifungal testing was performed to assess important differences in susceptibility between clinically relevant species/species complexes. We observed that no incorrect species complex identifications were made by MALDI-TOF MS, and 82.8% of the identifications were correct to the species level. This success rate was increased to 91% by lowering the cutoff for identification. Although the identification of the correct species complex member was not always guaranteed, antifungal susceptibility testing showed that discriminating between Fusarium species complexes can be important for treatment but is not necessarily required between members of a species complex. With this perspective, some Fusarium species complexes with closely related members can be considered as a whole, increasing the success rate of correct identifications to 97%. The application of our user-friendly MALDI-TOF MS identification approach resulted in a dramatic improvement in both time and accuracy compared to identification with the conventional method. A proof of principle of our MALDI-TOF MS approach in the clinical setting using recently isolated Fusarium strains demonstrated its validity. PMID:25411180

  19. Quantitative analysis of an oligomeric hindered amine light stabilizer in polypropylene by matrix-assisted laser desorption/ionization mass spectrometry using a solid sampling technique.

    Science.gov (United States)

    Taguchi, Yoshihiko; Ishida, Yasuyuki; Matsubara, Hideki; Ohtani, Hajime

    2006-01-01

    A small amount of an oligomeric hindered amine light stabilizer (HALS) (Adekastab LA-68LD) in polypropylene (PP) materials was directly determined by solid sampling matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) using an internal standard method. First the matrix reagent (dithranol), 20 mg, and the empirically selected internal standard, angiotensin I (MW = 1296.5), 5 microg, were premixed in the solid state. The matrix mixture was then co-ground with the PP sample containing the HALS in liquid nitrogen using a freezer mill. The powdered sample mixture was spotted on the sample plate, suspended in ion-exchanged water, dried to adhere on the plate, and subjected to MALDI-MS. Three series of the HALS components accompanied by the oxidized species were clearly observed as their molecular ions (M*+)) along with that of the internal standard in the mass spectra. A fairly good linear relationship (R2 = 0.9991) with a relative standard deviation of ca. 11% was observed between the relative peak intensities of the HALS components and the HALS contents ranging from 0.1-2.5 wt%, which could be used as the calibration line to determine the HALS content in PP composites directly by MALDI-MS. The UV-exposed PP composite samples were evaluated by this method to interpret the photostabilizing action of HALS in the PP materials based on the observed change in the relative abundances of the original and oxidized HALS components as a function of UV-exposure time. Copyright 2006 John Wiley & Sons, Ltd.

  20. High-throughput identification of bacteria and yeast by matrix-assisted laser desorption ionization-time of flight mass spectrometry in conventional medical microbiology laboratories.

    Science.gov (United States)

    van Veen, S Q; Claas, E C J; Kuijper, Ed J

    2010-03-01

    Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is suitable for high-throughput and rapid diagnostics at low costs and can be considered an alternative for conventional biochemical and molecular identification systems in a conventional microbiological laboratory. First, we evaluated MALDI-TOF MS using 327 clinical isolates previously cultured from patient materials and identified by conventional techniques (Vitek-II, API, and biochemical tests). Discrepancies were analyzed by molecular analysis of the 16S genes. Of 327 isolates, 95.1% were identified correctly to genus level, and 85.6% were identified to species level by MALDI-TOF MS. Second, we performed a prospective validation study, including 980 clinical isolates of bacteria and yeasts. Overall performance of MALDI-TOF MS was significantly better than conventional biochemical systems for correct species identification (92.2% and 83.1%, respectively) and produced fewer incorrect genus identifications (0.1% and 1.6%, respectively). Correct species identification by MALDI-TOF MS was observed in 97.7% of Enterobacteriaceae, 92% of nonfermentative Gram-negative bacteria, 94.3% of staphylococci, 84.8% of streptococci, 84% of a miscellaneous group (mainly Haemophilus, Actinobacillus, Cardiobacterium, Eikenella, and Kingella [HACEK]), and 85.2% of yeasts. MALDI-TOF MS had significantly better performance than conventional methods for species identification of staphylococci and genus identification of bacteria belonging to HACEK group. Misidentifications by MALDI-TOF MS were clearly associated with an absence of sufficient spectra from suitable reference strains in the MALDI-TOF MS database. We conclude that MALDI-TOF MS can be implemented easily for routine identification of bacteria (except for pneumococci and viridans streptococci) and yeasts in a medical microbiological laboratory.

  1. Accurate Identification of Common Pathogenic Nocardia Species: Evaluation of a Multilocus Sequence Analysis Platform and Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry

    Science.gov (United States)

    Chen, Sharon C-A.; Fan, Xin; Zhang, Li; Li, Hai-Xia; Hou, Xin; Cheng, Jing-Wei; Kong, Fanrong; Zhao, Yu-Pei; Xu, Ying-Chun

    2016-01-01

    Species identification of Nocardia is not straightforward due to rapidly evolving taxonomy, insufficient discriminatory power of conventional phenotypic methods and also of single gene locus analysis including 16S rRNA gene sequencing. Here we evaluated the ability of a 5-locus (16S rRNA, gyrB, secA1, hsp65 and rpoB) multilocus sequence analysis (MLSA) approach as well as that of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) in comparison with sequencing of the 5’-end 606 bp partial 16S rRNA gene to provide identification of 25 clinical isolates of Nocardia. The 5’-end 606 bp 16S rRNA gene sequencing successfully assigned 24 of 25 (96%) clinical isolates to species level, namely Nocardia cyriacigeorgica (n = 12, 48%), N. farcinica (n = 9, 36%), N. abscessus (n = 2, 8%) and N. otitidiscaviarum (n = 1, 4%). MLSA showed concordance with 16S rRNA gene sequencing results for the same 24 isolates. However, MLSA was able to identify the remaining isolate as N. wallacei, and clustered N. cyriacigeorgica into three subgroups. None of the clinical isolates were correctly identified to the species level by MALDI-TOF MS analysis using the manufacturer-provided database. A small “in-house” spectral database was established incorporating spectra of five clinical isolates representing the five species identified in this study. After complementation with the “in-house” database, of the remaining 20 isolates, 19 (95%) were correctly identified to species level (score ≥ 2.00) and one (an N. abscessus strain) to genus level (score ≥ 1.70 and Nocardia. MALDI-TOF MS can provide rapid and accurate identification but is reliant on a robust mass spectra database. PMID:26808813

  2. Trypsin functionalization and zirconia coating of mesoporous silica nanotubes for matrix-assisted laser desorption/ionization mass spectrometry analysis of phosphoprotein.

    Science.gov (United States)

    Zhang, Xiaoli; Wang, Fei; Xia, Yan

    2013-09-06

    Trypsin functionalized mesoporous silica nanotubes bioreactor (TEMSN) and zirconia layer coated mesoporous silica nanotubes (ZrO2-MSN) were developed to deal with the long in-solution digestion time of phosphoprotein and detection difficulty of phosphorylated peptides, respectively. Trypsin was immobilized on the mesoporous silica nanotubes via epoxy group and TEMSN were used as a bioreactor for digestion of α-casein within 3min. ZrO2-MSN were performed to enrich phosphopeptides selectively from in-solution digested peptide mixture of β-casein to demonstrate that ZrO2-MSN possessed remarkable selectivity for phosphorylated peptides even at 100/1 molar ratio of BSA/β-casein. The selective ability of ZrO2-MSN was also investigated in comparison to ZrO2 nanoparticles (ZrO2 NP). Moreover, phosphorylated peptides at the femtomole (2.5fmol) level can also be detected with high S/N (signal-to-noise) ratio. Phosphopeptides enriched from TEMSN-bioreactor digested peptide mixture of α-casein was also performed to evaluate the cooperative performance of TEMSN and ZrO2-MSN platform. The experimental results indicated that TEMSN-bioreactor digestion changed the distribution of relative abundance of phosphopeptides and improved the relative intensity of partial phosphopeptides. This analytical strategy has also been applied to the identification of phosphopeptides isolated from non-fat bovine milk and got a comparable results compared with other materials cited from the literature. By matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF MS), TEMSN and ZrO2-MSN were combined together for the rapid and comprehensive analysis of phosphoprotein. Copyright © 2013 Elsevier B.V. All rights reserved.

  3. Identification and Subtyping of Clinically Relevant Human and Ruminant Mycoplasmas by Use of Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

    Science.gov (United States)

    Renaudin, H.; Cauvin, E.; Del Prá Netto Machado, L.; Tricot, A.; Benoit, F.; Treilles, M.; Bébéar, C.

    2013-01-01

    Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) recently emerged as a technology for the identification of bacteria. In this study, we aimed to evaluate its applicability to human and ruminant mycoplasmal identification, which can be demanding and time-consuming when using phenotypic or molecular methods. In addition, MALDI-TOF MS was tested as a subtyping tool for certain species. A total of 29 main spectra (MSP) from 10 human and 13 ruminant mycoplasma (sub)species were included in a mycoplasma MSP database to complete the Bruker MALDI Biotyper database. After broth culture and protein extraction, MALDI-TOF MS was applied for the identification of 119 human and 143 ruminant clinical isolates that were previously identified by antigenic or molecular methods and for subcultures of 73 ruminant clinical specimens that potentially contained several mycoplasma species. MALDI-TOF MS resulted in accurate (sub)species-level identification with a score of ≥1.700 for 96% (251/262) of the isolates. The phylogenetically closest (sub)species were unequivocally distinguished. Although mixtures of the strains were reliably detected up to a certain cellular ratio, only the predominant species was identified from the cultures of polymicrobial clinical specimens. For typing purposes, MALDI-TOF MS proved to cluster Mycoplasma bovis and Mycoplasma agalactiae isolates by their year of isolation and genome profiles, respectively, and Mycoplasma pneumoniae isolates by their adhesin P1 type. In conclusion, MALDI-TOF MS is a rapid, reliable, and cost-effective method for the routine identification of high-density growing mycoplasmal species and shows promising prospects for its capacity for strain typing. PMID:23903545

  4. Evaluation of matrix-assisted laser desorption ionization-time of flight mass spectrometry for species identification of Acinetobacter strains isolated from blood cultures.

    Science.gov (United States)

    Kishii, K; Kikuchi, K; Matsuda, N; Yoshida, A; Okuzumi, K; Uetera, Y; Yasuhara, H; Moriya, K

    2014-05-01

    The clinical relevance of Acinetobacter species, other than A. baumannii, as human pathogens has not been sufficiently assessed owing to the insufficiency of simple phenotypic clinical diagnostic laboratory tests. Infections caused by these organisms have different impacts on clinical outcome and require different treatment and management approaches. It is therefore important to correctly identify Acinetobacter species. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has been introduced to identify a wide range of microorganisms in clinical laboratories, but only a few studies have examined its utility for identifying Acinetobacter species, particularly those of the non-Acinetobacter baumannii complex. We therefore evaluated MALDI-TOF MS for identification of Acinetobacter species by comparing it with sequence analysis of rpoB using 123 isolates of Acinetobacter species from blood. Of the isolates examined, we identified 106/123 (86.2%) to species, and 16/123 (13.0%) could only be identified as acinetobacters. The identity of one isolate could not be established. Of the 106 species identified, 89/106 (84.0%) were confirmed by rpoB sequence analysis, and 17/106 (16.0%) were discordant. These data indicate correct identification of 89/123 (72.4%) isolates. Surprisingly, all blood culture isolates were identified as 13 species of Acinetobacter, and the incidence of Acinetobacter pittii was unexpectedly high (42/123; 34.1%) and exceeded that of A. baumannii (22/123; 17.9%). Although the present identification rate using MALDI-TOF MS is not acceptable for species-level identification of Acinetobacter, further expansion of the database should remedy this situation. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  5. Imaging Mass Spectrometry by Matrix-Assisted Laser Desorption/Ionization and Stress-Strain Measurements in Iontophoresis Transepithelial Corneal Collagen Cross-Linking

    Directory of Open Access Journals (Sweden)

    Paolo Vinciguerra

    2014-01-01

    Full Text Available Purpose. To compare biomechanical effect, riboflavin penetration and distribution in transepithelial corneal collagen cross-linking with iontophoresis (I-CXL, with standard cross linking (S-CXL and current transepithelial protocol (TE-CXL. Materials and Methods. The study was divided into two different sections, considering, respectively, rabbit and human cadaver corneas. In both sections corneas were divided according to imbibition protocols and irradiation power. Imaging mass spectrometry by matrix-assisted laser desorption/ionization (MALDI-IMS and stress-strain measurements were used. Forty-eight rabbit and twelve human cadaver corneas were evaluated. Results. MALDI-IMS showed a deep riboflavin penetration throughout the corneal layers with I-CXL, with a roughly lower concentration in the deepest layers when compared to S-CXL, whereas with TE-CXL penetration was considerably less. In rabbits, there was a significant increase (by 71.9% and P=0.05 in corneal rigidity after I-CXL, when compared to controls. In humans, corneal rigidity increase was not significantly different among the subgroups. Conclusions. In rabbits, I-CXL induced a significant increase in corneal stiffness as well as better riboflavin penetration when compared to controls and TE-CXL but not to S-CXL. Stress-strain in human corneas did not show significant differences among techniques, possibly because of the small sample size of groups. In conclusion, I-CXL could be a valid alternative to S-CXL for riboflavin delivery in CXL, preserving the epithelium.

  6. Application and evaluation of solvent-free matrix-assisted laser desorption/ionization mass spectrometry for the analysis of derivatized fullerenes.

    Science.gov (United States)

    Kotsiris, Sotirios G; Vasil'ev, Yury V; Streletskii, Alexey V; Han, Ming; Mark, Lewis P; Boltalina, Olga V; Chronakis, Nikos; Orfanopoulos, Michael; Hungerbühler, Hartmut; Drewello, Thomas

    2006-01-01

    A variety of derivatized fullerenes have been studied by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. Of particular emphasis has been the evaluation of a recently introduced solvent-free sample/target preparation method. Solvent-free MALDI is particularly valuable in overcoming adverse solvent-related effects, such as insolubility and/or degradation of the sample. The method was applied to fullerene derivatives susceptible to decomposition under insufficiently "soft" MALDI conditions. Analytes included the hydrofullerene: C(60)H(36), fluorofullerenes: C(60)F(x) where x = 18, 36, 46, 48 and C(70)F(x) where x = 54, 56, methano-bridged amphiphilic ligand adducts to C(60) and the [4 + 2] cycloadduct of tetracene to C(60). The new solvent-free sample preparation is established as an exceedingly valuable addition to the repertoire of preparation protocols within MALDI. The MALDI mass spectra were of very high quality throughout, providing a testimony that "soft" MALDI conditions could be achieved. Using the [4 + 2] cycloadduct of tetracene to C(60) as the model analyte for direct comparison with solvent-based MALDI, the solvent-free approach led to less fragmentation and more abundant analyte ions. Applying solvent-free sample preparation, different matrix compounds have been examined for use in the MALDI of derivatized fullerenes, including sulfur, tetracyanoquinodimethane (TCNQ), 9-nitroanthracene (9-NA) and trans-2-[3-(4-tert-butylphenyl)-2-methyl-2- propenylidene]malononitrile (DCTB). DCTB was confirmed as the best performing matrix, reducing unwanted decomposition and suppression effects.

  7. Evaluation of the Vitek MS Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry System for Identification of Clinically Relevant Filamentous Fungi

    Science.gov (United States)

    McMullen, Allison R.; Wallace, Meghan A.; Pincus, David H.; Wilkey, Kathy

    2016-01-01

    Invasive fungal infections have a high rate of morbidity and mortality, and accurate identification is necessary to guide appropriate antifungal therapy. With the increasing incidence of invasive disease attributed to filamentous fungi, rapid and accurate species-level identification of these pathogens is necessary. Traditional methods for identification of filamentous fungi can be slow and may lack resolution. Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) has emerged as a rapid and accurate method for identification of bacteria and yeasts, but a paucity of data exists on the performance characteristics of this method for identification of filamentous fungi. The objective of our study was to evaluate the accuracy of the Vitek MS for mold identification. A total of 319 mold isolates representing 43 genera recovered from clinical specimens were evaluated. Of these isolates, 213 (66.8%) were correctly identified using the Vitek MS Knowledge Base, version 3.0 database. When a modified SARAMIS (Spectral Archive and Microbial Identification System) database was used to augment the version 3.0 Knowledge Base, 245 (76.8%) isolates were correctly identified. Unidentified isolates were subcultured for repeat testing; 71/319 (22.3%) remained unidentified. Of the unidentified isolates, 69 were not in the database. Only 3 (0.9%) isolates were misidentified by MALDI-TOF MS (including Aspergillus amoenus [n = 2] and Aspergillus calidoustus [n = 1]) although 10 (3.1%) of the original phenotypic identifications were not correct. In addition, this methodology was able to accurately identify 133/144 (93.6%) Aspergillus sp. isolates to the species level. MALDI-TOF MS has the potential to expedite mold identification, and misidentifications are rare. PMID:27225405

  8. Evaluation of the Vitek MS Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry System for Identification of Clinically Relevant Filamentous Fungi.

    Science.gov (United States)

    McMullen, Allison R; Wallace, Meghan A; Pincus, David H; Wilkey, Kathy; Burnham, C A

    2016-08-01

    Invasive fungal infections have a high rate of morbidity and mortality, and accurate identification is necessary to guide appropriate antifungal therapy. With the increasing incidence of invasive disease attributed to filamentous fungi, rapid and accurate species-level identification of these pathogens is necessary. Traditional methods for identification of filamentous fungi can be slow and may lack resolution. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has emerged as a rapid and accurate method for identification of bacteria and yeasts, but a paucity of data exists on the performance characteristics of this method for identification of filamentous fungi. The objective of our study was to evaluate the accuracy of the Vitek MS for mold identification. A total of 319 mold isolates representing 43 genera recovered from clinical specimens were evaluated. Of these isolates, 213 (66.8%) were correctly identified using the Vitek MS Knowledge Base, version 3.0 database. When a modified SARAMIS (Spectral Archive and Microbial Identification System) database was used to augment the version 3.0 Knowledge Base, 245 (76.8%) isolates were correctly identified. Unidentified isolates were subcultured for repeat testing; 71/319 (22.3%) remained unidentified. Of the unidentified isolates, 69 were not in the database. Only 3 (0.9%) isolates were misidentified by MALDI-TOF MS (including Aspergillus amoenus [n = 2] and Aspergillus calidoustus [n = 1]) although 10 (3.1%) of the original phenotypic identifications were not correct. In addition, this methodology was able to accurately identify 133/144 (93.6%) Aspergillus sp. isolates to the species level. MALDI-TOF MS has the potential to expedite mold identification, and misidentifications are rare. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  9. Direct Identification of Urinary Tract Pathogens From Urine Samples Using the Vitek MS System Based on Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry.

    Science.gov (United States)

    Kim, Yeongsic; Park, Kang Gyun; Lee, Kyungwon; Park, Yeon-Joon

    2015-07-01

    We evaluated the coincidence rate between Vitek MS system (bioMérieux, France) and Vitek 2 in identifying uropathogens directly from urine specimens. Urine specimens submitted to our microbiology laboratory between July and September 2013 for Gram staining and bacterial culture were analyzed. Bacterial identification was performed by using the conventional method. Urine specimens showing a single morphotype by Gram staining were processed by culturing and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Of 2,370 urine specimens, 251 showed a single morphotype on Gram staining, and among them, 202 were available for MALDI-TOF MS. In these 202 specimens, colony growth was observed in 189 specimens, and 145 specimens had significant growth of single-colony morphotype in culture. One hundred and ten (75.9%) of them had colony counts of ≥10(5) colony-forming units (CFU)/mL and included 71 enteric gram-negative bacteria (GNB), 5 glucose-non-fermenting GNB, 9 gram-positive cocci (GPC), and 25 yeasts. Furthermore, 70 (98.6%), 3 (60.0%), 4 (44.4%), and 5 (20.0%), respectively, of these were correctly identified by Vitek MS. Thirty-one specimens (21.4%; 11 GNB, 7 GPC, 12 yeasts, and 1 gram-positive bacillus) had colony counts of 10(4)-10(5) CFU/mL. Four specimens (2.8%) yielded colony counts of 10(3)-10(4) CFU/mL. Vitek MS showed high rate of accuracy for the identification of GNB in urine specimens (≥10(5) CFU/mL). This could become a rapid and accurate diagnostic method for urinary tract infection caused by GNB. However, for the identification of GPC and yeasts, further studies on appropriate pre-treatment are warranted.

  10. Direct identification of pathogens from positive blood cultures using matrix-assisted laser desorption-ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Rodríguez-Sánchez, B; Sánchez-Carrillo, C; Ruiz, A; Marín, M; Cercenado, E; Rodríguez-Créixems, M; Bouza, E

    2014-07-01

    In recent years, matrix-assisted laser desorption-ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) has proved a rapid and reliable method for the identification of bacteria and yeasts that have already been isolated. The objective of this study was to evaluate this technology as a routine method for the identification of microorganisms directly from blood culture bottles (BCBs), before isolation, in a large collection of samples. For this purpose, 1000 positive BCBs containing 1085 microorganisms have been analysed by conventional phenotypic methods and by MALDI-TOF MS. Discrepancies have been resolved using molecular methods: the amplification and sequencing of the 16S rRNA gene or the Superoxide Dismutase gene (sodA) for streptococcal isolates. MALDI-TOF predicted a species- or genus-level identification of 81.4% of the analysed microorganisms. The analysis by episode yielded a complete identification of 814 out of 1000 analysed episodes (81.4%). MALDI-TOF identification is available for clinicians within hours of a working shift, as oppose to 18 h later when conventional identification methods are performed. Moreover, although further improvement of sample preparation for polymicrobial BCBs is required, the identification of more than one pathogen in the same BCB provides a valuable indication of unexpected pathogens when their presence may remain undetected in Gram staining. Implementation of MALDI-TOF identification directly from the BCB provides a rapid and reliable identification of the causal pathogen within hours. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  11. Coumarins as new matrices for matrix-assisted laser-desorption/ionization Fourier transform ion cyclotron resonance mass spectrometric analysis of hydrophobic compounds

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Hang, E-mail: hangwang@sjtu.edu.cn [Instrumental Analysis Center, Shanghai Jiao Tong University, Dongchuan Road 800, Shanghai 200240 (China); Dai, Bona [Instrumental Analysis Center, Shanghai Jiao Tong University, Dongchuan Road 800, Shanghai 200240 (China); Liu, Bin [Key Laboratory of Kidney Disease Pathogenesis and Intervention of Hubei Province, College of Medicine, Hubei Polytechnic University, Huangshi, Hubei 435003 (China); Lu, Han [Department of Anesthesiology, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine (SJTU-SM), 197, Rui Jin Er Road, Shanghai 200025 (China)

    2015-07-02

    Highlights: • Coumarins were used as new MALDI matrices. • Coumarins were used for MALDI-FT ICR MS detection of hydrophobic compounds. • DCA had improvement in detection sensitivity, stability, selectivity and reproducibility. • DCA was applied to sterols detection in yeast cells. - Abstract: Hydrophobic compounds with hydroxyl, aldehyde or ketone groups are generally difficult to detect using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), because these compounds have low proton affinity and are poorly ionized by MALDI. Herein, coumarins have been used as new matrices for MALDI-MS analysis of a variety of hydrophobic compounds with low ionization efficiency, including steroids, coenzyme Q10, a cyclic lipopeptide and cholesterol oleate. Five coumarins, including coumarin, umbelliferone, esculetin, 7-hydroxycoumarin-3-carboxylic acid (HCA) and 6,7-dihydroxycoumarin-3-carboxylic acid (DCA), were compared with the conventional matrices of 2,5-dihydroxybenzoic acid (DHB) and α-cyano-4-hydroxycinnamic acid (CHCA). Coumarins with hydroxyl or carboxylic acid groups enabled detection. Taking DCA as an example, this matrix proved to be superior to DHB or CHCA in detection sensitivity, stability, spot-to-spot and sample-to-sample reproducibility, and accuracy. DCA increased the stability of the target compounds and decreased the loss of water. The [M + Na]{sup +} peaks were observed for all target compounds by adding NaCl as an additive, and the [M − H{sub 2}O + H]{sup +} and [M + H]{sup +} peaks decreased. DCA was selected for the identification of sterols in yeast cells, and thirteen sterols were detected by Fourier transform ion cyclotron resonance (FT ICR) mass spectrometry. This work demonstrates the potential of DCA as a new matrix for detection of hydrophobic molecules by MALDI-MS and provides an alternative tool for screening sterols in antifungal research.

  12. Reliable and reproducible method for rapid identification of Nocardia species by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Toyokawa, Masahiro; Kimura, Keigo; Nishi, Isao; Sunada, Atsuko; Ueda, Akiko; Sakata, Tomomi; Asari, Seishi

    2013-01-01

    Recently, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been challenged for the identification of Nocardia species. However, the standard ethanol-formic acid extraction alone is insufficient in allowing the membrane proteins of Nocardia species to be ionized by the matrix. We therefore aimed to establish our new extraction method for the MALDI-TOF MS-based identification of Nocardia species isolates. Our modified extraction procedure is through dissociation in 0.5% Tween-20 followed by bacterial heat-inactivation, mechanical breaking of the cell wall by acid-washed glass beads and protein extraction with formic acid and acetonitrile. As reference methods for species identification, full-length 16S rRNA gene sequencing and some phenotypical tests were used. In a first step, we made our own Nocardia database by analyzing 13 strains (13 different species including N. elegans, N. otitidiscaviarum, N. asiatica, N. abscessus, N. brasiliensis, N. thailandica, N. farcinica, N. nova, N. mikamii, N. cyriacigeorgica, N. asteroids, Nocardiopsis alba, and Micromonospora sp.) and registered to the MALDI BioTyper database. Then we established our database. The analysis of 12 challenge strains using the our database gave a 100% correct identification, including 8 strains identified to the species level and 4 strains to the genus level (N. elegans, N. nova, N. farcinica, Micromonospora sp.) according to the manufacture's log score specifications. In the estimation of reproducibility of our method intended for 4 strains, both within-run and between-run reproducibility were excellent. These data indicates that our method for rapid identification of Nocardia species is with reliability, reproducibility and cost effective.

  13. Application of flowerlike MgO for highly sensitive determination of lead via matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Hou, Jian; Chen, Suming; Cao, Changyan; Liu, Huihui; Xiong, Caiqiao; Zhang, Ning; He, Qing; Song, Weiguo; Nie, Zongxiu

    2016-08-01

    Matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) is a high-throughput method to achieve fast and accurate identification of lead (Pb) exposure, but is seldom used because of low ionization efficiency and insufficient sensitivity. Nanomaterials applied in MS are a promising technique to overcome the obstacles of MALDI. Flowerlike MgO nanostructures are applied for highly sensitive lead profiling in real samples. They can be used in two ways: (a) MgO is mixed with N-naphthylethylenediamine dihydrochloride (NEDC) as a novel matrix MgO/NEDC; (b) MgO is applied as an absorbent to enrich Pb ions in very dilute solution. The signal intensities of lead by MgO/NEDC were ten times higher than the NEDC matrix. It also shows superior anti-interference ability when analyzing 10 μmol/L Pb ions in the presence of organic substances or interfering metal ions. By applying MgO as adsorbent, the LOD of lead before enrichment is 1 nmol/L. Blood lead test can be achieved using this enrichment process. Besides, MgO can play the role of internal standard to achieve quantitative analysis. Flowerlike MgO nanostructures were applied for highly sensitive lead profiling in real samples. The method is helpful to prevent Pb contamination in a wide range. Further, the combination of MgO with MALDI MS could inspire more nanomaterials being applied in highly sensitive profiling of pollutants. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  14. Accurate Identification of Common Pathogenic Nocardia Species: Evaluation of a Multilocus Sequence Analysis Platform and Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry.

    Science.gov (United States)

    Xiao, Meng; Pang, Lu; Chen, Sharon C-A; Fan, Xin; Zhang, Li; Li, Hai-Xia; Hou, Xin; Cheng, Jing-Wei; Kong, Fanrong; Zhao, Yu-Pei; Xu, Ying-Chun

    2016-01-01

    Species identification of Nocardia is not straightforward due to rapidly evolving taxonomy, insufficient discriminatory power of conventional phenotypic methods and also of single gene locus analysis including 16S rRNA gene sequencing. Here we evaluated the ability of a 5-locus (16S rRNA, gyrB, secA1, hsp65 and rpoB) multilocus sequence analysis (MLSA) approach as well as that of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) in comparison with sequencing of the 5'-end 606 bp partial 16S rRNA gene to provide identification of 25 clinical isolates of Nocardia. The 5'-end 606 bp 16S rRNA gene sequencing successfully assigned 24 of 25 (96%) clinical isolates to species level, namely Nocardia cyriacigeorgica (n = 12, 48%), N. farcinica (n = 9, 36%), N. abscessus (n = 2, 8%) and N. otitidiscaviarum (n = 1, 4%). MLSA showed concordance with 16S rRNA gene sequencing results for the same 24 isolates. However, MLSA was able to identify the remaining isolate as N. wallacei, and clustered N. cyriacigeorgica into three subgroups. None of the clinical isolates were correctly identified to the species level by MALDI-TOF MS analysis using the manufacturer-provided database. A small "in-house" spectral database was established incorporating spectra of five clinical isolates representing the five species identified in this study. After complementation with the "in-house" database, of the remaining 20 isolates, 19 (95%) were correctly identified to species level (score ≥ 2.00) and one (an N. abscessus strain) to genus level (score ≥ 1.70 and Nocardia. MALDI-TOF MS can provide rapid and accurate identification but is reliant on a robust mass spectra database.

  15. Matrix-assisted laser desorption/ionization mass spectrometry imaging of cell cultures for the lipidomic analysis of potential lipid markers in human breast cancer invasion.

    Science.gov (United States)

    Wang, Shujuan; Chen, Xiaowu; Luan, Hemi; Gao, Dan; Lin, Shuhai; Cai, Zongwei; Liu, Jianjun; Liu, Hongxia; Jiang, Yuyang

    2016-02-28

    Breast cancer is the leading cause of cancer death among women worldwide. Identification of lipid targets that play a role in breast cancer invasion may advance our understanding of the rapid progression of cancer and may lead to the development of new biomarkers for the disease. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) was applied for the lipidomic profiling of two poorly invasive and two highly invasive breast cancer cell lines to identify the differentially accumulated lipids related to the invasive phenotype. The four cell lines were individually grown on indium tin oxide (ITO)-coated glass slides, analyzed as cell cultures. The raster width and matrix for detection were optimized to improve detection sensitivity. Optimized MSI measurements were performed directly on the cell culture with 9-aminoacridine as matrix, resulting in 215 endogenous compounds detected in positive ion mode and 267 endogenous compounds in negative ion mode in all the four cell lines, representing the largest group of analytes that have been analyzed from cells by a single MSI study. In highly invasive cell lines, 31 lipids including phosphatidylglycerol (PG) and phosphatidic acids were found upregulated and eight lipids including sphingomyelin (SM) downregulated in negative ion mode. The products of de novo fatty acid synthesis incorporated into membrane phospholipids, like oleic-acid-containing PG, may be involved in mitochondrial dysfunction and thus affect the invasion of breast cancer cells. The deficiency of SM may be related to the disruption of apoptosis in highly invasive cancer cells. This work uncovered more analytes in cells by MSI than previous reports, providing a better visualization and novel insights to advance our understanding of the relationship between rapid progression of breast cancer and lipid metabolism. The most altered lipids may aid the discovery of diagnostic markers and therapeutic targets of breast cancer. Copyright

  16. Routine identification of microorganisms by matrix-assisted laser desorption ionization time-of-flight mass spectrometry: Success rate, economic analysis, and clinical outcome.

    Science.gov (United States)

    Ge, Mao-Cheng; Kuo, An-Jing; Liu, Kuei-Lan; Wen, Ying-Hao; Chia, Ju-Hsin; Chang, Pi-Yueh; Lee, Ming-Hsun; Wu, Tsu-Lan; Chang, Shih-Cheng; Lu, Jang-Jih

    2017-10-01

    Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been widely used in microbial identification. This study evaluated the performance of MALDI-TOF MS and investigated the economic and medical impact of MALDI-TOF MS implementation. A total of 12,202 clinical isolates collected from April to September 2013 were identified using MALDI-TOF MS, and the success rates in identifying isolates were analyzed. The differences in the processing time, cost of consumables, weight of waste, and clinical impact between MALDI-TOF MS and biochemical reaction were compared. MALDI-TOF MS successfully identified 96% of 12,202 isolates, including 96.8% of 10,502 aerobes, 90.5% of 1481 anaerobes, 93.8% of 81 yeasts, and 90.6% of 138 nontuberculous mycobacteria at the genus level. By using MALDI-TOF MS, the processing time for aerobes decreased from 32.5 hours to 4.1 hours, and that for anaerobes decreased from 71.5 hours to 46 hours. For detection of aerobes and anaerobes, the cost of consumables was estimated to decrease by US$0.9 per isolate, thus saving US$94,500 in total annual isolation. Furthermore, the weight of waste decreased six-fold, resulting in a reduction of 350 kg/month or 4.2 tons/year. MALDI-TOF MS also increased the percentage of correct antibiotics treatment for Escherichia coli and Klebsiella pneumonia from 56.1% to 75% and shortened the initiation time of the correct antibiotic action from 3.3 hours to 2.5 hours. MALDI-TOF MS is a rapid, reliable, economical, and environmentally friendly method for routine microbial identification and may contribute to early appropriate antibiotic treatment in clinical settings. Copyright © 2016. Published by Elsevier B.V.

  17. Matrix-assisted laser desorption/ionisation-time of flight mass spectrometry: rapid identification of bacteria isolated from patients with cystic fibrosis.

    Science.gov (United States)

    Baillie, S; Ireland, K; Warwick, S; Wareham, D; Wilks, M

    2013-01-01

    Despite extensive research into the diagnosis and management of cystic fibrosis (CF) over the past decades, sufferers still have a median life expectancy of less than 37 years. Respiratory tract infections have a significant role in increasing the morbidity and mortality of patients with CF via a progressive decline in lung function. Rapid identification of organisms recovered from CF sputum is necessary for effective management of respiratory tract infections; however, standard techniques of identification are slow, technically demanding and expensive. The aim of this study is to asses the suitability of matrix-assisted laser desorption/ionisation-time of flight mass spectrometry (MALDI-TOF MS) in identifying bacteria isolated from the respiratory tract of patients with CF, and is assessed by testing the accuracy of MALDI-TOF MS in identifying samples from a reference collection of rare CF strains in conjunction with comparing MALDI-TOF MS and standard techniques in identifying clinical isolates from sputum samples of CF patients. MALDI-TOF MS accurately identified 100% of isolates from the reference collection of rare CF pathogens (EuroCare CF collection). The isolate identification given by MALDI-TOF MS agreed with that given by standard techniques for 479/481 (99.6%) clinical isolates obtained from respiratory samples provided by patients with CE In two (0.4%) of 481 samples there was a discrepancy in identification between MALDI-TOF MS and standard techniques. One organism was identified as Pseudomonas aeruginosa by MALDI-TOF but could only be identified by the laboratory's standard methods as of the Pseudomonas genus. The second organism was identified as P. beteli by MALDI-TOF MS and Stenotrophomonas maltophilia by standard methods. This study shows that MALDI-TOF MS is superior to standard techniques in providing cheap, rapid and accurate identification of CF sputum isolates.

  18. Direct bacterial identification in positive blood cultures by use of two commercial matrix-assisted laser desorption ionization-time of flight mass spectrometry systems.

    Science.gov (United States)

    Chen, Jonathan H K; Ho, Pak-Leung; Kwan, Grace S W; She, Kevin K K; Siu, Gilman K H; Cheng, Vincent C C; Yuen, Kwok-Yung; Yam, Wing-Cheong

    2013-06-01

    Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for the identification of bacteria and fungi was recently introduced in microbiology laboratories. This technology could greatly improve the clinical management of patients and guidance for chemotherapy. In this study, we used a commercial MALDI Sepsityper extraction method to evaluate the performance of two commercial MALDI-TOF MS systems, the Vitek MS IVD (bioMérieux) and the Microflex LT Biotyper (Bruker Daltonics) for direct bacterial identification in positive blood cultures. In 181 monomicrobial cultures, both systems generated genus to species level identifications for >90% of the specimens (Biotyper, 177/181 [97.8%]; Vitek MS IVD, 167/181 [92.3%]). Overall, the Biotyper system generated significantly more accurate identifications than the Vitek MS IVD system (P = 0.016; 177 versus 167 out of 181 specimens). The Biotyper system identified the minority species among polymicrobial blood cultures. We also compared the performance of an in-house extraction method with that of the Sepsityper on both MALDI-TOF MS systems. The in-house method generated more correct identifications at the genus level than the Sepsityper (96.7% versus 93.5%) on the Biotyper system, whereas the two methods exhibited the same performance level (88.0% versus 88.0%) on the Vitek MS IVD system. Our study confirmed the practical advantages of MALDI-TOF MS, and our in-house extraction method reduced the reagent cost to $1 per specimen, with a shorter turnaround time of 3 h, which is highly cost-effective for a diagnostic microbiology service.

  19. Efficient Detection of Carbapenemase Activity in Enterobacteriaceae by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry in Less Than 30 Minutes.

    Science.gov (United States)

    Lasserre, Camille; De Saint Martin, Luc; Cuzon, Gaelle; Bogaerts, Pierre; Lamar, Estelle; Glupczynski, Youri; Naas, Thierry; Tandé, Didier

    2015-07-01

    The recognition of carbapenemase-producing Enterobacteriaceae (CPE) isolates is a major laboratory challenge, and their inappropriate or delayed detection may have negative impacts on patient management and on the implementation of infection control measures. We describe here a matrix-assisted laser desorption ionization-time of flight (MALDI-TOF)-based method to detect carbapenemase activity in Enterobacteriaceae. After a 20-min incubation of the isolate with 0.5 mg/ml imipenem at 37°C, supernatants were analyzed by MALDI-TOF in order to identify peaks corresponding to imipenem (300 Da) and an imipenem metabolite (254 Da). A total of 223 strains, 77 CPE (OXA-48 variants, KPC, NDM, VIM, IMI, IMP, and NMC-A) and 146 non-CPE (cephalosporinases, extended-spectrum β-lactamases [ESBLs], and porin defects), were tested and used to calculate a ratio of imipenem hydrolysis: mass spectrometry [MS] ratio = metabolite/(imipenem + metabolite). An MS ratio cutoff was statistically determined to classify strains as carbapenemase producers (MS ratio of ≥0.82). We validated this method first by testing 30 of our 223 isolates (15 CPE and 15 non-CPE) 10 times to calculate an intraclass correlation coefficient (ICC of 0.98), showing the excellent repeatability of the method. Second, 43 strains (25 CPE and 18 non-CPE) different from the 223 strains used to calculate the ratio cutoff were used as external controls and blind tested. They yielded sensitivity and specificity of 100%. The total cost per test is time-saving, cost-efficient, and highly reliable and might be used in any routine laboratory, given the availability of mass spectrometry, to detect CPE. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  20. Matrix-assisted laser desorption ionization time of flight mass spectrometry and diagnostic testing for prosthetic joint infection in the clinical microbiology laboratory.

    Science.gov (United States)

    Peel, Trisha N; Cole, Nicolynn C; Dylla, Brenda L; Patel, Robin

    2015-03-01

    Identification of pathogen(s) associated with prosthetic joint infection (PJI) is critical for patient management. Historically, many laboratories have not routinely identified organisms such as coagulase-negative staphylococci to the species level. The advent of matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) has enhanced clinical laboratory capacity for accurate species-level identification. The aim of this study was to describe the species-level identification of microorganisms isolated from periprosthetic tissue and fluid specimens using MALDI-TOF MS alongside other rapid identification tests in a clinical microbiology laboratory. Results of rapid identification of bacteria isolated from periprosthetic joint fluid and/or tissue specimens were correlated with clinical findings at Mayo Clinic, Rochester, Minnesota, between May 2012 and May 2013. There were 178 PJI and 82 aseptic failure (AF) cases analyzed, yielding 770 organisms (median, 3/subject; range, 1-19/subject). MALDI-TOF MS was employed for the identification of 455 organisms (59%) in 197 subjects (123 PJIs and 74 AFs), with 89% identified to the species level using this technique. Gram-positive bacteria accounted for 68% and 93% of isolates in PJI and AF, respectively. However, the profile of species associated with infection compared to specimen contamination differed. Staphylococcus aureus and Staphylococcus caprae were always associated with infection, Staphylococcus epidermidis and Staphylococcus lugdunensis were equally likely to be a pathogen or a contaminant, whereas the other coagulase-negative staphylococci were more frequently contaminants. Most streptococcal and Corynebacterium isolates were pathogens. The likelihood that an organism was a pathogen or contaminant differed with the prosthetic joint location, particularly in the case of Propionibacterium acnes. MALDI-TOF MS is a valuable tool for the identification of bacteria isolated from patients

  1. Impact of rapid microbial identification directly from positive blood cultures using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry on patient management.

    Science.gov (United States)

    Martiny, D; Debaugnies, F; Gateff, D; Gérard, M; Aoun, M; Martin, C; Konopnicki, D; Loizidou, A; Georgala, A; Hainaut, M; Chantrenne, M; Dediste, A; Vandenberg, O; Van Praet, S

    2013-12-01

    For septic patients, delaying the initiation of antimicrobial therapy or choosing an inappropriate antibiotic can considerably worsen their prognosis. This study evaluated the impact of rapid microbial identification (RMI) from positive blood cultures on the management of patients with suspected sepsis. During a 6-month period, RMI by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was performed for all new episodes of bacteraemia. For each patient, the infectious disease specialist was contacted and questioned about his therapeutic decisions made based on the Gram staining and the RMI. This information was collected to evaluate the number of RMIs that led to a therapeutic change or to a modification of the patient's general management (e.g. fast removal of infected catheters). During the study period, 277 new episodes of bacteraemia were recorded. In 71.12% of the cases, MALDI-TOF MS resulted in a successful RMI (197/277). For adult and paediatric patients, 13.38% (21/157) and 2.50% (1/40) of the RMIs, respectively, resulted in modification of the treatment regimen, according to the survey. In many other cases, the MALDI-TOF MS was a helpful tool for infectious disease specialists because it confirmed suspected cases of contamination, especially in the paediatric population (15/40 RMIs, 37.50%), or suggested complementary diagnostic testing. This study emphasizes the benefits of RMI from positive blood cultures. Although the use of this technique represents an extra cost for the laboratory, RMI using MALDI-TOF MS has been implemented in our daily practice. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  2. Preparation of various lysogangliosides including lyso-fucosyl GM1 and delayed extraction matrix-assisted laser desorption ionization time-of-flight mass spectrometric analysis.

    Science.gov (United States)

    Taketomi, T; Hara, A; Uemura, K; Kurahashi, H; Sugiyama, E

    1997-02-01

    Our rapid method of microwave-mediated saponification for preparing lysoglycosphingolipids from their parent glycosphingolipids was also able to prepare lysogangliosides or modified lysogangliosides, which were identified by delayed extraction matrix-assisted laser desorption ionization time-of-flight mass spectrometric (DE MALDI-TOF MS) analysis. When GM3, GM2, and GM1 isolated from adult human brain gangliosides were subjected to the saponification, GM3 was found to give rise to only lyso-GM3 containing de-N-acetylneuraminic acid (de-N-acetyl lyso-GM3), whereas the GM2 produced both lyso-GM2 and the de-N-acetyl compound, and GM1 also gave both lyso-GM1 and the de-N-acetyl compound. In the saponification of GM1 and GDla, isolated from rat brain gangliosides, GM1 similarly produced both lyso-GM1 and the de-N-acetyl compound, but GDla was found to give rise to both dehydrated de-N-monoacetyl and dehydrated de-N-diacetyl lyso-GDla. However, the saponification of the GM1 fraction isolated from porcine brain gangliosides gave rise not only to both lyso-GM1 and the de-N-acetyl compound, but also unexpectedly to both lyso-fucosyl GM1 and its de-N-acetyl compound. The untreated GM1 fraction was examined by TLC and DE MALDI-TOF mass spectrometry, and proved to contain fucosyl-GM1. The DE MALDI-TOF MS analysis of the prepared lyso-gangliosides showed that their long chain bases consisted of d18:1 and d20:1 sphingosines in various ratios reflecting those of the different mammalian brain gangliosides.

  3. Application of matrix-assisted laser desorption ionization time-of-flight mass spectrometry with delayed ion extraction to ganglioside analyses.

    Science.gov (United States)

    Sugiyama, E; Hara, A; Uemura, K; Taketomi, T

    1997-07-01

    Various monosialo- and disialo-gangliosides and their derivatives were examined by delayed ion extraction matrix-assisted laser desorption ionization time-of-flight mass spectrometry (DE MALDI-TOF MS) in the reflector mode with alpha-cyano-4-hydroxycinnamic acid or 2,5-dihydroxybenzoic acid used as the matrix. Native gangliosides were generally found to give good spectra in the negative ion mode. 2,5-Dihydroxybenzoic acid was a better matrix for gangliosides than alpha-cyano-4-hydroxycinnamic acid, because this matrix seemed to minimize loss of sialic acid and carbon dioxide of gangliosides. About 1 pmol of ganglioside was able to be detected with this matrix. When "A-series" gangliosides such as GD1a and GalNAc-GD1a gave undesirable extra peaks probably due to loss of sialic acid besides molecule-related ion peaks, the methyl-esterification of the gangliosides at the carboxyl groups of sialic acids was found to be necessary to obtain good DE MALDI-TOF mass spectra in the positive ion mode. In contrast, "B-series" gangliosides such as GD1b, GD2, and GD3 gave rise to major dehydrated molecule-related ion [M-H2O-H]- peaks in the negative ion mode without the pretreatment of methyl-esterification. The DE MALDI-TOF mass spectrometric analysis enabled us to distinguish between GD1a and GD1b, which have the same molecular weight. It was also found that not only a purified sample, but also a mixed sample of various gangliosides was amenable to the identification of them by DE MALDI-TOF MS.

  4. Comparison of Vitek Matrix-assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry Versus Conventional Methods in Candida Identification.

    Science.gov (United States)

    Keçeli, Sema Aşkın; Dündar, Devrim; Tamer, Gülden Sönmez

    2016-02-01

    Candida species are generally identified by conventional methods such as germ tube or morphological appearance on corn meal agar, biochemical methods using API kits and molecular biological methods. Alternative to these methods, rapid and accurate identification methods of microorganisms called matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDİ-TOF MS) has recently been described. In this study, Candida identification results by API Candida kit, API 20C AUX kit and identifications on corn meal agar (CMA) are compared with the results obtained on Vitek-MS. All results were confirmed by sequencing internal transcribed spacer (ITS) regions of rDNA. Totally, 97 Candida strains were identified by germ tube test, CMA, API and Vitek-MS. Vitek-MS results were compatible with 74.2 % of API 20C AUX and 81.4 % of CMA results. The difference between the results of API Candida and API 20C AUX was detected. The ratio of discrepancy between Vitek-MS and API 20C AUX was 25.8 %. Candida species mostly identified as C. famata or C. tropicalis by and not compatible with API kits were identified as C. albicans by Vitek-MS. Sixteen Candida species having discrepant results with Vitek-MS, API or CMA were randomly chosen, and ITS sequence analysis was performed. The results of sequencing were compatible 56.2 % with API 20C AUX, 50 % with CMA and 93.7 % with Vitek-MS. When compared with conventional identification methods, MS results are more reliable and rapid for Candida identification. MS system may be used as routine identification method in clinical microbiology laboratories.

  5. Cost Analysis of Implementing Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry Plus Real-Time Antimicrobial Stewardship Intervention for Bloodstream Infections.

    Science.gov (United States)

    Patel, Twisha S; Kaakeh, Rola; Nagel, Jerod L; Newton, Duane W; Stevenson, James G

    2017-01-01

    Studies evaluating rapid diagnostic testing plus stewardship intervention have consistently demonstrated improved clinical outcomes for patients with bloodstream infections. However, the cost of implementing new rapid diagnostic testing can be significant, and such testing usually does not generate additional revenue. There are minimal data evaluating the impact of adding matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for rapid organism identification and dedicating pharmacy stewardship personnel time on the total hospital costs. A cost analysis was performed utilizing patient data generated from the hospital cost accounting system and included additional costs of MALDI-TOF equipment, supplies and personnel, and dedicated pharmacist time for blood culture review and of making interventions to antimicrobial therapy. The cost analysis was performed from a hospital perspective for 3-month blocks before and after implementation of MALDI-TOF plus stewardship intervention. A total of 480 patients with bloodstream infections were included in the analysis: 247 in the preintervention group and 233 in the intervention group. Thirty-day mortality was significantly improved in the intervention group (12% versus 21%, P cost per bloodstream infection was lower in the intervention group ($42,580 versus $45,019). Intensive care unit cost per bloodstream infection accounted for the largest share of the total costs in each group and was also lower in the intervention group ($10,833 versus $13,727). Implementing MALDI-TOF plus stewardship review and intervention decreased mortality for patients with bloodstream infections. Despite the additional costs of implementing MALDI-TOF and of dedicating pharmacy stewardship personnel time to interventions, the total hospital costs decreased by $2,439 per bloodstream infection, for an approximate annual cost savings of $2.34 million. Copyright © 2016 American Society for Microbiology.

  6. Use of matrix-assisted laser desorption/ionisation-time of flight mass spectrometry analyser in a diagnostic microbiology laboratory in a developing country

    Directory of Open Access Journals (Sweden)

    Atang Bulane

    2017-02-01

    Full Text Available Background: Rapid and accurate identification of pathogens is of utmost importance for management of patients. Current identification relies on conventional phenotypic methods which are time consuming. Matrix-assisted laser desorption/ionisation-time of flight mass spectrometry (MALDI-TOF MS is based on proteomic profiling and allows for rapid identification of pathogens.Objective: We compared MALDI-TOF MS against two commercial systems, MicroScan Walkaway and VITEK 2 MS.Methods: Over a three-month period from July 2013 to September 2013, a total of 227 bacteria and yeasts were collected from an academic microbiology laboratory (N = 121; 87 Gramnegatives, seven Gram-positives, 27 yeasts and other laboratories (N = 106; 35 Gram-negatives, 34 Gram-positives, 37 yeasts. Sixty-five positive blood cultures were initially processed with Bruker Sepsityper kit for direct identification.Results: From the 65 blood culture bottles, four grew more than one bacterial pathogen and MALDI-TOF MS identified only one isolate. The blood cultures yielded 21 Gram-negatives, 43 Gram-positives and one Candida. There were 21 Escherirchia coli isolates which were reported by the MALDI-TOF MS as E. coli/Shigella. Of the total 292 isolates, discrepant results were found for one bacterial and three yeast isolates. Discrepant results were resolved by testing with the API system with MALDI-TOF MS showing 100% correlation.Conclusion: The MALDI-TOF MS proved to be very useful for rapid and reliable identification of bacteria and yeasts directly from blood cultures and after culture of other specimens. The difference in time to identification was significant for all isolates. However, for positive blood cultures with minimal sample preparation time there was a massive difference in turn-around time with great appreciation by clinicians.

  7. Chemical analysis of pharmaceuticals and explosives in fingermarks using matrix-assisted laser desorption ionization/time-of-flight mass spectrometry.

    Science.gov (United States)

    Kaplan-Sandquist, Kimberly; LeBeau, Marc A; Miller, Mark L

    2014-02-01

    Chemical analysis of latent fingermarks, "touch chemistry," has the potential of providing intelligence or forensically relevant information. Matrix-assisted laser desorption ionization/time-of-flight mass spectrometry (MALDI/TOF MS) was used as an analytical platform for obtaining mass spectra and chemical images of target drugs and explosives in fingermark residues following conventional fingerprint development methods and MALDI matrix processing. There were two main purposes of this research: (1) develop effective laboratory methods for detecting drugs and explosives in fingermark residues and (2) determine the feasibility of detecting drugs and explosives after casual contact with pills, powders, and residues. Further, synthetic latent print reference pads were evaluated as mimics of natural fingermark residue to determine if the pads could be used for method development and quality control. The results suggest that artificial amino acid and sebaceous oil residue pads are not suitable to adequately simulate natural fingermark chemistry for MALDI/TOF MS analysis. However, the pads were useful for designing experiments and setting instrumental parameters. Based on the natural fingermark residue experiments, handling whole or broken pills did not transfer sufficient quantities of drugs to allow for definitive detection. Transferring drugs or explosives in the form of powders and residues was successful for preparing analytes for detection after contact with fingers and deposition of fingermark residue. One downfall to handling powders was that the analyte particles were easily spread beyond the original fingermark during development. Analyte particles were confined in the original fingermark when using transfer residues. The MALDI/TOF MS was able to detect procaine, pseudoephedrine, TNT, and RDX from contact residue under laboratory conditions with the integration of conventional fingerprint development methods and MALDI matrix. MALDI/TOF MS is a nondestructive

  8. Identification of microorganisms by FilmArray and matrix-assisted laser desorption ionization-time of flight mass spectrometry prior to positivity in the blood culture system.

    Science.gov (United States)

    Almuhayawi, Mohammed; Altun, Osman; Strålin, Kristoffer; Ozenci, Volkan

    2014-09-01

    In this study, we investigated the performance of the FilmArray and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) in identifying microorganisms from blood culture (BC) bottles prior to positivity. First, we used simulated BacT/Alert FA Plus BC bottles with five each for Escherichia coli and Staphylococcus aureus isolates. The FilmArray identified all 10 isolates before BC positivity with 9/10 at 5 h and 1 at 7.5 h after incubation in the BC system. MALDI-TOF MS failed to identify the isolates prior to positivity. When the bottles were incubated for 2.5 h at room temperature (RT) before we put them into the BC system, the FilmArray identified 6/10 at 2.5 h and the remaining 4 at 5 h. Finally, we tested simulated BC bottles after incubation at RT. Interestingly, 9/10 isolates were identified with the FilmArray after 8 h of incubation at RT. Second, we studied clinical BC bottles in quadruplicate. When three-fourths of the parallel bottles signaled positive, the FilmArray was run on the fourth nonsignaled bottle and was found to be positive in 14/15 such cases. Third, we analyzed the performance of the FilmArray in the identification of microorganisms from clinical BC bottles before incubation in the system. Two milliliters of broth from 400 BC bottles was collected after arrival at the laboratory and stored at -70°C. Sixteen bottles later signaled positive in the system. When the frozen broth from these bottles was analyzed, the FilmArray identified all the microorganisms in 8/16 bottles prior to incubation in the BC system. This study shows that the FilmArray can identify microorganisms from BC bottles prior to positivity and in some cases even prior to incubation in the BC system. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  9. Identification of bovine-associated coagulase-negative staphylococci by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry using a direct transfer protocol.

    Science.gov (United States)

    Cameron, M; Barkema, H W; De Buck, J; De Vliegher, S; Chaffer, M; Lewis, J; Keefe, G P

    2017-03-01

    This study evaluated matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-ToF MS) for the identification of bovine-associated coagulase-negative staphylococci (CNS), a heterogeneous group of different species. Additionally, we aimed to expand the MALDI-ToF MS database with new reference spectra as required to fill the gaps within the existing commercial spectral library. A total of 258 isolates of CNS were used in the study, covering 16 different CNS species. The majority of the isolates were previously identified by rpoB gene sequencing (n = 219), and the remainder were identified by sequencing of 16S rRNA, hsp60, or both rpoB and hsp60. The genotypic identification was considered the gold standard identification. All MALDI-ToF MS identifications were carried out using the direct transfer method. In a preliminary evaluation (n = 32 isolates; 2 of each species) with the existing commercial database, MALDI-ToF MS showed a typeability of 81% (26/32) and an accuracy of 96% (25/26). In the main evaluation (n = 226 isolates), MALDI-ToF MS with the existing commercial Biotyper (Bruker Daltonics Inc., Billerica, MA) database achieved a typeability of 92.0% (208/226) and an accuracy of 99.5% (207/208). Based on the assessment of the existing commercial database and prior knowledge of the species, a total of 13 custom reference spectra, covering 8 species, were created and added to the commercial database. Using the custom reference spectra expanded database, isolates were identified by MALDI-ToF MS with 100% typeability and 100% accuracy. Whereas the MALDI-ToF MS manufacturer's cutoff for species-level identification is 2.000, the reduction of the species level cutpoint to ≥1.700 improved the species-level identification rates (from 64 to 92% for the existing commercial database) when classifying CNS isolates. Overall, MALDI-ToF MS using the direct transfer method was shown to be a highly reliable tool for the identification of bovine

  10. Rapid and reliable species identification of wild mushrooms by matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Sugawara, Ryota; Yamada, Sayumi; Tu, Zhihao; Sugawara, Akiko; Suzuki, Kousuke; Hoshiba, Toshihiro; Eisaka, Sadao; Yamaguchi, Akihiro

    2016-08-31

    Mushrooms are a favourite natural food in many countries. However, some wild species cause food poisoning, sometimes lethal, due to misidentification caused by confusing fruiting bodies similar to those of edible species. The morphological inspection of mycelia, spores and fruiting bodies have been traditionally used for the identification of mushrooms. More recently, DNA sequencing analysis has been successfully applied to mushrooms and to many other species. This study focuses on a simpler and more rapid methodology for the identification of wild mushrooms via protein profiling based on matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF MS). A preliminary study using 6 commercially available cultivated mushrooms suggested that a more reproducible spectrum was obtained from a portion of the cap than from the stem of a fruiting body by the extraction of proteins with a formic acid-acetonitrile mixture (1 + 1). We used 157 wild mushroom-fruiting bodies collected in the centre of Hokkaido from June to November 2014. Sequencing analysis of a portion of the ribosomal RNA gene provided 134 identifications of mushrooms by genus or species, however 23 samples containing 10 unknown species that had lower concordance rate of the nucleotide sequences in a BLAST search (less than 97%) and 13 samples that had unidentifiable poor or mixed sequencing signals remained unknown. MALDI-TOF MS analysis yielded a reproducible spectrum (frequency of matching score ≥ 2.0 was ≥6 spectra from 12 spectra measurements) for 114 of 157 samples. Profiling scores that matched each other within the database gave correct species identification (with scores of ≥2.0) for 110 samples (96%). An in-house prepared database was constructed from 106 independent species, except for overlapping identifications. We used 48 wild mushrooms that were collected in autumn 2015 to validate the in-house database. As a result, 21 mushrooms were identified at the species level with

  11. Rapid identification of Burkholderia mallei and Burkholderia pseudomallei by intact cell Matrix-assisted Laser Desorption/Ionisation mass spectrometric typing

    Science.gov (United States)

    2012-01-01

    Background Burkholderia (B.) pseudomallei and B. mallei are genetically closely related species. B. pseudomallei causes melioidosis in humans and animals, whereas B. mallei is the causative agent of glanders in equines and rarely also in humans. Both agents have been classified by the CDC as priority category B biological agents. Rapid identification is crucial, because both agents are intrinsically resistant to many antibiotics. Matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-TOF MS) has the potential of rapid and reliable identification of pathogens, but is limited by the availability of a database containing validated reference spectra. The aim of this study was to evaluate the use of MALDI-TOF MS for the rapid and reliable identification and differentiation of B. pseudomallei and B. mallei and to build up a reliable reference database for both organisms. Results A collection of ten B. pseudomallei and seventeen B. mallei strains was used to generate a library of reference spectra. Samples of both species could be identified by MALDI-TOF MS, if a dedicated subset of the reference spectra library was used. In comparison with samples representing B. mallei, higher genetic diversity among B. pseudomallei was reflected in the higher average Eucledian distances between the mass spectra and a broader range of identification score values obtained with commercial software for the identification of microorganisms. The type strain of B. pseudomallei (ATCC 23343) was isolated decades ago and is outstanding in the spectrum-based dendrograms probably due to massive methylations as indicated by two intensive series of mass increments of 14 Da specifically and reproducibly found in the spectra of this strain. Conclusions Handling of pathogens under BSL 3 conditions is dangerous and cumbersome but can be minimized by inactivation of bacteria with ethanol, subsequent protein extraction under BSL 1 conditions and MALDI-TOF MS analysis being faster than

  12. Matrix-assisted laser desorption-ionization-time-of-flight mass spectrometry as a reliable proteomic method for characterization ofEscherichia coliandSalmonellaisolates.

    Science.gov (United States)

    Shell, Waleed S; Sayed, Mahmoud Lotfy; Allah, Fatma Mohamed Gad; Gamal, Fatma Elzahraa Mohamed; Khder, Afaf Ahmed; Samy, A A; Ali, Abdel Hakam M

    2017-09-01

    Identification of pathogenic clinical bacterial isolates is mainly dependent on phenotypic and genotypic characteristics of the microorganisms. These conventional methods are costive, time-consuming, and need special skills and training. An alternative, mass spectral (proteomics) analysis method for identification of clinical bacterial isolates has been recognized as a rapid, reliable, and economical method for identification. This study was aimed to evaluate and compare the performance, sensitivity and reliability of traditional bacteriology, phenotypic methods and matrix-assisted laser desorption-ionization-time-of-flight mass spectrometry (MALDI-TOF MS) in the identification of clinical Escherichia coli and Salmonella isolates recovered from chickens. A total of 110 samples (cloacal, liver, spleen, and/or gall bladder) were collected from apparently healthy and diseased chickens showing clinical signs as white chalky diarrhea, pasty vent, and decrease egg production as well as freshly dead chickens which showing postmortem lesions as enlarged liver with congestion and enlarged gall bladder from different poultry farms. Depending on colonial characteristics and morphological characteristics, E. coli and Salmonella isolates were recovered and detected in only 42 and 35 samples, respectively. Biochemical identification using API 20E identification system revealed that the suspected E. coli isolates were 33 out of 42 of colonial and morphological identified E. coli isolates where Salmonella isolates were represented by 26 out of 35 of colonial and morphological identified Salmonella isolates. Serological identification of isolates revealed that the most predominant E. coli serotypes were O1 and O78 while the most predominant Salmonella serotype of Salmonella was Salmonella Pullorum. All E. coli and Salmonella isolates were examined using MALDI-TOF MS. In agreement with traditional identification, MADI-TOF MS identified all clinical bacterial samples with valid

  13. Teaching Microbial Identification with Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS and Bioinformatics Tools

    Directory of Open Access Journals (Sweden)

    Wenfa Ng

    2013-01-01

    Full Text Available Ever since the first observation of “animalcules” under a microscope, and the subsequent discovery of microorganisms of myriad size, shape, pigmentation and motility modes, classification in aid of microbial identification is key to understanding inter-relationships between diverse microbes. Combining universal applicability with robustness, 16S rRNA sequencing is the gold standard for microbial typing; however, recent developments in clinical diagnostics have called attention to a shift towards PCR-independent instrumentation and methods given PCR’s requirement for expensive and complex sample preparation. Using ribosomal proteins as biomarkers for evolutionary relatedness, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS - originally developed for the soft ionization of proteins and peptides in proteomics studies - has been successfully applied to identifying bacteria, archaea, fungi and viruses to the species, and, on occasions, sub-species level. Though experimentally proven and increasingly adopted in the clinic, the relatively low-cost (on a per sample basis and rapid MALDI-TOF MS microbial identification technique, along with its theoretical principles and methodology, is a conspicuous absentee in contemporary microbiology curricula. Motivated by a desire to close the curriculum gap, this article describes a discovery-based activity for teaching microbial identification - using MALDI-TOF MS in combination with open-source genomics and proteomics search tools – while providing tips on mass spectra interpretation and activity implementation for lowering the barrier for classroom adoption. Infused with inquiry-based learning concepts guiding students in identifying microbes from environmental water samples with unknown species diversity, the activity spurs students’ learning by igniting their spirit of inquiry, which leads to better mastery of concepts; a significant departure from

  14. Exploring three-dimensional matrix-assisted laser desorption/ionization imaging mass spectrometry data: three-dimensional spatial segmentation of mouse kidney.

    Science.gov (United States)

    Trede, Dennis; Schiffler, Stefan; Becker, Michael; Wirtz, Stefan; Steinhorst, Klaus; Strehlow, Jan; Aichler, Michaela; Kobarg, Jan Hendrik; Oetjen, Janina; Dyatlov, Andrey; Heldmann, Stefan; Walch, Axel; Thiele, Herbert; Maass, Peter; Alexandrov, Theodore

    2012-07-17

    Three-dimensional (3D) imaging has a significant impact on many challenges of life sciences. Three-dimensional matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI-IMS) is an emerging label-free bioanalytical technique capturing the spatial distribution of hundreds of molecular compounds in 3D by providing a MALDI mass spectrum for each spatial point of a 3D sample. Currently, 3D MALDI-IMS cannot tap its full potential due to the lack efficient computational methods for constructing, processing, and visualizing large and complex 3D MALDI-IMS data. We present a new pipeline of efficient computational methods, which enables analysis and interpretation of a 3D MALDI-IMS data set. Construction of a MALDI-IMS data set was done according to the state-of-the-art protocols and involved sample preparation, spectra acquisition, spectra preprocessing, and registration of serial sections. For analysis and interpretation of 3D MALDI-IMS data, we applied the spatial segmentation approach which is well-accepted in analysis of two-dimensional (2D) MALDI-IMS data. In line with 2D data analysis, we used edge-preserving 3D image denoising prior to segmentation to reduce strong and chaotic spectrum-to-spectrum variation. For segmentation, we used an efficient clustering method, called bisecting k-means, which is optimized for hierarchical clustering of a large 3D MALDI-IMS data set. Using the proposed pipeline, we analyzed a central part of a mouse kidney using 33 serial sections of 3.5 μm thickness after the PAXgene tissue fixation and paraffin embedding. For each serial section, a 2D MALDI-IMS data set was acquired following the standard protocols with the high spatial resolution of 50 μm. Altogether, 512 495 mass spectra were acquired that corresponds to approximately 50 gigabytes of data. After registration of serial sections into a 3D data set, our computational pipeline allowed us to reveal the 3D kidney anatomical structure based on mass spectrometry

  15. Rapid identification of Burkholderia mallei and Burkholderia pseudomallei by intact cell Matrix-assisted Laser Desorption/Ionisation mass spectrometric typing.

    Science.gov (United States)

    Karger, Axel; Stock, Rüdiger; Ziller, Mario; Elschner, Mandy C; Bettin, Barbara; Melzer, Falk; Maier, Thomas; Kostrzewa, Markus; Scholz, Holger C; Neubauer, Heinrich; Tomaso, Herbert

    2012-10-10

    Burkholderia (B.) pseudomallei and B. mallei are genetically closely related species. B. pseudomallei causes melioidosis in humans and animals, whereas B. mallei is the causative agent of glanders in equines and rarely also in humans. Both agents have been classified by the CDC as priority category B biological agents. Rapid identification is crucial, because both agents are intrinsically resistant to many antibiotics. Matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-TOF MS) has the potential of rapid and reliable identification of pathogens, but is limited by the availability of a database containing validated reference spectra. The aim of this study was to evaluate the use of MALDI-TOF MS for the rapid and reliable identification and differentiation of B. pseudomallei and B. mallei and to build up a reliable reference database for both organisms. A collection of ten B. pseudomallei and seventeen B. mallei strains was used to generate a library of reference spectra. Samples of both species could be identified by MALDI-TOF MS, if a dedicated subset of the reference spectra library was used. In comparison with samples representing B. mallei, higher genetic diversity among B. pseudomallei was reflected in the higher average Eucledian distances between the mass spectra and a broader range of identification score values obtained with commercial software for the identification of microorganisms. The type strain of B. pseudomallei (ATCC 23343) was isolated decades ago and is outstanding in the spectrum-based dendrograms probably due to massive methylations as indicated by two intensive series of mass increments of 14 Da specifically and reproducibly found in the spectra of this strain. Handling of pathogens under BSL 3 conditions is dangerous and cumbersome but can be minimized by inactivation of bacteria with ethanol, subsequent protein extraction under BSL 1 conditions and MALDI-TOF MS analysis being faster than nucleic amplification methods. Our

  16. Rapid identification of Burkholderia mallei and Burkholderia pseudomallei by intact cell Matrix-assisted Laser Desorption/Ionisation mass spectrometric typing

    Directory of Open Access Journals (Sweden)

    Karger Axel

    2012-10-01

    Full Text Available Abstract Background Burkholderia (B. pseudomallei and B. mallei are genetically closely related species. B. pseudomallei causes melioidosis in humans and animals, whereas B. mallei is the causative agent of glanders in equines and rarely also in humans. Both agents have been classified by the CDC as priority category B biological agents. Rapid identification is crucial, because both agents are intrinsically resistant to many antibiotics. Matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-TOF MS has the potential of rapid and reliable identification of pathogens, but is limited by the availability of a database containing validated reference spectra. The aim of this study was to evaluate the use of MALDI-TOF MS for the rapid and reliable identification and differentiation of B. pseudomallei and B. mallei and to build up a reliable reference database for both organisms. Results A collection of ten B. pseudomallei and seventeen B. mallei strains was used to generate a library of reference spectra. Samples of both species could be identified by MALDI-TOF MS, if a dedicated subset of the reference spectra library was used. In comparison with samples representing B. mallei, higher genetic diversity among B. pseudomallei was reflected in the higher average Eucledian distances between the mass spectra and a broader range of identification score values obtained with commercial software for the identification of microorganisms. The type strain of B. pseudomallei (ATCC 23343 was isolated decades ago and is outstanding in the spectrum-based dendrograms probably due to massive methylations as indicated by two intensive series of mass increments of 14 Da specifically and reproducibly found in the spectra of this strain. Conclusions Handling of pathogens under BSL 3 conditions is dangerous and cumbersome but can be minimized by inactivation of bacteria with ethanol, subsequent protein extraction under BSL 1 conditions and MALDI-TOF MS

  17. Identification of clinical isolates of Aspergillus, including cryptic species, by matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Vidal-Acuña, M Reyes; Ruiz-Pérez de Pipaón, Maite; Torres-Sánchez, María José; Aznar, Javier

    2017-12-08

    An expanded library of matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been constructed using the spectra generated from 42 clinical isolates and 11 reference strains, including 23 different species from 8 sections (16 cryptic plus 7 noncryptic species). Out of a total of 379 strains of Aspergillus isolated from clinical samples, 179 strains were selected to be identified by sequencing of beta-tubulin or calmodulin genes. Protein spectra of 53 strains, cultured in liquid medium, were used to construct an in-house reference database in the MALDI-TOF MS. One hundred ninety strains (179 clinical isolates previously identified by sequencing and the 11 reference strains), cultured on solid medium, were blindy analyzed by the MALDI-TOF MS technology to validate the generated in-house reference database. A 100% correlation was obtained with both identification methods, gene sequencing and MALDI-TOF MS, and no discordant identification was obtained. The HUVR database provided species level (score of ≥2.0) identification in 165 isolates (86.84%) and for the remaining 25 (13.16%) a genus level identification (score between 1.7 and 2.0) was obtained. The routine MALDI-TOF MS analysis with the new database, was then challenged with 200 Aspergillus clinical isolates grown on solid medium in a prospective evaluation. A species identification was obtained in 191 strains (95.5%), and only nine strains (4.5%) could not be identified at the species level. Among the 200 strains, A. tubingensis was the only cryptic species identified. We demonstrated the feasibility and usefulness of the new HUVR database in MALDI-TOF MS by the use of a standardized procedure for the identification of Aspergillus clinical isolates, including cryptic species, grown either on solid or liquid media. © The Author 2017. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For

  18. Detection of Hanganutziu-Deicher antigens in O-glycans from pig heart tissues by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Jeong, Hee-Jin; Adhya, Mausumi; Park, Hae-Min; Kim, Yun-Gon; Kim, Byung-Gee

    2013-01-01

    In the α1,3-galactosyltransferase knockout (α-GalT KO) pig era, identification of the non-Gal epitopes is necessary for successful pig-to-human xenotransplantation. Recently, we successfully detected α-Gal epitopes as well as Hanganutziu-Deicher (H-D) antigens from the N-glycans in the pig heart tissues, which have been considered as promising non-Gal antigens. However, the profiling of O-glycan from pig heart tissues had not been performed owing to the difficulty of O-glycan preparation. In this study, we established the simple and sensitive method to profile O-glycans from pig heart aortic valve, aortic wall, pulmonary valve, pulmonary wall, and cardiac muscle tissues. To liberate O-glycans from the pig heart tissues, we used non-reductive β-elimination reagent and subsequently purified the glycans. After permethylation, the glycans were qualitatively analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The comprehensive O-glycan analysis method was successfully validated using model glycoproteins such as bovine serum fetuin (BSF) and bovine submaxillary gland mucin (BSM) glycoproteins, and their O-glycan profiles were in accordance with the data of previous studies. Next, we applied the method for O-glycan release and characterization to analysis of various pig heart tissues. As a result, total 39, 33, 24, 36, and 25 of O-glycans were detected from aortic valve, aortic wall, pulmonary valve, pulmonary wall, and cardiac muscle, respectively. Furthermore, four in aortic valve, one in aortic wall, one in pulmonary valve, one in pulmonary wall, and one in cardiac muscle were particularly determined as terminally N-glycolylneuraminic acid-linked O-glycans, which is considered to be the H-D antigens. Here, we initially described the O-glycan structures of various pig heart tissues, and additionally, the existence of H-D antigen type O-glycans was firstly identified. These results will be fundamental information

  19. Analyses of the in vitro non-enzymatic glycation of peptides/proteins by matrix-assisted laser desorption/ionization mass spectrometry

    Science.gov (United States)

    Lee, Bao-Shiang; Krishnanchettiar, Sangeeth; Lateef, Syed Salman; Gupta, Shalini

    2007-01-01

    Non-enzymatic glycation of proteins with the reducing agent glucose is implicated to be responsible for diabetes-derived complications, food browning, and aging. However, the non-enzymatic glycation process of peptides/proteins is not well understood and further research is needed to gain an understanding of the underlying principles involved in diabetes-related complications. In this study, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry is used to analyze the in vitro glycation of peptides/proteins. In addition to the physiological conditions, harsh conditions (higher concentration of glucose, higher or lower pH, and higher temperature) are also used in this study. Peptides/proteins are reacted with glucose for up to 120 h at 4 [degree sign]C, 37 [degree sign]C, or 65 [degree sign]C. Single and/or multiple glycations are observed using broad pH conditions (from 10% TFA with pKa of 0.5 to pH 10) at various glucose concentrations (from 0.01 M to 1 M). Data suggest that glucose reacts readily with both peptides and proteins, and the efficiency of the glycation increases with higher temperature, higher pH, higher glucose concentration, or longer incubation time. However, influence of the buffer pH on the efficiency of the glycation of peptides is less pronounced compared to that of proteins. This effect could result from denaturation of proteins at higher pH and the resultant exposure of potential glycation sites. This data could lead to the inference that the glycation process of peptides/proteins would occur but proceed very slowly under the diabetes conditions in vivo (37 [degree sign]C, ~neutral pH, ~0.007 M glucose). Postsource decay and MS/MS results of singly glycated angiotensin I, P14R (PPPPPPPPPPPPPPR), and human adrenocorticotropic hormone (ATCH) fragments 1-13 indicate that glucose reacts with the amino group of the N-terminal of ATCH 1-13 and the guanidino group of the arginine residue of both angiotensin I and P14R.

  20. Rapid Detection and Identification of Candidemia by Direct Blood Culturing on Solid Medium by Use of Lysis-Centrifugation Method Combined with Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Idelevich, Evgeny A; Grünastel, Barbara; Becker, Karsten

    2017-01-01

    Candida sepsis is a life-threatening condition with increasing prevalence. In this study, direct blood culturing on solid medium using a lysis-centrifugation procedure enabled successful Candida species identification by matrix-assisted laser desorption-ionization time of flight mass spectrometry on average 3.8 h (Sabouraud agar) or 7.4 h (chocolate agar) before the positivity signal for control samples in Bactec mycosis-IC/F or Bactec Plus aerobic/F bottles, respectively. Direct culturing on solid medium accelerated candidemia diagnostics compared to that with automated broth-based systems. Copyright © 2016 American Society for Microbiology.

  1. Misidentification of Saprochaete clavata as Magnusiomyces capitatus in clinical isolates: utility of internal transcribed spacer sequencing and matrix-assisted laser desorption ionization-time of flight mass spectrometry and importance of reliable databases.

    Science.gov (United States)

    Desnos-Ollivier, Marie; Blanc, Catherine; Garcia-Hermoso, Dea; Hoinard, Damien; Alanio, Alexandre; Dromer, Françoise

    2014-06-01

    Saprochaete clavata and Magnusiomyces capitatus are human pathogens that are frequently mistaken for each other due to their similar phenotypes and erroneous or limited databases. Based on internal transcribed spacer (ITS) sequences, we propose species-specific carbon assimilation patterns and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) fingerprints that enable the identification of S. clavata, M. capitatus, and Galactomyces candidus to the species level. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  2. Localization of an O-glycosylated site in the recombinant barley alpha-amylase 1 produced in yeast and correction of the amino acid sequence using matrix-assisted laser desorption/ionization mass spectrometry of peptide mixtures

    DEFF Research Database (Denmark)

    Andersen, Jens S.; Søgaard, M; Svensson, B

    1994-01-01

    degradation of two selected peptides isolated from the endoproteinase Lys-C digest corrected the sequence to be Val instead of Ala in position 284 and confirmed the O-glycosylation. These results demonstrate that the direct peptide mixture analysis by MALDI-MS is a rapid and sensitive method for protein......Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) of peptide mixtures was used to characterize recombinant barley alpha-amylase 1, produced in yeast. Three peptide mixtures were generated by cleavage with CNBr, digestion with endoproteinase Lys-C and Asp-N, respectively...

  3. Substrate-Enhanced Micro Laser Desorption Ionization Mass Spectrometry Project

    Data.gov (United States)

    National Aeronautics and Space Administration — Aerodyne Research, Inc. and the University of Massachusetts at Amherst will collaborate to develop laser desorption ionization (LDI) mass spectrometric analysis of...

  4. Analysis of native milk oligosaccharides directly from thin-layer chromatography plates by matrix-assisted laser desorption/ionization orthogonal-time-of-flight mass spectrometry with a glycerol matrix.

    Science.gov (United States)

    Dreisewerd, Klaus; Kölbl, Stefanie; Peter-Katalinić, Jasna; Berkenkamp, Stefan; Pohlentz, Gottfried

    2006-02-01

    We have recently presented a new method for direct coupling of high-performance thin-layer chromatography (HPTLC) with matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), illustrated by the analysis of a complex ganglioside mixture. In the current communication, an adaptation of this procedure to mixtures of native oligosaccharides from human and from elephant milk is described. The key features in this method are (1) glycerol as a liquid matrix, to provide a homogeneous wetting of the silica gel and a simple and fast MALDI preparation protocol, (2) an infrared (IR) laser for volume material ablation and particular soft desorption/ionization conditions, and (3) an orthogonal time-of-flight mass spectrometer for a high mass accuracy, independent of any irregularity of the silica gel surface. Chromatographic "mobility profiles" were determined by scanning the laser beam across the analyte bands. The current limit of detection for the MS analysis was determined to approximately 10 pmol of individual oligosaccharides spotted for chromatography. A liquid composite matrix, containing glycerol and the ultraviolet (UV-)MALDI matrix alpha-cyano-4-hydroxycinnamic acid, allows a direct HPTLC-MALDI-MS analysis with a 337 nm-UV laser as well. Compared to the IR-MALDI mode, the analytical sensitivity in UV-MALDI was found to be lower by one order of magnitude, whereas unspecific analyte ion fragmentation as well as adduct formation was found to be more extensive.

  5. Rapid identification of bacillus anthracis spores in suspicious powder samples by using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS)

    NARCIS (Netherlands)

    Dybwad, M.; Laaken, A.L. van der; Blatny, J.M.; Paauw, A.

    2013-01-01

    Rapid and reliable identification of Bacillus anthracis spores in suspicious powders is important to mitigate the safety risks and economic burdens associated with such incidents. The aim of this study was to develop and validate a rapid and reliable laboratory- based matrix-assisted laser

  6. Characterization of O-glycosylated precursors of insulin-like growth factor II by matrix-assisted laser desorption/ionization mass spectrometry

    NARCIS (Netherlands)

    Jespersen, S.; Koedam, J.A.; Hoogerbrugge, C.M.; Tjaden, U.R.; Greef, J. van der; Brande, J.L. van den

    1996-01-01

    High molecular weight precursors of insulin-like growth factor II (IGF-II) were isolated from Cohn fraction IV of human plasma by ultrafiltration, affinity chromatography and reversed-phase high-performance liquid chromatography. Molecular weight determination by matrix-assisted laser

  7. Late Periprosthetic Joint Infection due to Staphylococcus lugdunensis Identified by Matrix-Assisted Laser Desorption/Ionisation Time of Flight Mass Spectrometry: A Case Report and Review of the Literature

    Science.gov (United States)

    Szabados, Florian; Anders, Agnes; Kaase, Martin; Marlinghaus, Lennart; Gatermann, Sören G.; Teske, Wolfram; Lichtinger, Thomas

    2011-01-01

    Staphylococcus lugdunensis, member to the group of coagulase-negative staphylococci, is previously thought to be rarely isolated. Recently other staphylococci have been described, which were supposedly related to S. lugdunensis, such as Staphylococcus pseudolugdunensis and Staphylococcus pettenkoferi. To decrease the rate misidentifications, an accurate identification method, such as matrix-assisted laser desorption ionization time of flight mass spectrometry or molecular methods, should be used. S. lugdunensis is usually associated with severe infections similar to those caused by S. aureus. Moreover, it has been described that skin infections due to S. lugdunensis are severely underreported and could be also underreported in periprosthetic joint infections. Ours is the first case of a late periprosthetic infection of the hip due to S. lugdunensis, identified by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry. A periprosthetic infection due to S. lugdunensis should be treated according to protocols of S. aureus periprosthetic infections, and therefore an accurate species identification is desirable. PMID:21776276

  8. Evaluation of a matrix-assisted laser desorption ionization-time of flight mass spectrometry assisted, selective broth method to screen for vancomycin-resistant enterococci in patients at high risk.

    Science.gov (United States)

    Huang, Tsi-Shu; Lee, Susan Shin-Jung; Lee, Chia-Chien; Chen, Chiu-Yen; Chen, Fang-Chen; Chen, Bao-Chen; Sy, Cheng Len; Wu, Kuan-Sheng

    2017-01-01

    Bile esculin azide with vancomycin (BEAV) medium is a sensitive, but slightly less specific method for vancomycin-resistant enterococci (VRE) screening. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a rapid method for identification of clinical pathogens. This study aimed to assess the performance of a novel combination screening test for VRE, using BEAV broth combined with MALDI-TOF MS. Clinical specimens were collected from patients at risk of VRE carriage, and tested by the novel combination method, using selective BEAV broth culture method followed by MALDI-TOF MS identification (SBEAVM). The reference method used for comparison was the ChromID VRE agar method. A total of 135 specimens were collected from 78 patients, and 63 specimens tested positive for VRE positive using the ChromID VRE method (positive rate 46.7%). The sensitivity, specificity, positive predictive value, and negative predictive value of SBEAVM method after an incubation period of 28 hours were 93.7%, 90.3%, 89.4%, and 94.2%, respectively. The SBEAVM method when compared to the ChromID VRE method had a shorter turnaround time (29 vs. 48-72 hours) and lower laboratory cost ($2.11 vs. $3.23 per test). This study demonstrates that SBEAVM is a rapid, inexpensive, and accurate method for use in VRE screening.

  9. Metabolomic profiling of prostate cancer by matrix assisted laser desorption/ionization-Fourier transform ion cyclotron resonance mass spectrometry imaging using Matrix Coating Assisted by an Electric Field (MCAEF).

    Science.gov (United States)

    Wang, Xiaodong; Han, Jun; Hardie, Darryl B; Yang, Juncong; Pan, Jingxi; Borchers, Christoph H

    2017-07-01

    In this work, we combined the use of two MALDI matrices (quercetin and 9-aminoacridine), a recently developed new matrix coating technique - matrix coating assisted by an electric field (MCAEF), and matrix-assisted laser desorption/ionization - Fourier transform ion cyclotron resonance mass spectrometry (MALDI-FTICRMS) to detect and image endogenous compounds in the cancerous and non-cancerous regions of three human prostate cancer (stage II) tissue specimens. After three rounds of imaging data acquisitions (i.e., quercetin for positive and negative ion detection and 9-aminoacridine for negative ion detection), and metabolite identification, a total of 1091 metabolites including 1032 lipids and 59 other metabolites were routinely detected and successfully localized. Of these compounds, 250 and 217 were only detected in either the cancerous or the non-cancerous regions respectively, although we cannot rule out the presence of these metabolites at concentrations below the detection limit. In addition, 152 of the other 624 metabolites showed differential distributions (p<0.05, t-test) between the two regions of the tissues. Further studies on a larger number of clinical specimens will need to be carried out to confirm this large number of apparently cancer-related metabolites. The successful determination of the spatial locations and abundances of these endogenous biomolecules indicated significant metabolism abnormalities - e.g., increased energy charge and under-expression of neutral acyl glycerides, in the prostate cancer samples. To our knowledge, this work has resulted in MALDI-MS imaging of the largest group of metabolites in prostate cancer thus far and demonstrated the importance of using complementary matrices for comprehensive metabolomic imaging by MALDI-MS. This article is part of a Special Issue entitled: MALDI Imaging, edited by Dr. Corinna Henkel and Prof. Peter Hoffmann. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. Comparative Proteomic Analysis of Three Gelatinous Chinese Medicines and Their Authentications by Tryptic-digested Peptides Profiling using Matrix-assisted Laser Desorption/Ionization-time of Flight/Time of Flight Mass Spectrometry.

    Science.gov (United States)

    Yang, Huan; Zheng, Jie; Wang, Hai-Yan; Li, Nan; Yang, Ya-Ya; Shen, Yu-Ping

    2017-01-01

    Gelatinous Chinese medicines (GCMs) including Asini Corii Colla, Testudinis Carapacis ET Plastri Colla, and Cervi Cornus Colla, were made from reptile shell or mammalian skin or deer horn, and consumed as a popular tonic, as well as hemopoietic and hemostatic agents. Misuse of them would not exert their functions, and fake or adulterate products have caused drug market disorder and affected food and drug safety. GCMs are rich in denatured proteins, but insufficient in available DNA fragments, hence commonly used cytochrome c oxidase I barcoding was not successful for their authentication. In this study, we performed comparative proteomic analysis of them and their animal origins to identify the composition of intrinsic proteins for the first time. A reliable and convenient approach was proposed for their authentication, by the incorporation of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, two-dimensional electrophoresis, and matrix-assisted laser desorption/ionization-time of flight/time of flight mass spectrometry (MALDI-TOF/TOF-MS). A total of 26 proteins were identified from medicinal parts of original animals, and GCMs proteins presented in a dispersive manner in electrophoresis analyses due to complicated changes in the structure of original proteins caused by long-term decoction and the addition of ingredients during their manufacturing. In addition, by comparison of MALDI-TOF/TOF-MS profiling, 19 signature peptide fragments originated from the protein of GCM products were selected according to criteria. These could assist in the discrimination and identification of adulterates of GCMs and other ACMs for their form of raw medicinal material, the pulverized, and even the complex. Comparative proteomic analysis of three gelatinous Chinese medicines was conducted, and their authentications were based on tryptic-digested peptides profiling using matrix-assisted laser desorption/ionization-time of flight/time of flight mass spectrometry. Abbreviations

  11. Laser Desorption/Ionization-Time of Flight (LDI-TOF and Matrix-Assisted Laser Desorption/Ionization - Time of Flight (MALDI – TOF mass spectrometry of an Algerian asphaltene

    Directory of Open Access Journals (Sweden)

    T. Fergoug

    2017-09-01

    Full Text Available Both LDI-TOF and MALDI-TOF Mass spectroscopy experiments of an Algerian asphaltene derived from a deposit were performed. LDI mass experiments were conducted for both linear and reflectron modes under laser wavelength/attenuation variation. The different LDI-Mass spectra show that mass distribution depends on experimental condition for masses below 1000 amu and that the average molecular weight is around 650 for the polar fraction and beyond 1000 amu for non-polar ones. The use of different matrices as CHCA, HABA and Dithranol changes slightly the aspect of the spectra.

  12. MoS2/Ag nanohybrid: A novel matrix with synergistic effect for small molecule drugs analysis by negative-ion matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Zhao, Yaju; Deng, Guoqing; Liu, Xiaohui; Sun, Liang; Li, Hui; Cheng, Quan; Xi, Kai; Xu, Danke

    2016-09-21

    This paper reports a facile synthesis of molybdenum disulfide nanosheets/silver nanoparticles (MoS2/Ag) hybrid and its use as an effective matrix in negative ion matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The nanohybrid exerts a strong synergistic effect, leading to high performance detection of small molecule analytes including amino acids, peptides, fatty acids and drugs. The enhancement of laser desorption/ionization (LDI) efficiency is largely attributed to the high surface roughness and large surface area for analyte adsorption, better dispersibility, increased thermal conductivity and enhanced UV energy absorption as compared to pure MoS2. Moreover, both Ag nanoparticles and the edge of the MoS2 layers function as deprotonation sites for proton capture, facilitating the charging process in negative ion mode and promoting formation of negative ions. As a result, the MoS2/Ag nanohybrid proves to be a highly attractive matrix in MALDI-TOF MS, with desired features such as high desorption/ionization efficiency, low fragmentation interference, high salt tolerance, and no sweet-spots for mass signal. These characteristic properties allowed for simultaneous analysis of eight different drugs and quantification of acetylsalicylic acid in the spiked human serum. This work demonstrates for the first time the fabrication and application of a novel MoS2/Ag hybrid, and provides a new platform for use in the rapid and high throughput analysis of small molecules by mass spectrometry. Copyright © 2016. Published by Elsevier B.V.

  13. Evaluation of the Andromas matrix-assisted laser desorption ionization-time of flight mass spectrometry system for identification of aerobically growing Gram-positive bacilli.

    Science.gov (United States)

    Farfour, E; Leto, J; Barritault, M; Barberis, C; Meyer, J; Dauphin, B; Le Guern, A-S; Leflèche, A; Badell, E; Guiso, N; Leclercq, A; Le Monnier, A; Lecuit, M; Rodriguez-Nava, V; Bergeron, E; Raymond, J; Vimont, S; Bille, E; Carbonnelle, E; Guet-Revillet, H; Lécuyer, H; Beretti, J-L; Vay, C; Berche, P; Ferroni, A; Nassif, X; Join-Lambert, O

    2012-08-01

    Matrix-associated laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is a rapid and simple microbial identification method. Previous reports using the Biotyper system suggested that this technique requires a preliminary extraction step to identify Gram-positive rods (GPRs), a technical issue that may limit the routine use of this technique to identify pathogenic GPRs in the clinical setting. We tested the accuracy of the MALDI-TOF MS Andromas strategy to identify a set of 659 GPR isolates representing 16 bacterial genera and 72 species by the direct colony method. This bacterial collection included 40 C. diphtheriae, 13 C. pseudotuberculosis, 19 C. ulcerans, and 270 other Corynebacterium isolates, 32 L. monocytogenes and 24 other Listeria isolates, 46 Nocardia, 75 Actinomyces, 18 Actinobaculum, 11 Propionibacterium acnes, 18 Propionibacterium avidum, 30 Lactobacillus, 21 Bacillus, 2 Rhodococcus equi, 2 Erysipelothrix rhusiopathiae, and 38 other GPR isolates, all identified by reference techniques. Totals of 98.5% and 1.2% of non-Listeria GPR isolates were identified to the species or genus level, respectively. Except for L. grayi isolates that were identified to the species level, all other Listeria isolates were identified to the genus level because of highly similar spectra. These data demonstrate that rapid identification of pathogenic GPRs can be obtained without an extraction step by MALDI-TOF mass spectrometry.

  14. Evaluation of the Andromas Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry System for Identification of Aerobically Growing Gram-Positive Bacilli

    Science.gov (United States)

    Farfour, E.; Leto, J.; Barritault, M.; Barberis, C.; Meyer, J.; Dauphin, B.; Le Guern, A.-S.; Leflèche, A.; Badell, E.; Guiso, N.; Leclercq, A.; Le Monnier, A.; Lecuit, M.; Rodriguez-Nava, V.; Bergeron, E.; Raymond, J.; Vimont, S.; Bille, E.; Carbonnelle, E.; Guet-Revillet, H.; Lécuyer, H.; Beretti, J.-L.; Vay, C.; Berche, P.; Ferroni, A.; Nassif, X.

    2012-01-01

    Matrix-associated laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) is a rapid and simple microbial identification method. Previous reports using the Biotyper system suggested that this technique requires a preliminary extraction step to identify Gram-positive rods (GPRs), a technical issue that may limit the routine use of this technique to identify pathogenic GPRs in the clinical setting. We tested the accuracy of the MALDI-TOF MS Andromas strategy to identify a set of 659 GPR isolates representing 16 bacterial genera and 72 species by the direct colony method. This bacterial collection included 40 C. diphtheriae, 13 C. pseudotuberculosis, 19 C. ulcerans, and 270 other Corynebacterium isolates, 32 L. monocytogenes and 24 other Listeria isolates, 46 Nocardia, 75 Actinomyces, 18 Actinobaculum, 11 Propionibacterium acnes, 18 Propionibacterium avidum, 30 Lactobacillus, 21 Bacillus, 2 Rhodococcus equi, 2 Erysipelothrix rhusiopathiae, and 38 other GPR isolates, all identified by reference techniques. Totals of 98.5% and 1.2% of non-Listeria GPR isolates were identified to the species or genus level, respectively. Except for L. grayi isolates that were identified to the species level, all other Listeria isolates were identified to the genus level because of highly similar spectra. These data demonstrate that rapid identification of pathogenic GPRs can be obtained without an extraction step by MALDI-TOF mass spectrometry. PMID:22692743

  15. Amazonian vegetable oils and fats: fast typification and quality control via triacylglycerol (TAG) profiles from dry matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry fingerprinting.

    Science.gov (United States)

    Saraiva, Sérgio A; Cabral, Elaine C; Eberlin, Marcos N; Catharino, Rodrigo R

    2009-05-27

    Amazonian oils and fats display unique triacylglycerol (TAG) profiles and, because of their economic importance as renewable raw materials and use by the cosmetic and food industries, are often subject to adulteration and forgery. Representative samples of these oils (andiroba, Brazil nut, buriti, and passion fruit) and fats (cupuaçu, murumuru, and ucuúba) were characterized without pre-separation or derivatization via dry (solvent-free) matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Characteristic profiles of TAG were obtained for each oil and fat. Dry MALDI-TOF MS provides typification and direct and detailed information, via TAG profiles, of their variable combinations of fatty acids. A database from spectra could be developed and may be used for their fast and reliable typification, application screening, and quality control.

  16. Development of a Rapid and Accurate Identification Method for Citrobacter Species Isolated from Pork Products Using a Matrix-Assisted Laser-Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Kwak, Hye-Lim; Han, Sun-Kyung; Park, Sunghoon; Park, Si Hong; Shim, Jae-Yong; Oh, Mihwa; Ricke, Steven C; Kim, Hae-Yeong

    2015-09-01

    Previous detection methods for Citrobacter are considered time consuming and laborious. In this study, we have developed a rapid and accurate detection method for Citrobacter species in pork products, using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry (MS). A total of 35 Citrobacter strains were isolated from 30 pork products and identified by both MALDI-TOF MS and 16S rRNA gene sequencing approaches. All isolates were identified to the species level by the MALDI-TOF MS, while 16S rRNA gene sequencing results could not discriminate them clearly. These results confirmed that MALDI-TOF MS is a more accurate and rapid detection method for the identification of Citrobacter species.

  17. Proteome-based bacterial identification using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS): A revolutionary shift in clinical diagnostic microbiology.

    Science.gov (United States)

    Nomura, Fumio

    2015-06-01

    Rapid and accurate identification of microorganisms, a prerequisite for appropriate patient care and infection control, is a critical function of any clinical microbiology laboratory. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is a quick and reliable method for identification of microorganisms, including bacteria, yeast, molds, and mycobacteria. Indeed, there has been a revolutionary shift in clinical diagnostic microbiology. In the present review, the state of the art and advantages of MALDI-TOF MS-based bacterial identification are described. The potential of this innovative technology for use in strain typing and detection of antibiotic resistance is also discussed. This article is part of a Special Issue entitled: Medical Proteomics. Copyright © 2014 Elsevier B.V. All rights reserved.

  18. Recognition of Streptococcus pseudoporcinus Colonization in Women as a Consequence of Using Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry for Group B Streptococcus Identification.

    Science.gov (United States)

    Suwantarat, Nuntra; Grundy, Maureen; Rubin, Mayer; Harris, Renee; Miller, Jo-Anne; Romagnoli, Mark; Hanlon, Ann; Tekle, Tsigereda; Ellis, Brandon C; Witter, Frank R; Carroll, Karen C

    2015-12-01

    During a 14-month period of using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for group B streptococcus (GBS) identification, we recovered 32 (1%) Streptococcus pseudoporcinus isolates from 3,276 GBS screening cultures from female genital sources (25 isolates from pregnant women and 7 from nonpregnant women). An additional two S. pseudoporcinus isolates were identified from a urine culture and a posthysterectomy wound culture. These isolates were found to cross-react with three different GBS antigen agglutination kits, PathoDx (Remel) (93%), Prolex (Pro-Lab Diagnostics) (38%), and Streptex (Remel) (53%). New approaches to bacterial identification in routine clinical microbiology laboratories may affect the prevalence of S. pseudoporcinus. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  19. Application of Matrix-assisted Laser Desorption Ionization Time-of-flight Mass Spectrometry for Identification of Coagulase-negative Staphylococci Isolated from Milk of Cows with Subclinical Mastitis.

    Science.gov (United States)

    Banach, T; Bochniarz, M; Łyp, P; Adaszek, Ł; Wawron, W; Furmaga, B; Skrzypczak, M; Ziętek, J; Winiarczyk, S

    2016-09-01

    The aim of this study was to use matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) for the identification of coagulase-negative staphylococci (CNS) isolated from the milk of cows with subclinical mastitis. The study material consisted of 33 isolates of CNS, identified by the results of API Staph tests, obtained from the milk of cows with subclinical mastitis. Based on the spectra analyses, MALDI-TOF MS tests of 33 bacterial samples allowed identification of the microorganisms in 27 cases (81.8%). The most frequent cause of subclinical mastitis was found to be Staphylococcus sciuri (39%), while S. vitulinus was detected in 15% of the milk samples. The results obtained indicate that MALDI-TOF MS can be used for the identification of CNS isolated from bovine mastitis as a method supplementary to biochemical tests.

  20. Direct detection of the plant pathogens Burkholderia glumae, Burkholderia gladioli pv. gladioli, and Erwinia chrysanthemi pv. zeae in infected rice seedlings using matrix assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Kajiwara, Hideyuki

    2016-01-01

    The plant pathogens Burkholderia glumae, Burkholderia gladioli pv. gladioli, and Erwinia chrysanthemi pv. zeae were directly detected in extracts from infected rice seedlings by matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). This method did not require culturing of the pathogens on artificial medium. In the MALDI-TOF MS analysis, peaks originating from bacteria were found in extracts from infected rice seedlings. The spectral peaks showed significantly high scores, in spite of minor differences in spectra. The spectral peaks originating from host plant tissues did not affect this direct MALDI-TOF MS analysis for the rapid identification of plant pathogens. Copyright © 2015 Elsevier B.V. All rights reserved.

  1. Matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) mass spectrometry as a tool for rapid diagnosis of potentially toxigenic Corynebacterium species in the laboratory management of diphtheria-associated bacteria.

    Science.gov (United States)

    Konrad, R; Berger, A; Huber, I; Boschert, V; Hörmansdorfer, S; Busch, U; Hogardt, M; Schubert, S; Sing, A

    2010-10-28

    The rapid identification of the potentially toxigenic Corynebacterium species, C. diphtheriae, C. ulcerans and C. pseudotuberculosis is essential for diagnosis and treatment of diphtheria and diphtheria-like diseases. We used matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDIT-OF MS) in comparison with classical microbiological and molecular methods on 116 Corynebacterium strains. All 90 potentially toxigenic Corynebacterium strains collected by the German National Consiliary Laboratory on Diphtheria in a period of more than ten years were correctly identified by MALDI-TOF MS. We propose an algorithm for fast and reliable diagnosis of diphtheria incorporating MALDI-TOF MS, real-time tox PCR and Elek testing.

  2. Culture conditions and sample preparation methods affect spectrum quality and reproducibility during profiling of Staphylococcus aureus with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Goldstein, J E; Zhang, L; Borror, C M; Rago, J V; Sandrin, T R

    2013-08-01

    Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has emerged as a promising tool to rapidly characterize Staphylococcus aureus. Different protocols have been employed, but effects of experimental factors, such as culture condition and sample preparation, on spectrum quality and reproducibility have not been rigorously examined. We applied MALDI-TOF MS to characterize a model system consisting of five methicillin-sensitive (MSSA) and five methicillin-resistant S. aureus isolates (MRSA) under two culture conditions (agar and broth) and using two sample preparation methods [intact cell method and protein extraction method (PEM)]. The effects of these treatments on spectrum quality and reproducibility were quantified. PEM facilitated increases in the number of peaks and mass range width. Broth cultures further improved spectrum quality in terms of increasing the number of peaks. In addition, PEM increased reproducibility in samples prepared using identical culture conditions. MALDI imaging data suggested that the improvement in reproducibility may result from a more homogeneous distribution of sample associated with the broth/PEM treatment. Broth/PEM treatment also yielded the highest rate (96%) of correct classification for MRSA. Taken together, these results suggest that broth/PEM maximizes the performance of MALDI-TOF MS to characterize S. aureus. Two culture conditions (agar or broth) and two sample preparation methods (intact cell or protein extraction) were evaluated for their effects on profiling of Staphylococcus aureus using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Results indicated that MALDI-enabled profiling of S. aureus is most effective when cultures are grown in broth and processed using a protein extraction-based approach. These findings should enhance future efforts to maximize the performance of this approach to characterize strains of S. aureus. © 2013

  3. Analysis of gangliosides directly from thin-layer chromatography plates by infrared matrix-assisted laser desorption/ionization orthogonal time-of-flight mass spectrometry with a glycerol matrix.

    Science.gov (United States)

    Dreisewerd, Klaus; Müthing, Johannes; Rohlfing, Andreas; Meisen, Iris; Vukelić, Zeljka; Peter-Katalinić, Jasna; Hillenkamp, Franz; Berkenkamp, Stefan

    2005-07-01

    A novel method is presented for direct coupling of high-performance thin-layer chromatography (HPTLC) with matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) for the analysis of biomolecules. A first key feature is the use of a liquid matrix (glycerol), which provides a homogeneous wetting of the silica gel and a simple and fast MALDI preparation protocol. A second is the use of an Er:YAG infrared laser, which ablates layers of approximately 10-microm thickness of analyte-loaded silica gel and provides a soft desorption/ionization of even very labile analyte molecules. The orthogonal time-of-flight mass spectrometer employed in this study, finally provides a high accuracy of the mass determination, which is independent of any irregularity of the silica gel surface. The analytical potential of the method is demonstrated by the compositional mapping of a native GM3 (II(3)-alpha-Neu5Ac-LacCer) ganglioside mixture from cultured Chinese hamster ovary cells. The analysis is characterized by a high relative sensitivity, allowing the simultaneous detection of various major and minor GM3 species directly from individual HPTLC analyte bands. The lateral resolution of the direct HPTLC-MALDI-MS analysis is defined by the laser focus diameter of currently approximately 200 microm. This allows one to determine mobility profiles of individual species with a higher resolution than by reading off the chromatogram by optical absorption. The fluorescent dye primuline was, furthermore, successfully tested as a nondestructive, MALDI-compatible staining agent.

  4. Rapid Profiling of Bovine and Human Milk Gangliosides by Matrix-Assisted Laser Desorption/Ionization Fourier Transform Ion Cyclotron Resonance Mass Spectrometry.

    Science.gov (United States)

    Lee, Hyeyoung; An, Hyun Joo; Lerno, Larry A; German, J Bruce; Lebrilla, Carlito B

    2011-08-15

    Gangliosides are anionic glycosphingolipids widely distributed in vertebrate tissues and fluids. Their structural and quantitative expression patterns depend on phylogeny and are distinct down to the species level. In milk, gangliosides are exclusively associated with the milk fat globule membrane. They may participate in diverse biological processes but more specifically to host-pathogen interactions. However, due to the molecular complexities, the analysis needs extensive sample preparation, chromatographic separation, and even chemical reaction, which makes the process very complex and time-consuming. Here, we describe a rapid profiling method for bovine and human milk gangliosides employing matrix-assisted desorption/ionization (MALDI) Fourier transform ion cyclotron resonance (FTICR) mass spectrometry (MS). Prior to the analyses of biological samples, milk ganglioside standards GM3 and GD3 fractions were first analyzed in order to validate this method. High mass accuracy and high resolution obtained from MALDI FTICR MS allow for the confident assignment of chain length and degree of unsaturation of the ceramide. For the structural elucidation, tandem mass spectrometry (MS/MS), specifically as collision-induced dissociation (CID) and infrared multiphoton dissociation (IRMPD) were employed. Complex ganglioside mixtures from bovine and human milk were further analyzed with this method. The samples were prepared by two consecutive chloroform/methanol extraction and solid phase extraction. We observed a number of differences between bovine milk and human milk. The common gangliosides in bovine and human milk are NeuAc-NeuAc-Hex-Hex-Cer (GD3) and NeuAc-Hex-Hex-Cer (GM3); whereas, the ion intensities of ganglioside species are different between two milk samples. Kendrick mass defect plot yields grouping of ganglioside peaks according to their structural similarities. Gangliosides were further probed by tandem MS to confirm the compositional and structural assignments

  5. Recent progress in application of carbon nanomaterials in laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Wang, Jing; Liu, Qian; Liang, Yong; Jiang, Guibin

    2016-04-01

    Carbon nanomaterials have attracted great interest over past decades owing to their unique physical properties, versatile functionalization chemistry, and biological compatibility. In this article, we review recent progress in application of carbon nanomaterials in laser desorption/ionization mass spectrometry (LDI MS). Various types of carbon nanomaterials, including fullerenes, carbon nanotubes, graphene, carbon nanodots, nanodiamond, nanofibers, nanohorns, and their derivative forms, are involved. The applications of these materials as new matrices or probes in matrix-assisted or surface-enhanced laser desorption/ionization mass spectrometry (MALDI or SELDI MS) are discussed. Finally, we summarize current challenges and give our perspectives on the future of applications of carbon nanomaterials in LDI MS.

  6. Bruker Biotyper Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry System for Identification of Nocardia, Rhodococcus, Kocuria, Gordonia, Tsukamurella, and Listeria Species

    Science.gov (United States)

    Lee, Tai-Fen; Du, Shin-Hei; Teng, Shih-Hua; Liao, Chun-Hsing; Sheng, Wang-Hui; Teng, Lee-Jene

    2014-01-01

    We evaluated whether the Bruker Biotyper matrix-associated laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) system provides accurate species-level identifications of 147 isolates of aerobically growing Gram-positive rods (GPRs). The bacterial isolates included Nocardia (n = 74), Listeria (n = 39), Kocuria (n = 15), Rhodococcus (n = 10), Gordonia (n = 7), and Tsukamurella (n = 2) species, which had all been identified by conventional methods, molecular methods, or both. In total, 89.7% of Listeria monocytogenes, 80% of Rhodococcus species, 26.7% of Kocuria species, and 14.9% of Nocardia species (n = 11, all N. nova and N. otitidiscaviarum) were correctly identified to the species level (score values, ≥2.0). A clustering analysis of spectra generated by the Bruker Biotyper identified six clusters of Nocardia species, i.e., cluster 1 (N. cyriacigeorgica), cluster 2 (N. brasiliensis), cluster 3 (N. farcinica), cluster 4 (N. puris), cluster 5 (N. asiatica), and cluster 6 (N. beijingensis), based on the six peaks generated by ClinProTools with the genetic algorithm, i.e., m/z 2,774.477 (cluster 1), m/z 5,389.792 (cluster 2), m/z 6,505.720 (cluster 3), m/z 5,428.795 (cluster 4), m/z 6,525.326 (cluster 5), and m/z 16,085.216 (cluster 6). Two clusters of L. monocytogenes spectra were also found according to the five peaks, i.e., m/z 5,594.85, m/z 6,184.39, and m/z 11,187.31, for cluster 1 (serotype 1/2a) and m/z 5,601.21 and m/z 11,199.33 for cluster 2 (serotypes 1/2b and 4b). The Bruker Biotyper system was unable to accurately identify Nocardia (except for N. nova and N. otitidiscaviarum), Tsukamurella, or Gordonia species. Continuous expansion of the MALDI-TOF MS databases to include more GPRs is necessary. PMID:24759706

  7. Bruker biotyper matrix-assisted laser desorption ionization-time of flight mass spectrometry system for identification of Nocardia, Rhodococcus, Kocuria, Gordonia, Tsukamurella, and Listeria species.

    Science.gov (United States)

    Hsueh, Po-Ren; Lee, Tai-Fen; Du, Shin-Hei; Teng, Shih-Hua; Liao, Chun-Hsing; Sheng, Wang-Hui; Teng, Lee-Jene

    2014-07-01

    We evaluated whether the Bruker Biotyper matrix-associated laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) system provides accurate species-level identifications of 147 isolates of aerobically growing Gram-positive rods (GPRs). The bacterial isolates included Nocardia (n = 74), Listeria (n = 39), Kocuria (n = 15), Rhodococcus (n = 10), Gordonia (n = 7), and Tsukamurella (n = 2) species, which had all been identified by conventional methods, molecular methods, or both. In total, 89.7% of Listeria monocytogenes, 80% of Rhodococcus species, 26.7% of Kocuria species, and 14.9% of Nocardia species (n = 11, all N. nova and N. otitidiscaviarum) were correctly identified to the species level (score values, ≥ 2.0). A clustering analysis of spectra generated by the Bruker Biotyper identified six clusters of Nocardia species, i.e., cluster 1 (N. cyriacigeorgica), cluster 2 (N. brasiliensis), cluster 3 (N. farcinica), cluster 4 (N. puris), cluster 5 (N. asiatica), and cluster 6 (N. beijingensis), based on the six peaks generated by ClinProTools with the genetic algorithm, i.e., m/z 2,774.477 (cluster 1), m/z 5,389.792 (cluster 2), m/z 6,505.720 (cluster 3), m/z 5,428.795 (cluster 4), m/z 6,525.326 (cluster 5), and m/z 16,085.216 (cluster 6). Two clusters of L. monocytogenes spectra were also found according to the five peaks, i.e., m/z 5,594.85, m/z 6,184.39, and m/z 11,187.31, for cluster 1 (serotype 1/2a) and m/z 5,601.21 and m/z 11,199.33 for cluster 2 (serotypes 1/2b and 4b). The Bruker Biotyper system was unable to accurately identify Nocardia (except for N. nova and N. otitidiscaviarum), Tsukamurella, or Gordonia species. Continuous expansion of the MALDI-TOF MS databases to include more GPRs is necessary. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  8. Adsorption and laser-induced desorption of dimethylcadmium from silicon

    Science.gov (United States)

    Simonov, Alexander P.; Varakin, Vladimir N.

    1990-10-01

    The dynamics of Cd (cH ) 2 chemisorption and spontaneous decomposit ion on n-type 51(100) with native surface oxide the pathwa and efficiencies of the adsorbate desorption due to the absorption of the XeC1 laser radiation by silicon have been examined using laser-induced desorption miss spectrorrtry (LIDMS ) . The k inetics of these sur face processes has been found to depend on the preceding laser irradiation of the surface. The extremely fast chemisorption and efficient decomposition of the parent rrlecules have been observed on the irradiated silicon surface. The competition between intact dissociative and recombination desorption pathways was responsible for the observed laser fluence dependences of the desorption of Cd(CI-6) and i ts fragments. 1 . INTROOIJCTIct4 1 . 1 . Laser chemical vapour depos ition (LCVD). Laser-induced deposition of thin filme on solid surfaces by using volatile organometallic precursors has been the subject of numerous investigations in the 8Os2. Due to the spatial/temporal localization of laser radiation and the resonant nature of laser-rr1ecule interaction this deposition technique has such attractive features as submicrometer resolution of deposits high film growth rate and high quality lowtemperature processing. The deposition process can be controlled by varying the laser parameters (wavelength fluence beam spot on the substrate surface scanning speed ). A var iety of mater ials can be depos I ted using LCVD. Of special interest for microelectronics is the deposition of

  9. MoS{sub 2}/Ag nanohybrid: A novel matrix with synergistic effect for small molecule drugs analysis by negative-ion matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Yaju, E-mail: daisy19900911@hotmail.com [State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing, 210023 (China); Deng, Guoqing, E-mail: denggqq@sina.com [Department of Polymer Science and Engineering, Nanjing University, Nanjing, 210023 (China); Liu, Xiaohui, E-mail: lcswyh@126.com [State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing, 210023 (China); Sun, Liang, E-mail: sunliang@nju.edu.cn [State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing, 210023 (China); Li, Hui, E-mail: lihui@nju.edu.cn [State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing, 210023 (China); Cheng, Quan, E-mail: quan.cheng@ucr.edu [Department of Chemistry, University of California, Riverside, CA, 92521 (United States); Xi, Kai, E-mail: xikai@nju.edu.cn [Department of Polymer Science and Engineering, Nanjing University, Nanjing, 210023 (China); Xu, Danke, E-mail: xudanke@nju.edu.cn [State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing, 210023 (China)

    2016-09-21

    This paper reports a facile synthesis of molybdenum disulfide nanosheets/silver nanoparticles (MoS{sub 2}/Ag) hybrid and its use as an effective matrix in negative ion matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The nanohybrid exerts a strong synergistic effect, leading to high performance detection of small molecule analytes including amino acids, peptides, fatty acids and drugs. The enhancement of laser desorption/ionization (LDI) efficiency is largely attributed to the high surface roughness and large surface area for analyte adsorption, better dispersibility, increased thermal conductivity and enhanced UV energy absorption as compared to pure MoS{sub 2}. Moreover, both Ag nanoparticles and the edge of the MoS{sub 2} layers function as deprotonation sites for proton capture, facilitating the charging process in negative ion mode and promoting formation of negative ions. As a result, the MoS{sub 2}/Ag nanohybrid proves to be a highly attractive matrix in MALDI-TOF MS, with desired features such as high desorption/ionization efficiency, low fragmentation interference, high salt tolerance, and no sweet-spots for mass signal. These characteristic properties allowed for simultaneous analysis of eight different drugs and quantification of acetylsalicylic acid in the spiked human serum. This work demonstrates for the first time the fabrication and application of a novel MoS{sub 2}/Ag hybrid, and provides a new platform for use in the rapid and high throughput analysis of small molecules by mass spectrometry. - Highlights: • MoS{sub 2}/Ag nanohybrid was applied as a novel matrix in negative-ion MALDI-TOF MS. • The MoS{sub 2}/Ag nanohybrid exerted synergistic effect on the detection of small molecules. • The MoS{sub 2}/Ag nanohybrid showed good signal reproducibility and low background interferences comparing to organic matrices. • MoS{sub 2}/Ag allows simultaneous analysis of multiple drugs and quantification of

  10. Desorption by Femtosecond Laser Pulses : An Electron-Hole Effect?

    OpenAIRE

    D. M., NEWNS; T. F., HEINZ; J. A., MISEWICH; IBM Research Division, T. J. Watson Research Center; IBM Research Division, T. J. Watson Research Center; IBM Research Division, T. J. Watson Research Center

    1992-01-01

    Desorption of molecules from metal surfaces induced by femtosecond visible laser pulses has been reported. Since the lattice temperature rise is insufficient to explain desorption, an electronic mechanism is clearly responsible. It is shown that a theory based on direct coupling between the center-of-mass degree of freedom of the adsorbate and the electron-hole excitations of the substrate provides a satisfactory explanation of the various experimental findings.

  11. Femtosecond laser pulse induced desorption: A molecular dynamics simulation

    Energy Technology Data Exchange (ETDEWEB)

    Lončarić, Ivor, E-mail: ivor.loncaric@gmail.com [Centro de Física de Materiales CFM/MPC (CSIC-UPV/EHU), P. Manuel de Lardizabal 5, 20018 San Sebastián (Spain); Alducin, Maite [Centro de Física de Materiales CFM/MPC (CSIC-UPV/EHU), P. Manuel de Lardizabal 5, 20018 San Sebastián (Spain); Donostia International Physics Center DIPC, P. Manuel de Lardizabal 4, 20018 San Sebastián (Spain); Saalfrank, Peter [Institut für Chemie, Universität Potsdam, Karl-Liebknecht-Strasse 24-25, D-14476 Potsdam (Germany); Donostia International Physics Center DIPC, P. Manuel de Lardizabal 4, 20018 San Sebastián (Spain); Juaristi, J. Iñaki [Departamento de Física de Materiales, Facultad de Químicas, Universidad del País Vasco (UPV/EHU), Apartado 1072, 20080 San Sebastián (Spain); Centro de Física de Materiales CFM/MPC (CSIC-UPV/EHU), P. Manuel de Lardizabal 5, 20018 San Sebastián (Spain); Donostia International Physics Center DIPC, P. Manuel de Lardizabal 4, 20018 San Sebastián (Spain)

    2016-09-01

    In recent simulations of femtosecond laser induced desorption of molecular oxygen from the Ag(110) surface, it has been shown that depending on the properties (depth and electronic environment) of the well in which O{sub 2} is adsorbed, the desorption can be either induced dominantly by hot electrons or via excitations of phonons. In this work we explore whether the ratios between the desorption yields from different adsorption wells can be tuned by changing initial surface temperature and laser pulse properties. We show that the initial surface temperature is an important parameter, and that by using low initial surface temperatures the electronically mediated process can be favored. In contrast, laser properties seem to have only a modest influence on the results.

  12. Characterizing and optimizing a laser-desorption molecular beam source

    Science.gov (United States)

    Teschmit, Nicole; Długołecki, Karol; Gusa, Daniel; Rubinsky, Igor; Horke, Daniel A.; Küpper, Jochen

    2017-10-01

    The design and characterization of a new laser-desorption molecular beam source, tailored for use in x-ray free-electron laser and ultrashort-pulse laser imaging experiments, is presented. It consists of a single mechanical unit containing all source components, including the molecular-beam valve, the sample, and the fiber-coupled desorption laser, which is movable in five axes, as required for experiments at central facilities. Utilizing strong-field ionization, we characterize the produced molecular beam and evaluate the influence of desorption laser pulse energy, relative timing of valve opening and desorption laser, sample bar height, and which part of the molecular packet is probed on the sample properties. Strong-field ionization acts as a universal probe and allows detecting all species present in the molecular beam, and hence enables us to analyze the purity of the produced molecular beam, including molecular fragments. We present optimized experimental parameters for the production of the purest molecular beam, containing the highest yield of intact parent ions, which we find to be very sensitive to the placement of the desorbed-molecule plumes within the supersonic expansion.

  13. Laser-assisted electrochemistry

    Energy Technology Data Exchange (ETDEWEB)

    Glenn, D.F.

    1995-05-01

    The effect of laser irradiation on electrodeposition processes has been investigated. These studies demonstrated that the addition of laser irradiation to an electroplating process can dramatically enhance plating rates and current efficiencies, as well as improve the morphology of the resultant electrodeposit. During the course of these investigations, the mechanism for the laser enhancement of electrodeposition processes was determined. Experimental evidence was obtained to show that laser irradiation of the substrate results in increased metal ion concentrations at the surface of the electrode due to a laser-induced Soret effect. The laser-induced Soret effect has important implications for laser-assisted electrochemical processing. The increase in the surface concentration of ions allows efficient electrodeposition from dilute solutions. As such, laser- assisted electrodeposition may develop into an environmentally conscious manufacturing process by reducing waste and limiting worker exposure to toxic chemicals.

  14. Improvement of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry identification of difficult-to-identify bacteria and its impact in the workflow of a clinical microbiology laboratory.

    Science.gov (United States)

    Rodríguez-Sánchez, Belén; Marín, Mercedes; Sánchez-Carrillo, Carlos; Cercenado, Emilia; Ruiz, Adrián; Rodríguez-Créixems, Marta; Bouza, Emilio

    2014-05-01

    This study evaluates matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) capability for the identification of difficult-to-identify microorganisms. A total of 150 bacterial isolates inconclusively identified with conventional phenotypic tests were further assessed by 16S rRNA sequencing and by MALDI-TOF MS following 2 methods: a) a simplified formic acid-based, on-plate extraction and b) performing a tube-based extraction step. Using the simplified method, 29 isolates could not be identified. For the remaining 121 isolates (80.7%), we obtained a reliable identification by MALDI-TOF: in 103 isolates, the identification by 16S rRNA sequencing and MALDI TOF coincided at the species level (68.7% from the total 150 analyzed isolates and 85.1% from the samples with MALDI-TOF result), and in 18 isolates, the identification by both methods coincided at the genus level (12% from the total and 14.9% from the samples with MALDI-TOF results). No discordant results were observed. The performance of the tube-based extraction step allowed the identification at the species level of 6 of the 29 unidentified isolates by the simplified method. In summary, MALDI-TOF can be used for the rapid identification of many bacterial isolates inconclusively identified by conventional methods. Copyright © 2014 Elsevier Inc. All rights reserved.

  15. Heterotrophic monitoring at a drinking water treatment plant by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry after different drinking water treatments.

    Science.gov (United States)

    Sala-Comorera, Laura; Blanch, Anicet R; Vilaró, Carles; Galofré, Belén; García-Aljaro, Cristina

    2017-10-01

    The aim of this work was to assess the suitability of matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) for routine heterotrophic monitoring in a drinking water treatment plant. Water samples were collected from raw surface water and after different treatments during two campaigns over a 1-year period. Heterotrophic bacteria were studied and isolates were identified by MALDI-TOF MS. Moreover, the diversity index and the coefficient of population similarity were also calculated using biochemical fingerprinting of the populations studied. MALDI-TOF MS enabled us to characterize and detect changes in the bacterial community composition throughout the water treatment plant. Raw water showed a large and diverse population which was slightly modified after initial treatment steps (sand filtration and ultrafiltration). Reverse osmosis had a significant impact on the microbial diversity, while the final chlorination step produced a shift in the composition of the bacterial community. Although MALDI-TOF MS could not identify all the isolates since the available MALDI-TOF MS database does not cover all the bacterial diversity in water, this technique could be used to monitor bacterial changes in drinking water treatment plants by creating a specific protein profile database for tracking purposes.

  16. Optimization of experimental and modelling parameters for the differentiation of beverage spoiling yeasts by Matrix-Assisted-Laser-Desorption/Ionization-Time-of-Flight Mass Spectrometry (MALDI-TOF MS) in response to varying growth conditions.

    Science.gov (United States)

    Usbeck, Julia C; Kern, Carola C; Vogel, Rudi F; Behr, Jürgen

    2013-12-01

    The growth of spoiling yeasts in beverages results in reduced quality, economic and image losses. Therefore, biochemical and DNA-based identification methods have been developed but are mostly time-consuming and laborious. Matrix-Assisted-Laser-Desorption/Ionization-Time-Of-Flight Mass Spectrometry (MALDI-TOF MS) could deliver discriminative peptide mass fingerprints within minutes and could thus be a rapid and reliable tool for identification and differentiation. However, routine analysis of yeasts by MALDI-TOF MS is yet impaired by low reproducibility and effects of different physiological states of organisms on the reliability of the identification method are still controversial. The aim of this study was to optimize sample preparation and measurement parameterization using three spoilage yeasts (Saccharomyces cerevisiae var. diastaticus, Wickerhamomyces anomalus and Debaryomyces hansenii). The influence of environmental or physiological parameters including oxygen availability, different nutrients, cell density and growth phase were analysed and revealed small differences in mass fingerprints. Yeasts grown in the presence or absence of oxygen were precisely differentiated along these differences in mass fingerprints and a crude classification of growth phase was possible. Cell concentration did not affect the spectra distinctly, neither qualitatively nor quantitatively, and an influence of available nutrients could not be measured in each case. However, core mass peaks remained constant under all tested conditions enabling reliable identification. Copyright © 2013 Elsevier Ltd. All rights reserved.

  17. Comparative evaluation of two matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) systems for the identification of clinically significant yeasts.

    Science.gov (United States)

    Jamal, W Y; Ahmad, S; Khan, Z U; Rotimi, V O

    2014-09-01

    To prospectively evaluate the performance of two matrix-assisted laser desorption/ionization time-of-flight mass spectrometry systems (MALDI-TOF MS) for the identification of clinically significant yeast isolates compared to the VITEK 2 system. One hundred and eighty-eight consecutive yeast isolates were analyzed by Bruker Biotyper and VITEK MS. The results were compared with the conventional VITEK 2 yeast identification system. Discrepant results were resolved by direct sequencing of rDNA. Accurate identification by VITEK 2, VITEK MS, and Bruker Biotyper MS was 94.1% (177/188), 93.0% (175/188), and 92.6% (174/188), respectively. Three isolates were not identified by VITEK MS, while nine Candida orthopsilosis were misidentified as Candida parapsilosis, as this species is not present in its database. Eleven isolates were not identified or were wrongly identified by Bruker Biotyper and although another 14 were correctly identified, the score was unreliable at MALDI-TOF MS systems was essentially comparable to that of the conventional VITEK 2 yeast identification system. However, future expansion of the databases may further improve the outcome and accuracy of identification of yeast species. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

  18. Characterization of low-molecular weight iodine-terminated polyethylenes by gas chromatography/mass spectrometry and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry with the use of derivatization.

    Science.gov (United States)

    Zaikin, Vladimir G; Borisov, Roman S; Polovkov, Nikolai Yu; Zhilyaev, Dmitry I; Vinogradov, Aleksei A; Ivanyuk, Aleksei V

    2013-01-01

    Gas chromatography/mass spectrometry (GC/MS) and matrix-assisted laser desorption/ionization time-of-flight (MALDI-ToF) mass spectrometry, in conjunction with various derivatization approaches, have been applied to structure determination of individual oligomers and molecular-mass distributions (MMD) in low-molecular mass polyethylene having an iodine terminus. Direct GC/MS analysis has shown that the samples under investigation composed of polyethyelene-iodides (major components) and n-alkanes. Exchange reaction with methanol in the presence of NaOH gave rise to methoxy-derivatives and n-alkenes. Electron ionization mass spectra have shown that the former contained terminal methoxy groups indicating the terminal position of the iodine atom in the initial oligomers. MMD parameters have been determined with the aid of MALDI mass spectrometry followed by preliminary derivatization-formation of covalently bonded charge through the reaction of iodides with triphenylphosphine, trialkylamines, pyridine or quinoline. The mass spectra revealed well-resolved peaks for cationic parts of derivatized oligomers allowing the determination of MMD. The latter values have been compared with those calculated from GC/MS data.

  19. Evaluation of repetitive-PCR and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for rapid strain typing of Bacillus coagulans.

    Science.gov (United States)

    Sato, Jun; Nakayama, Motokazu; Tomita, Ayumi; Sonoda, Takumi; Hasumi, Motomitsu; Miyamoto, Takahisa

    2017-01-01

    In order to establish rapid and accurate typing method for Bacillus coagulans strains which is important for controlling in some canned foods and tea-based beverages manufacturing because of the high-heat resistance of the spores and high tolerance of the vegetative cells to catechins and chemicals, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and repetitive-PCR (rep-PCR) were evaluated. For this purpose, 28 strains of B. coagulans obtained from various culture collections were tested. DNA sequence analyses of the genes encoding 16S rRNA and DNA gyrase classified the test strains into two and three groups, respectively, regardless of their phenotypes. Both MALDI-TOF MS and rep-PCR methods classified the test strains in great detail. Strains classified in each group showed similar phenotypes, such as carbohydrate utilization determined using API 50CH. In particular, the respective two pairs of strains which showed the same metabolic characteristic were classified into the same group by both MALDI-TOF MS and rep-PCR methods separating from the other strains. On the other hand, the other strains which have the different profiles of carbohydrate utilization were separated into different groups by these methods. These results suggested that the combination of MALDI-TOF MS and rep-PCR analyses was advantageous for the rapid and detailed typing of bacterial strains in respect to both phenotype and genotype.

  20. Top-Down Proteomic Identification of Shiga Toxin 2 Subtypes from Shiga Toxin-Producing Escherichia coli by Matrix-Assisted Laser Desorption Ionization–Tandem Time of Flight Mass Spectrometry

    Science.gov (United States)

    Zaragoza, William J.; Sultan, Omar; Woo, Nathan; Quiñones, Beatriz; Cooley, Michael B.; Mandrell, Robert E.

    2014-01-01

    We have analyzed 26 Shiga toxin-producing Escherichia coli (STEC) strains for Shiga toxin 2 (Stx2) production using matrix-assisted laser desorption ionization (MALDI)–tandem time of flight (TOF-TOF) tandem mass spectrometry (MS/MS) and top-down proteomic analysis. STEC strains were induced to overexpress Stx2 by overnight culturing on solid agar supplemented with either ciprofloxacin or mitomycin C. Harvested cells were lysed by bead beating, and unfractionated bacterial cell lysates were ionized by MALDI. The A2 fragment of the A subunit and the mature B subunit of Stx2 were analyzed by MS/MS. Sequence-specific fragment ions were used to identify amino acid subtypes of Stx2 using top-down proteomic analysis using software developed in-house at the U.S. Department of Agriculture (USDA). Stx2 subtypes (a, c, d, f, and g) were identified on the basis of the mass of the A2 fragment and the B subunit as well as from their sequence-specific fragment ions by MS/MS (postsource decay). Top-down proteomic identification was in agreement with DNA sequencing of the full Stx2 operon (stx2) for all strains. Top-down results were also compared to a bioassay using a Vero-d2EGFP cell line. Our results suggest that top-down proteomic identification is a rapid, highly specific technique for distinguishing Stx2 subtypes. PMID:24584253

  1. Two tools for applying chromatographic retention data to the mass-based identification of peptides during hydrogen/deuterium exchange experiments by nano-liquid chromatography/matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Gershon, P D

    2010-12-15

    Two tools are described for integrating LC elution position with mass-based data in hydrogen-deuterium exchange (HDX) experiments by nano-liquid chromatography/matrix-assisted laser desorption/ionization mass spectrometry (nanoLC/MALDI-MS, a novel approach to HDX-MS). The first of these, 'TOF2H-Z Comparator', highlights peptides in HDX experiments that are potentially misidentified on the basis of mass alone. The program first calculates normalized values for the organic solvent concentration responsible for the elution of ions in nanoLC/MALDI HDX experiments. It then allows the solvent gradients for the multiple experiments contributing to an MS/MS-confirmed peptic peptide library to be brought into mutual alignment by iteratively re-modeling variables among LC parameters such as gradient shape, solvent species, fraction duration and LC dead time. Finally, using the program, high-probability chromatographic outliers can be flagged within HDX experimental data. The role of the second tool, 'TOF2H-XIC Comparator', is to normalize the LC chromatograms corresponding to all deuteration timepoints of all HDX experiments of a project, to a common reference. Accurate normalization facilitates the verification of chromatographic consistency between all ions whose spectral segments contribute to particular deuterium uptake plots. Gradient normalization in this manner revealed chromatographic inconsistencies between ions whose masses were either indistinguishable or separated by precise isotopic increments. Copyright © 2010 John Wiley & Sons, Ltd.

  2. Quantitative determination of nicotinic acid in micro liter volume of urine sample by drop-to-drop solvent microextraction coupled to matrix assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Shrivas, Kamlesh; Patel, Devesh Kumar

    2011-01-01

    Drop-to-drop solvent microextraction (DDSME) coupled with matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS) for quantitative determination of nicotinic acid in one drop of urine sample has been proposed. All parameters, such as type of organic solvent, extraction time, exposure volume solvent, pH of the sample solution that affecting the separation and preconcentration of nicotinic acid were investigated. Under the optimal conditions, the detection limit of the method was 20 ng mL(-1) and the relative standard deviations (RSD) for determination of the nicotinic acid were in the range of 8.0-12.5%. The calculated calibration curves gave linearity in the range of 80-1000 ng mL(-1). The main advantages of the proposed method are simple, fast, and small amount of sample solution is used for separation and preconcentration of nicotinic acid. This method could be also useful for the analysis of other interested analytes in small volume of biological samples, like plasma, saliva and urine, where the availability of samples are limited. Copyright © 2010 Elsevier B.V. All rights reserved.

  3. Top-down proteomic identification of Shiga toxin 2 subtypes from Shiga toxin-producing Escherichia coli by matrix-assisted laser desorption ionization-tandem time of flight mass spectrometry.

    Science.gov (United States)

    Fagerquist, Clifton K; Zaragoza, William J; Sultan, Omar; Woo, Nathan; Quiñones, Beatriz; Cooley, Michael B; Mandrell, Robert E

    2014-05-01

    We have analyzed 26 Shiga toxin-producing Escherichia coli (STEC) strains for Shiga toxin 2 (Stx2) production using matrix-assisted laser desorption ionization (MALDI)-tandem time of flight (TOF-TOF) tandem mass spectrometry (MS/MS) and top-down proteomic analysis. STEC strains were induced to overexpress Stx2 by overnight culturing on solid agar supplemented with either ciprofloxacin or mitomycin C. Harvested cells were lysed by bead beating, and unfractionated bacterial cell lysates were ionized by MALDI. The A2 fragment of the A subunit and the mature B subunit of Stx2 were analyzed by MS/MS. Sequence-specific fragment ions were used to identify amino acid subtypes of Stx2 using top-down proteomic analysis using software developed in-house at the U.S. Department of Agriculture (USDA). Stx2 subtypes (a, c, d, f, and g) were identified on the basis of the mass of the A2 fragment and the B subunit as well as from their sequence-specific fragment ions by MS/MS (postsource decay). Top-down proteomic identification was in agreement with DNA sequencing of the full Stx2 operon (stx2) for all strains. Top-down results were also compared to a bioassay using a Vero-d2EGFP cell line. Our results suggest that top-down proteomic identification is a rapid, highly specific technique for distinguishing Stx2 subtypes.

  4. Role of the UV absorber as a matrix in matrix-assisted laser desorption/ionization mass spectrometric analysis of a mixture of a UV absorber and a stabilizer.

    Science.gov (United States)

    Im, Song-Hee; Choi, Sung-Seen

    2010-09-01

    A mixture of a UV absorber (Tinuvin 234 or Tinuvin 329) and a UV stabilizer (Tinuvin 770) was analyzed using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) without any matrix. Fragmentation patterns of the UV absorbers and stabilizer were also investigated. The mass spectra showed the [M+H](+) ions and some fragment ions. Tinuvin 234, Tinuvin 329, and Tinuvin 770 generated three (m/z 119, 370, 432), one (m/z 252), and two (m/z 124 and 140) fragment ions, respectively. These fragment ions can be used to identify the chemical structures of the UV absorbers and stabilizer. Since the UV absorber performed a role as the matrix, the ion abundance of the UV stabilizer was enhanced by mixing with the UV absorber. When organic materials extracted from polypropylene (PP) containing the UV absorber and stabilizer were directly analyzed using MALDI-MS without any matrix, the protonated molecule of the UV stabilizer was detected in abundance but the product ions of the UV absorber were not observed. When 2,5-dihydroxybenzoic acid was used as a matrix, the protonated molecule of the UV absorber was observed. 2010 John Wiley & Sons, Ltd.

  5. Graphene/TiO2 nanocomposite based solid-phase extraction and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for lipidomic profiling of avocado (Persea americana Mill.).

    Science.gov (United States)

    Shen, Qing; Yang, Mei; Li, Linqiu; Cheung, Hon-Yeung

    2014-12-10

    Phospholipids possess important physiological, structural and nutritional functions in biological systems. This study described a solid-phase extraction (SPE) method, employing graphene and titanium dioxide (G/TiO2) nanocomposite as sorbent, for the selective isolation and enrichment of phospholipids from avocado (Persea americana Mill.). Based on the principal that the phosphoryl group in the phospholipid can interact with TiO2 via a bridging bidentate mode, an optimum condition was established for SPE, and was successfully applied to prepare avocado samples. The extracts were monitored by matrix-assisted laser desorption ionization time-of-flight/tandem mass spectrometry (MALDI-TOF/MS) in both positive-ion and negative-ion modes. Results showed that phospholipids could be efficiently extracted in a clean manner by G/TiO2 based SPE. In addition, the signals of phospholipids were enhanced while the noise was reduced. Some minor peaks became more obvious. In conclusion, the nanocomposite material of G/TiO2 was proved to be a promising sorbent for selective separation of phospholipids from crude lipid extract. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

  6. Synthesis and matrix-assisted laser desorption/ionization time-of-flight characterization of bisphenol-A copolyformals containing nickel(II)/Schiff base, eicosane and 2-butene units in the main chain.

    Science.gov (United States)

    Vitalini, Daniele; Spina, Emanuela; Rapisardi, Roberto; Scamporrino, Emilio; Mineo, Placido

    2006-01-01

    Some bisphenol-A copolyformals, containing in the main chain different amounts of a Ni-diimine nonlinear optical (NLO)-chromophore, eicosane and/or 2-butene units, were synthesized by condensation reaction between dibromomethane and suitable mixtures of Ni(II)/Schiff base complex, 1,20-di(bisphenol-A)ether-eicosane and/or 1,4-di(bisphenol-A)ether-2-butene. Structural composition and thermal properties of polymeric materials were inferred by analyses by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS), nuclear magnetic resonance (NMR), gel permeation chromatography (GPC), differential scanning calorimetry (DSC) and thermogravimetry (TG). MALDI-TOF data show that both Ni-diimine and unsaturated units are present in the copolyformals with a homogeneous arrangement in all the polymer mass range (GPC data). It has also been ascertained that the glass transition temperature (Tg) of the copolymers changes as a consequence of the abundance of aliphatic units in the macromolecules (DSC data). Cross-linking experiments of the copolymer at temperatures near the Tg value and under UV irradiation were also performed. Copyright 2006 John Wiley & Sons, Ltd.

  7. Identification and Quantification of N-Acyl Homoserine Lactones Involved in Bacterial Communication by Small-Scale Synthesis of Internal Standards and Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry

    Science.gov (United States)

    Leipert, Jan; Treitz, Christian; Leippe, Matthias; Tholey, Andreas

    2017-08-01

    N-acyl homoserine lactones (AHL) are small signal molecules involved in the quorum sensing of many gram-negative bacteria, and play an important role in biofilm formation and pathogenesis. Present analytical methods for identification and quantification of AHL require time-consuming sample preparation steps and are hampered by the lack of appropriate standards. By aiming at a fast and straightforward method for AHL analytics, we investigated the applicability of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Suitable MALDI matrices, including crystalline and ionic liquid matrices, were tested and the fragmentation of different AHL in collision-induced dissociation MS/MS was studied, providing information about characteristic marker fragments ions. Employing small-scale synthesis protocols, we established a versatile and cost-efficient procedure for fast generation of isotope-labeled AHL standards, which can be used without extensive purification and yielded accurate standard curves. Quantitative analysis was possible in the low pico-molar range, with lower limits of quantification reaching from 1 to 5 pmol for different AHL. The developed methodology was successfully applied in a quantitative MALDI MS analysis of low-volume culture supernatants of Pseudomonas aeruginosa. [Figure not available: see fulltext.

  8. Identification of blood culture isolates directly from positive blood cultures by use of matrix-assisted laser desorption ionization-time of flight mass spectrometry and a commercial extraction system: analysis of performance, cost, and turnaround time.

    Science.gov (United States)

    Lagacé-Wiens, Philippe R S; Adam, Heather J; Karlowsky, James A; Nichol, Kimberly A; Pang, Paulette F; Guenther, Jodi; Webb, Amanda A; Miller, Crystal; Alfa, Michelle J

    2012-10-01

    Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry represents a revolution in the rapid identification of bacterial and fungal pathogens in the clinical microbiology laboratory. Recently, MALDI-TOF has been applied directly to positive blood culture bottles for the rapid identification of pathogens, leading to reductions in turnaround time and potentially beneficial patient impacts. The development of a commercially available extraction kit (Bruker Sepsityper) for use with the Bruker MALDI BioTyper has facilitated the processing required for identification of pathogens directly from positive from blood cultures. We report the results of an evaluation of the accuracy, cost, and turnaround time of this method for 61 positive monomicrobial and 2 polymicrobial cultures representing 26 species. The Bruker MALDI BioTyper with the Sepsityper gave a valid (score, >1.7) identification for 85.2% of positive blood cultures with no misidentifications. The mean reduction in turnaround time to identification was 34.3 h (P turnaround time to identification.

  9. Identification of Blood Culture Isolates Directly from Positive Blood Cultures by Use of Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry and a Commercial Extraction System: Analysis of Performance, Cost, and Turnaround Time

    Science.gov (United States)

    Adam, Heather J.; Karlowsky, James A.; Nichol, Kimberly A.; Pang, Paulette F.; Guenther, Jodi; Webb, Amanda A.; Miller, Crystal; Alfa, Michelle J.

    2012-01-01

    Matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) mass spectrometry represents a revolution in the rapid identification of bacterial and fungal pathogens in the clinical microbiology laboratory. Recently, MALDI-TOF has been applied directly to positive blood culture bottles for the rapid identification of pathogens, leading to reductions in turnaround time and potentially beneficial patient impacts. The development of a commercially available extraction kit (Bruker Sepsityper) for use with the Bruker MALDI BioTyper has facilitated the processing required for identification of pathogens directly from positive from blood cultures. We report the results of an evaluation of the accuracy, cost, and turnaround time of this method for 61 positive monomicrobial and 2 polymicrobial cultures representing 26 species. The Bruker MALDI BioTyper with the Sepsityper gave a valid (score, >1.7) identification for 85.2% of positive blood cultures with no misidentifications. The mean reduction in turnaround time to identification was 34.3 h (P turnaround time to identification. PMID:22875888

  10. Rapid detection of meticillin-resistant Staphylococcus aureus bacteraemia using combined three-hour short-incubation matrix-assisted laser desorption/ionization time-of-flight MS identification and Alere Culture Colony PBP2a detection test.

    Science.gov (United States)

    Delport, Johannes Andries; Mohorovic, Ivor; Burn, Sandi; McCormick, John Kenneth; Schaus, David; Lannigan, Robert; John, Michael

    2016-07-01

    Meticillin-resistant Staphylococcus aureus (MRSA) bloodstream infection is responsible for significant morbidity, with mortality rates as high as 60 % if not treated appropriately. We describe a rapid method to detect MRSA in blood cultures using a combined three-hour short-incubation BRUKER matrix-assisted laser desorption/ionization time-of-flight MS BioTyper protocol and a qualitative immunochromatographic assay, the Alere Culture Colony Test PBP2a detection test. We compared this combined method with a molecular method detecting the nuc and mecA genes currently performed in our laboratory. One hundred and seventeen S. aureus blood cultures were tested of which 35 were MRSA and 82 were meticillin-sensitive S. aureus (MSSA). The rapid combined test correctly identified 100 % (82/82) of the MSSA and 85.7 % (30/35) of the MRSA after 3 h. There were five false negative results where the isolates were correctly identified as S. aureus, but PBP2a was not detected by the Culture Colony Test. The combined method has a sensitivity of 87.5 %, specificity of 100 %, a positive predictive value of 100 % and a negative predictive value of 94.3 % with the prevalence of MRSA in our S. aureus blood cultures. The combined rapid method offers a significant benefit to early detection of MRSA in positive blood cultures.

  11. Accuracy of PCR targeting different markers for Staphylococcus aureus identification: a comparative study using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry as the gold standard.

    Science.gov (United States)

    Saraiva, Mauro M; De Leon, Candice M; Santos, Silvana C; Stipp, Danilo T; Souza, Miliane M; Santos Filho, Lauro; Gebreyes, Wondwossen A; Oliveira, Celso J

    2018-03-01

    Staphylococcus aureus is considered a major pathogen in veterinary and human medicine, and the emergence of multidrug-resistant strains, such as livestock-associated methicillin-resistant S. aureus, means that reliable, inexpensive, and fast methods are required to identify S. aureus obtained from animal sources. We tested the accuracy of a PCR targeting the genes femA, nuc, and coa in identifying S. aureus from animals. A total of 157 Staphylococcus spp. isolates were examined by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry; 18 different Staphylococcus species were identified. Of 68 S. aureus isolates, the genes femA, nuc, and coa were found in 61, 53, and 32 isolates, respectively. Considering MALDI-TOF as the gold standard, the PCR assays targeting all 3 genes showed 100% specificity; the sensitivity values were 89.7, 77.9, and 47.0% for femA, nuc, and coa, respectively. Sensitivity was 100% when femA and nuc markers were targeted simultaneously. These results confirm PCR as an accurate method to identify S. aureus species from animal sources and strongly suggest the simultaneous use of primers targeting femA and nuc genes.

  12. N-phosphorylation labeling for analysis of twenty natural amino acids and small peptides by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Gao, Xiang; Bi, Xin; Wei, Juntong; Peng, Zhimin; Liu, Hongxia; Jiang, Yuyang; Wei, Wei; Cai, Zongwei

    2013-05-07

    N-phosphorylation labeling was utilized to analyze the low molecular weight compounds by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). A wide range of natural amino acids and short peptides was successfully analyzed by MALDI-TOF MS without matrix background interferences. The N-phosphorylation labeling reaction was carried out easily within 30 min in a one-pot reaction under mild reaction conditions. The phosphoryl derivatization reaction is a global labeling approach with high selectivity and high specificity with targeting only on the N-terminal and ε-amino group of Lys. The incorporation of a neutral phosphoryl group with high gas-phase affinity of protons not only improves the ionization efficiency of target molecules and simultaneously decreases the ion suppression effects in MALDI-TOF MS analysis, but also greatly reduces or eliminates the matrix background interferences by suppressing the matrix signals and increasing the molecular weight of the targeted compounds. By applying the N-phosphorylation labeling approach, many amino acids could be detected in serum samples by using MALDI-TOF MS.

  13. Evaluation of matrix-assisted laser desorption ionization-time of flight mass spectrometry-based VITEK MS system for the identification of Acinetobacter species from blood cultures: comparison with VITEK 2 and MicroScan systems.

    Science.gov (United States)

    Lee, Seung Yeob; Shin, Jong Hee; Kim, Soo Hyun; Shin, Myung Geun; Suh, Soon Pal; Ryang, Dong Wook

    2015-01-01

    Acinetobacter species are the leading cause of bloodstream infection (BSI), but their correct identification is challenging. We evaluated the matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS)-based VITEK MS (bioMérieux, France), and two automated systems, VITEK 2 (bioMérieux) and MicroScan (Siemens, USA) for identification of Acinetobacter BSI isolates. A total of 187 BSI isolates recovered at a university hospital in Korea between 2010 and 2012 were analyzed. The identification results obtained using VITEK MS and two automated systems were compared with those of rpoB sequencing. Of 187 isolates analyzed, 176 were identified to the species level by rpoB sequencing: the Acinetobacter baumannii group (ABG; 101 A. baumannii, 43 A. nosocomialis, 10 A. pittii isolates) was most commonly identified (82.4%), followed by Acinetobacter genomic species 13BJ/14TU (5.3%), A. ursingii (2.1%), A. soli (2.1%), A. bereziniae (1.1%), and A. junii (1.1%). Correct identification rates to the species group (ABG) level or the species level was comparable among the three systems (VITEK MS, 90.3%; VITEK 2, 89.2%; MicroScan, 86.9%). However, VITEK MS generated fewer misidentifications (0.6%) than VITEK 2 (10.8%) and MicroScan (13.1%) (PAcinetobacter BSI isolates, with fewer misidentifications and better discrimination between the ABG and non-ABG isolates.

  14. Microorganisms in cryopreserved semen and culture media used in the in vitro production (IVP) of bovine embryos identified by matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS).

    Science.gov (United States)

    Zampieri, Dávila; Santos, Vanessa G; Braga, Patrícia A C; Ferreira, Christina R; Ballottin, Daniela; Tasic, Ljubica; Basso, Andréa C; Sanches, Bruno V; Pontes, José H F; da Silva, Bárbara Pereira; Garboggini, Fabiana Fantinatti; Eberlin, Marcos N; Tata, Alessandra

    2013-09-01

    Commercial cattle breeders produce their own herd offspring for the dairy and beef market using artificial insemination. The procedure involves sanitary risks associated with the collection and commercialization of the germplasm, and the in vitro production and transfer of the bovine embryos must be monitored by strict health surveillance. To avoid the spreading of infectious diseases, one must rely on using controlled and monitored germplasm, media, and reagents that are guaranteed free of pathogens. In this article, we investigated the use of a new mass spectrometric approach for fast and accurate identification of bacteria and fungi in bovine semen and in culture media employed in the embryo in vitro production process. The microorganisms isolated from samples obtained in a commercial bovine embryo IVP setting were identified in a few minutes by their conserved peptide/protein profile, obtained applying matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS), matched against a commercial database. The successful microorganisms MS identification has been confirmed by DNA amplification and sequencing. Therefore, the MS technique seems to offer a powerful tool for rapid and accurate microorganism identification in semen and culture media samples. Copyright © 2013 Elsevier Inc. All rights reserved.

  15. Methylobacterium Species Promoting Rice and Barley Growth and Interaction Specificity Revealed with Whole-Cell Matrix-Assisted Laser Desorption/Ionization-Time-of-Flight Mass Spectrometry (MALDI-TOF/MS) Analysis.

    Science.gov (United States)

    Tani, Akio; Sahin, Nurettin; Fujitani, Yoshiko; Kato, Akiko; Sato, Kazuhiro; Kimbara, Kazuhide

    2015-01-01

    Methylobacterium species frequently inhabit plant surfaces and are able to utilize the methanol emitted from plants as carbon and energy sources. As some of the Methylobacterium species are known to promote plant growth, significant attention has been paid to the mechanism of growth promotion and the specificity of plant-microbe interactions. By screening our Methylobacterium isolate collection for the high growth promotion effect in vitro, we selected some candidates for field and pot growth tests for rice and barley, respectively. We found that inoculation resulted in better ripening of rice seeds, and increased the size of barley grains but not the total yield. In addition, using whole-cell matrix-assister laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF/MS) analysis, we identified and classified Methylobacterium isolates from Methylobacterium-inoculated rice plants. The inoculated species could not be recovered from the rice plants, and in some cases, the Methylobacterium community structure was affected by the inoculation, but not with predomination of the inoculated species. The isolates from non-inoculated barley of various cultivars grown in the same field fell into just two species. These results suggest that there is a strong selection pressure at the species level of Methylobacterium residing on a given plant species, and that selection of appropriate species that can persist on the plant is important to achieve growth promotion.

  16. Direct identification of microorganisms from positive blood cultures using the lysis-filtration technique and matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS): a multicentre study.

    Science.gov (United States)

    Farina, Claudio; Arena, Fabio; Casprini, Patrizia; Cichero, Paola; Clementi, Massimo; Cosentino, Marina; Degl'Innocenti, Roberto; Giani, Tommaso; Luzzaro, Francesco; Mattei, Romano; Mauri, Carola; Nardone, Maria; Rossolini, Gian Maria; Serna Ortega, Paula Andrea; Vailati, Francesca

    2015-04-01

    Microbial identification from blood cultures is essential to institute optimal antibiotic therapy and improve survival possibilities. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has been successfully applied to identify bacteria and yeasts from positive blood cultures broths. The aim of this multicentre study was to evaluate the reliability of the lysis-filtration technique associated with MALDI-TOF MS to directly identify microorganisms from 765 positive blood cultures collected in six Italian hospitals. Overall, 675/765 (78.1%) blood isolates were correctly identified at the species level, with significant differences between Gram-negative and Gram-positive bacteria (92.6%, and 69.8%, respectively). Some difficulties arise in identifying Streptococcus pneumoniae, Staphylococcus aureus, yeasts and anaerobes. The lysis-filtration protocol is a suitable procedure in terms of performance in identifying microorganisms, but it is quite expensive and technically time-consuming since the time of filtration is not regular for all the samples. The application of the MALDI-TOF MS technique to the direct microbial identification from positive blood cultures is a very promising approach, even if more experience must be gained to minimize errors and costs.

  17. Coupling of metal-organic frameworks-containing monolithic capillary-based selective enrichment with matrix-assisted laser desorption ionization-time-of-flight mass spectrometry for efficient analysis of protein phosphorylation.

    Science.gov (United States)

    Li, Daojin; Yin, Danyang; Chen, Yang; Liu, Zhen

    2017-05-19

    Protein phosphorylation is a major post-translational modification, which plays a vital role in cellular signaling of numerous biological processes. Mass spectrometry (MS) has been an essential tool for the analysis of protein phosphorylation, for which it is a key step to selectively enrich phosphopeptides from complex biological samples. In this study, metal-organic frameworks (MOFs)-based monolithic capillary has been successfully prepared as an effective sorbent for the selective enrichment of phosphopeptides and has been off-line coupled with matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) for efficient analysis of phosphopeptides. Using š-casein as a representative phosphoprotein, efficient phosphorylation analysis by this off-line platform was verified. Phosphorylation analysis of a nonfat milk sample was also demonstrated. Through introducing large surface areas and highly ordered pores of MOFs into monolithic column, the MOFs-based monolithic capillary exhibited several significant advantages, such as excellent selectivity toward phosphopeptides, superb tolerance to interference and simple operation procedure. Because of these highly desirable properties, the MOFs-based monolithic capillary could be a useful tool for protein phosphorylation analysis. Copyright © 2016 Elsevier B.V. All rights reserved.

  18. High-sensitivity matrix-assisted laser desorption/ionization Fourier transform mass spectrometry analyses of small carbohydrates and amino acids using oxidized carbon nanotubes prepared by chemical vapor deposition as matrix.

    Science.gov (United States)

    Wang, Cui-hong; Li, Jian; Yao, Sheng-jun; Guo, Yin-long; Xia, Xing-hua

    2007-12-05

    In matrix-assisted laser desorption/ionization (MALDI) Fourier transform mass spectrometry (FTMS) analyses of small oligosaccharides and amino acids, high sensitivities for oligosaccharides (10 fmol) were obtained by introducing oxidized carbon nanotubes (CNTs) with short and open-end structure as valuable matrix. The CNTs were deposited in porous anodic alumina (PAA) templates by chemical vapor deposition. Transmission electron microscopy (TEM) images show that those CNTs include low levels of amorphous carbon. Thus, the background interference signals generally caused by amorphous carbon powder in CNTs can be reduced effectively. Experiments also confirmed that the FTMS signal intensity of CNTs prepared in PAA template is much lower than that of commercial multi-wall carbon nanotubes (MCNTs). Moreover, the purified process for CNTs with mixed acid (H2SO4 and HNO3) also contributed to the minimization of background. Intense signals corresponding to alkali cation adduct of neutral carbohydrates and amino acids have been acquired. In addition, reliable quantitative analyses for urine and corn root were also achieved successfully. The present work will open a new way to the application of oxidized CNTs as an effective matrix in MALDI MS research.

  19. Comparison of phenotypic methods and matrix-assisted laser desorption ionisation time-of-flight mass spectrometry for the identification of aero-tolerant Actinomyces spp. isolated from soft-tissue infections.

    Science.gov (United States)

    Ng, L S Y; Sim, J H C; Eng, L C; Menon, S; Tan, T Y

    2012-08-01

    Aero-tolerant Actinomyces spp. are an under-recognised cause of cutaneous infections, in part because identification using conventional phenotypic methods is difficult and may be inaccurate. Matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF MS) is a promising new technique for bacterial identification, but with limited data on the identification of aero-tolerant Actinomyces spp. This study evaluated the accuracy of a phenotypic biochemical kit, MALDI-TOF MS and genotypic identification methods for the identification of this problematic group of organisms. Thirty aero-tolerant Actinomyces spp. were isolated from soft-tissue infections over a 2-year period. Species identification was performed by 16 s rRNA sequencing and genotypic results were compared with results obtained by API Coryne and MALDI-TOF MS. There was poor agreement between API Coryne and genotypic identification, with only 33% of isolates correctly identified to the species level. MALDI-TOF MS correctly identified 97% of isolates to the species level, with 33% of identifications achieved with high confidence scores. MALDI-TOF MS is a promising new tool for the identification of aero-tolerant Actinomyces spp., but improvement of the database is required in order to increase the confidence level of identification.

  20. Creation of an In-House Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry Corynebacterineae Database Overcomes Difficulties in Identification of Nocardia farcinica Clinical Isolates.

    Science.gov (United States)

    Paściak, Mariola; Dacko, Władysław; Sikora, Joanna; Gurlaga, Danuta; Pawlik, Krzysztof; Miękisiak, Grzegorz; Gamian, Andrzej

    2015-08-01

    Nocardiosis is a rare disease that is caused by Gram-positive actinobacteria of the Nocardia genus and affects predominantly immunocompromised patients. In its disseminated form, it has a predilection for the central nervous system and is associated with high mortality rates. Therefore, prompt identification of the pathogen is critical. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry is a relatively novel technique used for identification of microorganisms. In this work, an upgraded MALDI-TOF Biotyper database containing Corynebacterineae representatives of strains deposited in the Polish Collection of Microorganisms was created and used for identification of the strain isolated from a nocardial brain abscess, mimicking a brain tumor, in an immunocompetent patient. Testing with the API Coryne system initially incorrectly identified Rhodococcus sp., while chemotaxonomic tests, especially mycolic acid analysis, enabled correct Nocardia identification only at the genus level. Subsequent sequence analysis of 16S rRNA and secA1 genes confirmed the identification. To improve the accuracy of the results, an in-house database was constructed using optimized parameters; with the use of the database, the strain was eventually identified as Nocardia farcinica. Clinical laboratories processing various clinical strains can upgrade a commercial database to improve and to accelerate the results obtained. This is especially important in the case of Nocardia, for which valid microbial diagnosis remains challenging; reference laboratories are often required to identify and to survey these rare actinobacteria. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  1. Creation of an In-House Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry Corynebacterineae Database Overcomes Difficulties in Identification of Nocardia farcinica Clinical Isolates

    Science.gov (United States)

    Dacko, Władysław; Sikora, Joanna; Gurlaga, Danuta; Pawlik, Krzysztof; Miękisiak, Grzegorz; Gamian, Andrzej

    2015-01-01

    Nocardiosis is a rare disease that is caused by Gram-positive actinobacteria of the Nocardia genus and affects predominantly immunocompromised patients. In its disseminated form, it has a predilection for the central nervous system and is associated with high mortality rates. Therefore, prompt identification of the pathogen is critical. Matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) mass spectrometry is a relatively novel technique used for identification of microorganisms. In this work, an upgraded MALDI-TOF Biotyper database containing Corynebacterineae representatives of strains deposited in the Polish Collection of Microorganisms was created and used for identification of the strain isolated from a nocardial brain abscess, mimicking a brain tumor, in an immunocompetent patient. Testing with the API Coryne system initially incorrectly identified Rhodococcus sp., while chemotaxonomic tests, especially mycolic acid analysis, enabled correct Nocardia identification only at the genus level. Subsequent sequence analysis of 16S rRNA and secA1 genes confirmed the identification. To improve the accuracy of the results, an in-house database was constructed using optimized parameters; with the use of the database, the strain was eventually identified as Nocardia farcinica. Clinical laboratories processing various clinical strains can upgrade a commercial database to improve and to accelerate the results obtained. This is especially important in the case of Nocardia, for which valid microbial diagnosis remains challenging; reference laboratories are often required to identify and to survey these rare actinobacteria. PMID:26041903

  2. Bacillus subtilis chitinase identified by matrix-assisted laser desorption/ionization time-of flight/time of flight mass spectrometry has insecticidal activity against Spodoptera litura Fab.

    Science.gov (United States)

    Chandrasekaran, Rajamanickam; Revathi, Kannan; Thanigaivel, Annamalai; Kirubakaran, Suyambulingam Arunachalam; Senthil-Nathan, Sengottayan

    2014-11-01

    An extracellular chitinase was identified and purified (CS1 and CS2) from Bacillus subtilis. The 16S rRNA sequencing was submitted in GenBank (accession numbers KC336487 and KC412256). The purified crude enzymes were identified through matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry (MALDI-TOF MS) analysis. The peptide sequences were matched with chitinase sequences. The peak m/z with 1297. 592 and 3094.570 mascot search resulted sequence was blasted with NCBI protein sequences and confirmed that it is a chitinase enzyme. The effects of chitinase on gut enzymes lactate dehydrogenase, acid phosphatase, alkaline phosphatase and adenosine triphosphatase of the tobacco cutworm Spodoptera litura larvae were investigated. At all concentrations tested, chitinase decreased the activities of these gut enzymes relative to the control. When chitinase treated leaves were fed to larvae in bioassays, gut tissue and gut enzymes were affected. The histological study clearly shows the chitinase treated larval gut, peritrophic membrane and epithelial cells were affected significantly. Chitinase isolated from B. subtilis has effectively reduced the gut enzyme activity and growth of S. litura. The chitin based bioformulation may serve as an effective biocide against the polyphagous pest like S. litura. Copyright © 2014 Elsevier Inc. All rights reserved.

  3. Characterization of Klebsiella isolates by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and determination of antimicrobial resistance with VITEK 2 advanced expert system (AES).

    Science.gov (United States)

    Karagöz, Alper; Acar, Sümeyra; Körkoca, Hanifi

    2015-01-01

    The purpose of the study was to evaluate the performance of the VITEK mass spectrometry (MS) (bioMérieux, France) system for the identification of Klebsiella spp. isolated from different sources. Moreover, while assessing the ability of the VITEK 2 automated expert system (AES) to recognize antimicrobial resistance patterns, the researchers have extended the study to compare VITEK 2 with the routine antimicrobial susceptibility testing method. This study tested 51 Klebsiella spp. isolates that were isolated from environmental examples and clinical examples. Results of conventional methods and the matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS were compared. Then, any differing results were compared against a reference 16S rRNA gene sequence, and when indicated, a recA sequencing analysis was done. VITEK MS correctly identified 100% of the Klebsiella spp. isolates. There were two K. oxytoca isolates incorrectly identified to the species level with conventional methods according to the 16S rRNA gene sequencing analysis. In addition, a VITEK 2 AST-N261 card was used for the detection of extended spectrum beta-lactamases (ESBL). Using the VITEK 2 AES, ESBL positivity was found at the rate of 16.3% whereas this rate was 4.08% using the disk diffusion method. MALDI-TOF MS is a rapid and accurate method for the identification of Klebsiella spp. Moreover, the bioMérieux AES provides a useful laboratory tool for the interpretation of susceptibility results.

  4. Global proteomic analysis of plasma from mice infected with Plasmodium berghei ANKA using two dimensional gel electrophoresis and matrix assisted laser desorption ionization-time of flight mass spectrometry

    Directory of Open Access Journals (Sweden)

    Kokwaro Gilbert O

    2011-07-01

    Full Text Available Abstract Background A global proteomic strategy was used to identify proteins, which are differentially expressed in the murine model of severe malaria in the hope of facilitating future development of novel diagnostic, disease monitoring and treatment strategies. Methods Mice (4-week-old CD1 male mice were infected with Plasmodium berghei ANKA strain, and infection allowed to establish until a parasitaemia of 30% was attained. Total plasma and albumin depleted plasma samples from infected and control (non-infected mice were separated by two-dimensional gel electrophoresis (2-DE. After staining, the gels were imaged and differential protein expression patterns were interrogated using image analysis software. Spots of interest were then digested using trypsin and the proteins identified using matrix-assisted laser desorption and ionization-time of flight (MALDI-TOF mass spectrometry (MS and peptide mass fingerprinting software. Results Master gels of control and infected mice, and the corresponding albumin depleted fractions exhibited distinctly different 2D patterns comparing control and infected plasma, respectively. A wide range of proteins demonstrated altered expression including; acute inflammatory proteins, transporters, binding proteins, protease inhibitors, enzymes, cytokines, hormones, and channel/receptor-derived proteins. Conclusions Malaria-infection in mice results in a wide perturbation of the host serum proteome involving a range of proteins and functions. Of particular interest is the increased secretion of anti-inflammatory and anti apoptotic proteins.

  5. Identification and Quantification of N-Acyl Homoserine Lactones Involved in Bacterial Communication by Small-Scale Synthesis of Internal Standards and Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry

    Science.gov (United States)

    Leipert, Jan; Treitz, Christian; Leippe, Matthias; Tholey, Andreas

    2017-12-01

    N-acyl homoserine lactones (AHL) are small signal molecules involved in the quorum sensing of many gram-negative bacteria, and play an important role in biofilm formation and pathogenesis. Present analytical methods for identification and quantification of AHL require time-consuming sample preparation steps and are hampered by the lack of appropriate standards. By aiming at a fast and straightforward method for AHL analytics, we investigated the applicability of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Suitable MALDI matrices, including crystalline and ionic liquid matrices, were tested and the fragmentation of different AHL in collision-induced dissociation MS/MS was studied, providing information about characteristic marker fragments ions. Employing small-scale synthesis protocols, we established a versatile and cost-efficient procedure for fast generation of isotope-labeled AHL standards, which can be used without extensive purification and yielded accurate standard curves. Quantitative analysis was possible in the low pico-molar range, with lower limits of quantification reaching from 1 to 5 pmol for different AHL. The developed methodology was successfully applied in a quantitative MALDI MS analysis of low-volume culture supernatants of Pseudomonas aeruginosa. [Figure not available: see fulltext.

  6. Evaluation of the Bruker Biotyper Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry System for Identification of Aspergillus Species Directly from Growth on Solid Agar Media

    Directory of Open Access Journals (Sweden)

    Ying Li

    2017-06-01

    Full Text Available We evaluated the accuracy of the Bruker Biotyper matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS system at identifying clinical isolates of Aspergillus species that were grown on agar media. A total of 381 non-duplicate Aspergillus isolates representing 21 different Aspergillus species identified by molecular analysis were included in this study. The Bruker Biotyper MALDI-TOF MS system was able to identify 30.2% (115/381 of the isolates to the species level (score values of ≥2.000 and 49.3% to the genus level (score values of 1.700–1.999. When the identification cutoff value was lowered from ≥2.000 to ≥1.700, the species-level identification rate increased to 79.5% with a slight rise of false identification from 2.6 to 5.0%. From another aspect, a correct species-level identification rate of 89% could be reached by the Bruker Biotyper MALDI-TOF MS system regardless of the score values obtained. The Bruker Biotyper MALDI-TOF MS system had a moderate performance in identification of Aspergillus directly inoculated on solid agar media. Continued expansion of the Bruker Biotyper MALDI-TOF MS database and adoption of alternative cutoff values for interpretation are required to improve the performance of the system for identifying highly diverse species of clinically encountered Aspergillus isolates.

  7. Development of a novel matrix-assisted laser desorption/ionization time-of-flight mass spectrum (MALDI-TOF-MS)-based typing method to identify meticillin-resistant Staphylococcus aureus clones.

    Science.gov (United States)

    Ueda, O; Tanaka, S; Nagasawa, Z; Hanaki, H; Shobuike, T; Miyamoto, H

    2015-06-01

    Mass spectrum analysis enables species- and subspecies-level identification, and can be used as an epidemiological tool in outbreak management. However, its reliability at clonal level has yet to be established. To establish a matrix-assisted laser desorption/ionization time-of-flight mass-spectrum-based method that enables bacterial clone identification with accuracy equivalent to pulsed-field gel electrophoresis/phage open-reading frame typing (PFGE/POT). Meticillin-resistant Staphylococcus aureus (MRSA) was used in this study. Mass spectra were obtained from a standard strain of S. aureus (ATCC29213) and 57 clinically isolated strains, categorized according to POT. Peaks associated with MRSA clone identification (N = 67) were extracted. Based on this peak information, the feasibility of MRSA clone identification was examined by cluster analysis. In addition to the 58 strains used for peak extraction, mass spectrum analysis of 24 clinically isolated outbreak strains revealed that peak data could be used for successful identification of clones. These typing results were fully consistent with the PFGE and POT results. This novel method enables simple and rapid typing with accuracy equivalent to PFGE/POT. This method would be suited to rapid outbreak analysis, offering accurate information to combat infectious diseases. Copyright © 2015 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.

  8. An approach to glycobiology from glycolipidomics: ganglioside molecular scanning in the brains of patients with Alzheimer's disease by TLC-blot/matrix assisted laser desorption/ionization-time of flight MS.

    Science.gov (United States)

    Taki, Takao

    2012-01-01

    We have established a new approach to glyco- and lipidomics using a thin layer chromatography (TLC)-Blot/matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MS) system. This new approach consists of a combination of a method for transferring lipids separated on a TLC-plate to a poly-vinylidene difluoride (PVDF) membrane and direct mass spectrometric analysis of the individual lipids on the membrane by ion trap-type MALDI-TOF MS. This technology was applied to the analysis of individual lipids from the human brain. Then, based on the results of this analysis, ganglioside molecular species in neural diseases were analyzed. The levels of gangliosides GD1b and GT1b were lower in the hippocampal gray matter of patients with Alzheimer's disease than in the hippocampal gray matter of patients with Parkinson's disease or the control patients. The molecular scanning of individual ganglioside molecular species showed a significant reduction of d20 : 1/C18 : 0 ceramide-containing gangliosides in patients with Alzheimer's disease. These findings suggest that Alzheimer's disease is a kind of ganglioside metabolic disease affecting the hippocampal area. A new approach to glycobiology by the TLC-Blot/MALDI-TOF MS system is proposed.

  9. Method for simultaneous imaging of endogenous low molecular weight metabolites in mouse brain using TiO2 nanoparticles in nanoparticle-assisted laser desorption/ionization-imaging mass spectrometry.

    Science.gov (United States)

    Shrivas, Kamlesh; Hayasaka, Takahiro; Sugiura, Yuki; Setou, Mitsutoshi

    2011-10-01

    We report the detection of a group of endogenous low molecular weight metabolites (LMWM) in mouse brain (80-500 Da) using TiO(2) nanoparticles (NPs) in nanoparticle-assisted laser desorption/ionization-imaging mass spectrometry (Nano-PALDI-IMS) without any washing and separation step prior to MS analysis. The identification of metabolites using TiO(2) NPs was compared with a conventional organic matrix 2,5-dihydroxybenzoic acid (DHB) where signals of 179 molecules were specific to TiO(2) NPs, 4 were specific to DHB, and 21 were common to both TiO(2) NPs and DHB. The use of TiO(2) NPs enabled the detection of a higher number of LMWM as compared to DHB and gold NPs as a matrix. This approach is a simple, inexpensive, washing, and separation free for imaging and identification of LMWM in mouse brain. We believe that the biochemical information from distinct regions of the brain using a Nano-PALDI-IMS will be helpful in elucidating the imbalances linked with diseases in biomedical samples.

  10. Evaluation of repetitive-PCR and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS for rapid strain typing of Bacillus coagulans.

    Directory of Open Access Journals (Sweden)

    Jun Sato

    Full Text Available In order to establish rapid and accurate typing method for Bacillus coagulans strains which is important for controlling in some canned foods and tea-based beverages manufacturing because of the high-heat resistance of the spores and high tolerance of the vegetative cells to catechins and chemicals, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS and repetitive-PCR (rep-PCR were evaluated. For this purpose, 28 strains of B. coagulans obtained from various culture collections were tested. DNA sequence analyses of the genes encoding 16S rRNA and DNA gyrase classified the test strains into two and three groups, respectively, regardless of their phenotypes. Both MALDI-TOF MS and rep-PCR methods classified the test strains in great detail. Strains classified in each group showed similar phenotypes, such as carbohydrate utilization determined using API 50CH. In particular, the respective two pairs of strains which showed the same metabolic characteristic were classified into the same group by both MALDI-TOF MS and rep-PCR methods separating from the other strains. On the other hand, the other strains which have the different profiles of carbohydrate utilization were separated into different groups by these methods. These results suggested that the combination of MALDI-TOF MS and rep-PCR analyses was advantageous for the rapid and detailed typing of bacterial strains in respect to both phenotype and genotype.

  11. Examination of the skin barrier repair/wound healing process using a living skin equivalent (LSE) model and matrix-assisted laser desorption-ionization-mass spectrometry imaging (MALDI-MSI).

    Science.gov (United States)

    Lewis, E E L; Barrett, M R T; Freeman-Parry, L; Bojar, R A; Clench, M R

    2018-01-21

    Examination of the skin barrier repair/wound healing process using a living skin equivalent (LSE) model and matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI) to identify lipids directly involved as potential biomarkers. These biomarkers may be used to determine whether an in vivo wound is going to heal for example if infected. An in vitro LSE model was wounded with a scalpel blade and assessed at day 4 post wounding by histology and MALDI-MSI. Samples were sectioned at wound site and were either formalin fixed paraffin embedded (FFPE) for histology or snapped frozen (FF) for MSI analysis. The combination of using an in vitro wounded skin model with MSI allowed the identification of lipids involved in the skin barrier repair/wound healing process. The technique was able to highlight lipids directly in the wound site and distinguish differences in lipid distribution between the epidermis and wound site. This novel method of coupling an in vitro LSE with MSI allowed in depth molecular analysis of the skin barrier repair/wound healing process. The technique allowed the identification of lipids directly involved in the skin barrier repair/wound healing process, indicating these biomarkers may be potentially be used within clinic. These biomarkers will help determine, which stage of the skin barrier repair/wound healing process the wound is in to provide the best treatment. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  12. Methylobacterium Species Promoting Rice and Barley Growth and Interaction Specificity Revealed with Whole-Cell Matrix-Assisted Laser Desorption/Ionization-Time-of-Flight Mass Spectrometry (MALDI-TOF/MS Analysis.

    Directory of Open Access Journals (Sweden)

    Akio Tani

    Full Text Available Methylobacterium species frequently inhabit plant surfaces and are able to utilize the methanol emitted from plants as carbon and energy sources. As some of the Methylobacterium species are known to promote plant growth, significant attention has been paid to the mechanism of growth promotion and the specificity of plant-microbe interactions. By screening our Methylobacterium isolate collection for the high growth promotion effect in vitro, we selected some candidates for field and pot growth tests for rice and barley, respectively. We found that inoculation resulted in better ripening of rice seeds, and increased the size of barley grains but not the total yield. In addition, using whole-cell matrix-assister laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF/MS analysis, we identified and classified Methylobacterium isolates from Methylobacterium-inoculated rice plants. The inoculated species could not be recovered from the rice plants, and in some cases, the Methylobacterium community structure was affected by the inoculation, but not with predomination of the inoculated species. The isolates from non-inoculated barley of various cultivars grown in the same field fell into just two species. These results suggest that there is a strong selection pressure at the species level of Methylobacterium residing on a given plant species, and that selection of appropriate species that can persist on the plant is important to achieve growth promotion.

  13. In situ analysis of plant tissue underivatized carbohydrates and on-probe enzymatic degraded starch by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry by using carbon nanotubes as matrix.

    Science.gov (United States)

    Gholipour, Yousef; Nonami, Hiroshi; Erra-Balsells, Rosa

    2008-12-15

    Underivatized carbohydrates of tulip bulb and leaf tissues were characterized in situ by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) by using carbon nanotubes (CNTs) as matrix. Two sample preparation methods--(i) depositing CNTs on the fresh tissue slices placed on the probe and (ii) locating semitransparent tissues on a dried layer of CNTs on the probe--were examined. Furthermore, practicability of in situ starch analysis by MALDI-TOF MS was examined by detection of glucose originated from on-probe amyloglucosidase-catalyzed degradation of starch on the tissue surface. Besides, CNTs could efficiently desorb/ionize natural mono-, di-, and oligosaccharides extracted from tulip bulb tissues as well as glucose resulting from starch enzymatic degradation in vitro. These results were compared with those obtained by in situ MALDI-TOF MS analysis of similar tissues. Positive ion mode showed superior signal reproducibility. CNTs deposited under semitransparent tissue could also desorb/ionize neutral carbohydrates, leading to nearly complete elimination of matrix cluster signals but with an increase in tissue-originated signals. Furthermore, several experiments were carried out to compare the efficiency of 2,5-dihydroxybenzoic acid, nor-harmane, alpha-cyano-4-hydroxycinnamic acid, and CNTs as matrices for MALDI of neutral carbohydrates from the intact plant tissue surface and for enzymatic tissue starch degradation; these results are discussed in brief. Among matrices studied, the lowest laser power was needed to acquire carbohydrate signals with high signal-to-noise ratio and resolution when CNTs were used.

  14. Superbasic alkyl-substituted bisphosphazene proton sponges: a new class of deprotonating matrices for negative ion matrix-assisted ionization/laser desorption mass spectrometry of low molecular weight hardly ionizable analytes.

    Science.gov (United States)

    Calvano, C D; Cataldi, T R I; Kögel, J F; Monopoli, A; Palmisano, F; Sundermeyer, J

    2016-07-30

    Here hardly ionizable and low molecular weight compounds are detected in negative ion mode by using novel superbasic proton sponges based on 1,8-bisphosphazenylnaphthalene (PN) as MALDI matrices. Among the selected proton sponges, 1,8-bis(trispyrrolidinophosphazenyl)naphthalene (TPPN) has shown the best behaviour as matrix since it allows the direct detection of intact cholesterol without derivatization also in real challenging samples. Very weakly acidic compounds such as sterols, steroids, fatty alcohols and saccharides were detected in reflectron negative ion mode by a MALDI TOF/TOF system equipped with a neodymium-doped yttrium lithium fluoride (Nd:YLF) laser (345 nm) with typical mass accuracy of 10 ppm. MS/MS experiments were performed by using ambient air as the collision gas. Contrary to traditional MALDI matrices, superbasic proton sponges allowed the easy deprotonation of an alcohol functional group without a previous chemical derivatization step. Experimental evidence indicates that analyte deprotonation is achieved in the condensed phase, i.e. PN superbasic proton sponges operate according to a recently proposed model named matrix assisted ionization/laser desorption (MAILD). A detection limit of 3 pmol/spot of cholesterol (model compound) with a signal-to-noise ratio ≥ 10 was typically obtained. For the first time, the usefulness of novel superbasic proton sponges is demonstrated for MALDI detection of hardly ionizable compounds such as sterols, steroids, fatty alcohols and saccharides. The leading candidate TPPN has been successfully applied for negative ion MAILD-MS analysis of cholesterol, fatty acids and phospholipids in egg yolk and brain tissue extracts. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  15. Direct bacterial identification from positive blood cultures using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry: A systematic review and meta-analysis.

    Science.gov (United States)

    Ruiz-Aragón, Jesús; Ballestero-Téllez, Mónica; Gutiérrez-Gutiérrez, Belén; de Cueto, Marina; Rodríguez-Baño, Jesús; Pascual, Álvaro

    2017-10-27

    The rapid identification of bacteraemia-causing pathogens could assist clinicians in the timely prescription of targeted therapy, thereby reducing the morbidity and mortality of this infection. In recent years, numerous techniques that rapidly and directly identify positive blood cultures have been marketed, with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) being one of the most commonly used. The aim of this systematic review and meta-analysis was to evaluate the accuracy of MALDI-TOF (Bruker®) for the direct identification of positive blood culture bottles. A meta-analysis was performed to summarize the results of the 32 studies evaluated. The overall quality of the studies was moderate. For Gram-positive bacteria, overall rates of correct identification of the species ranged from 0.17 to 0.98, with a cumulative rate (random-effects model) of 0.72 (95% CI: 0.64-0.80). For Gram-negative bacteria, correct identification rates ranged from 0.66 to 1.00, with a cumulative effect of 0.92 (95% CI: 0.88-0.95). For Enterobacteriaceae, the rate was 0.96 (95% CI: 0.94-0.97). MALDI-TOF mass spectrometry shows high accuracy for the correct identification of Gram-negative bacteria, particularly Enterobacteriaceae, directly from positive blood culture bottles, and moderate accuracy for the identification of Gram-positive bacteria (low for some species). Copyright © 2017 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

  16. Diamond, titanium dioxide, titanium silicon oxide, and barium strontium titanium oxide nanoparticles as matrixes for direct matrix-assisted laser desorption/ionization mass spectrometry analysis of carbohydrates in plant tissues.

    Science.gov (United States)

    Gholipour, Yousef; Giudicessi, Silvana L; Nonami, Hiroshi; Erra-Balsells, Rosa

    2010-07-01

    Nanoparticles (NPs) of diamond, titanium dioxide, titanium silicon oxide, barium strontium titanium oxide, and silver (Ag) were examined for their potential as MALDI matrixes for direct laser desorption/ionization of carbohydrates, especially fructans, from plant tissue. Two sample preparation methods including solvent-assisted and solvent-free (dry) NPs deposition were performed and compared. All examined NPs except for Ag could desorb/ionize standard sucrose and fructans in positive and in negative ion mode. Ag NPs yielded good signals only for nonsalt-doped samples that were measured in the negative ion mode. In the case of in vivo studies, except for Ag, all NPs studied could desorb/ionize carbohydrates from tissue in both the positive and negative ion modes. Furthermore, compared to the results obtained with soluble sugars extracted from plant tissues, fructans with higher molecular weight intact molecular ions could be detected when the plant tissues were directly profiled. The limit of detection (LOD) of fructans and the ratios between signal intensities and fructan concentrations were analyzed. NPs had similar LODs for standard fructan triose (1-kestose) in the positive ion mode and better LODs in the negative ion mode when compared with the common crystalline organic MALDI matrixes used for carbohydrates (2,5-dihydroxybenzoic acid and nor-harmane) or carbon nanotubes. Solvent-free NP deposition on tissues partially improves the signal acquisition. Although lower signal-to-noise ratio sugar signals were acquired from the tissues when compared to the solvent-assisted method, the reproducibility averaged over all sample was more uniform.

  17. Laser desorption mass spectrometry for fast DNA analysis

    Energy Technology Data Exchange (ETDEWEB)

    Chen, C.H.; Ch`ang, L.Y.; Taranenko, N.I.; Allman, S.L.; Tang, K.; Matteson, K.J.

    1995-09-01

    During the past few years, major effort has been directed toward developing mass spectrometry to measure biopolymers because of the great potential benefit to biomedical research. Hellenkamp and his co-workers were the first to report that large polypeptide molecules can be ionized and detected without significant fragmentation when a greater number of nicotinic acid molecules are used as a matrix. This method is now well known as matrix-assisted laser desorption/ionization (MALDI). Since then, various groups have reported measurements of very large proteins by MALDI. Reliable protein analysis by MALDI is more or less well established. However, the application of MALDI to nucleic acids analysis has been found to be much more difficult. Most research on the measurement of nucleic acid by MALDI were stimulated by the Human Genome Project. Up to now, the only method for reliable routine analysis of nucleic acid is gel electrophoresis. Different sizes of nucleic acids can be separated in gel medium when a high electric field is applied to the gel. However, the time needed to separate different sizes of DNA segments usually takes from several minutes to several hours. If MALDI can be successfully used for nucleic acids analysis, the analysis time can be reduced to less than I millisecond. In addition, no tagging with radioactive materials or chemical dyes is needed. In this work, we will review recent progress related to MALDI for DNA analysis.

  18. Graphene coated silica applied for high ionization matrix assisted laser desorption/ionization mass spectrometry: A novel approach for environmental and biomolecule analysis.

    Science.gov (United States)

    Nasser Abdelhamid, Hani; Wu, Bo-Sgum; Wu, Hui-Fen

    2014-08-01

    The integration of nanotechnology with mass spectrometry for sensitive and selective detection of molecules is a hot/important field of research. Synthesis of graphene (G) coated with mesoporous silica (SiO2, G@SiO2) for mass spectrometric application has been demonstrated. For the first time, we proposed the significant role of surfactant that used during the synthesis of mesorporous silicate (SiO2) in mass spectrometry. It was noticed that G could initiate SiO2 via surfactants which work as initiators for further ionization. The porosity of SiO2 trapped the analytes that was released and ionized with the surfactant fragments. Undoubtedly, strong background interferences were present in the case of organic matrix, which greatly obscured the detection of low molecular weight compounds. G@SiO2 nanocomposite affords several advantages, such as the ability to detect small molecules (silica mesoporosity, and high ionization efficiency over than G or conventional matrices. The high performance of G@SiO2 is not only due to the large surface area but also due to high desorption/ionization efficiency of inevitably surfactant (cetyltrimethylammonium chloride, CATB). Unlike the conventional MALDI-MS, the G@SiO2-MS is capable of generating multiply charged polysaccharides. The present method was validated to detect surfactants with low limits of detection. Copyright © 2014 Elsevier B.V. All rights reserved.

  19. Polylactosaminoglycan Glycomics: Enhancing the Detection of High-molecular-weight N-glycans in Matrix-assisted Laser Desorption Ionization Time-of-flight Profiles by Matched Filtering*

    Science.gov (United States)

    Bern, Marshall; Brito, Alejandro E.; Pang, Poh-Choo; Rekhi, Angad; Dell, Anne; Haslam, Stuart M.

    2013-01-01

    For over 30 years, protocols based on the mass spectrometry (MS) of permethylated derivatives, complemented by enzymatic degradations, have underpinned glycomic experiments aimed at defining the structures of individual glycans present in the complex mixtures that are characteristic of biological samples. Both MS instrumentation and sample handling have improved markedly in recent years, enabling greater sensitivity and better signal-to-noise ratios, thereby facilitating the detection of glycans at much higher masses than could be achieved in the past. The latter is especially important for the characterization of the biologically important class of N-glycans that carry polylactosaminoglycan chains. Such advances in data acquisition heighten the need for informatics tools to assist in glycan structure assignment. Here, utilizing mouse lung tissue as a model system, we present evidence of polylactosaminoglycan-containing N-glycans with permethylated molecular weights exceeding 13 kDa. We show that antennae branching patterns and lengths can be successfully determined at these high masses via MS/MS experiments, even when MS ion counts are very low. We also describe the development and application of a matched filtering algorithm for assisting high-molecular-weight glycan detection and structure assignment. PMID:23325768

  20. Assessment of Time to Clinical Response in Patients with Sepsis Treated Before and After Implementation of a Matrix-Assisted Laser Desorption Ionization Time-of-Flight Blood Culture Identification Algorithm.

    Science.gov (United States)

    Carreno, Joseph J; Lomaestro, Ben M; Jacobs, Apryl L; Meyer, Rachel E; Evans, Ann; Montero, Clemente I

    2016-08-01

    OBJECTIVE To evaluate time to clinical response before and after implementation of rapid blood culture identification technologies. DESIGN Before-and-after trial. SETTING Large, tertiary, urban, academic health-sciences center. PATIENTS Patients >18 years old with sepsis and concurrent bacteremia or fungemia were included in the study; patients who were pregnant, had polymicrobial septicemia, or were transferred from an outside hospital were excluded. INTERVENTION Prior to the intervention, polymerase chain reaction was used to identify Staphylococcus species from positive blood cultures, and traditional laboratory techniques were used to identify non-staphylococcal species. After the intervention, matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) assay and FilmArray were also used to identify additional species. During both periods, the antimicrobial stewardship team provided prospective audit and feedback for all patients on antibiotics. RESULTS A total of 219 patients were enrolled in the study: 115 patients prior to the intervention and 104 after the intervention. The median time to clinical response was statistically significantly shorter in the postintervention group than in the preintervention group (2 days vs 4 days, respectively; P=.002). By Cox regression, the implementation of MALDI-TOF and FilmArray was associated with shorter time to clinical response (hazard ratio [HR], 1.360; 95% confidence interval [CI], 1.018-1.816). After controlling for potential confounders, the study group was not independently associated with clinical response (adjusted HR, 1.279; 95% CI, 0.955-1.713). Mortality was numerically, but not statistically significantly, lower in the postintervention group than in the preintervention group (7.6% vs 11.4%; P=.342). CONCLUSIONS In the setting of an existing antimicrobial stewardship program, implementation of MALDI-TOF and FilmArray was associated with improved time to clinical response. Further research is needed

  1. Feasibility of using atmospheric pressure matrix-assisted laser desorption/ionization with ion trap mass spectrometry in the analysis of acetylated xylooligosaccharides derived from hardwoods and Arabidopsis thaliana.

    Science.gov (United States)

    Chong, Sun-Li; Nissilä, Teemu; Ketola, Raimo A; Koutaniemi, Sanna; Derba-Maceluch, Marta; Mellerowicz, Ewa J; Tenkanen, Maija; Tuomainen, Päivi

    2011-11-01

    The atmospheric pressure matrix-assisted laser desorption/ionization with ion trap mass spectrometry (AP-MALDI-ITMS) was investigated for its ability to analyse plant-derived oligosaccharides. The AP-MALDI-ITMS was able to detect xylooligosaccharides (XOS) with chain length of up to ten xylopyranosyl residues. Though the conventional MALDI-time-of-flight/mass spectrometry (TOF/MS) showed better sensitivity at higher mass range (>m/z 2,000), the AP-MALDI-ITMS seems to be more suitable for detection of acetylated XOS, and the measurement also corresponded better than the MALDI-TOF/MS analysis to the actual compositions of the pentose- and hexose-derived oligosaccharides in a complex sample. The structures of two isomeric aldotetrauronic acids and a mixture of acidic XOS were elucidated by AP-MALDI-ITMS using multi-stages mass fragmentation up to MS(3). Thus, the AP-MALDI-ITMS demonstrated an advantage in determining both mass and structures of plant-derived oligosaccharides. In addition, the method of combining the direct endo-1,4-β-D-xylanase hydrolysis of plant material, and then followed by AP-MALDI-ITMS detection, was shown to recognize the substitution variations of glucuronoxylans in hardwood species and in Arabidopsis thaliana. To our knowledge, this is the first report to demonstrate the acetylation of glucuronoxylan in A. thaliana. The method, which requires only a small amount of plant material, such as 1 to 5 mg for the A. thaliana stem material, can be applied as a high throughput fingerprinting tool for the fast comparison of glucuronoxylan structures among plant species or transformants that result from in vivo cell wall modification.

  2. Analysis of matrix-assisted laser desorption/ionization quadrupole time-of-flight collision-induced dissociation spectra of simple precursor ions and isobaric oligosaccharide ion mixtures based on product ion intensities and pattern recognition.

    Science.gov (United States)

    Jovanović, Marko; Tyldesley-Worster, Richard

    2017-05-30

    Qualitative analysis of glycomic tandem mass spectrometry (MS/MS) data based on m/z values of product ions alone is widely used, and often sufficient for analysis of single analytes. However, most complex glycomic mixtures contain multiple isobaric oligosaccharides, in which case this approach is often limited. Here we show how ion intensity information can be used in order to enhance MS/MS data analysis, and extract both qualitative and semiquantitative information from complex glycomic MS/MS datasets. A matrix-assisted laser desorption/ionization quadrupole time-of-flight (MALDI QTOF) mass spectrometer was used in this study. We compared the intensities of product ions within a single product ion series, determined by their length, across the whole glycomic MS/MS dataset. In order to detect discernable patterns, the intensity data was normalized to the intensity of each product ion within the series. In most cases, normalized data yielded easily discernable patterns, relevant either for analysis of specific glycomic structure types, or mechanistic MS studies. We used our approach on a glycomic sample consisting of human milk oligosaccharides. The approach yielded useful results for both qualitative and semiquantitative analyses. All normalizations performed were not equally rich in information and the information content of generated tables was not possible to predict. These analyses were shown to be independent of instrument manufacturer. Our approach enabled more detailed qualitative analysis of MS/MS spectra of precursor ions containing isobaric oligosaccharide structures. While limited semiquantitative information could be extracted from the raw data as well, the accuracy of this method should be significantly enhanced when standard calibration mixtures can be prepared. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

  3. Rapid Differentiation of Haemophilus influenzae and Haemophilus haemolyticus by Use of Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry with ClinProTools Mass Spectrum Analysis.

    Science.gov (United States)

    Chen, Jonathan H K; Cheng, Vincent C C; Wong, Chun-Pong; Wong, Sally C Y; Yam, Wing-Cheong; Yuen, Kwok-Yung

    2017-09-01

    Haemophilus influenzae is associated with severe invasive disease, while Haemophilus haemolyticus is considered part of the commensal flora in the human respiratory tract. Although the addition of a custom mass spectrum library into the matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) system could improve identification of these two species, the establishment of such a custom database is technically complicated and requires a large amount of resources, which most clinical laboratories cannot afford. In this study, we developed a mass spectrum analysis model with 7 mass peak biomarkers for the identification of H. influenzae and H. haemolyticus using the ClinProTools software. We evaluated the diagnostic performance of this model using 408 H. influenzae and H. haemolyticus isolates from clinical respiratory specimens from 363 hospitalized patients and compared the identification results with those obtained with the Bruker IVD MALDI Biotyper. The IVD MALDI Biotyper identified only 86.9% of H. influenzae (311/358) and 98.0% of H. haemolyticus (49/50) clinical isolates to the species level. In comparison, the ClinProTools mass spectrum model could identify 100% of H. influenzae (358/358) and H. haemolyticus (50/50) clinical strains to the species level and significantly improved the species identification rate (McNemar's test, P mass spectrometry to handle closely related bacterial species when the proprietary spectrum library failed. This approach should be useful for the differentiation of other closely related bacterial species. Copyright © 2017 American Society for Microbiology.

  4. Rapid discrimination of Haemophilus influenzae, H. parainfluenzae, and H. haemolyticus by fluorescence in situ hybridization (FISH) and two matrix-assisted laser-desorption-ionization time-of-flight mass spectrometry (MALDI-TOF-MS) platforms.

    Science.gov (United States)

    Frickmann, Hagen; Christner, Martin; Donat, Martina; Berger, Anja; Essig, Andreas; Podbielski, Andreas; Hagen, Ralf Matthias; Poppert, Sven

    2013-01-01

    Due to considerable differences in pathogenicity, Haemophilus influenzae, H. parainfluenzae and H. haemolyticus have to be reliably discriminated in routine diagnostics. Retrospective analyses suggest frequent misidentifications of commensal H. haemolyticus as H. influenzae. In a multi-center approach, we assessed the suitability of fluorescence in situ hybridization (FISH) and matrix-assisted laser-desorption-ionization time-of-flight mass-spectrometry (MALDI-TOF-MS) for the identification of H. influenzae, H. parainfluenzae and H. haemolyticus to species level. A strain collection of 84 Haemophilus spp. comprising 50 H. influenzae, 25 H. parainfluenzae, 7 H. haemolyticus, and 2 H. parahaemolyticus including 77 clinical isolates was analyzed by FISH with newly designed DNA probes, and two different MALDI-TOF-MS systems (Bruker, Shimadzu) with and without prior formic acid extraction. Among the 84 Haemophilus strains analyzed, FISH led to 71 correct results (85%), 13 uninterpretable results (15%), and no misidentifications. Shimadzu MALDI-TOF-MS resulted in 59 correct identifications (70%), 19 uninterpretable results (23%), and 6 misidentifications (7%), using colony material applied directly. Bruker MALDI-TOF-MS with prior formic acid extraction led to 74 correct results (88%), 4 uninterpretable results (5%) and 6 misidentifications (7%). The Bruker MALDI-TOF-MS misidentifications could be resolved by the addition of a suitable H. haemolyticus reference spectrum to the system's database. In conclusion, no analyzed diagnostic procedure was free of errors. Diagnostic results have to be interpreted carefully and alternative tests should be applied in case of ambiguous test results on isolates from seriously ill patients.

  5. Early identification of microorganisms in blood culture prior to the detection of a positive signal in the BACTEC FX system using matrix-assisted laser desorption/ionization-time of flight mass spectrometry.

    Science.gov (United States)

    Wang, Ming-Cheng; Lin, Wei-Hung; Yan, Jing-Jou; Fang, Hsin-Yi; Kuo, Te-Hui; Tseng, Chin-Chung; Wu, Jiunn-Jong

    2015-08-01

    Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) is a valuable method for rapid identification of blood stream infection (BSI) pathogens. Integration of MALDI-TOF MS and blood culture system can speed the identification of causative BSI microorganisms. We investigated the minimal microorganism concentrations of common BSI pathogens required for positive blood culture using BACTEC FX and for positive identification using MALDI-TOF MS. The time to detection with positive BACTEC FX and minimal incubation time with positive MALDI-TOF MS identification were determined for earlier identification of common BSI pathogens. The minimal microorganism concentrations required for positive blood culture using BACTEC FX were >10(7)-10(8) colony forming units/mL for most of the BSI pathogens. The minimal microorganism concentrations required for identification using MALDI-TOF MS were > 10(7) colony forming units/mL. Using simulated BSI models, one can obtain enough bacterial concentration from blood culture bottles for successful identification of five common Gram-positive and Gram-negative bacteria using MALDI-TOF MS 1.7-2.3 hours earlier than the usual time to detection in blood culture systems. This study provides an approach to earlier identification of BSI pathogens prior to the detection of a positive signal in the blood culture system using MALDI-TOF MS, compared to current methods. It can speed the time for identification of BSI pathogens and may have benefits of earlier therapy choice and on patient outcome. Copyright © 2013. Published by Elsevier B.V.

  6. Characteristics of glycation and glycation sites of lysozyme by matrix-assisted laser desorption/ionization time of flight/time-of-flight mass spectrometry and Liquid chromatography-electrospray ionization tandem mass spectrometry.

    Science.gov (United States)

    Ruan, Eric Dongliang; Wang, Hui; Ruan, Yuanyuan; Juáreza, Manuel

    2014-01-01

    Protein glycation with reducing sugars through the Maillard reaction is regarded as one of the most important reactions in food chem- istry. Amadori rearrangement products [ARPs] are produced at the initial stage of the Maillard reaction and then advanced glycation products may be formed. We report here that using matrix-assisted laser desorption/ionization mass spectrometry with time-of-flight detection [MALDI-TOF-MS] and electrospray ionization mass spectrometry (ESI-MSJ to monitor the glycation process in lysozyme and the D-glucose model system. MALDI-TOF-MS displayed a heterogeneous distribution of glycation via a total mass shift in spectra. However electrospray ionization mass spectrometry [ESI-MS] data showed that a total of four molecules of glucose reacted with Lysozyme at an increase in molecular weight by a 162 Da unit. Further, we identified the glycation sites of lysozyme by using MALDI-TOF/TOF-MS and Liquid chromatography [LC]-ESI-MS/MS. Besides the two glycation sites of Lys1 and Lys97 identified by MALDI-TOF/TOF-MS, the other two glycation sites of Lys13 and Lys116 were characterized unambiguously by LC-ESI-MS/MS. Both MALDI-TOF/TOF-MS and LC-ESI-MS/ MS provided confidence in the study of the glycation by restricting the number of possible residues through (un]modified ions. The study is useful to monitor and characterize glycation in protein systems based on both MALDI-TOF-MS and ESI-MS. Comparatively, LC-ESI-MS/MS provides more fragments with better recovery for the identification of glycation than MALDI-TOF/TOF-MS.

  7. Ribosomal subunit protein typing using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) for the identification and discrimination of Aspergillus species.

    Science.gov (United States)

    Nakamura, Sayaka; Sato, Hiroaki; Tanaka, Reiko; Kusuya, Yoko; Takahashi, Hiroki; Yaguchi, Takashi

    2017-04-26

    Accurate identification of Aspergillus species is a very important subject. Mass spectral fingerprinting using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) is generally employed for the rapid identification of fungal isolates. However, the results are based on simple mass spectral pattern-matching, with no peak assignment and no taxonomic input. We propose here a ribosomal subunit protein (RSP) typing technique using MALDI-TOF MS for the identification and discrimination of Aspergillus species. The results are concluded to be phylogenetic in that they reflect the molecular evolution of housekeeping RSPs. The amino acid sequences of RSPs of genome-sequenced strains of Aspergillus species were first verified and compared to compile a reliable biomarker list for the identification of Aspergillus species. In this process, we revealed that many amino acid sequences of RSPs (about 10-60%, depending on strain) registered in the public protein databases needed to be corrected or newly added. The verified RSPs were allocated to RSP types based on their mass. Peak assignments of RSPs of each sample strain as observed by MALDI-TOF MS were then performed to set RSP type profiles, which were then further processed by means of cluster analysis. The resulting dendrogram based on RSP types showed a relatively good concordance with the tree based on β-tubulin gene sequences. RSP typing was able to further discriminate the strains belonging to Aspergillus section Fumigati. The RSP typing method could be applied to identify Aspergillus species, even for species within section Fumigati. The discrimination power of RSP typing appears to be comparable to conventional β-tubulin gene analysis. This method would therefore be suitable for species identification and discrimination at the strain to species level. Because RSP typing can characterize the strains within section Fumigati, this method has potential as a powerful and reliable tool in

  8. Analyses of black fungi by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS): species-level identification of clinical isolates of Exophiala dermatitidis.

    Science.gov (United States)

    Kondori, Nahid; Erhard, Marcel; Welinder-Olsson, Christina; Groenewald, Marizeth; Verkley, Gerard; Moore, Edward R B

    2015-01-01

    Conventional mycological identifications based on the recognition of morphological characteristics can be problematic. A relatively new methodology applicable for the identification of microorganisms is based on the exploitation of taxon- specific mass patterns recorded from abundant cell proteins directly from whole-cell preparations, using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). This study reports the application of MALDI-TOF MS for the differentiation and identifications of black yeasts, isolated from the respiratory tracts of patients with cystic fibrosis (CF). Initial phenotypic and DNA sequence-based analyses identified these isolates to be Exophiala dermatitidis. The type strains of E. dermatitidis (CBS 207.35(T)) and other species of Exophiala were included in the MALDI-TOF MS analyses to establish the references for comparing the mass spectra of the clinical isolates of Exophiala. MALDI-TOF MS analyses exhibited extremely close relationships among the clinical isolates and with the spectra generated from the type strain of E. dermatitidis. The relationships observed between the E. dermatitidis strains from the MALDI-TOF MS profiling analyses were supported by DNA sequence-based analyses of the rRNA ITS1 and ITS2 regions. These data demonstrated the applicability of MALDI-TOF MS as a reliable, rapid and cost-effective method for the identification of isolates of E. dermatitidis and other clinically relevant fungi and yeasts that typically are difficult to identify by conventional methods. © FEMS 2014. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  9. Source-identifying biomarker ions between environmental and clinical Burkholderia pseudomallei using whole-cell matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Niyompanich, Suthamat; Jaresitthikunchai, Janthima; Srisanga, Kitima; Roytrakul, Sittiruk; Tungpradabkul, Sumalee

    2014-01-01

    Burkholderia pseudomallei is the causative agent of melioidosis, which is an endemic disease in Northeast Thailand and Northern Australia. Environmental reservoirs, including wet soils and muddy water, serve as the major sources for contributing bacterial infection to both humans and animals. The whole-cell matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (whole-cell MALDI-TOF MS) has recently been applied as a rapid, accurate, and high-throughput tool for clinical diagnosis and microbiological research. In this present study, we employed a whole-cell MALDI-TOF MS approach for assessing its potency in clustering a total of 11 different B. pseudomallei isolates (consisting of 5 environmental and 6 clinical isolates) with respect to their origins and to further investigate the source-identifying biomarker ions belonging to each bacterial group. The cluster analysis demonstrated that six out of eleven isolates were grouped correctly to their sources. Our results revealed a total of ten source-identifying biomarker ions, which exhibited statistically significant differences in peak intensity between average environmental and clinical mass spectra using ClinProTools software. Six out of ten mass ions were assigned as environmental-identifying biomarker ions (EIBIs), including, m/z 4,056, 4,214, 5,814, 7,545, 7,895, and 8,112, whereas the remaining four mass ions were defined as clinical-identifying biomarker ions (CIBIs) consisting of m/z 3,658, 6,322, 7,035, and 7,984. Hence, our findings represented, for the first time, the source-specific biomarkers of environmental and clinical B. pseudomallei.

  10. The Technical and Biological Reproducibility of Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) Based Typing: Employment of Bioinformatics in a Multicenter Study.

    Science.gov (United States)

    Oberle, Michael; Wohlwend, Nadia; Jonas, Daniel; Maurer, Florian P; Jost, Geraldine; Tschudin-Sutter, Sarah; Vranckx, Katleen; Egli, Adrian

    2016-01-01

    The technical, biological, and inter-center reproducibility of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI TOF MS) typing data has not yet been explored. The aim of this study is to compare typing data from multiple centers employing bioinformatics using bacterial strains from two past outbreaks and non-related strains. Participants received twelve extended spectrum betalactamase-producing E. coli isolates and followed the same standard operating procedure (SOP) including a full-protein extraction protocol. All laboratories provided visually read spectra via flexAnalysis (Bruker, Germany). Raw data from each laboratory allowed calculating the technical and biological reproducibility between centers using BioNumerics (Applied Maths NV, Belgium). Technical and biological reproducibility ranged between 96.8-99.4% and 47.6-94.4%, respectively. The inter-center reproducibility showed a comparable clustering among identical isolates. Principal component analysis indicated a higher tendency to cluster within the same center. Therefore, we used a discriminant analysis, which completely separated the clusters. Next, we defined a reference center and performed a statistical analysis to identify specific peaks to identify the outbreak clusters. Finally, we used a classifier algorithm and a linear support vector machine on the determined peaks as classifier. A validation showed that within the set of the reference center, the identification of the cluster was 100% correct with a large contrast between the score with the correct cluster and the next best scoring cluster. Based on the sufficient technical and biological reproducibility of MALDI-TOF MS based spectra, detection of specific clusters is possible from spectra obtained from different centers. However, we believe that a shared SOP and a bioinformatics approach are required to make the analysis robust and reliable.

  11. Evaluation of the Bruker Matrix-Assisted Laser Desorption-Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) System for the Identification of Clinically Important Dermatophyte Species.

    Science.gov (United States)

    Karabıçak, Nilgün; Karatuna, Onur; İlkit, Macit; Akyar, Işın

    2015-10-01

    Dermatophytes can invade the stratum corneum of the skin and other keratinized tissues and are responsible for a broad diversity of diseases of skin, nails and hair. Although the standard identification of dermatophytoses depends on macroscopic and microscopic characterization of the colonies grown on special media, there are a number of limitations owing to intraspecies morphological variability, atypical morphology or interspecies morphological similarity which entails improvement in the identification methods. Matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a novel method which proved to be effective for rapid and reliable identification of dermatophytes grown in cultures when compared to conventional methods. We evaluated the performance of Bruker MALDI-TOF MS System (Bruker Daltonics, Germany) for identification of clinically relevant dermatophytes. In order to increase the identification capacity of the system, we created supplemental spectral database entries using ten reference dermatophyte strains (ten species in two genera). The utility of the generated database was then challenged using a total of 126 dermatophytes (115 clinical isolates and 11 additional reference strains). The results were evaluated by both manufacturer-recommended and lowered cutoff scores. MALDI-TOF MS provided correct identification in 122 (96.8 %) and 113 (89.7 %) of the isolates with the lowered scores and using the supplemented database, respectively, versus 65 (51.6 %) and 17 (13.5 %) correct identifications obtained by the unmodified database and recommended scores at the genus and species levels, respectively. Our results support the potential utility of MALDI-TOF MS as a routine tool for accurate and reliable identification of dermatophytes.

  12. The Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) identification versus biochemical tests: a study with enterobacteria from a dairy cattle environment.

    Science.gov (United States)

    Rodrigues, Naiara Miranda Bento; Bronzato, Greiciane França; Santiago, Gabrielli Stefaninni; Botelho, Larissa Alvarenga Batista; Moreira, Beatriz Meurer; Coelho, Irene da Silva; Souza, Miliane Moreira Soares de; Coelho, Shana de Mattos de Oliveira

    Mastitis adversely affects milk production and in general cows do not regain their full production levels post recovery, leading to considerable economic losses. Moreover the percentage decrease in milk production depends on the specific pathogen that caused the infection and enterobacteria are responsible for this greater reduction. Phenotypic tests are among the currently available methods used worldwide to identify enterobacteria; however they tend to misdiagnose the species despite the multiple tests carried out. On the other hand The Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) technique has been attracting attention for its precise identification of several microorganisms at species level. In the current study, 183 enterobacteria were detected in milk (n=47) and fecal samples (n=94) from cows, and samples from water (n=23) and milk lines (n=19). All these samples were collected from a farm in Rio de Janeiro with the specific purpose of presenting the MALDI-TOF MS technique as an efficient methodology to identify Enterobacteriaceae from bovine environments. The MALDI-TOF MS technique results matched the biochemical test results in 92.9% (170/183) of the enterobacteria species and the gyrB sequencing confirmed 100% of the proteomic technique results. The amino acid decarboxylation test made the most misidentifications and Enterobacter spp. was the most misidentified genus (76.9%, 10/13). These results aim to clarify the current biochemical errors in enterobacteria identification, considering isolates from a bovine environment, and show the importance for more careful readings of phenotypic tests which are often used in veterinary microbiology laboratories. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

  13. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF) directly from positive blood culture flasks allows rapid identification of bloodstream infections in immunosuppressed hosts.

    Science.gov (United States)

    Egli, A; Osthoff, M; Goldenberger, D; Halter, J; Schaub, S; Steiger, J; Weisser, M; Frei, R

    2015-06-01

    In immunosuppressed hosts, rapid identification of microorganisms of bloodstream infections is crucial to ensuring effective antimicrobial therapy. Conventional culture requires up to 72 h from sample collection to pathogen identification. We used the SepsiTyper Kit and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF; Microflex, Bruker) directly from positive blood culture (BacT/ALERT 3D, FN/FA vials; bioMérieux) in comparison to standard culture methodology (VITEK 2; bioMérieux) for species identification. A total of 62 consecutive positive blood cultures from immunosuppressed patients (solid organ or hematopoietic transplant recipients, or with febrile neutropenia) were analyzed. Culture yielded gram-negative bacteria (GNB) in 27/62 (43.5%) and gram-positive (GPB) in 35/62 (56.5%) vials. For GNB, the predominant species identified by MALDI-TOF and confirmed by VITEK were Escherichia coli (16/16 correctly identified) and Enterobacter cloacae (4/4), with a sensitivity and specificity of 92.6% and 100%, respectively. For GPB, predominant species were Staphylococcus aureus (3/3), coagulase-negative staphylococci (12/24), and Enterococcus faecium (6/6) with a sensitivity of 100%, 60%, and 100%, respectively. The median time from blood collection to species identification was 27.4 h with MALDI-TOF identification and 46.6 h with conventional methodology. Using MALDI-TOF directly from positive blood cultures allowed a shorter time to identification with high sensitivity and specificity in immunosuppressed patients. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  14. Comparison of peptide nucleic acid fluorescence in situ hybridization assays with culture-based matrix-assisted laser desorption/ionization-time of flight mass spectrometry for the identification of bacteria and yeasts from blood cultures and cerebrospinal fluid cultures.

    Science.gov (United States)

    Calderaro, A; Martinelli, M; Motta, F; Larini, S; Arcangeletti, M C; Medici, M C; Chezzi, C; De Conto, F

    2014-08-01

    Peptide nucleic acid fluorescence in situ hybridization (PNA FISH) is a molecular diagnostic tool for the rapid detection of pathogens directly from liquid media. The aim of this study was to prospectively evaluate PNA FISH assays in comparison with culture-based matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) identification, as a reference method, for both blood and cerebrospinal fluid (CSF) cultures, during a 1-year investigation. On the basis of the Gram stain microscopy results, four different PNA FISH commercially available assays were used ('Staphylococcus aureus/CNS', 'Enterococcus faecalis/OE', 'GNR Traffic Light' and 'Yeasts Traffic Light' PNA FISH assays, AdvanDx). The four PNA FISH assays were applied to 956 positive blood cultures (921 for bacteria and 35 for yeasts) and 11 CSF cultures. Among the 921 blood samples positive for bacteria, PNA FISH gave concordant results with MALDI-TOF MS in 908/921 (98.64%) samples, showing an agreement of 99.4% in the case of monomicrobial infections. As regards yeasts, the PNA FISH assay showed a 100% agreement with the result obtained by MALDI-TOF MS. When PNA FISH assays were tested on the 11 CSF cultures, the results agreed with the reference method in all cases (100%). PNA FISH assays provided species identification at least one work-day before the MALDI-TOF MS culture-based identification. PNA FISH assays showed an excellent efficacy in the prompt identification of main pathogens, yielding a significant reduction in reporting time and leading to more appropriate patient management and therapy in cases of sepsis and severe infections. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  15. Universal protocol for the rapid automated detection of carbapenem-resistant Gram-negative bacilli directly from blood cultures by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF/MS).

    Science.gov (United States)

    Oviaño, Marina; Sparbier, Katrin; Barba, Maria José; Kostrzewa, Markus; Bou, Germán

    2016-12-01

    Detection of carbapenemase-producing bacteria directly from blood cultures is a major challenge, as patients with bacteraemia are critically ill. Early detection can be helpful for selection of the most appropriate antibiotic therapy as well as adequate control of outbreaks. In the current study, a novel matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF)-based method was developed for the rapid, automated detection of carbapenemase-producing Enterobacteriaceae, Pseudomonas aeruginosa and Acinetobacter baumannii directly from blood cultures. Carbapenemase activity was determined in 30 min by measuring hydrolysis of imipenem (0.31 mg/mL) in blood cultures spiked with a series of 119 previously characterised isolates, 81 of which carried a carbapenemase enzyme (10 blaKPC, 10 blaVIM, 10 blaNDM, 10 blaIMP, 26 blaOXA-48-type, 9 blaOXA-23, 1 blaOXA-237, 3 blaOXA-24 and 2 blaOXA-58). Twenty blood cultures obtained from bacteraemic patients carrying blaOXA-48-producing isolates were also analysed using the same protocol. Analysis was performed using MALDI-TOF Biotyper® Compass software, which automatically provides a result of sensitivity or resistance, calculated as the logRQ or ratio of hydrolysis of the antibiotic. This assay is simple to perform, inexpensive, time saving, universal for Gram-negative bacilli, and highly reliable (overall sensitivity and specificity of 98% and 100%, respectively). Moreover, the protocol could be established as a standardised method in clinical laboratories as it does not require specialised training in mass spectrometry. Copyright © 2016 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

  16. Qualitative and quantitative analysis of Potentilla fulgens roots by NMR, matrix-assisted laser desorption/ionisation with time-of-flight MS, electrospray ionisation MS/MS and HPLC/UV.

    Science.gov (United States)

    Choudhary, Alka; Radhika, Manukonda; Chatterjee, Anupam; Banerjee, Uttam Chand; Singh, Inder Pal

    2015-01-01

    Potentilla fulgens is a commonly used folk medicine by natives of northeast India, Nepal and Bhutan and is rich in polyphenolic and triterpene constituents. To identify chemomarkers in the roots of P. fulgens by an interplay of (13)C-NMR, matrix-assisted laser desorption/ionisation with time-of-flight (MALDI/TOF) MS, electrospray ionisation (ESI) MS/MS and HPLC/UV. The (13)C-NMR spectrum of crude methanolic extract was recorded in deuterated dimethyl sulphoxide. For MALDI/TOF/MS analysis, 2,5-dihydroxybenzoic acid was used as the matrix. For determination of chemical constituents, two independent simple isocratic HPLC/UV methods for monomeric/oligomeric flavanols and triterpene acids were developed and validated. The (13)C-NMR spectrum of the methanolic extract indicated the presence of B-type oligomeric polyphenolics containing mainly epicatechin/catechin (epicat/cat) and epiafzelechin/afzelechin (epiafz/afz) as the monomeric units. Several isobaric monomeric and oligomeric flavanols and triterpenoids were tentatively identified by MALDI/TOF/MS and ESI/MS/MS. Fourteen compounds (four monomeric and five dimeric flavanols and five triterpene acids) were isolated using repeated column chromatography and semi-preparative HPLC, and were quantitated using HPLC/UV. It is evident from these analyses that roots of P. fulgens contain flavans, including oligomeric flavanols, as major constituents followed by triterpene acids. The methods described can be applied to other Potentilla species to identify their constituents. Copyright © 2014 John Wiley & Sons, Ltd.

  17. Genotyping for Glycophorin GYP(B-A-B) Hybrid Genes Using a Single Nucleotide Polymorphism-Based Algorithm by Matrix-Assisted Laser Desorption/Ionisation, Time-of-Flight Mass Spectrometry.

    Science.gov (United States)

    Wei, Ling; Lopez, Genghis H; Ji, Yanli; Condon, Jennifer A; Irwin, Darryl L; Luo, Guangping; Hyland, Catherine A; Flower, Robert L

    2016-10-01

    The genetic basis for five GP(B-A-B) MNS system hybrid glycophorin blood group antigens results from rearrangement between the homologous GYPA and GYPB genes. Each hybrid glycophorin displays a characteristic profile of antigens. Currently, no commercial serological reagents are currently available to serologically type for these antigens. The aim of this study was to develop a single nucleotide polymorphism (SNP) mapping genotyping technique to allow characterisation of various GYP(B-A-B) hybrid alleles. Matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) mass spectrometry (MS) assays were designed to genotype five GYP(B-A-B) hybrid alleles. Eight nucleotide positions were targeted and incorporated into the SNP mapping protocol. The allelic frequencies were calculated using peak areas. Sanger sequencing was performed to resolve a GYP*Hop 3' breakpoint. Observed allelic peak area ratios either coincided with the expected ratio or were skewed (above or below) from the expected ratio with switching occurring at and after the expected break point to generate characteristic mass spectral plots for each hybrid. Sequencing showed that the GYP*Hop crossover in the intron 3 region, for this example, was identical to that for GYP*Bun reference sequence. An analytical algorithm using MALDI-TOF MS genotyping platform defined GYPA inserts for five GYP(B-A-B) hybrids. The SNP mapping technique described here demonstrates proof of concept that this technology is viable for genotyping hybrid glycophorins, GYP(A-B-A), GYP(A-B) and GYP(B-A), and addresses the gap in current typing technologies.

  18. Bacterial rapid identification with matrix assisted laser desorption/ionization time-of-flight mass spectrometry: development of an 'in-house method' and comparison with Bruker Sepsityper(®) kit.

    Science.gov (United States)

    Frédéric Ric, S; Antoine, M; Bodson, A; Lissoir, B

    2015-10-01

    The objective of this study was to compare an in-house matrix-assisted laser desorption ionization with time of flight (MALDI-TOF) method and a commercial MALDI-TOF kit (Sepsityper(®) kit) for direct bacterial identification in positive blood cultures. We also evaluated the time saved and the cost associated with the rapid identification techniques. We used the BACTEC(®) automated system for detecting positive blood cultures. Direct identification using Sepsityper kit and the in-house method were compared with conventional identification by MALDI-TOF using pure bacterial culture on the solid phase. We also evaluated different cut-off scores for rapid bacterial identification. In total, 127 positive blood vials were selected. The rate of rapid identification with the MALDI Sepsityper kit was 25.2% with the standard cut-off and 33.9% with the enlarged cut-off, while the results for the in-house method were 44.1 and 61.4%, respectively. Error rates with the enlarged cut-off were 6.98 (n = 3) and 2.56% (n = 2) for Sepsityper and the in-house method, respectively. Identification rates were higher for gram-negative bacteria. Direct bacterial identification succeeded in supplying rapid identification of the causative organism in cases of sepsis. The time taken to obtain a result was nearly 24  hours shorter for the direct bacterial identification methods than for conventional MALDI-TOF on solid phase culture. Compared with the Sepsityper kit, the in-house method offered better results and fewer errors, was more cost-effective and easier to use.

  19. Comparison of the Bruker Biotyper and VITEK MS Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry Systems Using a Formic Acid Extraction Method to Identify Common and Uncommon Yeast Isolates.

    Science.gov (United States)

    Lee, Hyun Seung; Shin, Jong Hee; Choi, Min Ji; Won, Eun Jeong; Kee, Seung Jung; Kim, Soo Hyun; Shin, Myung Geun; Suh, Soon Pal

    2017-05-01

    Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) allows rapid and accurate identification of clinical yeast isolates. In-tube formic acid/acetonitrile (FA/ACN) extraction is recommended prior to the analysis with MALDI Biotyper, but the direct on-plate FA extraction is simpler. We compared the Biotyper with the VITEK MS for the identification of various clinically relevant yeast species, focusing on the use of the FA extraction method. We analyzed 309 clinical isolates of 42 yeast species (four common Candida species, Cryptococcus neoformans, and 37 uncommon yeast species) using the Biotyper and VITEK MS systems. FA extraction was used initially for all isolates. If 'no identification' result was obtained following the initial FA extraction, these samples were then retested by using FA (both systems, additive FA) or FA/ACN (Biotyper only, additive FA/ACN) extraction. These results were compared with those obtained by sequence-based identification. Both systems correctly identified all 158 isolates of the four common Candida species after the initial FA extraction. The Biotyper correctly identified 8.7%, 30.4%, and 100% of 23 C. neoformans isolates after performing initial FA, additive FA, and FA/ACN extractions, respectively, while VITEK MS identified all C. neoformans isolates after the initial FA extraction. Both systems had comparable identification rates of 37 uncommon yeast species (128 isolates), following the initial FA (Biotyper, 74.2%; VITEK MS, 73.4%) or additive FA (Biotyper, 82.0%; VITEK MS, 73.4%). The identification rate of most common and uncommon yeast isolates is comparable between simple FA extraction/Biotyper method and VITEK MS methods, but FA/ACN extraction is necessary for C. neoformans identification by Biotyper.

  20. A relative and absolute quantification of neutral N-linked oligosaccharides using modification with carboxymethyl trimethylammonium hydrazide and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Gil, Geun-Cheol; Kim, Yun-Gon; Kim, Byung-Gee

    2008-08-01

    Quantification of oligosaccharides is of great importance to investigate variations or changes in the glycans of glycoconjugates. Mass spectrometry (MS) has been widely applied to identification and structural analysis of complex oligosaccharides. However, quantification using MS alone is still quite challenging due to heterogeneous charge states and different ionization efficiency of various types of oligosaccharides. To overcome such shortcomings, derivatization with carboxymethyl trimethylammonium hydrazide (Girard's reagent T [GT]) was introduced to generate a permanent cationic charge at the reducing end of neutral oligosaccharides, resulting in mainly [M](+) ion using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), so that the ambiguities caused by metal adduct peaks such as [M+K](+) and [M+Na](+) were avoided. To verify our method, the relative and absolute quantification of neutral glycans from human immunoglobulin G (IgG) and ovalbumin with internal standards of dextran ladders using MALDI-TOF MS were compared with those performed by conventional normal-phase high-performance liquid chromatography (NP-HPLC) profiling. The quantification using GT derivatization and MALDI-TOF MS agreed well with the HPLC profiling data and showed excellent reliability and reproducibility with better resolution and sensitivity. This method was further applied to quantify the enzymatically desialylated N-glycans from miniature pig kidney membrane proteins. The results showed that the low-abundance structures that could not be resolved by NP-HPLC were quantified with high sensitivity. Thus, this novel method of using modification of neutral sugars with GT is quite powerful for neutral glycan analysis, especially to quantify minute glycan samples with undetectable levels using HPLC.

  1. Use of matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry for bacterial monitoring in routine analysis at a drinking water treatment plant.

    Science.gov (United States)

    Sala-Comorera, Laura; Vilaró, Carles; Galofré, Belén; Blanch, Anicet R; García-Aljaro, Cristina

    2016-10-01

    The study of bacterial communities throughout a drinking water treatment plant could provide a basic understanding of the effects of water processing that could then be used to improve the management of such plants. However, it is necessary to develop new analytical techniques that are sufficiently efficient, robust and fast for their effective and useful application in routine analysis. The aim of this study is therefore to assess the performance of matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS), as compared to the PhenePlate™ system, for routine analysis in a drinking water treatment plant. To this end we studied a total of 277 colonies isolated in different seasons and from different points throughout the water treatment process, including: raw water, sand filtration, ultrafiltration, reverse osmosis and chlorination. The colonies were analysed using MALDI-TOF MS by direct deposition of the cells on the plate. The colonies were also biochemically fingerprinted using the PhenePlate™ system, clustered according to their similarity and a representative strain was selected for 16S rRNA gene sequencing and API® gallery-based identification. The use of MALDI-TOF MS was reliable compared to the PhenePlate™ system and has the advantage of being faster and relatively cheap. Bacteria typing by MALDI-TOF MS is therefore a promising method to replace conventional routine phenotypic methods for the identification of bacteria in drinking water laboratories, thanks to its robustness. The major limiting factor for MALDI-TOF MS is the lack of a suitable mass spectra database; although each laboratory can develop its own library. This methodology will provide a tracking tool for companies to use in risk management and the detection of possible failures in both the water treatment processes and the distribution network, as well as offering characterization of the intrinsic microbial populations. Copyright © 2016 Elsevier Gmb

  2. Matrix-assisted laser desorption/ionization mass spectrometric analysis of poly(3,4-ethylenedioxythiophene) in solid-state dye-sensitized solar cells: comparison of in situ photoelectrochemical polymerization in aqueous micellar and organic media.

    Science.gov (United States)

    Zhang, Jinbao; Ellis, Hanna; Yang, Lei; Johansson, Erik M J; Boschloo, Gerrit; Vlachopoulos, Nick; Hagfeldt, Anders; Bergquist, Jonas; Shevchenko, Denys

    2015-04-07

    Solid-state dye-sensitized solar cells (sDSCs) are devoid of such issues as electrolyte evaporation or leakage and electrode corrosion, which are typical for traditional liquid electrolyte-based DSCs. Poly(3,4-ethylenedioxythiophene) (PEDOT) is one of the most popular and efficient p-type conducting polymers that are used in sDSCs as a solid-state hole-transporting material. The most convenient way to deposit this insoluble polymer into the dye-sensitized mesoporous working electrode is in situ photoelectrochemical polymerization. Apparently, the structure and the physicochemical properties of the generated conducting polymer, which determine the photovoltaic performance of the corresponding solar cell, can be significantly affected by the preparation conditions. Therefore, a simple and fast analytical method that can reveal information on polymer chain length, possible chemical modifications, and impurities is strongly required for the rapid development of efficient solar energy-converting devices. In this contribution, we applied matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) for the analysis of PEDOT directly on sDSCs. It was found that the PEDOT generated in aqueous micellar medium possesses relatively shorter polymeric chains than the PEDOT deposited from an organic medium. Furthermore, the micellar electrolyte promotes a transformation of one of the thiophene terminal units to thiophenone. The introduction of a carbonyl group into the PEDOT molecule impedes the growth of the polymer chain and reduces the conductivity of the final polymer film. Both the simplicity of sample preparation (only application of the organic matrix onto the solar cell is needed) and the rapidity of analysis hold the promise of making MALDI MS an essential tool for the physicochemical characterization of conducting polymer-based sDSCs.

  3. Evaluation of dermal extracellular matrix and epidermal-dermal junction modifications using matrix-assisted laser desorption/ionization mass spectrometric imaging, in vivo reflectance confocal microscopy, echography, and histology: effect of age and peptide applications.

    Science.gov (United States)

    Mondon, Philippe; Hillion, Mélanie; Peschard, Olivier; Andre, Nada; Marchand, Thibault; Doridot, Emmanuel; Feuilloley, Marc Gj; Pionneau, Cédric; Chardonnet, Solenne

    2015-06-01

    This study was conducted to establish a new methodology for evaluating elements of dermal extracellular matrix (ECM), of epidermal-dermal junction (EDJ), and effects of molecules which can modulate their synthesis. This methodology is based on matrix-assisted laser desorption/ionization mass spectrometric imaging (MALDI-MSI). In vivo reflectance confocal microscopy (in vivo RCM) and echography were also used. Using immunohistochemistry methods on explants, age-related modification data were obtained for selected dermal ECM and EDJ proteins (collagen I, collagen IV, collagen VII, collagen XVII, nidogen I, decorin/decorunt) and used as reference for MALDI-MSI studies. A methodology was developed with MALDI-MSI to map epidermis and dermis proteins. Then MALDI-MSI was used to study age modifications. In vivo RCM and high-frequency ultrasounds were used to evaluate ECM and EDJ undulation modifications caused by aging. Anti-aging molecule evaluations were performed with a blend of palmitoyl oligopeptide and palmitoyl tetrapeptide-7. Immunohistochemistry studies demonstrated that the selected proteins were found to be less abundant in aged group explants vs. young group except for decorin. MALDI-MSI studies correlated the results obtained for decorin. In vivo RCM measurements indicated a decrease of EDJ undulation depth with age and ECM modifications in the upper part of dermis. Echography demonstrated that the peptide blend reduced subepidermal low-echogenic band thickness and improved its density. In vivo RCM studies indicated that the peptides improved the ECM structure vs. placebo. This preliminary MALDI-MSI study raised some technical difficulties that were overcome. Further studies will be conducted to identify more proteins and to demonstrate the interest of this method for cosmetic evaluations. © 2015 Wiley Periodicals, Inc.

  4. Species level identification of coagulase negative Staphylococcus spp. from buffalo using matrix-assisted laser desorption ionization-time of flight mass spectrometry and cydB real-time quantitative PCR.

    Science.gov (United States)

    Pizauro, Lucas J L; de Almeida, Camila C; Soltes, Glenn A; Slavic, Durda; Rossi-Junior, Oswaldo D; de Ávila, Fernando A; Zafalon, Luiz F; MacInnes, Janet I

    2017-05-01

    Incorrect identification of Staphylococcus spp. can have serious clinical and zoonotic repercussions. Accordingly, the aim of this study was to determine if matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and/or cydB real- time quantitative PCR (qPCR) could be used to accurately identify coagulase negative Staphylococcus spp. (CoNS) obtained from buffalo milk and milking environment samples. Seventy-five of 84 CoNS isolates could be identified to the species level (score value >1.99) using MALDI-TOF MS. However, as determined by cytochrome d ubiquinol oxidase subunit II (cydB) qPCR and by 16S RNA and cydB gene sequencing, 10S. agnetis strains were wrongly identified as S. hyicus by MALDI-TOF MS. In addition, 9 isolates identified by MALDI-TOF only to the genus level (score values between 1.70 and 1.99) could be identified to species by cydB qPCR. Our findings suggest that MALDI-TOF MS is a reliable method for rapid identification of S. chromogenes and S. epidermidis (species of interest both in human and veterinary medicine) and may be able to correctly identify other Staphylococcus spp. However, at present not all Staphylococcus spp. found in buffalo milk can be accurately identified by MALDI-TOF MS and for these organisms, the cydB qPCR developed in the current study may provide a reliable alternative method for rapid identification of CoNS species. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. The Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS identification versus biochemical tests: a study with enterobacteria from a dairy cattle environment

    Directory of Open Access Journals (Sweden)

    Naiara Miranda Bento Rodrigues

    Full Text Available Abstract Mastitis adversely affects milk production and in general cows do not regain their full production levels post recovery, leading to considerable economic losses. Moreover the percentage decrease in milk production depends on the specific pathogen that caused the infection and enterobacteria are responsible for this greater reduction. Phenotypic tests are among the currently available methods used worldwide to identify enterobacteria; however they tend to misdiagnose the species despite the multiple tests carried out. On the other hand The Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS technique has been attracting attention for its precise identification of several microorganisms at species level. In the current study, 183 enterobacteria were detected in milk (n = 47 and fecal samples (n = 94 from cows, and samples from water (n = 23 and milk lines (n = 19. All these samples were collected from a farm in Rio de Janeiro with the specific purpose of presenting the MALDI-TOF MS technique as an efficient methodology to identify Enterobacteriaceae from bovine environments. The MALDI-TOF MS technique results matched the biochemical test results in 92.9% (170/183 of the enterobacteria species and the gyrB sequencing confirmed 100% of the proteomic technique results. The amino acid decarboxylation test made the most misidentifications and Enterobacter spp. was the most misidentified genus (76.9%, 10/13. These results aim to clarify the current biochemical errors in enterobacteria identification, considering isolates from a bovine environment, and show the importance for more careful readings of phenotypic tests which are often used in veterinary microbiology laboratories.

  6. Evaluation of the Bruker Biotyper matrix-assisted laser desorption/ionization time-of-flight mass spectrometry system for identification of clinical and environmental isolates of Burkholderia pseudomallei

    Directory of Open Access Journals (Sweden)

    He eWang

    2016-04-01

    Full Text Available Burkholderia pseudomallei is not represented in the current version of Bruker Biotyper matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS system. A total of 66 isolates of B. pseudomallei, including 30 clinical isolates collected from National Taiwan University Hospital (NTUH, n=27 and Peking Union Medical College Hospital (PUMCH, n=3, and 36 isolates of genetically confirmed strains, including 13 from clinical samples and 23 from environmental samples, collected from southern Taiwan were included in this study. All these isolates were identified by partial 16S rDNA gene sequencing analysis and the Bruker Biotyper MALDI-TOF MS system. Among the 30 isolates initially identified as B. pseudomallei by conventional identification methods, one was identified as B. cepacia complex (NTUH and three were identified as B. putida (PUMCH by partial 16S rDNA gene sequencing analysis and Bruker Biotyper MALDI-TOF MS system. The Bruker Biotyper MALDI-TOF MS system misidentified 62 genetically confirmed B. pseudomallei isolates as B. thailandensis or Burkholderia species (score values, 1.803-2.063 when the currently available database (DB 5627 was used. However, using a newly created MALDI-TOF MS database (including B. pseudomallei NTUH-3 strain, all isolates were correctly identified as B. pseudomallei (score values >2.000, 100%. An additional 60 isolates of genetically confirmed B. cepacia complex and B. putida were also evaluated by the Bruker Biotyper MALDI-TOF MS system using the newly created database and none of these isolates were identified as B. pseudomallei. MALDI-TOF MS is a versatile and robust tool for the rapid identification of B. pseudomallei using the enhanced database.

  7. 2,5-dihydroxybenzoic acid salts for matrix-assisted laser desorption/ionization time-of-flight mass spectrometric lipid analysis: simplified spectra interpretation and insights into gas-phase fragmentation.

    Science.gov (United States)

    Jaskolla, Thorsten W; Onischke, Kristin; Schiller, Jürgen

    2014-06-30

    In the last decades the interest in lipids as important components of membranes has considerably increased. Nowadays, lipids are often routinely analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). In this regard, many relevant aspects are so far unknown, e.g., gas-phase stabilities, adduct formation and fragmentation. To fill this gap, MALDI matrix salts are presented which allow for simplified lipid analysis and elucidation of the underlying gas-phase fragmentation mechanisms. MALDI-TOF MS was used due to its beneficial properties for lipid investigations, e.g., high sensitivity, simple sample preparations, and a high tolerance to contaminants. The lipid hydrolysis, ionization and fragmentation properties of synthesized near neutral Na(+) and NH4 (+) salts of the commonly used MALDI matrix 2,5-dihydroxybenzoic acid were compared to that of DHB free acid itself as well as to base addition to DHB during dried-droplet sample preparation. Many lipid classes such as sterols, triacylglycerols, phosphatidylcholines and -ethanolamines undergo initial protonation with subsequent prompt partial up to quantitative fragmentation when analyzed with classical acidic matrices by MALDI-TOF MS. Neutral matrix salts can prevent initial analyte fragmentation by suppression of analyte protonation. Additionally, intramolecular gas-phase fragmentation reactions can be inhibited due to analyte stabilization by cation chelation. Base addition during sample preparation leads not only to in situ generation of matrix salts but also to analyte hydrolysis. Neutral DHB salts avoid separation of lipid species into several ionization states when used as matrices in MALDI-TOF MS. This allows for simplified lipid spectra interpretation. Due to the high cationization efficiency of DHB matrix salts, certain lipid classes become detectable which cannot be analyzed easily using standard acidic DHB. Copyright © 2014 John Wiley & Sons, Ltd.

  8. Off-line hyphenation of boronate affinity monolith-based extraction with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for efficient analysis of glycoproteins/glycopeptides.

    Science.gov (United States)

    Bie, Zijun; Chen, Yang; Li, Hengye; Wu, Ronghu; Liu, Zhen

    2014-06-27

    Boronate affinity materials have attracted increasing attentions as sample enrichment platforms for glycoproteomic analysis in recent years. However, most of the boronate affinity materials that have already employed for proteomic analysis are suffering from apparent disadvantages, such as alkaline pH for binding, weak affinity, and relatively poor selectivity. Benzoboroxoles are a unique class of boronic acids which have showed excellent binding properties for the recognition of cis-diol-containing compounds. Recently, a 3-carboxy-benzoboroxole-functionalized monolithic column had been reported and it had exhibited the best selectivity and affinity as well as the lowest binding pH among all reported boronate affinity monolithic columns. In this study, an off-line hyphenation of this boronate affinity monolithic column-based extraction with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was developed and the powerfulness of this hyphenated approach in the analysis of glycoproteins and glycopeptides in complex samples was investigated. The approach was first applied to the analysis of glycopeptides in the tryptic digest of horseradish peroxidase (HRP). Totally 22 glycopeptides were identified. To the best of our knowledge, this is the best performance among all the boronic acid-functionalized materials. We further employed this approach to the analysis of intact proteins in human saliva. Totally 6 intact glycoproteins were successfully identified. As comparison, when the samples were analyzed without extraction, only a few glycopeptides were identified from the tryptic digest of HRP while no glycoproteins were found from the saliva samples. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. Thymol treatment of bacteria prior to matrix-assisted laser desorption/ionization time-of-flight mass spectrometric analysis aids in identifying certain bacteria at the subspecies level.

    Science.gov (United States)

    Holland, Ricky D; Wilkes, Jon G; Cooper, Willie M; Alusta, Pierre; Williams, Anna; Pearce, Bruce; Beaudoin, Michael; Buzatu, Dan

    2014-12-15

    The identification of bacteria based on mass spectra produced by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) has become routine since its introduction in 1996. The major drawback is that bacterial patterns produced by MALDI are dependent on sample preparation prior to analysis. This results in poor reproducibility in identifying bacterial types and between laboratories. The need for a more broadly applicable and useful sample handling procedure is warranted. Thymol was added to the suspension solvent of bacteria prior to MALDI analysis. The suspension solvent consisted of ethanol, water and TFA. The bacterium was added to the thymol suspension solvent and heated. An aliquot of the bacterial suspension was mixed directly with the matrix solution at a 9:1 ratio, matrix/bacteria solution, respectively. The mixture was then placed on the MALDI plate and allowed to air dry before MALDI analysis. The thymol method improved the quality of spectra and number of peaks when compared to other sample preparation procedures studied. The bacterium-identifying biomarkers assigned to four strains of E. coli were statistically 95% reproducible analyzed on three separate days. The thymol method successfully differentiated between the four E. coli strains. In addition, the thymol procedure could identify nine out of ten S. enterica serovars over a 3-day period and nine S. Typhimurium strains from the other ten serovars 90% of the time over the same period. The thymol method can identify certain bacteria at the sub-species level and yield reproducible results over time. It improves the quality of spectra by increasing the number of peaks when compared to the other sample preparation methods assessed in this study. Published in 2014. This article is a U.S. Government work and is in the public domain in the USA. Published in 2014. This article is a U.S. Government work and is in the public domain in the USA.

  10. A clean-up technology for the simultaneous determination of lysophosphatidic acid and sphingosine-1-phosphate by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry using a phosphate-capture molecule, Phos-tag.

    Science.gov (United States)

    Morishige, Jun-ichi; Urikura, Mai; Takagi, Haruko; Hirano, Kaoru; Koike, Tohru; Tanaka, Tamotsu; Satouchi, Kiyoshi

    2010-04-15

    Lysophosphatidic acid (LPA) and sphingosine-1-phosphate (S1P) are growth factor-like lipids having a phosphate group. The concentrations of these mediator lipids in blood are considered to be potential biomarkers for early detection of cancer or vascular diseases. Here, we report a method for simultaneous determination of LPA and S1P using Phos-tag, a zinc complex that specifically binds to a phosphate-monoester group. Although both LPA and S1P are hydrophilic compounds, we found that they acquire hydrophobic properties when they form complexes with Phos-tag. Based on this finding, we developed a method for the enrichment of LPA and S1P from biological samples. The first partition in a two-phase solvent system consisting of chloroform/methanol/water (1:1:0.9, v/v/v) is conducted for the removal of lipids. LPA and S1P are specifically extracted as Phos-tag complexes at the second partition by adding Phos-tag. The Phos-tag complexes of LPA and S1P are detectable by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) and quantifiable based on the relative intensities of ions using 17:0 LPA and C17 S1P as internal standards. The protocol was validated by analyses of these mediator lipids in calf serum, a rat brain and a lung. The clean-up protocol is rapid, requires neither thin-layer chromatography (TLC) nor liquid chromatography (LC), and is applicable to both blood and solid tissue samples. We believe that our protocol will be useful for a routine analysis of LPA and S1P in many clinical samples. 2010 John Wiley & Sons, Ltd.

  11. Species-Level Identification of Actinomyces Isolates Causing Invasive Infections: Multiyear Comparison of Vitek MS (Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry) to Partial Sequencing of the 16S rRNA Gene

    Science.gov (United States)

    Gregson, D.; Church, D. L.

    2016-01-01

    Actinomyces species are uncommon but important causes of invasive infections. The ability of our regional clinical microbiology laboratory to report species-level identification of Actinomyces relied on molecular identification by partial sequencing of the 16S ribosomal gene prior to the implementation of the Vitek MS (matrix-assisted laser desorption ionization–time of flight mass spectrometry [MALDI-TOF MS]) system. We compared the use of the Vitek MS to that of 16S rRNA gene sequencing for reliable species-level identification of invasive infections caused by Actinomyces spp. because limited data had been published for this important genera. A total of 115 cases of Actinomyces spp., either alone or as part of a polymicrobial infection, were diagnosed between 2011 and 2014. Actinomyces spp. were considered the principal pathogen in bloodstream infections (n = 17, 15%), in skin and soft tissue abscesses (n = 25, 22%), and in pulmonary (n = 26, 23%), bone (n = 27, 23%), intraabdominal (n = 16, 14%), and central nervous system (n = 4, 3%) infections. Compared to sequencing and identification from the SmartGene Integrated Database Network System (IDNS), Vitek MS identified 47/115 (41%) isolates to the correct species and 10 (9%) isolates to the correct genus. However, the Vitek MS was unable to provide identification for 43 (37%) isolates while 15 (13%) had discordant results. Phylogenetic analyses of the 16S rRNA sequences demonstrate high diversity in recovered Actinomyces spp. and provide additional information to compare/confirm discordant identifications between MALDI-TOF and 16S rRNA gene sequences. This study highlights the diversity of clinically relevant Actinomyces spp. and provides an important typing comparison. Based on our analysis, 16S rRNA gene sequencing should be used to rapidly identify Actinomyces spp. until MALDI-TOF databases are optimized. PMID:26739153

  12. Species-Level Identification of Actinomyces Isolates Causing Invasive Infections: Multiyear Comparison of Vitek MS (Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry) to Partial Sequencing of the 16S rRNA Gene.

    Science.gov (United States)

    Lynch, T; Gregson, D; Church, D L

    2016-03-01

    Actinomyces species are uncommon but important causes of invasive infections. The ability of our regional clinical microbiology laboratory to report species-level identification of Actinomyces relied on molecular identification by partial sequencing of the 16S ribosomal gene prior to the implementation of the Vitek MS (matrix-assisted laser desorption ionization-time of flight mass spectrometry [MALDI-TOF MS]) system. We compared the use of the Vitek MS to that of 16S rRNA gene sequencing for reliable species-level identification of invasive infections caused by Actinomyces spp. because limited data had been published for this important genera. A total of 115 cases of Actinomyces spp., either alone or as part of a polymicrobial infection, were diagnosed between 2011 and 2014. Actinomyces spp. were considered the principal pathogen in bloodstream infections (n = 17, 15%), in skin and soft tissue abscesses (n = 25, 22%), and in pulmonary (n = 26, 23%), bone (n = 27, 23%), intraabdominal (n = 16, 14%), and central nervous system (n = 4, 3%) infections. Compared to sequencing and identification from the SmartGene Integrated Database Network System (IDNS), Vitek MS identified 47/115 (41%) isolates to the correct species and 10 (9%) isolates to the correct genus. However, the Vitek MS was unable to provide identification for 43 (37%) isolates while 15 (13%) had discordant results. Phylogenetic analyses of the 16S rRNA sequences demonstrate high diversity in recovered Actinomyces spp. and provide additional information to compare/confirm discordant identifications between MALDI-TOF and 16S rRNA gene sequences. This study highlights the diversity of clinically relevant Actinomyces spp. and provides an important typing comparison. Based on our analysis, 16S rRNA gene sequencing should be used to rapidly identify Actinomyces spp. until MALDI-TOF databases are optimized. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  13. Mass spectrometry based lipid(ome) analyzer and molecular platform: a new software to interpret and analyze electrospray and/or matrix-assisted laser desorption/ionization mass spectrometric data of lipids: a case study from Mycobacterium tuberculosis.

    Science.gov (United States)

    Sabareesh, Varatharajan; Singh, Gurpreet

    2013-04-01

    Mass Spectrometry based Lipid(ome) Analyzer and Molecular Platform (MS-LAMP) is a new software capable of aiding in interpreting electrospray ionization (ESI) and/or matrix-assisted laser desorption/ionization (MALDI) mass spectrometric data of lipids. The graphical user interface (GUI) of this standalone programme is built using Perl::Tk. Two databases have been developed and constituted within MS-LAMP, on the basis of Mycobacterium tuberculosis (M. tb) lipid database (www.mrl.colostate.edu) and that of Lipid Metabolites and Pathways Strategy Consortium (LIPID MAPS; www.lipidmaps.org). Different types of queries entered through GUI would interrogate with a chosen database. The queries can be molecular mass(es) or mass-to-charge (m/z) value(s) and molecular formula. LIPID MAPS identifier also can be used to search but not for M. tb lipids. Multiple choices have been provided to select diverse ion types and lipids. Satisfying to input parameters, a glimpse of various lipid categories and their population distribution can be viewed in the output. Additionally, molecular structures of lipids in the output can be seen using ChemSketch (www.acdlabs.com), which has been linked to the programme. Furthermore, a version of MS-LAMP for use in Linux operating system is separately available, wherein PyMOL can be used to view molecular structures that result as output from General Lipidome MS-LAMP. The utility of this software is demonstrated using ESI mass spectrometric data of lipid extracts of M. tb grown under two different pH (5.5 and 7.0) conditions. Copyright © 2013 John Wiley & Sons, Ltd.

  14. Yersinia enterocolitica in Diagnostic Fecal Samples from European Dogs and Cats: Identification by Fourier Transform Infrared Spectroscopy and Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

    Science.gov (United States)

    Stamm, Ivonne; Hailer, Mandy; Depner, Barbara; Kopp, Peter A.

    2013-01-01

    Yersinia enterocolitica is the main cause of yersiniosis in Europe, one of the five main bacterial gastrointestinal diseases of humans. Beside pigs, companion animals, especially dogs and cats, were repeatedly discussed in the past as a possible source of pathogenic Y. enterocolitica. To investigate the presence and types of Y. enterocolitica in companion animals, a total of 4,325 diagnostic fecal samples from dogs and 2,624 samples from cats were tested. The isolates obtained were differentiated by using matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) and Fourier transform infrared spectroscopy (FT-IR). Isolated Y. enterocolitica strains were bioserotyped. The detection of the ail gene by PCR and confirmation by FT-IR were used as a pathogenicity marker. Y. enterocolitica strains were isolated from 198 (4.6%) of the dog and 8 (0.3%) of the cat fecal samples investigated. One hundred seventy-nine isolates from dogs were analyzed in detail. The virulence factor Ail was detected in 91.6% of isolates. Isolates of biotype 4 (54.7%) and, to a lesser extent, biotypes 2 (23.5%), 3 (11.2%), and 5 (2.2%) were detected. The remaining 8.4% of strains belonged to the ail-negative biotype 1A. All 7 isolates from cats that were investigated in detail were ail positive. These results indicate that companion animals could be a relevant reservoir for a broad range of presumptively human-pathogenic Y. enterocolitica types. MALDI-TOF MS and FT-IR proved to be valuable methods for the rapid identification of Y. enterocolitica, especially in regard to the large number of samples that were investigated in a short time frame. PMID:23284028

  15. Yersinia enterocolitica in diagnostic fecal samples from European dogs and cats: identification by fourier transform infrared spectroscopy and matrix-assisted laser desorption ionization-time of flight mass spectrometry.

    Science.gov (United States)

    Stamm, Ivonne; Hailer, Mandy; Depner, Barbara; Kopp, Peter A; Rau, Jörg

    2013-03-01

    Yersinia enterocolitica is the main cause of yersiniosis in Europe, one of the five main bacterial gastrointestinal diseases of humans. Beside pigs, companion animals, especially dogs and cats, were repeatedly discussed in the past as a possible source of pathogenic Y. enterocolitica. To investigate the presence and types of Y. enterocolitica in companion animals, a total of 4,325 diagnostic fecal samples from dogs and 2,624 samples from cats were tested. The isolates obtained were differentiated by using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and Fourier transform infrared spectroscopy (FT-IR). Isolated Y. enterocolitica strains were bioserotyped. The detection of the ail gene by PCR and confirmation by FT-IR were used as a pathogenicity marker. Y. enterocolitica strains were isolated from 198 (4.6%) of the dog and 8 (0.3%) of the cat fecal samples investigated. One hundred seventy-nine isolates from dogs were analyzed in detail. The virulence factor Ail was detected in 91.6% of isolates. Isolates of biotype 4 (54.7%) and, to a lesser extent, biotypes 2 (23.5%), 3 (11.2%), and 5 (2.2%) were detected. The remaining 8.4% of strains belonged to the ail-negative biotype 1A. All 7 isolates from cats that were investigated in detail were ail positive. These results indicate that companion animals could be a relevant reservoir for a broad range of presumptively human-pathogenic Y. enterocolitica types. MALDI-TOF MS and FT-IR proved to be valuable methods for the rapid identification of Y. enterocolitica, especially in regard to the large number of samples that were investigated in a short time frame.

  16. Impact of matrix-assisted laser desorption ionization time-of-flight mass spectrometry on the clinical management of patients with Gram-negative bacteremia: a prospective observational study.

    Science.gov (United States)

    Clerc, Olivier; Prod'hom, Guy; Vogne, Christelle; Bizzini, Alain; Calandra, Thierry; Greub, Gilbert

    2013-04-01

    Early identification of pathogens from blood cultures using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry may optimize the choice of empirical antibiotic therapy in the setting of bloodstream infections. We aimed to assess the impact of this new technology on the use of antibiotic treatment in patients with gram-negative bacteremia. We conducted a prospective observational study from January to December 2010 to evaluate the sequential and separate impacts of Gram stain reporting and MALDI-TOF bacterial identification performed on blood culture pellets in patients with gram-negative bacteremia. The primary outcome was the impact of MALDI-TOF on empirical antibiotic choice. Among 202 episodes of gram-negative bacteremia, Gram stain reporting had an impact in 42 cases (20.8%). MALDI-TOF identification led to a modification of empirical therapy in 71 of all 202 cases (35.1%), and in 16 of 27 cases (59.3%) of monomicrobial bacteremia caused by AmpC-producing Enterobacteriaceae. The most frequently observed impact was an early appropriate broadening of the antibiotic spectrum in 31 of 71 cases (43.7%). In total, 143 of 165 episodes (86.7%) of monomicrobial bacteremia were correctly identified at genus level by MALDI-TOF. In a low prevalence area for extended spectrum betalactamases (ESBL) and multiresistant gram-negative bacteria, MALDI-TOF performed on blood culture pellets had an impact on the clinical management of 35.1% of all gram-negative bacteremia cases, demonstrating a greater impact than Gram stain reporting. Thus, MALDI-TOF could become a vital second step beside Gram stain in guiding the empirical treatment of patients with bloodstream infection.

  17. Evaluation of matrix-assisted laser desorption ionization-time-of-flight mass spectrometry in comparison to 16S rRNA gene sequencing for species identification of nonfermenting bacteria.

    Science.gov (United States)

    Mellmann, A; Cloud, J; Maier, T; Keckevoet, U; Ramminger, I; Iwen, P; Dunn, J; Hall, G; Wilson, D; Lasala, P; Kostrzewa, M; Harmsen, D

    2008-06-01

    Nonfermenting bacteria are ubiquitous environmental opportunists that cause infections in humans, especially compromised patients. Due to their limited biochemical reactivity and different morphotypes, misidentification by classical phenotypic means occurs frequently. Therefore, we evaluated the use of matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) for species identification. By using 248 nonfermenting culture collection strains composed of 37 genera most relevant to human infections, a reference database was established for MALDI-TOF MS-based species identification according to the manufacturer's recommendations for microflex measurement and MALDI BioTyper software (Bruker Daltonik GmbH, Leipzig, Germany), i.e., by using a mass range of 2,000 to 20,000 Da and a new pattern-matching algorithm. To evaluate the database, 80 blind-coded clinical nonfermenting bacterial strains were analyzed. As a reference method for species designation, partial 16S rRNA gene sequencing was applied. By 16S rRNA gene sequencing, 57 of the 80 isolates produced a unique species identification (>or=99% sequence similarity); 11 further isolates gave ambiguous results at this threshold and were rated as identified to the genus level only. Ten isolates were identified to the genus level (>or=97% similarity); and two isolates had similarity values below this threshold, were counted as not identified, and were excluded from further analysis. MALDI-TOF MS identified 67 of the 78 isolates (85.9%) included, in agreement with the results of the reference method; 9 were misidentified and 2 were unidentified. The identities of 10 randomly selected strains were 100% correct when three different mass spectrometers and four different cultivation media were used. Thus, MALDI-TOF MS-based species identification of nonfermenting bacteria provided accurate and reproducible results within 10 min without any substantial costs for consumables.

  18. Ammonia-treated N-(1-naphthyl) ethylenediamine dihydrochloride as a novel matrix for rapid quantitative and qualitative determination of serum free fatty acids by matrix-assisted laser desorption/ionization-Fourier transform ion cyclotron resonance mass spectrometry.

    Science.gov (United States)

    Zhang, Yaping; Wang, Yanmin; Guo, Shuai; Guo, Yumei; Liu, Hui; Li, Zhili

    2013-09-10

    The blood free fatty acids (FFAs), which provide energy to the cell and act as substrates in the synthesis of fats, lipoproteins, liposaccharides, and eicosanoids, involve in a number of important physiological processes. In the present study, matrix-assisted laser desorption/ionization-Fourier transform ion cyclotron resonance mass spectrometry (MALDI-FTICR MS) with ammonia-treated N-(1-naphthyl) ethylenediamine dihydrochloride (ATNEDC) as a novel MALDI matrix in a negative ion mode was employed to directly quantify serum FFAs. Multiple point internal standard calibration curves between the concentration ratios of individual fatty acids to internal standard (IS, C17:0) versus their corresponding intensity ratios were constructed for C14:0, C16:1, C16:0, C18:0, C18:1, C18:2, C18:3, C20:4, and C22:6, respectively, in their mixture, with correlation coefficients between 0.991 and 0.999 and limits of detection (LODs) between 0.2 and 5.4μM, along with the linear dynamic range of more than two orders of magnitude. The results indicate that the multiple point internal standard calibration could reduce the impact of ion suppression and improve quantification accuracy in the MALDI mode. The quantitative results of nine FFAs from 339 serum samples, including 161 healthy controls, 118 patients with hyperglycemia and 60 patients without hyperglycemia show that FFAs levels in hyperglycemic patient sera are significantly higher than those in healthy controls and patients without hyperglycemia, and elevated FFA levels are also associated with increased levels of fasting blood glucose (FBG) in hyperglycemic patient sera. Serum FFAs were identified on the basis of the observed accurate molecular masses and reliable isotope distributions obtained by MALDI-FTICR MS. Copyright © 2013 Elsevier B.V. All rights reserved.

  19. A silicon nanomembrane detector for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) of large proteins.

    Science.gov (United States)

    Park, Jonghoo; Blick, Robert H

    2013-10-11

    We describe a MALDI-TOF ion detector based on freestanding silicon nanomembrane technology. The detector is tested in a commercial MALDI-TOF mass spectrometer with equimolar mixtures of proteins. The operating principle of the nanomembrane detector is based on phonon-assisted field emission from these silicon nanomembranes, in which impinging ion packets excite electrons in the nanomembrane to higher energy states. Thereby the electrons can overcome the vacuum barrier and escape from the surface of the nanomembrane via field emission. Ion detection is demonstrated of apomyoglobin (16,952 Da), aldolase (39,212 Da), bovine serum albumin (66,430 Da), and their equimolar mixtures. In addition to the three intact ions, a large number of fragment ions are also revealed by the silicon nanomembrane detector, which are not observable with conventional detectors.

  20. A Silicon Nanomembrane Detector for Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS of Large Proteins

    Directory of Open Access Journals (Sweden)

    Jonghoo Park

    2013-10-01

    Full Text Available We describe a MALDI-TOF ion detector based on freestanding silicon nanomembrane technology. The detector is tested in a commercial MALDI-TOF mass spectrometer with equimolar mixtures of proteins. The operating principle of the nanomembrane detector is based on phonon-assisted field emission from these silicon nanomembranes, in which impinging ion packets excite electrons in the nanomembrane to higher energy states. Thereby the electrons can overcome the vacuum barrier and escape from the surface of the nanomembrane via field emission. Ion detection is demonstrated of apomyoglobin (16,952 Da, aldolase (39,212 Da, bovine serum albumin (66,430 Da, and their equimolar mixtures. In addition to the three intact ions, a large number of fragment ions are also revealed by the silicon nanomembrane detector, which are not observable with conventional detectors.

  1. Laser Desorption Ionization Mass Spectrometry Imaging of Drosophila Brain Using Matrix Sublimation versus Modification with Nanoparticles.

    Science.gov (United States)

    Phan, Nhu T N; Mohammadi, Amir Saeid; Dowlatshahi Pour, Masoumeh; Ewing, Andrew G

    2016-02-02

    Laser desorption ionization mass spectrometry (LDI-MS) is used to image brain lipids in the fruit fly, Drosophila, a common invertebrate model organism in biological and neurological studies. Three different sample preparation methods, including sublimation with two common organic matrixes for matrix-assisted laser desorption ionization (MALDI) and surface-assisted laser desorption ionization (SALDI) using gold nanoparticles, are examined for sample profiling and imaging the fly brain. Recrystallization with trifluoroacetic acid following matrix deposition in MALDI is shown to increase the incorporation of biomolecules with one matrix, resulting in more efficient ionization, but not for the other matrix. The key finding here is that the mass fragments observed for the fly brain slices with different surface modifications are significantly different. Thus, these approaches can be combined to provide complementary analysis of chemical composition, particularly for the small metabolites, diacylglycerides, phosphatidylcholines, and triacylglycerides, in the fly brain. Furthermore, imaging appears to be beneficial using modification with gold nanoparticles in place of matrix in this application showing its potential for cellular and subcellular imaging. The imaging protocol developed here with both MALDI and SALDI provides the best and most diverse lipid chemical images of the fly brain to date with LDI.

  2. Coffee-ring effects in laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Hu, Jie-Bi; Chen, Yu-Chie; Urban, Pawel L

    2013-03-05

    This report focuses on the heterogeneous distribution of small molecules (e.g. metabolites) within dry deposits of suspensions and solutions of inorganic and organic compounds with implications for chemical analysis of small molecules by laser desorption/ionization (LDI) mass spectrometry (MS). Taking advantage of the imaging capabilities of a modern mass spectrometer, we have investigated the occurrence of "coffee rings" in matrix-assisted laser desorption/ionization (MALDI) and surface-assisted laser desorption/ionization (SALDI) sample spots. It is seen that the "coffee-ring effect" in MALDI/SALDI samples can be both beneficial and disadvantageous. For example, formation of the coffee rings gives rise to heterogeneous distribution of analytes and matrices, thus compromising analytical performance and reproducibility of the mass spectrometric analysis. On the other hand, the coffee-ring effect can also be advantageous because it enables partial separation of analytes from some of the interfering molecules present in the sample. We report a "hidden coffee-ring effect" where under certain conditions the sample/matrix deposit appears relatively homogeneous when inspected by optical microscopy. Even in such cases, hidden coffee rings can still be found by implementing the MALDI-MS imaging technique. We have also found that to some extent, the coffee-ring effect can be suppressed during SALDI sample preparation. Copyright © 2013 Elsevier B.V. All rights reserved.

  3. Laser desorption and time-of-flight mass spectrometry. Fundamentals .Applications; Desorption laser et spectrometrie de masse par temps de vol. Aspects fondamentaux. Applications

    Energy Technology Data Exchange (ETDEWEB)

    Chaurand, P.

    1994-11-01

    Time-of-flight mass spectrometry is a very powerful technique for the analysis of heavy molecular ions (100 000 u and more). The ejection in the gas phase and the ionization of these molecules is now possible through the MALDI technique (Matrix Assisted Laser Desorption Ionization). This technique consists in mixing the heavy molecules to be analysed with a organic matrix which absorbs at the wavelength of the laser. The necessary irradiance are of the order of 10{sup 6} W/cm{sup 2}. In these conditions we have shown that the mass resolutions are optimum and that the relative mass accuracies are of the order of 10{sup -4}. We have also demonstrated that the emission angle of the molecular ions in MALDI depends on the incident angle of the laser light. During the desorption process, the molecular ions are emitted in the opposite direction of the incident laser light. This effect is particularly important for the design of the accelerating stage of the time-of-flight spectrometers. Problems relative to the detection of these heavy molecular ions have been studied in details between 0.5 10{sup 4} m/s and 10{sup 5} m/s. The velocity threshold of the electronic emission is lower than the value of 0.5 10{sup 4} m/s. The relation between the electronic emission and the projectile velocity is complex. Finally, examples on mass identification of C{sub 60} molecules and derivated C{sub 60} are presented. Desorption methods are compared. (author). 32 refs., 34 figs.

  4. Laser desorption/ionization mass spectrometric analysis of small molecules using fullerene-derivatized silica as energy-absorbing material.

    Science.gov (United States)

    Szabo, Zoltan; Vallant, Rainer M; Takátsy, Anikó; Bakry, Rania; Najam-ul-Haq, Muhammad; Rainer, Matthias; Huck, Christian W; Bonn, Günther K

    2010-05-01

    In spite of the growing acceptance of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for the analysis of a wide variety of compounds, including polymers and proteins, its use in analyzing low-molecular-weight molecules (silica particles with different pore sizes are applied as thin layer for laser desorption/ionization (LDI) mass spectrometric analysis. Thus, an interference of intrinsic matrix ions can be eliminated or minimized in comparison with the state-of-the-art weak organic acid matrices. The desorption/ionization ability of the developed fullerene-silica materials depends on the applied laser power, sample preparation and pore size of the silica particles. Thus, fullerene-silica serves as an LDI support for mass spectrometric analysis of molecules (silica is demonstrated by the mass analysis of variety of small molecules such as carbohydrates, amino acids, peptides, phospholipids and drugs. 2010 John Wiley & Sons, Ltd.

  5. Identification of Low Molecular Weight Glutenin Alleles by Matrix-Assisted Laser Desorption/Ionization Time-Of-Flight Mass Spectrometry (MALDI-TOF-MS) in Common Wheat (Triticum aestivum L.).

    Science.gov (United States)

    Wang, Aili; Liu, Li; Peng, Yanchun; Islam, Shahidul; Applebee, Marie; Appels, Rudi; Yan, Yueming; Ma, Wujun

    2015-01-01

    Low molecular weight glutenin subunits (LMW-GS) play an important role in determining dough properties and breadmaking quality. However, resolution of the currently used methodologies for analyzing LMW-GS is rather low which prevents an efficient use of genetic variations associated with these alleles in wheat breeding. The aim of the current study is to evaluate and develop a rapid, simple, and accurate method to differentiate LMW-GS alleles using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. A set of standard single LMW-GS allele lines as well as a suite of well documented wheat cultivars were collected from France, CIMMYT, and Canada. Method development and optimization were focused on protein extraction procedures and MALDI-TOF instrument settings to generate reproducible diagnostic spectrum peak profiles for each of the known wheat LMW-GS allele. Results revealed a total of 48 unique allele combinations among the studied genotypes. Characteristic MALDI-TOF peak patterns were obtained for 17 common LMW-GS alleles, including 5 (b, a or c, d, e, f), 7 (a, b, c, d or i, f, g, h) and 5 (a, b, c, d, f) patterns or alleles for the Glu-A3, Glu-B3, and Glu-D3 loci, respectively. In addition, some reproducible MALDI-TOF peak patterns were also obtained that did not match with any known alleles. The results demonstrated a high resolution and throughput nature of MALDI-TOF technology in analyzing LMW-GS alleles, which is suitable for application in wheat breeding programs in processing a large number of wheat lines with high accuracy in limited time. It also suggested that the variation of LMW-GS alleles is more abundant than what has been defined by the current nomenclature system that is mainly based on SDS-PAGE system. The MALDI-TOF technology is useful to differentiate these variations. An international joint effort may be needed to assign allele symbols to these newly identified alleles and determine their effects on end

  6. Matrix-assisted Laser Desorption Ionization-Time-of-Flight Mass Spectrometry (MALDI-TOF MS) as a Reliable Tool to Identify Species of Catalase-negative Gram-positive Cocci not Belonging to the Streptococcus Genus.

    Science.gov (United States)

    Almuzara, Marisa; Barberis, Claudia; Velázquez, Viviana Rojas; Ramirez, Maria Soledad; Famiglietti, Angela; Vay, Carlos

    2016-01-01

    To evaluate the performance of matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) by using 190 Catalase-negative Gram-Positive Cocci (GPC) clinical isolates. All isolates were identified by conventional phenotypic tests following the proposed scheme by Ruoff and Christensen and MALDI-TOF MS (Bruker Daltonics, BD, Bremen, Germany). Two different extraction methods (direct transfer formic acid method on spot and ethanol formic acid extraction method) and different cut-offs for genus/specie level identification were used. The score cut-offs recommended by the manufacturer (≥ 2.000 for species-level, 1.700 to 1.999 for genus level and MALDI-TOF MS identification was considered correct when the result obtained from MS database agreed with the phenotypic identification result. When both methods gave discordant results, the 16S rDNA or sodA genes sequencing was considered as the gold standard identification method. The results obtained by MS concordant with genes sequencing, although discordant with conventional phenotyping, were considered correct. MS results discordant with 16S or sodA identification were considered incorrect. Using the score cut-offs recommended by the manufacturer, 97.37% and 81.05% were correctly identified to genus and species level, respectively. On the other hand, using lower cut-off scores for identification, 97.89% and 94.21% isolates were correctly identified to genus and species level respectively by MALDI-TOF MS and no significant differences between the results obtained with two extraction methods were obtained. The results obtained suggest that MALDI-TOF MS has the potential of being an accurate tool for Catalase-negative GPC identification even for those species with difficult diagnosis as Helcococcus, Abiotrophia, Granulicatella, among others. Nevertheless, expansion of the library, especially including more strains with different spectra on the same species might overcome potential "intraspecies

  7. Comparison of ZnS semiconductor nanoparticles capped with various functional groups as the matrix and affinity probes for rapid analysis of cyclodextrins and proteins in surface-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Kailasa, Suresh Kumar; Kiran, Kamatam; Wu, Hui-Fen

    2008-12-15

    Zinc sulfide (ZnS) semiconductor nanoparticles (NPs) capped with a variety of functional groups including bare ZnS NPs, 3-mercaptopropanoic acid (ZnS-3-MPA), sodium citrate (ZnS-citrate), cysteamine (ZnS-Cys), and 2-mercaptoethane sulfonate (ZnS-2-MES) have been investigated as the matrix and affinity probes for analysis of alpha-, beta-, and gamma-cyclodextrins (CDs), ubiquitin, and insulin in biological samples by using surface-assisted laser desorption/ionization time-of-flight mass spectrometry (SALDI-TOF-MS). Various parameters that would influence the ionization efficiency and sensitivity of these ZnS NPs in SALDI-TOF-MS were examined including the effect of capping agents, sample pH, ion abundance, and concentration of ZnS NPs. Among these ZnS NPs, our results have demonstrated that ZnS-3-MPA exhibited the highest efficiency toward CDs, ubiquitin, and insulin for high-sensitivity detection in SALDI-TOF-MS. The detection limits were 20-55 nM for CDs, 91 nM for ubiquitin, and 85 nM for insulin. The applicability of the present method is demonstrated by detection of ubiquitin-like proteins in oyster mushroom and also in the analysis of analytes in biological samples such as human urine and plasma. To our best knowledge, this is the first time semiconductor NPs were used as the matrix and affinity probes for high-sensitivity detection of organic and biomolecules in SALDI-TOF-MS. This approach exhibits the advantages of being simple, rapid, efficient, and straightforward for direct analysis of organic and biological samples in SALDI-TOF-MS without the need for time-consuming separation processes, tedious washing steps, or further laborious purification. In addition, it also can provide a sensitive and reliable quantitative assay for small- and large-molecule analysis with the detectable mass up to 8500 Da. We believe that this novel ZnS nanoprobe is simple, efficient, lower cost (compared with Au, Ag, and Pt NPs), fast, and with the potential for high

  8. Ammonia-treated N-(1-naphthyl) ethylenediamine dihydrochloride as a novel matrix for rapid quantitative and qualitative determination of serum free fatty acids by matrix-assisted laser desorption/ionization-Fourier transform ion cyclotron resonance mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Yaping [Department of Biophysics and Structural Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and School of Basic Medicine, Peking Union Medical College, 5 Dongdan San Tiao, Beijing 100005 (China); Wang, Yanmin [Department of Clinical Laboratory, Heze Municipal Hospital, Shandong (China); Guo, Shuai; Guo, Yumei; Liu, Hui [Department of Biophysics and Structural Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and School of Basic Medicine, Peking Union Medical College, 5 Dongdan San Tiao, Beijing 100005 (China); Li, Zhili, E-mail: lizhili@ibms.pumc.edu.cn [Department of Biophysics and Structural Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and School of Basic Medicine, Peking Union Medical College, 5 Dongdan San Tiao, Beijing 100005 (China)

    2013-09-10

    Graphical abstract: -- Highlights: •A novel MALDI matrix for the detection of serum free fatty acids is ammonia-treated N-(1-naphthyl) ethylenediamine dihydrochloride. •Multiple point internal standard calibration curves were constructed for nine FFAs, respectively, with excellent correlation coefficients between 0.991 and 0.999. •The MALDI-MS approach was used to rapidly differentiate the patients with and without hyperglycemia and healthy controls. -- Abstract: The blood free fatty acids (FFAs), which provide energy to the cell and act as substrates in the synthesis of fats, lipoproteins, liposaccharides, and eicosanoids, involve in a number of important physiological processes. In the present study, matrix-assisted laser desorption/ionization-Fourier transform ion cyclotron resonance mass spectrometry (MALDI-FTICR MS) with ammonia-treated N-(1-naphthyl) ethylenediamine dihydrochloride (ATNEDC) as a novel MALDI matrix in a negative ion mode was employed to directly quantify serum FFAs. Multiple point internal standard calibration curves between the concentration ratios of individual fatty acids to internal standard (IS, C{sub 17:0}) versus their corresponding intensity ratios were constructed for C{sub 14:0}, C{sub 16:1}, C{sub 16:0}, C{sub 18:0}, C{sub 18:1}, C{sub 18:2}, C{sub 18:3}, C{sub 20:4}, and C{sub 22:6}, respectively, in their mixture, with correlation coefficients between 0.991 and 0.999 and limits of detection (LODs) between 0.2 and 5.4 μM, along with the linear dynamic range of more than two orders of magnitude. The results indicate that the multiple point internal standard calibration could reduce the impact of ion suppression and improve quantification accuracy in the MALDI mode. The quantitative results of nine FFAs from 339 serum samples, including 161 healthy controls, 118 patients with hyperglycemia and 60 patients without hyperglycemia show that FFAs levels in hyperglycemic patient sera are significantly higher than those in healthy

  9. "Bottom-up" in situ proteomic differentiation of human and non-human haemoglobins for forensic purposes by matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry.

    Science.gov (United States)

    Kamanna, S; Henry, J; Voelcker, N; Linacre, A; Kirkbride, K P

    2017-11-30

    The detection and identification of human blood on crime-related items are of particular relevance to many investigations because shed blood can provide evidence of violent contact between individuals. However, for any detection and identification technique, specificity is a critical performance characteristic to assess; that is, whether the technique has the capability to differentiate between human blood (which usually is of relevance to a criminal investigation) and non-human blood (which usually would not be associated with a crime but may be detected incidentally). Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) approaches using "top-down" (detection of intact proteins) and "bottom-up" (detection of tryptic peptide markers) were used to detect and identify haemoglobin in blood from humans and from a range of Australian native mammals; the technique could be carried out directly on blood stains without the need to extract proteins (i.e., in situ measurement). Imaging of haemoglobin was achieved in bloodied fingermarks, including those that had been enhanced using two "industry standard" fingermark enhancement processes. Differentiation of intact haemoglobin proteins in human and non-human blood using "top-down" MALDI-TOF-MS was difficult. However, in situ "bottom-up" approaches using tandem mass spectrometry (MS/MS) and de novo sequencing of tryptic digest peptides allowed unambiguous differentiation. Imaging mass spectrometry of human haemoglobin, even when it was mixed with animal blood, was achieved in bloodied fingermarks that had been enhanced using two common processes (staining with Amido Black or dusted with magnetic powder) and "lifted" using adhesive tape. The MALDI-TOF-MS-based in situ "bottom-up" proteomic methodology described here shows great promise for the detection of human blood and even imaging of blood in bloodied fingermarks. The approach is sensitive, can differentiate between human blood and

  10. Determination of spatial distribution of melamine-cyanuric acid crystals in rat kidney tissue by histology and imaging matrix-assisted laser desorption/ionization quadrupole time-of-flight mass spectrometry.

    Science.gov (United States)

    Kim, Chae-Wook; Yun, Jun-Won; Bae, Il-Hong; Lee, Joon-Seok; Kang, Hyun-Jin; Joo, Kyung-Mi; Jeong, Hye-Jin; Chung, Jin-Ho; Park, Young-Ho; Lim, Kyung-Min

    2010-01-01

    After the outbreak of acute renal failure associated with melamine-contaminated pet food, many attempts have been made to uncover the mechanism underlying the renal toxicity caused by melamine and melamine-related compounds. Using rat models, we investigated the renal crystal formation following the ingestion of a melamine-cyanuric acid mixture (M+CA, 1:1) to gain insight into the M+CA-induced renal toxicity. M+CA did not induce toxicity in precision-cut kidney slices, suggesting that M+CA does not have a direct nephrotoxicity. On the contrary, oral administration of M+CA for 3 days induced nephrotoxicity as determined by increased serum blood urea nitrogen and creatinine, reduced creatinine clearance, and enlarged kidneys in the animals treated with 50 mg/kg M+CA (melamine, 25 mg/kg, and cyanuric acid, 25 mg/kg; 2 of 10 animals) and 100 mg/kg M+CA (9 of 9 animals). While urine crystals were found in all animals treated with M+CA (25-100 mg/kg), histological examination revealed that renal crystals could be observed only in the kidneys of animals showing signs of nephrotoxicity. Remarkably, at 50 mg/kg M+CA, crystals were observed mainly in the medulla region of the kidney, while at 100 mg/kg, crystals were disseminated throughout the cortex and medulla regions. To further investigate the crystal formation by M+CA, matrix-assisted laser desorption/ionization quadrupole time-of-flight (MALDI-Q-TOF) imaging mass spectrometry detecting melamine distribution through monitoring the product ion (m/z 85, M + H) from melamine (m/z 127, M + H) was developed to directly obtain the image of melamine distribution in the kidney. The distribution image of melamine in kidney tissue confirmed that dense points of melamine were located only in the medulla region at 50 mg/kg M+CA, while at 100 mg/kg, they were disseminated widely from the cortex to medulla. These results demonstrated that M+CA ingestion could lead to crystal formation in kidney tubules along the osmotic gradient and

  11. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry combined with multidimensional scaling, binary hierarchical cluster tree and selected diagnostic masses improves species identification of Neolithic keratin sequences from furs of the Tyrolean Iceman Oetzi.

    Science.gov (United States)

    Hollemeyer, Klaus; Altmeyer, Wolfgang; Heinzle, Elmar; Pitra, Christian

    2012-08-30

    The identification of fur origins from the 5300-year-old Tyrolean Iceman's accoutrement is not yet complete, although definite identification is essential for the socio-cultural context of his epoch. Neither have all potential samples been identified so far, nor there has a consensus been reached on the species identified using the classical methods. Archaeological hair often lacks analyzable hair scale patterns in microscopic analyses and polymer chain reaction (PCR)-based techniques are often inapplicable due to the lack of amplifiable ancient DNA. To overcome these drawbacks, a matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) method was used exclusively based on hair keratins. Thirteen fur specimens from his accoutrement were analyzed after tryptic digest of native hair. Peptide mass fingerprints (pmfs) from ancient samples and from reference species mostly occurring in the Alpine surroundings at his lifetime were compared to each other using multidimensional scaling and binary hierarchical cluster tree analysis. Both statistical methods highly reflect spectral similarities among pmfs as close zoological relationships. While multidimensional scaling was useful to discriminate specimens on the zoological order level, binary hierarchical cluster tree reached the family or subfamily level. Additionally, the presence and/or absence of order, family and/or species-specific diagnostic masses in their pmfs allowed the identification of mammals mostly down to single species level. Red deer was found in his shoe vamp, goat in the leggings, cattle in his shoe sole and at his quiver's closing flap as well as sheep and chamois in his coat. Canid species, like grey wolf, domestic dog or European red fox, were discovered in his leggings for the first time, but could not be differentiated to species level. This is widening the spectrum of processed fur-bearing species to at least one member of the Canidae family. His fur cap was

  12. Applicability of an in-House Saponin-Based Extraction Method in Bruker Biotyper Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry System for Identification of Bacterial and Fungal Species in Positively Flagged Blood Cultures

    Directory of Open Access Journals (Sweden)

    Jung-Yien Chien

    2016-09-01

    Full Text Available We used an in-house saponin-based extraction method to evaluate the performance of the Bruker Biotyper matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/MS system for the identification of bacteria and fungi in 405 positively flagged blood culture bottles. Results obtained from MALDI-TOF/MS were compared with those obtained using conventional phenotypic identification methods. Of the 405 positively flagged blood culture bottles, 365 showed monomicrobal growth and were correctly identified to the species (72.1% or genus (89.6% level using the Bruker Biotyper system. The remaining 40 positively flagged blood culture bottles showed polymicrobial growth. Of them, 82.5% (n=33 of the isolates were correctly identified to the species level and 92.5% (n=37 to the genus level using the Bruker Biotyper system. The overall accuracy of identification to the genus level in flagged blood cultures was 89.5% for Gram-positive organisms, 93.5% for Gram-negative pathogens and 71.9% for fungi. Confidence scores were 1.500 for 307 (75.8% bottles, 1.700 for 249 (61.5% bottles and 2.000 for 142 (35.1% bottles. None of the yeast cultures yielded scores 1.700. Using an identification-score cutoff of 1.500, the MALDI Biotyper correctly identified 99.2% of Gram-positive bacteria, 97.6% of Gram-negative bacteria and 100% of yeast isolates to the genus level and 77.6% of Gram-positive bacteria, 87.1% of Gram-negative bacteria and 100.0% of yeast isolates to the species level. The overall rate of identification using our protocol was 89.9% (364/405 for genus level identification and 73.1% (296/405 for species level identification. Yeast isolates yielded the lowest confidence scores, which compromised the accuracy of identification. Further optimization of the protein extraction procedure in positive blood cultures is needed to improve the rate of identification.

  13. Molecular and epidemiological analysis of nosocomial carbapenem-resistant Klebsiella spp. using repetitive extragenic palindromic-polymerase chain reaction and matrix-assisted laser desorption/ionization-time of flight.

    Science.gov (United States)

    Treviño, Mercedes; Navarro, Daniel; Barbeito, Gema; García-Riestra, Carlos; Crespo, Carlos; Regueiro, Benito J

    2011-09-01

    Infections with carbapenem-resistant enterobacteria are an emerging threat. This study reports the microbiologic, clinical, and epidemiologic features and the therapeutic outcomes of the infections caused by carbapenem- and pandrug-resistant Klebsiella emerged in our hospital. Fingerprinting analyses by automated repetitive extragenic palindromic-polymerase chain reaction (rep-PCR) and matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry are also compared. Carbapenem-resistant Klebsiella spp. affecting 13 patients were investigated using automated rep-PCR (DiversiLab System) and MALDI-TOF. Species identification was performed by Vitek 2 System and MALDI-TOF. Antimicrobial susceptibility testing was made using Vitek 2 System and Etest. Screening for extended spectrum beta-lactamase (ESBL) and carbapenemase production was made by double disk synergy and Hodge tests, respectively. Synergy studies were performed using Etest. DNA array was used for detection of KPC and ESBLs. bla(VIM-1) gene was amplified by PCR and sequencing. Use of carbapenems in the hospital was studied. A total of 13 patients were found to be colonized/infected with carbapenem-resistant Klebsiella. All patients were previously submitted to surgery and/or presented with severe underlying disease. After carbapenem-resistant Klebsiella isolation, the majority of the patients were treated with amikacin plus carbapenem, tigecycline, or fosfomycin. All Klebsiella isolates (n = 14), except two, had the bla(VIM-1) gene and all Klebsiella pneumoniae also had bla(SHV) gene associated with ESBL production. DiversiLab system showed higher discriminatory power than MALDI-TOF for strain typing. The risk of a rapid dissemination and the persistence of these multidrug-resistant strains through the time determine the need to implement routine procedures for metallo-beta-lactamase detection and measures for prevention of the spread of these microorganisms. The combined use of

  14. Comparison of matrix-assisted laser desorption ionization-time-of-flight mass spectrometry assay with conventional methods for detection of IMP-6, VIM-2, NDM-1, SIM-1, KPC-1, OXA-23, and OXA-51 carbapenemase-producing Acinetobacter spp., Pseudomonas aeruginosa, and Klebsiella pneumoniae.

    Science.gov (United States)

    Lee, Wonmok; Chung, Hae-Sun; Lee, Yangsoon; Yong, Dongeun; Jeong, Seok Hoon; Lee, Kyungwon; Chong, Yunsop

    2013-11-01

    Matrix-assisted laser desorption ionization-time-of-flight mass spectrometry assay was able to detect carbapenemase producers, including SIM-1 or OXA-51, within 4 hours using 20 μL of 0.5 g/L ertapenem solution as a substrate. This assay is more rapid and accurate than the modified Hodge test and 3-dimensional extract bioassay. Hence, it can be used an alternative test to identify carbapenemase-mediated carbapenem resistance in Gram-negative bacteria. © 2013.

  15. Photodissociation of Gaseous Ions Formed by Laser Desorption.

    Science.gov (United States)

    1986-09-20

    produced by separate pathways from the (M-I)- ion or from consecutive photodissociations. Hesperidin : In the negative ion LD mass spectrum of this compound...an ion of m/z r𔃼 was produced from the sodium salt of hesperidin phosphoric acid ester. This ion was observed to dissociate by loss of the attached...Experimental conditions are same as in the top spectrum. Figure 8. Top. Negative ions formed by laser desorption from Na-salt of hesperidin phosphoric acid ester

  16. Identification of Bacillus spp. colonizing the nasal mucosa of healthy adults living in the suburban area using the matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF MS system

    Directory of Open Access Journals (Sweden)

    Kosikowska Urszula

    2014-09-01

    Full Text Available Bacillus spp. can be regarded as a rare component of the nasal mucosa microflora. The aim of this study was to identify Bacillus spp. from the nasal mucosa of healthy adults living in the suburban area near Lublin using the matrix-assisted laser desorptionionization time-of-flight mass spectrometry (MALDI-TOF MS system.

  17. Laser desorption ionization and peptide sequencing on laser induced silicon microcolumn arrays

    Science.gov (United States)

    Vertes, Akos [Reston, VA; Chen, Yong [San Diego, CA

    2011-12-27

    The present invention provides a method of producing a laser-patterned silicon surface, especially silicon wafers for use in laser desorption ionization (LDI-MS) (including MALDI-MS and SELDI-MS), devices containing the same, and methods of testing samples employing the same. The surface is prepared by subjecting a silicon substrate to multiple laser shots from a high-power picosecond or femtosecond laser while in a processing environment, e.g., underwater, and generates a remarkable homogenous microcolumn array capable of providing an improved substrate for LDI-MS.

  18. Identification of Microalgae by Laser Desorption/Ionization Mass Spectrometry Coupled with Multiple Nanomatrices.

    Science.gov (United States)

    Peng, Lung-Hsiang; Unnikrishnan, Binesh; Shih, Chi-Yu; Hsiung, Tung-Ming; Chang, Jeng; Hsu, Pang-Hung; Chiu, Tai-Chia; Huang, Chih-Ching

    2016-04-01

    In this study, we demonstrate a simple method to identify microalgae by surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS) using three different substrates: HgSe, HgTe, and HgTeSe nanostructures. The fragmentation/ionization processes of complex molecules in algae varied according to the heat absorption and transfer efficiency of the nanostructured matrices (NMs). Therefore, the mass spectra obtained for microalgae showed different patterns of m/z values for different NMs. The spectra contained both significant and nonsignificant peaks. Constructing a Venn diagram with the significant peaks obtained for algae when using HgSe, HgTe, and HgTeSe NMs in m/z ratio range 100-1000, a unique relationship among the three sets of values was obtained. This unique relationship of sets is different for each species of microalgae. Therefore, by observing the particular relationship of sets, we successfully identified different algae such as Isochrysis galbana, Emiliania huxleyi, Thalassiosira weissflogii, Nannochloris sp., Skeletonema cf. costatum, and Tetraselmis chui. This simple and cost-effective SALDI-MS analysis method coupled with multi-nanomaterials as substrates may be extended to identify other microalgae and microorganisms in real samples. Graphical Abstract Identification of microalgae by surface-assisted laser desorption/ionization mass spectrometry coupled with three different mercury-based nanosubstrates.

  19. Laser Desorption Ionization Quadrupole Ion Trap Time-of-Flight Mass Spectrometry of Au m Fe n +/- Clusters Generated from Gold-Iron Nanoparticles and their Giant Nanoflowers. Electrochemical and/or Plasma Assisted Synthesis

    Science.gov (United States)

    Mawale, Ravi Madhukar; Ausekar, Mayuri Vilas; Pavliňák, David; Galmiz, Oleksandr; Kubáček, Pavel; Havel, Josef

    2017-02-01

    Gold nanoparticles (NP) with average diameter 100 nm synthesized from tetrachloroauric acid solution using stainless steel as a reducing agent were found to contain iron. Applying simultaneously high frequency (HF) plasma discharge in solution during the electrochemical reduction, giant gold-iron nanoflowers with average size 1000-5000 nm were formed. Scanning electron microscopy (SEM) shows the morphology of the nanopowders produced as polygonal yet nearly spherical, whereas iron content in both products determined by energy dispersive X-ray analysis (EDX) was found to be at 2.5 at. %. Laser desorption ionization (LDI) of both nanomaterials and mass spectrometric analysis show the formation of Au m Fe n +/- ( m = 1-35; n = 1-3) clusters. Structure of few selected clusters in neutral or monocharged forms were computed by density functional theory (DFT) calculations and it was found that typical distances of an iron nucleus from adjacent gold nuclei lie in the interval 2.5 to 2.7 Å. Synthetized Au-Fe nanoparticles were found stable for at least 2 mo at room temperature (even in aqueous solution) without any stabilizing agent. Produced Au-Fe nanoparticles in combination with standard MALDI matrices enhance ionization of peptides and might find use in nanomedicine.

  20. Coupling of Phosphate-Imprinted Mesoporous Silica Nanoparticles-Based Selective Enrichment with Matrix-Assisted Laser Desorption Ionization-Time-of-Flight Mass Spectrometry for Highly Efficient Analysis of Protein Phosphorylation.

    Science.gov (United States)

    Chen, Yang; Li, Daojin; Bie, Zijun; He, Xinpei; Liu, Zhen

    2016-01-19

    Protein phosphorylation is a major post-translational modification and represents a ubiquitous mechanism for the cellular signaling of many different biological processes. Selective enrichment of phosphopeptides from the complex biological samples is a key step for the mass spectrometric (MS) analysis of protein phosphorylation. Herein, we present phosphate-imprinted mesoporous silica nanoparticles (MSNs) as an ideal sorbent for selective enrichment of phosphopeptides and an off-line combination with matrix-asisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) for highly efficient analysis of protein phosphorylation. The phosphate-imprinted MSNs were prepared according to a newly reported strategy called dual-template docking oriented molecular imprinting (DTD-OMI). The prepared molecularly imprinted mesoporous material exhibited several significant merits, such as excellent selectivity toward phosphopeptides, tolerance to interference, fast binding equilibrium, and large binding capacity, which made the molecularly imprinted mesoporous material an ideal sorbent for selective enrichment of phosphopeptides. Using β-casein as a representative phosphoprotein, highly efficient phosphorylation analysis by the off-line platform was verified. Phosphorylation analysis of a nonfat milk sample was also well demonstrated. Because of their highly desirable properties, the phosphate-imprinted MSNs could find more applications in the analysis of protein phosphorylation.

  1. Electronically driven adsorbate excitation mechanism in femtosecond-pulse laser desorption

    DEFF Research Database (Denmark)

    Brandbyge, Mads; Hedegård, Per; Heinz, T. F.

    1995-01-01

    Femtosecond-pulse laser desorption is a process in which desorption is driven by a subpicosecond temperature pulse of order 5000 K in the substrate-adsorbate electron system, whose energy is transferred into the adsorbate center-of-mass degrees of freedom by a direct coupling mechanism. We presen...

  2. Ambient femtosecond laser vaporization and nanosecond laser desorption electrospray ionization mass spectrometry.

    Science.gov (United States)

    Flanigan, Paul; Levis, Robert

    2014-01-01

    Recent investigations of ambient laser-based transfer of molecules into the gas phase for subsequent mass spectral analysis have undergone a renaissance resulting from the separation of vaporization and ionization events. Here, we seek to provide a snapshot of recent femtosecond (fs) duration laser vaporization and nanosecond (ns) duration laser desorption electrospray ionization mass spectrometry experiments. The former employs pulse durations of vaporization with little or no fragmentation. When coupled to electrospray ionization, femtosecond laser vaporization provides a universal, rapid mass spectral analysis method requiring no sample workup. Remarkably, laser pulses with intensities exceeding 10(13) W cm(-2) desorb intact macromolecules, such as proteins, and even preserve the condensed phase of folded or unfolded protein structures according to the mass spectral charge state distribution, as demonstrated for cytochrome c and lysozyme. Because of the ability to vaporize and ionize multiple components from complex mixtures for subsequent analysis, near perfect classification of explosive formulations, plant tissue phenotypes, and even the identity of the manufacturer of smokeless powders can be determined by multivariate statistics. We also review the more mature field of nanosecond laser desorption for ambient mass spectrometry, covering the wide range of systems analyzed, the need for resonant absorption, and the spatial imaging of complex systems like tissue samples.

  3. Site Specificity in Femtosecond Laser Desorption of Neutral H Atoms from Graphite(0001)

    DEFF Research Database (Denmark)

    Frigge, R.; Hoger, T.; Siemer, B.

    2010-01-01

    Femtosecond laser excitation and density functional theory reveal site and vibrational state specificity in neutral atomic hydrogen desorption from graphite induced by multiple electronic transitions. Multimodal velocity distributions witness the participation of ortho and para pair states of che...

  4. Ambient Femtosecond Laser Vaporization and Nanosecond Laser Desorption Electrospray Ionization Mass Spectrometry

    Science.gov (United States)

    Flanigan, Paul; Levis, Robert

    2014-06-01

    Recent investigations of ambient laser-based transfer of molecules into the gas phase for subsequent mass spectral analysis have undergone a renaissance resulting from the separation of vaporization and ionization events. Here, we seek to provide a snapshot of recent femtosecond (fs) duration laser vaporization and nanosecond (ns) duration laser desorption electrospray ionization mass spectrometry experiments. The former employs pulse durations of proteins, and even preserve the condensed phase of folded or unfolded protein structures according to the mass spectral charge state distribution, as demonstrated for cytochrome c and lysozyme. Because of the ability to vaporize and ionize multiple components from complex mixtures for subsequent analysis, near perfect classification of explosive formulations, plant tissue phenotypes, and even the identity of the manufacturer of smokeless powders can be determined by multivariate statistics. We also review the more mature field of nanosecond laser desorption for ambient mass spectrometry, covering the wide range of systems analyzed, the need for resonant absorption, and the spatial imaging of complex systems like tissue samples.

  5. Airborne laser-spark for ambient desorption/ionisation.

    Science.gov (United States)

    Bierstedt, Andreas; Riedel, Jens

    A novel direct sampling ionisation scheme for ambient mass spectrometry is presented. Desorption and ionisation are achieved by a quasi-continuous laser induced plasma in air. Since there are no solid or liquid electrodes involved the ion source does not suffer from chemical interferences or fatigue originating from erosive burning or from electrode consumption. The overall plasma maintains electro-neutrality, minimising charge effects and accompanying long term drift of the charged particles trajectories. In the airborne plasma approach the ambient air not only serves as the plasma medium but at the same time also slows down the nascent ions via collisional cooling. Ionisation of the analyte molecules does not occur in the plasma itself but is induced by interaction with nascent ionic fragments, electrons and/or far ultraviolet photons in the plasma vicinity. At each individual air-spark an audible shockwave is formed, providing new reactive species, which expands concentrically and, thus, prevents direct contact of the analyte with the hot region inside the plasma itself. As a consequence the interaction volume between plasma and analyte does not exceed the threshold temperature for thermal dissociation or fragmentation. Experimentally this indirect ionisation scheme is demonstrated to be widely unspecific to the chemical nature of the analyte and to hardly result in any fragmentation of the studied molecules. A vast ensemble of different test analytes including polar and non-polar hydrocarbons, sugars, low mass active ingredients of pharmaceuticals as well as natural biomolecules in food samples directly out of their complex matrices could be shown to yield easily accessible yet meaningful spectra. Since the plasma medium is humid air, the chemical reaction mechanism of the ionisation is likely to be similar to other ambient ionisation techniques. Wir stellen hier eine neue Ionisationsmethode für die Umgebungsionisation (ambient ionisation) vor. Sowohl die

  6. Laser Assisted Microsurgical Anastomosis.

    Science.gov (United States)

    1983-09-22

    our axoral transport platophysiological and ultravascular findings indicat to be inferior ll. to laser epineurial repair, conventional microsurgical...ventral motoneurons . Experimental Neurology, 21:41-51, 1968. 21. Marinacci, AA: Diagnosis of "all median hand". 22. Marinacci, AA: The problem of unusual

  7. Laser assisted graffiti paints removing

    Science.gov (United States)

    Novikov, B. Y.; Chikalev, Y. V.; Shakhno, E. A.

    2011-02-01

    It's hard to imagine a modern city view without some drawings and inscriptions, usually called "graffiti". Traditional cleaning methods do not suit modern requirements. Investigation of possibilities of laser assisted paints removing is described in this article. The conditions for removing different paints from different surfaces were defined.

  8. Laser desorption/ionization mass spectrometry for direct profiling and imaging of small molecules from raw biological materials

    Energy Technology Data Exchange (ETDEWEB)

    Cha, Sangwon [Iowa State Univ., Ames, IA (United States)

    2008-01-01

    Matrix-assisted laser desorption/ionization(MALDI) mass spectrometry(MS) has been widely used for analysis of biological molecules, especially macromolecules such as proteins. However, MALDI MS has a problem in small molecule (less than 1 kDa) analysis because of the signal saturation by organic matrixes in the low mass region. In imaging MS (IMS), inhomogeneous surface formation due to the co-crystallization process by organic MALDI matrixes limits the spatial resolution of the mass spectral image. Therefore, to make laser desorption/ionization (LDI) MS more suitable for mass spectral profiling and imaging of small molecules directly from raw biological tissues, LDI MS protocols with various alternative assisting materials were developed and applied to many biological systems of interest. Colloidal graphite was used as a matrix for IMS of small molecules for the first time and methodologies for analyses of small metabolites in rat brain tissues, fruits, and plant tissues were developed. With rat brain tissues, the signal enhancement for cerebroside species by colloidal graphite was observed and images of cerebrosides were successfully generated by IMS. In addition, separation of isobaric lipid ions was performed by imaging tandem MS. Directly from Arabidopsis flowers, flavonoids were successfully profiled and heterogeneous distribution of flavonoids in petals was observed for the first time by graphite-assisted LDI(GALDI) IMS.

  9. A revisit of high collision energy effects on collision-induced dissociation spectra using matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry (MALDI-LIFT-TOF/TOF): application to the sequencing of RNA/DNA chimeras.

    Science.gov (United States)

    Mauger, Florence; Tabet, Jean-Claude; Gut, Ivo G

    2014-07-15

    High-energy collision-induced dissociation (CID) spectra of isomeric RNA/DNA chimeras using matrix-assisted laser desorption/ionization time-of-flight LIFT mass spectrometry (MALDI-LIFT-TOF/TOF) can potentially be applied for an exhaustive fragment characterization in a nucleic acid sequencing scheme. These chimeras contain deoxynucleotides and at the 3'-end a ribonucleotide with a 3'-phosphate group. Deprotonated RNA/DNA chimeras of 4-, 5-, 7- and 10-mers are analyzed by CID. This enhances consecutive dissociations from both the precursor and prompt product anions generated by MALDI and metastable fragmentations prior to entering the LIFT cell. Gas-phase fragmentations of 4- and 5-mers produced many fragment ions, from base release prior to consecutive cleavage of the nucleotide phosphate bond linkage phosphate. The unusual a4(-) product ion is a specific and diagnostic dissociation of the 4-mer if the ribonucleotide contains cytosine. As the size of RNA/DNA chimeras increase, several abundant product ions are generated mainly from zwitterionic forms (deprotonated phosphate ester and protonated base sites): [(M-H)-BiH](-), [ai-BiH](-), wj(-), [wj, (ai-BiH)](-) (if Bi  ≠ T) as internal product ion, and more rarely [wj-BiH](-). The absence of the majority of the [ai-BiH](-) series although the wj (-) series suggested that the higher critical energy processes with a loose transition state are favored yielding the wj(-) series. A large number of abundant fragment ions are detected which enable each isomer to be sequenced. This sequencing method is high-throughput, accurate and could be used to sequence isomers of up to 10-mers and also oligonucleotides of unknown sequence. However, RNA/DNA chimeras without thymine must be sufficiently concentrated to reach desorption of deprotonated molecular species to be selected in LIFT to produce all fragment ions within measurable abundances. Copyright © 2014 John Wiley & Sons, Ltd.

  10. High fluence deposition of polyethylene glycol films at 1064 nm by matrix assisted pulsed laser evaporation (MAPLE)

    DEFF Research Database (Denmark)

    Purice, Andreea; Schou, Jørgen; Kingshott, P.

    2007-01-01

    microbalance. The laser fluence needed to produce PEG films turned out to be unexpectedly high with a threshold of 9 J/cm(2) and the deposition rate was much lower than that with laser light at 355 nm. Results from matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF...

  11. Laser Infrared Desorption Spectroscopy to Detect Complex Organic Molecules on Icy Planetary Surfaces

    Science.gov (United States)

    Sollit, Luke S.; Beegle, Luther W.

    2008-01-01

    Laser Desorption-Infrared Spectroscopy (LD-IR) uses an IR laser pulse to desorb surface materials while a spectrometer measures the emission spectrum of the desorbed materials (Figure 1). In this example, laser desorption operates by having the incident laser energy absorbed by near surface material (10 microns in depth). This desorption produces a plume that exists in an excited state at elevated temperatures. A natural analog for this phenomenon can be observed when comets approach the sun and become active and individual molecular emission spectra can be observed in the IR [1,2,3,4,5]. When this occurs in comets, the same species that initially emit radiation down to the ground state are free to absorb it, reducing the amount of d