Daiki Asakawa

University of Liège, Liège, WAL, Belgium

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Publications (41)119.84 Total impact

  • [show abstract] [hide abstract]
    ABSTRACT: Matrix-assisted laser desorption/ionization in-source decay produces highly informative fragments for the sequencing of peptides/proteins. Among amino acids, cysteine and proline residues were found to specifically influence the fragment yield. As they are both frequently found in small peptide structures for which de novo sequencing is mandatory, the understanding of their specific behaviors would allow useful fragmentation rules to be established. In the case of cysteine, a c•/w fragment pair originating from Xxx-Cys is formed by side-chain loss from the cysteine residue. The presence of a proline residue contributes to an increased yield of ISD fragments originating from N-Cα bond cleavage at Xxx1-Xxx2Pro, which is attributable to the cyclic structure of the proline residue. Our results suggest that the aminoketyl radical formed by MALDI-ISD generally induces the homolytic N-Cα bond cleavage located on the C-terminal side of the radical site. In contrast, N-Cα bond cleavage at Xxx-Pro produces no fragments and the N-Cα bond at the Xxx1-Xxx2Pro bond is alternatively cleaved via a heterolytic cleavage pathway.
    Journal of the American Society for Mass Spectrometry 04/2014; · 3.59 Impact Factor
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    ABSTRACT: Matrix-assisted laser desorption/ionization in-source decay (MALDI-ISD) is a useful method for top-down sequencing of proteins and preferentially produces the c'/z• fragment pair. Subsequently, radical z• fragments undergo a variety of radical reactions. This article focuses on the chemical properties of the 1,5-diaminonaphthalene (1,5-DAN) adduct on z fragment ions (zn*), which are abundant ions in MALDI-ISD spectra. Post-source decay (PSD) of the zn* fragments resulted in specific peptide bond cleavage adjacent to the binding site of 1,5-DAN, leading to the preferential formation of y'(n-1) fragments. The dominant loss of an amino acid with 1,5-DAN from zn* can be used in pseudo-MS3 mode to identify the C-terminal side fragments from a complex MALDI-ISD spectrum or to determine missed cleavage residues using MALDI-ISD. Although the N-Cα bond at the N-terminal side of Pro is not cleaved by MALDI-ISD, pseudo-MS3 via zn* can confirm the presence of a Pro residue.
    Analytical Chemistry 02/2014; · 5.70 Impact Factor
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    ABSTRACT: The type of ions detected after in-source decay (ISD) in a MALDI source differs according to the ion source pressure and on the mass analyzer used. We present the mechanism leading to the final ISD ions for a Fourier transform-ion cyclotron resonance mass spectrometer (FTICR MS). The MALDI ion source was operated at intermediate pressure to cool the resulting ions and increase their lifetime during the long residence times in the FTICR ion optics. This condition produces not only c', z' and w fragments, but also a, y' and d fragments. In particular, d ions help to identify isobaric amino acid residues present near the N-terminal amino acid. Desorbed ions collide with background gas during desorption, leading to proton mobilization from Arg residues to a less favored protonation site. As a result, in the case of ISD with MALDI FTICR, the influence of the Arg residue in ISD fragmentation is less straightforward than for TOF MS and the sequence coverage is thus improved. MALDI-ISD combined with FTICR MS appears to be a useful method for sequencing of peptides and proteins including discrimination of isobaric amino acid residues and site determination of phosphorylation. Additionally we also used new software for in silico elimination of MALDI matrix peaks from MALDI-ISD FTICR mass spectra. The combination of high resolving power of an FTICR analyzer and matrix subtraction software helps to interpret the low m/z region of MALDI-ISD spectra. Finally, several of these developed methods are applied in unison towards a MALDI ISD FTICR imaging experiment on mouse brain to achieve better results.
    Analytical Chemistry 07/2013; · 5.70 Impact Factor
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    ABSTRACT: Matrix-assisted laser desorption/ionization in-source decay (MALDI-ISD) is initiated by hydrogen transfer from matrix molecules to the carbonyl oxygen of peptide backbone with subsequent radical-induced cleavage leading to c'/z• fragments pair. MALDI-ISD is a very powerful method to obtain long sequence tags from proteins or to do de novo sequencing of peptides. Besides classical fragmentation, MALDI-ISD also shows specific fragments for which the mechanism of formation enlightened the MALDI-ISD process. In this study, the MALDI-ISD mechanism is reviewed, and a specific mechanism is studied in details: the N-terminal side of Cys residue (Xxx-Cys) is described to promote the generation of c' and w fragments in MALDI-ISD. Our data suggest that for sequences containing Xxx-Cys motifs, the N-Cα bond cleavage occurs following the hydrogen attachment to the thiol group of Cys side-chain. The c•/w fragments pair is formed by side-chain loss of the Cys residue with subsequent radical-induced cleavage at the N-Cα bond located at the left side (N-terminal direction) of the Cys residue. This fragmentation pathway preferentially occurs at free Cys residue and is suppressed when the cysteines are involved in disulfide bonds. Hydrogen attachment to alkylated Cys residues using iodoacetamide gives free Cys residue by the loss of •CH2 CONH2 radical. The presence of alkylated Cys residue also suppress the formation of c•/w fragments pair via the (Cβ )-centered radical, whereas w fragment is still observed as intense signal. In this case, the z• fragment formed by hydrogen attachment of carbonyl oxygen followed side-chain loss at alkylated Cys leads to a w fragment. Hydrogen attachment on peptide backbone and side-chain of Cys residue occurs therefore competitively during MALDI-ISD process. Copyright © 2013 John Wiley & Sons, Ltd.
    Biological Mass Spectrometry 03/2013; 48(3):352-360. · 3.41 Impact Factor
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    ABSTRACT: MALDI in-source decay (ISD) was first introduced in 1995 in a landmark paper by RS Brown and JJ Lenon (Anal. Chem. 1995, 67, 3990-3999). Although MALDI MS is considered a soft ionization technique which enables the analysis of high molecular weight proteins exceeding several hundred kDa in an intact manner, it became apparent that some analyte fragmentation occurs on a very short time scale during the desorption ionization process. ISD is the combination of laser induced fragmentation and rapidly occurring metastable decay. This phenomenon can be used to obtain primary sequence information on peptides and (small) proteins also referred as top down sequencing. Beyond its analytical utility, the ISD fragmentation mechanism is currently being studied by several groups. In this perspective special feature article, the MALDI-ISD mechanism is reviewed and a specific fragmentation mechanism is introduced and studied in details: the N-terminal side of Cys residue is described to promote the generation of c' and w ISD fragments. Dr Edwin De Pauw is Professor of Chemistry at Liege University in Belgium. One of the objectives of his mass spectrometry laboratory is to contribute to the development of new fundamental concepts and to enlighten their applications in the fields of analytical chemistry.
    Biological Mass Spectrometry 03/2013; 48(3):i. · 3.41 Impact Factor
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    ABSTRACT: The early mechanisms of matrix-assisted laser desorption/ionization in-source decay (MALDI-ISD) is described herein. MALDI-ISD is initiated by the hydrogen transfer from excited matrix molecules to the carbonyl oxygen of peptide backbone which is followed by a radical-induced cleavage producing c'/z• fragment pair. As expected, the use of 2,5-DHB or 1,5-DAN was efficient to induce MALDI-ISD, and the strongest intensity of MALDI-ISD fragments was observed when laser shots are performed on matrix crystals. In contrast, the hydrogen radical transfer reaction was suppressed by using ionic liquid and amorphous structure of 2,5-DHB and 1,5-DAN mixture as a matrix. Our results suggest that the hydrogen transfer occurs on the matrix crystal during the dissipation of the laser energy and before desorption, following ISD fragments formed in the MALDI plume.
    The Journal of Physical Chemistry B 01/2013; · 3.61 Impact Factor
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    ABSTRACT: MALDI mass spectrometry imaging (MALDI MSI) is a rapidly growing method in biomedical research allowing molecular mapping of proteins on histological sections. The images can be analyzed in terms of spectral pattern to define regions of interest. However, the identification and the differential quantitative analysis of proteins require off line or in situ proteomic methods using enzymatic digestion. The rapid identification of biomarkers holds great promise for diagnostic research but the major obstacle is the absence of rapid and direct method to detect and identify with a sufficient dynamic range a set of specific biomarkers. In the current work, we present a proof of concept for a method allowing identifying simultaneously a set of selected biomarkers on histological slices with minimal sample treatment using in-source decay (ISD) MSI and MALDI-Fourier transform ion cyclotron resonance (FTICR). In the proposed method, known biomarkers are spotted next to the tissue of interest, the whole MALDI plate being coated with 1,5-DAN matrix. The latter enhances MALDI radical-induced ISD, providing large tags of the amino acid sequences. Comparative analysis of ISD fragments between the reference spots and the specimen in imaging mode allows for unambiguous identification of the selected biomarker while preserving full spatial resolution. Moreover, the high resolution/high mass accuracy provided by FTICR mass spectrometry allows the identification of proteins. Well-resolved peaks and precise measurements of masses and mass differences allow the construction of reliable sequence tags for proteins identification. The method will allow the use MALDI-FTICR MSI as method for rapid targeted biomarker detection in complement to classical histology.
    Analytical Chemistry 01/2013; · 5.70 Impact Factor
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    ABSTRACT: MALDI in-source decay (ISD) has been used for top-down sequencing of proteins. The use of the matrix 1,5-diaminonapthalene (1,5-DAN) gave abundant w ions, which are formed from the unimolecular dissociation of z• radical fragments via α cleavage reaction and thus help identify which of the isobaric amino acids, Leu or Ile, is present. The high abundance of w ions in MALDI-ISD with 1,5-DAN results from the low collision rate in the MALDI plume. MALDI-ISD with 1,5-DAN appears to be an useful method for the top-down sequencing of proteins, including discrimination of Leu and Ile near the C-terminal end.
    Journal of the American Society for Mass Spectrometry 01/2013; · 3.59 Impact Factor
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    ABSTRACT: Matrix-assisted laser desorption/ionization (MALDI) is now a mature method allowing the identification and, more challenging, the quantification of biopolymers (proteins, nucleic acids, glycans, etc). MALDI spectra show mostly intact singly charged ions. To obtain fragments, the activation of singly charged precursors is necessary, but not efficient above 3.5 kDa, thus making MALDI MS/MS difficult for large species. In-source decay (ISD) is a prompt fragmentation reaction that can be induced thermally or by radicals. As fragments are formed in the source, precursor ions cannot be selected; however, the technique is not limited by the mass of the analyzed compounds and pseudo MS3 can be performed on intense fragments. The discovery of new matrices that enhance the ISD yield, combined with the high sensitivity of MALDI mass spectrometers, and software development, opens new perspectives. We first review the mechanisms involved in the ISD processes, then discuss ISD applications like top-down sequencing and post-translational modifications (PTMs) studies, and finally review MALDI-ISD tissue imaging applications.
    Topics in current chemistry 09/2012; · 8.46 Impact Factor
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    ABSTRACT: The use of specific matrixes allows enhancing the scope of in-source decay (ISD) applications in matrix-assisted laser desorption/ionization (MALDI) thanks to the specificity of analyte-matrix chemistry. The use of an oxidizing matrix, 5-nitrosalicylic acid (5-NSA), for MALDI-ISD of glycans is shown to promote fragmentation pathways involving radical precursors. Both glycosidic and cross-ring cleavages are promoted by hydrogen abstraction from hydroxyl group of glycans by 5-NSA molecules. Cross-ring cleavage ions are potentially useful in linkage analysis, one of the most critical steps of glycan characterization. Moreover, we show here that isobaric glycans could be distinguished by structure specific ISD ions and that the molar ratio of glycan isomers in the mixture can be estimated from their fragment ions abundance. The use of 5-NSA also opens the possibility to perform pseudo-MS(3) analysis of glycans. Therefore, MALDI-ISD with 5-NSA is a useful method for identification of glycans and semiquantitative analysis of mixture of glycan isomers.
    Analytical Chemistry 08/2012; 84(17):7463-8. · 5.70 Impact Factor
  • Daiki Asakawa, Mitsuo Takayama
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    ABSTRACT: The mechanism of in-source decay (ISD) in matrix-assisted laser desorption/ionization (MALDI) has been described. The MALDI-ISD with an oxidizing matrix is initiated by hydrogen abstraction from peptides to matrix molecules, leading to hydrogen-deficient peptide radicals. Subsequently, the C(α)-C and C(α)-H bonds are cleaved, forming the a•/x fragment pair and [M-2H], respectively. Those reactions competitively occur during MALDI-ISD processes. Our results suggest that the C(α)-H bond cleavage to form [M-2H] was induced by collisions between hydrogen-deficient peptide radicals and matrix molecules in the MALDI plume. In contrast, the C(α)-C bond cleavages occur via a unimolecular dissociation process and independently of the collision rate in the MALDI plume. The formation mechanism of the a-, b-, and d-series fragments are also described. We report 2,5-bis(2-hydroxyethoxy)-7,7,8,8-tetracyanoquinodimethane (bisHE-TCNQ), being known as an organic semiconductor and an electron acceptor, as a novel suitable matrix for the MALDI-ISD of peptides via hydrogen abstraction.
    The Journal of Physical Chemistry B 02/2012; 116(13):4016-23. · 3.61 Impact Factor
  • Daiki Asakawa, Mitsuo Takayama
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    ABSTRACT: Matrix-assisted laser desorption/ionization in-source decay (MALDI-ISD) has been used for characterization of a phosphorylated peptides and proteins because labile phosphate group is not lost during the MALDI-ISD process. The conventional MALDI-ISD is initiated by the hydrogen transfer from reducing matrix molecules to peptide backbone, leading to c'- and z'-series ions. In contrast, when an oxidizing chemical 5-nitrosalicylic acid (5-NSA) is served as the MALDI-ISD matrix, a- and x-series ions are specifically generated by hydrogen abstraction from peptide backbone to matrix molecule. The 5-NSA provides useful complementary information to the conventional MALDI-ISD for the analysis of amino acid sequencing and site localization of phosphorylation in peptides. The MALDI-ISD with reducing and oxidizing matrix could be a useful method for the de novo peptide sequencing.
    Biological Mass Spectrometry 02/2012; 47(2):180-7. · 3.41 Impact Factor
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    ABSTRACT: MALDI-ISD of peptides were studied using several salicylic acid derivatives, 2,5-dihydroxybenzoic acid (2,5-DHB), 5-aminosalicylic acid (5-ASA), 5-formylsalicylic acid (5-FSA), and 5-nitrosalicylic acid (5-NSA) as matrices. The difference in the nature of the functional group at the 5-position in the salicylic acid derivatives can dramatically affect the ISD products. The use of 2,5-DHB and 5-ASA leads to "hydrogen-abundant" peptide radicals and subsequent radical-induced N-Cα bonds cleavage. N-Cα bond cleavage gave a c'/z (·) fragment pair and radical z (·)-series fragments gain a hydrogen radical or react with a matrix radical. In contrast, the use of 5-NSA resulted in the production of a "hydrogen-deficient" peptide radical that contained a radical site on the amide nitrogen in the peptide backbone. Subsequently, the radical site on the amide nitrogen induces Cα-C bond dissociation, leading to a (·)/x fragment pair. The a (·)-series ions undergo further hydrogen abstraction to form a-series ions after Cα-C bond cleavage. Since the Pro residue does not contain a nitrogen-centered radical site, Cα-C bond cleavage does not occur. Alternatively, the specific cleavage of CO-N bonds leads to a b (·)/y fragment pair at Xxx-Pro which occurs via hydrogen abstraction from the Cα-H in the Pro residue. The use of 5-FSA generated both a (·)/x- and c'/z (·)-series fragment pairs. An oxidizing matrix provides useful complementary information in MALDI-ISD compared to a reducing matrix for the analysis of amino acid sequencing and site localization in cases of phosphopeptides. MALDI-ISD, when used in conjunction with both reducing and oxidizing matrices is a potentially useful method for de novo peptide sequencing.
    Mass spectrometry (Tokyo, Japan). 01/2012; 1(1):A0002.
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    ABSTRACT: The influence of arginine (Arg), lysine (Lys), and phenylalanine (Phe) residues and phosphorylation on the molecular ion yields of model peptides have been quantitatively studied using matrix-assisted laser desorption/ionization (MALDI) mass spectrometry in both positive- and negative-ion mode. The results obtained from these experiments have been interpreted from the standpoint of two different components, namely, desorption and ionization, on the basis of the physicochemical properties of constituent amino acids of the model peptides. The presence of basic residues such as Arg and Lys enhanced the ion yields of protonated molecules [M + H](+). An N-terminal rather than a C-terminal Arg residue was advantageous for the formation of both [M + H](+) and [M - H](-). The presence of the Phe residue resulted in the increase of the ion yields of both [M + H](+) and [M - H](-). In contrast, the presence of phosphate group(s) contributed to the suppression of the yields of both [M + H](+) and [M - H](-) due to the loss of phosphate group. The detection limits for both [M + H](+) and [M - H](-) of model peptides have been evaluated.
    Journal of the American Society for Mass Spectrometry 11/2011; 23(1):108-15. · 3.59 Impact Factor
  • Daiki Asakawa, Mitsuo Takayama
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    ABSTRACT: The use of 5-nitrosalicylic acid (5-NSA) as a matrix for in-source decay (ISD) of peptides during matrix-assisted laser desorption/ionization (MALDI) is described herein. Mechanistically, the decay process is initiated by a hydrogen abstraction from a peptide backbone amide nitrogen by 5-NSA. Hydrogen abstraction results in formation of an oxidized peptide containing a radical amide nitrogen. Subsequently, the C(α)-C bond N-terminal to the peptide bond is cleaved to form an a·/x fragment pair. The C(α)-C bonds C-terminal to Gly residues were less susceptible to cleavage than were those of other residues. C(α)-C bonds N-terminal to Pro and Sar residues were not cleaved by the aforementioned mechanism; instead, after hydrogen abstraction from a Pro or Sar C(α)-H bond, the peptide bond N-terminal to the Pro was cleaved yielding b- and y-series ions. We also show that fragments produced by MALDI 5-NSA-induced ISD were formed independently of the ionization process.
    Rapid Communications in Mass Spectrometry 09/2011; 25(17):2379-83. · 2.51 Impact Factor
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    Daiki Asakawa, Mitsuo Takayama
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    ABSTRACT: The use of 5-formylsalicylic acid (5-FSA) and 5-nitrosalicylic acid (5-NSA) as novel matrices for in-source decay (ISD) of peptides in matrix-assisted laser desorption/ionization (MALDI) is described. The use of 5-FSA and 5-NSA generated a- and x-series ions accompanied by oxidized peptides [M - 2 H + H](+). The preferential formation of a- and x-series ions was found to be dependent on the hydrogen-accepting ability of matrix. The hydrogen-accepting ability estimated from the ratio of signal intensity of oxidized product [M - 2 H + H](+) to that of non-oxidized protonated molecule [M + H](+) of peptide was of the order 5-NSA > 5-FSA > 5-aminosalicylic acid (5-ASA) ≒ 2,5-dihydroxyl benzoic acid (2,5-DHB) ≒ 0. The results suggest that the hydrogen transfer reaction from peptide to 5-FSA and 5-NSA occurs during the MALDI-ISD processes. The hydrogen abstraction from peptides results in the formation of oxidized peptides containing a radical site on the amide nitrogen with subsequent radical-induced cleavage at the Cα-C bond, leading to the formation of a- and x-series ions. The most significant feature of MALDI-ISD with 5-FSA and 5-NSA is the specific cleavage of the Cα-C bond of the peptide backbone without degradation of side-chain and post-translational modifications (PTM). The matrix provides a useful complementary method to conventional MALDI-ISD for amino acid sequencing and site localization of PTMs in peptides.
    Journal of the American Society for Mass Spectrometry 07/2011; 22(7):1224-33. · 3.59 Impact Factor
  • Daiki Asakawa, Kenzo Hiraoka
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    ABSTRACT: Electrospray droplet impact (EDI) secondary ion mass spectrometry (SIMS) is a desorption/ionization technique for mass spectrometry in which highly charged water clusters produced from an atmospheric-pressure electrospray are accelerated in vacuum by several kV and impact on the sample deposited on the metal substrate. The abundances of the secondary ions for C(60) and amino acids are measured as a function of the acceleration voltage of the primary charged water droplets. Two desorption/ionization mechanisms are suggested in the EDI ionization processes: low-energy and high-energy regimes. In the low-energy regime, the excess charges in the primary droplets play a role in the formation of secondary ions. In the high-energy regime, samples are ionized by the supersonic collision of the primary droplets with the sample. The yield of secondary ions increases by about three orders of magnitude with increase in the acceleration voltage of the primary droplets from 1.75 kV to 10 kV.
    Rapid Communications in Mass Spectrometry 03/2011; 25(5):655-60. · 2.51 Impact Factor
  • Daiki Asakawa, Kenzo Hiraoka
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    ABSTRACT: Electrospray droplet impact (EDI)/SIMS is a new desorption/ionization technique for mass spectrometry. EDI/SIMS utilizes large multiply charged water clusters produced by atmospheric pressure electrospray as primary projectiles. It was found to afford extremely soft desorption/ionization compared with conventional SIMS, and has been used for detection of peptides and proteins. In this study, EDI/SIMS was applied to the detection of peptide in a highly concentrated NaCl solution. The persistent appearance of peptide ions for 1 ppm peptides in NaCl is probably because of the segregation of peptides on the crystallized salts. The samples dried under vacuum gave better EDI/SIMS mass spectra than those under ambient atmospheric pressure. Copyright © 2011 John Wiley & Sons, Ltd.
    Surface and Interface Analysis 01/2011; 43(10):1341 - 1345. · 1.22 Impact Factor
  • Daiki Asakawa, Kenzo Hiraoka
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    ABSTRACT: Electrospray droplet impact (EDI)/secondary ion mass spectrometry (SIMS) is a new desorption/ionization technique for mass spectrometry in which highly charged water clusters produced from atmospheric-pressure electrospray are accelerated in vacuum by several kV and impact on the sample deposited on the metal substrate. In this study, we applied EDI/SIMS directly to fruits, such as bananas, strawberries, grapes and apples. The major components in the fruits--fructose, glucose, sucrose and organic acids--could be observed with strong signal intensities. EDI/SIMS was also applied to the analysis of different regions of strawberries and apples. Compared with matrix-assisted laser desorption/ionization (MALDI), ion signals with lower background signals could be obtained, particularly for the low molecular weight analytes.
    Rapid Communications in Mass Spectrometry 08/2010; 24(16):2431-8. · 2.51 Impact Factor
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    ABSTRACT: Electrospray droplet impact (EDI)/secondary ion mass spectrometry (SIMS) is a new desorption/ionization technique for mass spectrometry in which highly charged water clusters produced from the atmospheric-pressure electrospray are accelerated in vacuum by 10 kV and impact the sample deposited on the metal substrate. EDI/SIMS was shown to enhance intact molecular ion formation dramatically compared to conventional SIMS. EDI/SIMS has been successfully applied to the analysis of mouse brain without any sample preparation. Five types of lipids, i.e. phosphatidylcholine (PC), phosphatidylserine, phosphatidylinositol (PI), galactocerebroside (GC) and sulfatide (ST), were readily detected from mouse brain section. In addition, by EDI/SIMS, six different regions of the mouse brain (cerebral cortex, corpus callosum, striatum, medulla oblongata, cerebellar cortex and cerebellar medulla) were examined. While GCs and STs were found to be rich in white matter, PIs were rich in gray matter.
    Biological Mass Spectrometry 03/2010; 45(4):437-43. · 3.41 Impact Factor

Publication Stats

202 Citations
116 Downloads
2k Views
119.84 Total Impact Points

Institutions

  • 2012–2013
    • University of Liège
      • • Laboratoire de Spectrométrie de Masse (LSM)
      • • Department of Chemistry
      Liège, WAL, Belgium
  • 2011–2012
    • Yokohama City University
      • Graduate School of Nanobioscience
      Yokohama-shi, Kanagawa-ken, Japan
  • 2006–2011
    • University of Yamanashi
      • • Clean Energy Research Center
      • • Interdisciplinary Graduate School of Medicine and Engineering
      Kōfu-shi, Yamanashi-ken, Japan
  • 2008
    • Mitsubishi Chemical Group Science and Technology Research Center, Inc.
      Yokohama, Kanagawa, Japan