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1.
Opt Lett ; 44(22): 5533-5536, 2019 Nov 15.
Article in English | MEDLINE | ID: mdl-31730101

ABSTRACT

We present a method for recovery of narrow homogeneous spectral features out of a broad inhomogeneous overlapped profile based on second-derivative processing of the absorption spectra of alkali metal atomic vapor nanocells. The method is shown to preserve the frequency positions and amplitudes of spectral transitions, thus being applicable for quantitative spectroscopy. The proposed technique was successfully applied and tested for measurements of hyperfine splitting and atomic transition probabilities, development of an atomic frequency reference, determination of isotopic abundance, study of atom-surface interaction, and determination of magnetic-field-induced modification of atomic transition frequency and probability. The obtained experimental results are fully consistent with theoretical modeling.

2.
J Orthop Trauma ; 32(4): e139-e144, 2018 04.
Article in English | MEDLINE | ID: mdl-29558376

ABSTRACT

The mission of any academic orthopaedic training program can be divided into 3 general areas of focus: clinical care, academic performance, and research. Clinical care is evaluated on clinical volume, patient outcomes, patient satisfaction, and becoming increasingly focused on data-driven quality metrics. Academic performance of a department can be used to motivate individual surgeons, but objective measures are used to define a residency program. Annual in-service examinations serve as a marker of resident knowledge base, and board pass rates are clearly scrutinized. Research productivity, however, has proven harder to objectively quantify. In an effort to improve transparency and better account for conflicts of interest, bias, and self-citation, multiple bibliometric measures have been developed. Rather than using individuals' research productivity as a surrogate for departmental research, we sought to establish an objective methodology to better assess a residency program's ability to conduct meaningful research. In this study, we describe a process to assess the number and quality of publications produced by an orthopaedic residency department. This would allow chairmen and program directors to benchmark their current production and make measurable goals for future research investment. The main goal of the benchmarking system is to create an "h-index" for residency programs. To do this, we needed to create a list of relevant articles in the orthopaedic literature. We used the Journal Citation Reports. This publication lists all orthopaedic journals that are given an impact factor rating every year. When we accessed the Journal Citation Reports database, there were 72 journals included in the orthopaedic literature section. To ensure only relevant, impactful journals were included, we selected journals with an impact factor greater than 0.95 and an Eigenfactor Score greater than 0.00095. After excluding journals not meeting these criteria, we were left with 45 journals. We performed a Scopus search over a 10-year period of these journals and created a database of articles and their affiliated institutions. We performed several iterations of this to maximize the capture of articles attributed to institutions with multiple names. Based off of this extensive database, we were able to analyze all allopathic US residency programs based on their quality research productivity. We believe this as a novel methodology to create a system by which residency program chairmen and directors can assess progress over time and accurate comparison with other programs.


Subject(s)
Biomedical Research/statistics & numerical data , Efficiency , Internship and Residency , Orthopedics/education , Bibliometrics , Humans , Orthopedics/statistics & numerical data
3.
Eur J Neurosci ; 38(3): 2434-44, 2013 Aug.
Article in English | MEDLINE | ID: mdl-23668781

ABSTRACT

It is known that expectation of reward speeds up saccades. Past studies have also shown the presence of a saccadic velocity bias in the orbit, resulting from a biomechanical regulation over varying eccentricities. Nevertheless, whether and how reward expectation interacts with the biomechanical regulation of saccadic velocities over varying eccentricities remains unknown. We addressed this question by conducting a visually guided double-step saccade task. The role of reward expectation was tested in monkeys performing two consecutive horizontal saccades, one associated with reward prospect and the other not. To adequately assess saccadic velocity and avoid adaptation, we systematically varied initial eye positions, saccadic directions and amplitudes. Our results confirmed the existence of a velocity bias in the orbit, i.e., saccadic peak velocity decreased linearly as the initial eye position deviated in the direction of the saccade. The slope of this bias increased as saccadic amplitudes increased. Nevertheless, reward prospect facilitated velocity to a greater extent for saccades away from than for saccades toward the orbital centre, rendering an overall reduction in the velocity bias. The rate (slope) and magnitude (intercept) of reward modulation over this velocity bias were linearly correlated with amplitudes, similar to the amplitude-modulated velocity bias without reward prospect, which presumably resulted from a biomechanical regulation. Small-amplitude (≤ 5°) saccades received little modulation. These findings together suggest that reward expectation modulated saccadic velocity not as an additive signal but as a facilitating mechanism that interacted with the biomechanical regulation.


Subject(s)
Cognition/physiology , Reward , Saccades , Animals , Biomechanical Phenomena/physiology , Macaca mulatta
4.
Nanotechnology ; 20(35): 355303, 2009 Sep 02.
Article in English | MEDLINE | ID: mdl-19671965

ABSTRACT

In this paper, we report on a process to prepare gold nanoparticle stripes on SiO(2) by convective/capillary assembly without any patterning of the substrate. Electrical devices were then fabricated using stencil lithography in order to avoid any contamination. I(V) measurements at room temperature show that these stripes have an ohmic behavior between +/- 0.5 V with a resistivity ranging from one to two orders higher than the gold bulk value. Furthermore, I(V) and I(t) measurements reveal current fluctuations that were interpreted in terms of charging and discharging of nanoparticle islands leading to a very large electrostatic perturbation of current conduction paths. Unconventional relative amplitudes of up to 99% RTS fluctuations were observed.

5.
Akush Ginekol (Sofiia) ; 43(5): 11-8, 2004.
Article in Bulgarian | MEDLINE | ID: mdl-15518278

ABSTRACT

AIM OF THE STUDY: To assess the feasibility of first trimester ultrasound screening for structural and chromosomal fetal anomalies in multiple gestations. METHODS: An observational prospective follow up study was carried out in 32 cases of multiple pregnancies. Two scans were scheduled in each case--the first, between 6-9 weeks of gestation (w.g.) and the second, between 11-14 w.g. The aim was assessment of fetal number, viability, chorionicity/amnionicity and fetal biometry. In addition, nuchal translucency [NT] measurement, assessment of risk for chromosomal anomalies and fetal anatomy survey were always performed. Increased NT > or = 95 percentile and/or detection of structural anomaly were considered indications for invasive prenatal diagnosis and fetal karyotyping. Selective fetocide was considered in cases of chromosomal or structural anomalies and in high-order multiple gestations (> or = 3 fetuses). Pregnancy outcome was ascertained by the physical examination of the fetuses, placentas and membranes postpartum, the hospital records, the referring physicians or the parents. RESULTS: From 32 cases of multiple pregnancies included in the study, 28 were twins, and 4--triplets. 68% (19/28) of the twin pregnancies were bichorionic-biamniotic [Bi-Bi], 25% (7/28)--monochorionic-biamniotic [Mo-Bi] and 7% (2/28)--monochorionic-monoamniotic [Mo-Mo]. 4 cases of increased NT in one of the twins (1--associated with trisomy 21) were observed, as well as 2 cases of structural fetal anomalies (1--discordant for exencephaly, and 1--with conjoint twins), 2 cases of feto-fetal transfusion syndrome that developed in the second trimester (1--associated with increased NT between 11-14 w.g.), 1 case of TRAP syndrome [twin-reversed arterial perfusion] and 1 case of cord entanglement in monoamniotic twins. In addition, there were 4 cases of a vanishing twin in the first trimester, and in 2 other cases spontaneous miscarriage of both twins occurred before 24 w.g. In two of the triplet pregnancies selective fetocide was performed, one was successfully delivered at 33 w.g. and in the last case the parents chose to terminate the pregnancy. CONCLUSIONS: First trimester ultrasound is a method of choice for detection of major structural fetal anomalies in multiple gestations. Increased NT between 11-14 w.g. in multiple pregnancies is a useful screening tool for detection of chromosomal fetal anomalies, while in monochorionic twins its presence might predict the development of fetofetal transfusion syndrome. First trimester selective fetocide in high-order multiple gestations or in affected twins is one of the options in pregnancy management.


Subject(s)
Chromosome Disorders/diagnostic imaging , Congenital Abnormalities/diagnostic imaging , Nuchal Translucency Measurement , Pregnancy, Multiple , Abortion, Missed/diagnostic imaging , Abortion, Missed/embryology , Chromosome Disorders/embryology , Congenital Abnormalities/embryology , Female , Fetofetal Transfusion/diagnostic imaging , Fetofetal Transfusion/embryology , Follow-Up Studies , Gestational Age , Humans , Pregnancy , Pregnancy Trimester, First , Prospective Studies
6.
Akush Ginekol (Sofiia) ; 43(3): 3-10, 2004.
Article in Bulgarian | MEDLINE | ID: mdl-15341249

ABSTRACT

AIM OF THE STUDY: To assess the feasibility of ultrasound screening and diagnosis of structural fetal anomalies at the 11-14 week scan. METHODS: An observational prospective follow up study from March 2000 till May 2003 was performed at three referral centers by seven experienced sonographers with high-resolution ultrasound equipment. 1135 singleton pregnancies between 11+0 and 14+6 weeks gestation (w.g.) participated in the study. The first trimester scan included assessment of fetal number, viability and biometry, nuchal translucency [NT] measurement and fetal anatomy survey performed according to standardized published protocols. Increased NT > or = 95th centile and/or diagnosis of structural fetal anomaly was considered as indication for invasive prenatal diagnosis, early fetal echocardiogram and follow-up scans, including a detailed fetal anomaly scan at 18-22 w.g. and a third scan at 28-32 w.g. Pregnancy outcome was ascertained from hospital records, referring physicians or the patients themselves. RESULTS: The overall prevalence of structural fetal anomalies in the present study was 4.6% (53/1135). 22% (12/53) of the structural anomalies were detected between 11-14 w.g. 9 of those had normal karyotype, and 3 were associated with chromosomal anomalies. Furthermore, 10 cases of increased NT, with or without non-immune hydrops fetalis, were associated with congenital heart disease, rare genetic syndromes and adverse pregnancy outcome later in gestation. The ultrasound detection rate of structural fetal anomalies in the present study increased from 22% (12/53), to 69% (37/53) and 79% (42/53) for the first trimester scan, the first and second trimester scans, and the combination of all three scans, respectively. 21% (11/53) of all structural fetal anomalies were missed by prenatal ultrasound. CONCLUSIONS: The first trimester scan is a method of choice for the diagnosis of major structural fetal anomalies. NT measurement is a useful screening test for chromosomal anomalies. In cases with increased NT subsequent development of congenital heart disease, rare genetic syndromes or adverse pregnancy outcome should be ruled out. At present, the second trimester scan constitutes an indispensable tool for the detection of most structural abnormalities. Even in advanced gestation the prenatal diagnosis of certain anomalies is difficult and often unfeasible.


Subject(s)
Congenital Abnormalities/diagnostic imaging , Ultrasonography, Prenatal , Chromosome Aberrations , Congenital Abnormalities/embryology , Congenital Abnormalities/epidemiology , Congenital Abnormalities/genetics , Female , Follow-Up Studies , Gestational Age , Humans , Karyotyping , Mass Screening , Neck/diagnostic imaging , Neck/embryology , Predictive Value of Tests , Pregnancy , Prevalence , Prospective Studies
7.
Clin Exp Obstet Gynecol ; 30(2-3): 109-10, 2003.
Article in English | MEDLINE | ID: mdl-12854855

ABSTRACT

A case of bilateral cleft lip and palate associated with increased fetal nuchal translucency detected at 14 weeks of gestation in a cocaine abusing pregnant woman is presented. There were no other associated structural or chromosomal abnormalities. We propose that systematic examination in both the sagittal and parasagittal plane of the fetal profile and recognition of the characteristic ultrasound appearance of a premaxillary protruding echogenic mass should make detection of this type of cleft relatively easy at the moment of the first trimester scan. First trimester diagnosis of cleft lip and palate can facilitate the parental decision-making process on continuing or terminating the pregnancy and should open the perspective of fetal surgery.


Subject(s)
Cleft Lip/diagnostic imaging , Cleft Palate/diagnostic imaging , Cocaine-Related Disorders/complications , Gestational Age , Neck/embryology , Ultrasonography, Prenatal , Abortion, Induced , Adult , Female , Humans , Neck/diagnostic imaging , Pregnancy
8.
Biochem J ; 360(Pt 1): 179-88, 2001 Nov 15.
Article in English | MEDLINE | ID: mdl-11696006

ABSTRACT

Cytosolic and nuclear O-linked N-acetylglucosaminylation has been proposed to be involved in the nuclear transport of cytosolic proteins. We have isolated nuclear and cytosolic N-acetyl-d-glucosamine (GlcNAc)-specific lectins from adult rat liver by affinity chromatography on immobilized GlcNAc and identified these lectins, by a proteomic approach, as belonging to the heat-shock protein (HSP)-70 family (one of them being heat-shock cognate 70 stress protein). Two-dimensional electrophoresis indicated that the HSP-70 fraction contained three equally abundant proteins with molecular masses of 70, 65 and 55 kDa. The p70 and p65 proteins are phosphorylated and are themselves O-linked GlcNAc (O-GlcNAc)-modified. The HSP-70 associated into high molecular mass complexes that dissociated in the presence of reductive and chaotropic agents. The lectin(s) present in this complex was (were) able to recognize cytosolic and nuclear ligands, which have been isolated using wheat germ agglutinin affinity chromatography. These ligands are O-GlcNAc glycosylated as demonstrated by [(3)H]galactose incorporation and analysis of the products released by reductive beta-elimination. The isolated lectins specifically recognized ligands present in both the cytosol and the nucleus of human resting lymphocytes. These results demonstrated the existence of endogenous GlcNAc-specific lectins, identified as HSP-70 proteins, which could act as a shuttle for the nucleo-cytoplasmic transport of O-GlcNAc glycoproteins between the cytosol and the nucleus.


Subject(s)
Acetylglucosamine/metabolism , Cell Nucleus/metabolism , Cytosol/metabolism , Heat-Shock Proteins/metabolism , Lectins/chemistry , Liver/metabolism , Animals , Biotinylation , Blotting, Western , Electrophoresis, Gel, Two-Dimensional , Electrophoresis, Polyacrylamide Gel , Glycoproteins/metabolism , Isoelectric Focusing , Ligands , Lymphocytes/metabolism , Phosphorylation , Rats , Rats, Wistar , Sepharose/chemistry
9.
Glycobiology ; 11(8): 663-76, 2001 Aug.
Article in English | MEDLINE | ID: mdl-11479277

ABSTRACT

The fine structural motifs of sialic acids, a frequent terminal monosaccharide of glycans, seem to contain essential biological properties. To identify such subtle structural differences, a reliable method was developed for the qualitative and quantitative identification of sialic acids present in different tissues and fluids. This method involved, after liberation of sialic acids by mild acid hydrolysis, their methyl esterification using diazomethane in the presence of methanol and the formation of volatile derivatives using heptafluorobutyric anhydride. The derivatives were analyzed by gas chromatography coupled to mass spectrometry in the electron impact mode. This technique allowed the separation and identification of a large variety of sialic acids, including different O-acylated forms of N-acetyl and N-glycolyl neuraminic acids and of 3-deoxy-D-glycero-D-galacto-nonulosonic acid (Kdn). This method allowed also identifying 8-O-methylated and 8-O-sulfated derivatives, de-N-acetylated neuraminic acid, and 1,7-sialic acid lactones. Compounds present in very complex mixtures could be identified through their fragmentation patterns. Because of the stability of the heptafluorobutyrate derivatives, this method presents important improvements compared to the previous techniques, because it can be frequently applied on very small amounts of crude samples. This methodology will support progress in the field of the biology of sialic acids.


Subject(s)
Fluorocarbons/chemistry , Sialic Acids/chemistry , Acylation , Amines/chemistry , Animals , Gas Chromatography-Mass Spectrometry/methods , Lactones/chemistry , Methylation , N-Acetylneuraminic Acid/analogs & derivatives , N-Acetylneuraminic Acid/chemistry , Neuraminic Acids/chemistry , Propionates/metabolism , Sugar Acids/chemistry , Sulfates/chemistry
10.
Biochem J ; 357(Pt 1): 167-82, 2001 Jul 01.
Article in English | MEDLINE | ID: mdl-11415447

ABSTRACT

The free-living nematode Caenorhabditis elegans is a relevant model for studies on the role of glycoconjugates during development of multicellular organisms. Several genes coding for glycosyltransferases involved in the synthesis of N- and O-linked glycans have already been isolated, but, apart from repetitive dimers of glycosaminoglycans, no detailed structure of either type of component has been published so far. This study aimed to establish the structures of the major O-glycans synthesized by C. elegans to give an insight into the endogenous glycosyltransferase activities expressed in this organism. By the use of NMR and MS, we have resolved the sequence of seven of these components that present very unusual features. Most of them were characterized by the type-1 core substituted on Gal and/or GalNAc by (beta1-4)Glc and (beta1-6)Glc residues. Another compound exhibited the GalNAc(beta1-4)N-acetylglucosaminitol sequence in the terminal position, to which was attached a tetramer of beta-Gal substituted by both Fuc and 2-O-methyl-fucose residues. Our experimental procedure led also to the isolation of glycosaminoglycan-like components and oligomannosyl-type N-glycans. In particular, the data confirmed that C. elegans synthesizes the ubiquitous linker sequence GlcA(beta1-3)Gal(beta1-3)Gal(beta1-4)Xyl.


Subject(s)
Caenorhabditis elegans/metabolism , Glucose/metabolism , Oligosaccharides/chemistry , Polysaccharides/chemistry , Polysaccharides/chemical synthesis , Animals , Carbohydrate Conformation , Carbohydrate Sequence , Chondroitin/chemistry , Chromatography, High Pressure Liquid , Glucose/analysis , Methylation , Molecular Sequence Data , Monosaccharides/analysis , Mucins/chemistry , Nuclear Magnetic Resonance, Biomolecular , Polysaccharides/isolation & purification
11.
Anal Biochem ; 284(2): 201-16, 2000 Sep 10.
Article in English | MEDLINE | ID: mdl-10964402

ABSTRACT

In a previous work (Zanetta et al. Glycobiology 9, 255-266 (1999)), it was reported that all constituents of gangliosides could be obtained as heptafluorobutyrate derivatives after methanolysis in a single gas chromatography analysis. This report demonstrates that gas chromatography coupled with mass spectrometry in the electron impact mode allows identification and quantification of long-chain bases and fatty acids without interference from monosaccharides. On the basis of ions specific for families and for individual compounds, sphingosines, sphinganines, and phytosphingosines (including ramified, unsaturated, hydroxylated, and etherified compounds) can be identified. Fatty acid methyl esters, including linear, ramified, unsaturated, and hydroxylated species, are identified and quantified in the same way. Possible extensions of this method to the fatty moiety of other lipids (alkylacylglycerol and dimethyl acetal) are discussed.


Subject(s)
Fluorocarbons/chemistry , Gas Chromatography-Mass Spectrometry/methods , Glycolipids/analysis , Animals , Bacteria/chemistry , Esters/analysis , Fatty Acids/chemistry , Glycolipids/chemistry , Hydroxylation , Rats , Yeasts/chemistry
12.
Glycobiology ; 10(5): 511-9, 2000 May.
Article in English | MEDLINE | ID: mdl-10764840

ABSTRACT

Erythropoietin (Epo) is a 166 amino acids protein containing three N-glycosylation sites (Asn-24, Asn-38, and Asn-83) and 1 O- glycosylation site (Ser-126) and involved in the regulation of the level of red blood cells. Today, only one recombinant human Epo (rHuEpo), produced in CHO cell line, is extensively used in therapy to cure severe anemia. The structure of the glycan chains of this rHuEpo slightly differ of those of the urinary human Epo (uHuEpo), considered as the natural Epo molecule. In an attempt to produce a rHuEpo as close as possible to the uHuEpo, Epo gene was expressed in a human lymphoblastoid cell line, named RPMI 1788. In order to fully characterize the Epo-RPMI, structural characterizations of the protein skeleton as well as glycan chains were undergone. As expected, the amino acid sequence of the Epo-RPMI conformed to that of uHuEpo. Surprisingly, the structure of some N-glycan chains, as mainly determined by ESI-MS, revealed some unusual characteristics. Thus, 80% of N-glycans possess a bisecting GlcNAc residue, 25% bear a second fucose residue which is present, in a large part, in a sialyl Le(x)motif, and 13% contain more than three LacNAc repeats (up to five per molecule). Despite these unusual structural characteristics, the data concerning the in vitro and in vivo biological activities were not impaired when compared to Epo-CHO and uHuEpo.


Subject(s)
Erythropoietin/chemistry , Erythropoietin/metabolism , Lymphocytes/metabolism , Amino Acid Sequence , Animals , CHO Cells , Cell Line , Chemical Fractionation , Cricetinae , Erythropoietin/genetics , Glycopeptides/analysis , Glycopeptides/isolation & purification , Glycosylation , Humans , Methylation , Molecular Sequence Data , Monosaccharides/analysis , N-Acetylneuraminic Acid/metabolism , Oligosaccharides/analysis , Pharmacokinetics , Recombinant Proteins , Sialyl Lewis X Antigen , Structure-Activity Relationship
13.
Enzyme Microb Technol ; 26(5-6): 374-380, 2000 Mar 01.
Article in English | MEDLINE | ID: mdl-10713210

ABSTRACT

Peptide T has a sequence (Ala-Ser-Thr-Thr-Thr-Asn-Tyr-Thr) belonging to HIV envelope that is involved in the interaction with CD(4) receptor of T lymphocytes. Research of protease activities towards this peptide is very relevant for AIDS therapy. Characterization of specificity of subtilisin Carlsberg towards this very hydrophilic peptide is proposed by using high-performance liquid chromatography and mass spectrometry. Peptide T was totally hydrolysed by the protease after 24 h. Separation of hydrophilic fragments was perfected with an hydrophilic stationary phase and a reversed acetonitrile gradient. Peptide masses were determined by ion spray mass spectrometry. Four primary and one secondary hydrolysis products were found, corresponding to cleavage at the carboxylic side of threonine. Specifities of subtilisin Carlsberg towards the Segments 19 to 26 of bovine pancreatic ribonuclease A, an homologous fragment of peptide T, and peptide T were compared.

14.
Anal Biochem ; 267(2): 300-8, 1999 Feb 15.
Article in English | MEDLINE | ID: mdl-10036134

ABSTRACT

The most common method used for the liberation of monosaccharides from glycoprotein N-glycans involves anhydrous methanolysis because it liberates almost quantitatively monosaccharides as O-methylglycosides, which are resistant to further degradation. However, it is generally assumed that this method does not cleave quantitatively the N-glycosidic bonds. This paper demonstrates that classical methanolysis conditions quantitatively cleave the N-glycosidic bond (96%), liberating glucosamine (and not its O-methylglycosides) and other minor reaction products which were identified. Because other N-acetyl-d-glucosamine (GlcNAc) residues are quantitatively liberated as the O-methylglycosides of glucosamine, the GlcNAc residue involved in the N-glycosidic bond is separated from the others using gas chromatography of heptafluorobutyrate derivatives.


Subject(s)
Glycoproteins/analysis , Monosaccharides/analysis , Chromatography, Gas , Gas Chromatography-Mass Spectrometry , Magnetic Resonance Spectroscopy , Polysaccharides/analysis
15.
Glycobiology ; 9(3): 255-66, 1999 Mar.
Article in English | MEDLINE | ID: mdl-10024663

ABSTRACT

We have developed a method involving the formation of hepta-fluorobutyrate derivatives of O-methyl-glycosides liberated from glycoproteins and glycolipids following methanolysis. The stable derivatives of the most common monosaccharides of these glycoconjugates (Ara, Rha, Xyl, Fuc, Gal, Man, Glc, GlcNAc, GalNAc, Neu5Ac, KDN) can be separated and quantitatively and reproducibly determined with a high degree of sensitivity level (down to 25 pmol) in the presence of lysine as an internal standard. The GlcNAc residue bound to Asn in N-glycans is quantitatively recovered as two peaks. The latter were easily distinguished from the other GlcNAc residues of N-glycans, thus allowing a considerable improvement of the data on structure of N-glycans obtained from a single carbohydrate analysis. The most common contaminants present in buffers commonly used for the isolation of soluble or membrane-bound glycoproteins (SDS, Triton X-100, DOC, TRIS, glycine, and polyacrylamide or salts, as well as monosaccharide constituents of proteoglycans or degradation products of nucleic acids) do not interfere with these determinations. A carbohydrate analysis of glycoproteins isolated from a SDS/PAGE gel or from PDVF membranes can be performed on microgram amounts without significant interferences. Since fatty acid methyl esters and sphingosine derivatives are separated from the monosaccharide peaks, the complete composition of gangliosides can be achieved in a single step starting from less than 1 microg of the initial compound purified by preparative Silicagel TLC. Using electron impact ionization mass spectrometry, reporter ions for the different classes of O-methyl-glycosides (pentoses, deoxy-hexoses, hexoses, hexosamines, uronic acids, sialic acid, and KDN) allow the identification of these compounds in very complex mixtures. The mass of each compound can be determined in the chemical ionization mode and detection of positive or negative ions. This method presents a considerable improvement compared to those using TMS derivatives. Indeed the heptafluorobutyrate derivatives are stable, and acylation of amino groups is complete. Moreover, there is no interference with contaminants and the separation between fatty acid methyl-esters and O-methyl glycosides is achieved.


Subject(s)
Chromatography, Gas/methods , Fluorocarbons/chemistry , Glycolipids/chemistry , Glycoproteins/chemistry , Methylglycosides/chemistry , Monosaccharides/isolation & purification , Acylation , Carbohydrate Sequence , Gas Chromatography-Mass Spectrometry/methods , Molecular Sequence Data
16.
Glycoconj J ; 16(10): 617-27, 1999 Oct.
Article in English | MEDLINE | ID: mdl-10972140

ABSTRACT

A major impediment in the analysis of glycosaminoglycans is the difficulty to cleave quantitatively the glycosidic bonds because of the stabilisation of glycosidic bonds and of the relative instability of the liberated constituents. This manuscript describes a modified procedure of methanolysis in the presence of barium acetate, reducing the destruction of uronic acids and increasing the cleavage yield. The reaction products could be identified and analysed quantitatively by GC and GC/MS of the heptafluorobutyrate derivatives of O-methyl glycosides of monosaccharides (for keratan sulphate and chondroitin sulphate B), or as a mixture of O-methyl glycosides of monosaccharides and of disaccharides (for the other sulphated glycosaminoglycans). Quantitative molar ratio between the different monosaccharide constituents (including the linkage region constituents) could be obtained, even when proteoglycans also contain classical N-glycans or O-glycans.


Subject(s)
Fluorocarbons/metabolism , Glycosaminoglycans/chemistry , Glycosaminoglycans/metabolism , Methanol/metabolism , Proteoglycans/chemistry , Proteoglycans/metabolism , Acylation , Artifacts , Barium/metabolism , Chondroitin Sulfates/chemistry , Chondroitin Sulfates/metabolism , Chromatography, Thin Layer , Disaccharides/analysis , Gas Chromatography-Mass Spectrometry/methods , Heparin/chemistry , Heparin/metabolism , Keratan Sulfate/chemistry , Keratan Sulfate/metabolism , Methylglycosides/chemistry , Methylglycosides/metabolism , Monosaccharides/analysis , Polysaccharides/analysis , Polysaccharides/chemistry , Uronic Acids/metabolism
17.
J Biol Chem ; 273(50): 33644-51, 1998 Dec 11.
Article in English | MEDLINE | ID: mdl-9837949

ABSTRACT

The binding of Bandeiraea simplicifolia lectin-I isolectin B4 on the endogenous glycoproteins of different insect cell lines led us to characterize for the first time a UDP-Gal:Galbeta1-3GalNAc alpha1, 4-galactosyltransferase in a Mamestra brassicae cell line (Mb). The study of the acceptor specificity indicated that the Mb alpha-galactosyltransferase prefers Galbeta1-3-R as acceptor, and among such glycans, the relative substrate activity Vmax/Km was equal to 20 microliters.mg-1.h-1 for Galbetal-3GlcNAcbeta1-O-octyl and to 330 microliters.mg-1.h-1 for Galbeta1-3GalNAcalpha-1-O-benzyl, showing clearly that Galbeta1-3GalNAc disaccharide was the more suitable acceptor substrate for Mb alpha-galactosyltransferase activity. Nuclear magnetic resonance and mass spectrometry data allowed us to establish that the Mb alpha-galactosyltransferase synthesizes one unique product, Galalpha1-4Galbeta1-3GalNAcalpha1-O-benzyl. The Galbeta1-3GalNAc disaccharide is usually present on O-glycosylation sites of numerous asialoglycoproteins and at the nonreducing end of some glycolipids. We observed that Mb alpha1,4-galactosyltransferase catalyzed the transfer of galactose onto both natural acceptors. Finally, we demonstrated that the trisaccharide Galalpha1-4Galbeta1-3GalNAcalpha1-O-benzyl was able to inhibit anti-PK monoclonal antibody-mediated hemagglutination of human blood group PK1 and PK2 erythrocytes.


Subject(s)
Galactosyltransferases/metabolism , Moths/enzymology , Plant Lectins , Animals , Asialoglycoproteins/metabolism , Cell Line , Galactose/metabolism , Galactosyltransferases/chemistry , Glycolipids/metabolism , Humans , Lectins/metabolism , Moths/cytology , Protein Binding , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Spodoptera/cytology , Substrate Specificity
18.
Carbohydr Res ; 311(1-2): 51-9, 1998 Sep.
Article in English | MEDLINE | ID: mdl-9821266

ABSTRACT

A methodology for the determination of the sialylation pattern of N-glycans, extent of sialylation and the ratio between alpha-(2-->3) and alpha-(2-->6) sialyl linkages, is presented based on the labelling of the C-3 and C-6 hydroxyl groups of Gal residues obtained after permethylation, saponification, selective desialylation of sialylated oligosaccharides and methanolysis. Deuteromethylation and GC/MS analysis of Gal derivatives allow to determine the sialylation level of glycans. O-Ethyl ether labelling followed by GC analysis of the resulting Gal derivatives allows to obtain the ratio between alpha-(2-->3) and alpha-(2-->6) sialyl linkages. The method was applied to LNT (LcOse4: beta-D-Galp-(1-->3)-beta-D-GlcpNAc-(1-->3)-beta-D- Galp-(1-->4)-D-Glcp), LSTa (IV3NeuAcLcOse4: alpha-Neup5Ac-(2-->3)-beta-D-Galp-(1-->3)-beta-D- GlcpNac-(1-->3-beta-D-Galp-(1-->4)-D-Glcp), LSTc (IV6NeuAcn LcOse4: alpha-Neup5Ac-(2-->6)-beta-D-Galp-(1-->4)-beta-D-GlcpNAc-(1-->3)- beta-D-Galp-(1-->4)-D-Glcp) and a bisialylated biantennary N-glycan in which sialic acid is bound to Gal residues via an alpha-(2-->6) linkage. Using this method, it was found that 92.8% of N-glycans in bovine fetuin is sialylated and that the ratio of alpha-(2-->6) versus alpha-(2-->3) sialyl linkages was 31:19.


Subject(s)
N-Acetylneuraminic Acid/chemistry , Polysaccharides/chemistry , alpha-Fetoproteins/chemistry , Carbohydrate Conformation , Carbohydrate Sequence , Gas Chromatography-Mass Spectrometry , Methylation , Molecular Sequence Data
19.
Physiol Meas ; 19(2): 127-48, 1998 May.
Article in English | MEDLINE | ID: mdl-9626678

ABSTRACT

Near-field microwave radiometry and radiometric imaging are non-invasive techniques that are able to provide temperature information at a depth of up to several centimetres in subcutaneous tissues. They are based on the measurement of microwave electromagnetic thermal noise. This paper describes the basic principles, measurement methods and limitations of the techniques and the results of clinical studies, and it reviews recent progress.


Subject(s)
Microwaves , Radiometry/methods , Radiometry/trends , Skin Temperature , Hot Temperature , Humans , Radiometry/instrumentation
20.
Prep Biochem Biotechnol ; 27(1): 1-17, 1997 Feb.
Article in English | MEDLINE | ID: mdl-9090720

ABSTRACT

Two lipophosphoglycans (LPG) from two Leishmania mexicana (Soberano and Yucatan) strains, isolated in Mexico, were purified by affinity chromatography with the Con A lectin. The LPG from each strain, with a 10 kDa molecular weight, possesses two fractions: one with high mannose concentrations and the other with a more heterogeneous saccharidic composition; the mannose-rich fraction and the heterogeneous fraction from the Yucatan strain show a single band of identity with rabbit IgG against promastigotes from L. mexicana Yucatan. Antigen recognition was abolished by treating the high mannose LPGs with alpha-mannosidase. The presence of non reductor alpha-mannose sequences, as determinant epitopes in L. mexicana Soberano and Yucatan strains, was determined by mass spectrometry analysis and enzymatic cleavage.


Subject(s)
Glycosphingolipids/isolation & purification , Leishmania mexicana/chemistry , Animals , Chromatography, Affinity/methods , Chromatography, Gel , Concanavalin A , Female , Glycosphingolipids/chemistry , Hydrogen-Ion Concentration , Immunodiffusion , Mass Spectrometry , Rabbits
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