ltq orbitrapxl mass spectrometer  (Thermo Fisher)


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    Structured Review

    Thermo Fisher ltq orbitrapxl mass spectrometer
    LC-MS/MS retinal proteome analysis in R347 versus wt mice. Whole retina protein (n = 4) and retinal membrane protein (n = 4) extracts were prepared from one month-old R347 and wt mice. LC-MS/MS analysis was performed on an Ultimate3000 nano HPLC system coupled to a <t>LTQ</t> <t>OrbitrapXL</t> mass spectrometer. ( a ) Representative LC-MS/MS profile detected in the retinal samples. ( b ) Distribution of the 1895 identified protein IDs, which were mapped to 1446 gene IDs. 1337 gene IDs were present in the GO database (GO); GO: M refers to entries with membrane in their GO search terms. ( c ) Volcano plot representation of the identified protein IDs in R347 versus wt retinas. X-axis indicates difference in expression level on a log2 scale, whereas the y-axis represents corresponding p-values (Student’s t-Test) on a negative log scale. Red lines indicate 0.5-fold and 2-fold differences in protein expression and significance level of p = 0.05, respectively. Scaling was limited to -6 and 6 on the horizontal axis and 4 on the vertical axis. Entries outside the limiting values were set to the corresponding limiting values.
    Ltq Orbitrapxl Mass Spectrometer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 97/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ltq orbitrapxl mass spectrometer/product/Thermo Fisher
    Average 97 stars, based on 14 article reviews
    Price from $9.99 to $1999.99
    ltq orbitrapxl mass spectrometer - by Bioz Stars, 2020-02
    97/100 stars

    Images

    1) Product Images from "microRNA regulatory circuits in a mouse model of inherited retinal degeneration"

    Article Title: microRNA regulatory circuits in a mouse model of inherited retinal degeneration

    Journal: Scientific Reports

    doi: 10.1038/srep31431

    LC-MS/MS retinal proteome analysis in R347 versus wt mice. Whole retina protein (n = 4) and retinal membrane protein (n = 4) extracts were prepared from one month-old R347 and wt mice. LC-MS/MS analysis was performed on an Ultimate3000 nano HPLC system coupled to a LTQ OrbitrapXL mass spectrometer. ( a ) Representative LC-MS/MS profile detected in the retinal samples. ( b ) Distribution of the 1895 identified protein IDs, which were mapped to 1446 gene IDs. 1337 gene IDs were present in the GO database (GO); GO: M refers to entries with membrane in their GO search terms. ( c ) Volcano plot representation of the identified protein IDs in R347 versus wt retinas. X-axis indicates difference in expression level on a log2 scale, whereas the y-axis represents corresponding p-values (Student’s t-Test) on a negative log scale. Red lines indicate 0.5-fold and 2-fold differences in protein expression and significance level of p = 0.05, respectively. Scaling was limited to -6 and 6 on the horizontal axis and 4 on the vertical axis. Entries outside the limiting values were set to the corresponding limiting values.
    Figure Legend Snippet: LC-MS/MS retinal proteome analysis in R347 versus wt mice. Whole retina protein (n = 4) and retinal membrane protein (n = 4) extracts were prepared from one month-old R347 and wt mice. LC-MS/MS analysis was performed on an Ultimate3000 nano HPLC system coupled to a LTQ OrbitrapXL mass spectrometer. ( a ) Representative LC-MS/MS profile detected in the retinal samples. ( b ) Distribution of the 1895 identified protein IDs, which were mapped to 1446 gene IDs. 1337 gene IDs were present in the GO database (GO); GO: M refers to entries with membrane in their GO search terms. ( c ) Volcano plot representation of the identified protein IDs in R347 versus wt retinas. X-axis indicates difference in expression level on a log2 scale, whereas the y-axis represents corresponding p-values (Student’s t-Test) on a negative log scale. Red lines indicate 0.5-fold and 2-fold differences in protein expression and significance level of p = 0.05, respectively. Scaling was limited to -6 and 6 on the horizontal axis and 4 on the vertical axis. Entries outside the limiting values were set to the corresponding limiting values.

    Techniques Used: Liquid Chromatography with Mass Spectroscopy, Mass Spectrometry, Mouse Assay, High Performance Liquid Chromatography, Expressing

    2) Product Images from "Novel Mass Spectrometric Method for Phosphorylation Quantification Using Cerium Oxide Nanoparticles and Tandem Mass Tags"

    Article Title: Novel Mass Spectrometric Method for Phosphorylation Quantification Using Cerium Oxide Nanoparticles and Tandem Mass Tags

    Journal: Analytical chemistry

    doi: 10.1021/ac203248s

    Direct-infusion nanoESI LTQ-OrbitrapXL analysis of phosphorylation levels of a standard phosphopeptide (IKNLQpSLDPSH) mixed in varying amounts with its nonphosphorylated counterpart. Ratio of mixing of phosphorylated with unphosphorylated were 1:1, 1:9,
    Figure Legend Snippet: Direct-infusion nanoESI LTQ-OrbitrapXL analysis of phosphorylation levels of a standard phosphopeptide (IKNLQpSLDPSH) mixed in varying amounts with its nonphosphorylated counterpart. Ratio of mixing of phosphorylated with unphosphorylated were 1:1, 1:9,

    Techniques Used:

    3) Product Images from "Novel Protein Substrates of the Phospho-Form Modification System in Neisseria gonorrhoeae and Their Connection to O-Linked Protein Glycosylation"

    Article Title: Novel Protein Substrates of the Phospho-Form Modification System in Neisseria gonorrhoeae and Their Connection to O-Linked Protein Glycosylation

    Journal: Infection and Immunity

    doi: 10.1128/IAI.05920-11

    Identification of PE modification sites in NGO1043 by MS 2 . MS analyses were performed on an LTQ OrbitrapXL mass spectrometer. (A) MS 2 spectrum of the PE-modified peptide 27 EAAQAVESDVK 37 at m/z 635.29 [M + 2H] 2+ . A dehydroalanine (DHA) was detected at
    Figure Legend Snippet: Identification of PE modification sites in NGO1043 by MS 2 . MS analyses were performed on an LTQ OrbitrapXL mass spectrometer. (A) MS 2 spectrum of the PE-modified peptide 27 EAAQAVESDVK 37 at m/z 635.29 [M + 2H] 2+ . A dehydroalanine (DHA) was detected at

    Techniques Used: Modification, Mass Spectrometry

    Related Articles

    High Performance Liquid Chromatography:

    Article Title: NINL and DZANK1 Co-function in Vesicle Transport and Are Essential for Photoreceptor Development in Zebrafish
    Article Snippet: Mass spectrometry and data analysis LC-MS/MS analysis was performed on an Ultimate3000 nano RSLC system (Thermo Scientific) coupled to a LTQ Orbitrap Velos or to an LTQ OrbitrapXL mass spectrometer (Thermo Scientific) by a nano spray ion source. .. Tryptic peptide mixtures were automatically injected and loaded at a flow rate of 6 μl/min in 0.1% trifluoroacetic acid in HPLC-grade water onto a nano trap column (75 μm internal diameter (i.d.)

    Article Title: Direct Detection of Biotinylated Proteins by Mass Spectrometry
    Article Snippet: As peptides eluted from the microcapillary column, they were electrosprayed directly into an LTQ-OrbitrapXL mass spectrometer (ThermoFinnigan, Palo Alto, CA) with the application of a distal 2.4 kV spray voltage. .. Application of mass spectrometer scan functions and HPLC solvent gradients were controlled by the Xcaliber data system.

    Article Title: Deciphering Membrane-Associated Molecular Processes in Target Tissue of Autoimmune Uveitis by Label-Free Quantitative Mass Spectrometry *
    Article Snippet: .. LC-MSMS analysis was performed on an Ultimate3000 nano HPLC system (Dionex, Sunnyvale, CA) online coupled to a LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific) by a nano spray ion source. .. Samples originating from in-solution digests of membrane fractions of retinal tissue were acidified with TFA and automatically loaded to the HPLC system equipped with a nano trap column (100 μm i.d.

    Article Title: microRNA regulatory circuits in a mouse model of inherited retinal degeneration
    Article Snippet: .. LC-MS/MS analysis was carried out on an Ultimate3000 nano HPLC system (Dionex, Sunnyvale, CA, USA) coupled to a LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific Inc., Waltham, MA, USA) . .. MS spectra were acquired in OrbitrapXL and up to 10 of the most abundant peptide ions selected for fragmentation in the linear ion trap.

    Article Title: Novel Protein Substrates of the Phospho-Form Modification System in Neisseria gonorrhoeae and Their Connection to O-Linked Protein Glycosylation
    Article Snippet: The LC separation was performed on an Agilent 1200 series capillary high-performance liquid chromatography (HPLC) system. .. The LC system was connected to a nanoelectrospray source of a Thermo Scientific LTQ OrbitrapXL mass spectrometer operated by Xcalibur 2.0.

    Article Title: A QUICK Screen for Lrrk2 Interaction Partners - Leucine-rich Repeat Kinase 2 is Involved in Actin Cytoskeleton Dynamics *
    Article Snippet: .. LC-MS/MS analysis was performed on an Ultimate3000 nano high-pressure liquid chromatography (HPLC) system (Dionex, Sunnyvale, CA) coupled to a LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific) by a nano spray ion source. .. Tryptic peptide mixtures were automatically injected and loaded at a flow rate of 30 μl/min in 95% buffer C (2% acetonitrile, 0.1% trifluoroacetic acid in HPLC grade water) and 5% buffer B (98% acetonitrile, 0.1% formic acid in HPLC grade water) onto a nano trap column (100 μm i.d.

    Flow Cytometry:

    Article Title: NINL and DZANK1 Co-function in Vesicle Transport and Are Essential for Photoreceptor Development in Zebrafish
    Article Snippet: Mass spectrometry and data analysis LC-MS/MS analysis was performed on an Ultimate3000 nano RSLC system (Thermo Scientific) coupled to a LTQ Orbitrap Velos or to an LTQ OrbitrapXL mass spectrometer (Thermo Scientific) by a nano spray ion source. .. Tryptic peptide mixtures were automatically injected and loaded at a flow rate of 6 μl/min in 0.1% trifluoroacetic acid in HPLC-grade water onto a nano trap column (75 μm internal diameter (i.d.)

    Article Title: Probing the orientation of inhibitor and epoxy-eicosatrienoic acid binding in the active site of soluble epoxide hydrolase
    Article Snippet: Nano-RPLC tandem mass spectrometric analysis was performed on an LTQ-OrbitrapXL mass spectrometer (ThermoFisher Scientific, San Jose, CA) equipped with an ADVANCE nanospray ion source (Michrom Bioresources, Auburn, CA), a Surveyor MS pump (ThermoFisher), and a microautosampler (ThermoFisher). .. Then the sample was loaded for 10 min at 2% solvent B (0.1% (v/v) formic acid in ACN) and 98% solvent A (0.1% (v/v) formic acid in water) at a flow rate of 500 nL/min.

    Article Title: Deciphering Membrane-Associated Molecular Processes in Target Tissue of Autoimmune Uveitis by Label-Free Quantitative Mass Spectrometry *
    Article Snippet: LC-MSMS analysis was performed on an Ultimate3000 nano HPLC system (Dionex, Sunnyvale, CA) online coupled to a LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific) by a nano spray ion source. .. × 2 cm, packed with Acclaim PepMap100 C18, 5 μm, 100 Å, Dionex) at a flow rate of 30 μl/minute in 5% buffer B (80% acetonitrile, 0.1% FA in HPLC-grade water) and 95% buffer A (5% acetonitrile, 0.1% FA in HPLC-grade water).

    Article Title: Type IV Pilus Assembly Proficiency and Dynamics Influence Pilin Subunit Phospho-Form Macro- and Microheterogeneity in Neisseria gonorrhoeae
    Article Snippet: LC separation was achieved with a gradient from 5% to 55% of acetonitrile with 0.1% formic acid over 60 minutes employing a flow rate of 0.2 µl min−1 . .. Mass spectra were acquired in the positive-ion mode applying a data-dependent automatic switch between survey scan and MS2 acquisition on the Thermo Scientific LTQ OrbitrapXL mass spectrometer operated by Xcalibur 2.0.

    Article Title: Lack of Tyrosylprotein Sulfotransferase-2 Activity Results in Altered Sperm-Egg Interactions and Loss of ADAM3 and ADAM6 in Epididymal Sperm *
    Article Snippet: Flow-through and elution fractions were separated by SDS-PAGE and then silver-stained or transferred to Whatman Protan BA85 nitrocellulose (Fisher) for PSG2 immunoblotting. .. An LTQ-ORBITRAPXL mass spectrometer (ThermoFisher Scientific, San Jose, CA) equipped with a nano-HPLC(C18 ) system was used to collect the spectra.

    Article Title: Novel Protein Substrates of the Phospho-Form Modification System in Neisseria gonorrhoeae and Their Connection to O-Linked Protein Glycosylation
    Article Snippet: LC separation was achieved with a gradient from 5% to 60% of acetonitrile over 60 min employing a flow rate of 0.2 μl per min. .. The LC system was connected to a nanoelectrospray source of a Thermo Scientific LTQ OrbitrapXL mass spectrometer operated by Xcalibur 2.0.

    Article Title: A QUICK Screen for Lrrk2 Interaction Partners - Leucine-rich Repeat Kinase 2 is Involved in Actin Cytoskeleton Dynamics *
    Article Snippet: LC-MS/MS analysis was performed on an Ultimate3000 nano high-pressure liquid chromatography (HPLC) system (Dionex, Sunnyvale, CA) coupled to a LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific) by a nano spray ion source. .. Tryptic peptide mixtures were automatically injected and loaded at a flow rate of 30 μl/min in 95% buffer C (2% acetonitrile, 0.1% trifluoroacetic acid in HPLC grade water) and 5% buffer B (98% acetonitrile, 0.1% formic acid in HPLC grade water) onto a nano trap column (100 μm i.d.

    Article Title: HLA ligandomics identifies histone deacetylase 1 as target for ovarian cancer immunotherapy
    Article Snippet: Nanoflow LC-MS/MS Mass spectrometry was performed on an LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific, Bremen, Germany) equipped with a nanoelectron spray ion source and coupled to an Ultimate 3000 RSLC Nano UHPLC System (Dionex, Sunnyvale, CA, USA). .. After injection, samples were loaded with 3% of solvent B (20% H2 O, 80% acetonitrile and 0.04% formic acid) on a 2 cm PepMap 100 C18 Nanotrap column (Dionex) at a flowrate of 4 μL/min for 10 min. Peptides were then separated on a 50 cm PepMap C18 column with a particle size of 2 μm (Dionex) running at 45°C with a gradient ranging from 3 to 30% solvent B within 140 min and a flow rate of 300 nL/min.

    Selection:

    Article Title: Type IV Pilus Assembly Proficiency and Dynamics Influence Pilin Subunit Phospho-Form Macro- and Microheterogeneity in Neisseria gonorrhoeae
    Article Snippet: Mass spectra were acquired in the positive-ion mode applying a data-dependent automatic switch between survey scan and MS2 acquisition on the Thermo Scientific LTQ OrbitrapXL mass spectrometer operated by Xcalibur 2.0. .. The ion selection threshold was 500 counts and selected fragment ions were dynamically excluded for 180 seconds.

    Chromatography:

    Article Title: Direct Detection of Biotinylated Proteins by Mass Spectrometry
    Article Snippet: Paragraph title: Peptide Chromatography and MS Collection ... As peptides eluted from the microcapillary column, they were electrosprayed directly into an LTQ-OrbitrapXL mass spectrometer (ThermoFinnigan, Palo Alto, CA) with the application of a distal 2.4 kV spray voltage.

    Article Title: Novel Protein Substrates of the Phospho-Form Modification System in Neisseria gonorrhoeae and Their Connection to O-Linked Protein Glycosylation
    Article Snippet: Dried samples were redissolved in 0.1% formic acid prior to liquid chromatography-tandem mass spectrometry (LC-MS2 ) analyses. .. The LC system was connected to a nanoelectrospray source of a Thermo Scientific LTQ OrbitrapXL mass spectrometer operated by Xcalibur 2.0.

    Isolation:

    Article Title: Hydra Mesoglea Proteome Identifies Thrombospondin as a Conserved Component Active in Head Organizer Restriction
    Article Snippet: For laminin staining of isolated mesoglea the monoclonal mab52 antibody was used at 1:1000. .. The nano UPLC system was coupled online to an LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific).

    Injection:

    Article Title: NINL and DZANK1 Co-function in Vesicle Transport and Are Essential for Photoreceptor Development in Zebrafish
    Article Snippet: Mass spectrometry and data analysis LC-MS/MS analysis was performed on an Ultimate3000 nano RSLC system (Thermo Scientific) coupled to a LTQ Orbitrap Velos or to an LTQ OrbitrapXL mass spectrometer (Thermo Scientific) by a nano spray ion source. .. Tryptic peptide mixtures were automatically injected and loaded at a flow rate of 6 μl/min in 0.1% trifluoroacetic acid in HPLC-grade water onto a nano trap column (75 μm internal diameter (i.d.)

    Article Title: Type IV Pilus Assembly Proficiency and Dynamics Influence Pilin Subunit Phospho-Form Macro- and Microheterogeneity in Neisseria gonorrhoeae
    Article Snippet: A 5 µl peptide sample was injected onto the extraction column (Zorbax 300 SB-C18 of 5 µm particle size, 5 by 0.3 mm (Agilent)) and then washed with a mobile phase of 0.1% formic acid and 3% acetonitrile by the capillary pump with a flow rate of 4 µl min−1 . .. Mass spectra were acquired in the positive-ion mode applying a data-dependent automatic switch between survey scan and MS2 acquisition on the Thermo Scientific LTQ OrbitrapXL mass spectrometer operated by Xcalibur 2.0.

    Article Title: Novel Protein Substrates of the Phospho-Form Modification System in Neisseria gonorrhoeae and Their Connection to O-Linked Protein Glycosylation
    Article Snippet: Five-microliter samples were injected onto the extraction column (5 by 0.3 mm filled with Zorbax 300 SB-C18 of 5-μm particle size; Agilent), before peptides were eluted in the back-flush mode from the extraction column onto the C18 reverse-phase column (0.075 by 150 mm, GlycproSIL C18 -80 Å; Glycpromass, Stove, Germany). .. The LC system was connected to a nanoelectrospray source of a Thermo Scientific LTQ OrbitrapXL mass spectrometer operated by Xcalibur 2.0.

    Article Title: A QUICK Screen for Lrrk2 Interaction Partners - Leucine-rich Repeat Kinase 2 is Involved in Actin Cytoskeleton Dynamics *
    Article Snippet: LC-MS/MS analysis was performed on an Ultimate3000 nano high-pressure liquid chromatography (HPLC) system (Dionex, Sunnyvale, CA) coupled to a LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific) by a nano spray ion source. .. Tryptic peptide mixtures were automatically injected and loaded at a flow rate of 30 μl/min in 95% buffer C (2% acetonitrile, 0.1% trifluoroacetic acid in HPLC grade water) and 5% buffer B (98% acetonitrile, 0.1% formic acid in HPLC grade water) onto a nano trap column (100 μm i.d.

    Article Title: HLA ligandomics identifies histone deacetylase 1 as target for ovarian cancer immunotherapy
    Article Snippet: Nanoflow LC-MS/MS Mass spectrometry was performed on an LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific, Bremen, Germany) equipped with a nanoelectron spray ion source and coupled to an Ultimate 3000 RSLC Nano UHPLC System (Dionex, Sunnyvale, CA, USA). .. After injection, samples were loaded with 3% of solvent B (20% H2 O, 80% acetonitrile and 0.04% formic acid) on a 2 cm PepMap 100 C18 Nanotrap column (Dionex) at a flowrate of 4 μL/min for 10 min. Peptides were then separated on a 50 cm PepMap C18 column with a particle size of 2 μm (Dionex) running at 45°C with a gradient ranging from 3 to 30% solvent B within 140 min and a flow rate of 300 nL/min.

    Incubation:

    Article Title: Arhodomonas sp. Strain Seminole and Its Genetic Potential To Degrade Aromatic Compounds under High-Salinity Conditions
    Article Snippet: After incubation, the reducing buffer was replaced with 55 mM iodoacetamide in 50 mM ammonium bicarbonate, and samples were alkylated for 1 h at room temperature in the dark. .. After overnight digestion at 37°C, the trypsinolytic peptide products were extracted with 0.5% trifluoroacetic acid and analyzed by LC-MS/MS using an LTQ-OrbitrapXL mass spectrometer (ThermoFisher Scientific, MA) basically as described by Voruganti et al. ( ) but with a 40-cm column packed with 3-μm Magic C18 AQ particles (Bruker, MA).

    SDS Page:

    Article Title: Lack of Tyrosylprotein Sulfotransferase-2 Activity Results in Altered Sperm-Egg Interactions and Loss of ADAM3 and ADAM6 in Epididymal Sperm *
    Article Snippet: Flow-through and elution fractions were separated by SDS-PAGE and then silver-stained or transferred to Whatman Protan BA85 nitrocellulose (Fisher) for PSG2 immunoblotting. .. An LTQ-ORBITRAPXL mass spectrometer (ThermoFisher Scientific, San Jose, CA) equipped with a nano-HPLC(C18 ) system was used to collect the spectra.

    Article Title: Arhodomonas sp. Strain Seminole and Its Genetic Potential To Degrade Aromatic Compounds under High-Salinity Conditions
    Article Snippet: Protein extracts were prefractionated on 12% SDS-PAGE gels and stained with Coomassie blue, and each lane was excised into 3 slices. .. After overnight digestion at 37°C, the trypsinolytic peptide products were extracted with 0.5% trifluoroacetic acid and analyzed by LC-MS/MS using an LTQ-OrbitrapXL mass spectrometer (ThermoFisher Scientific, MA) basically as described by Voruganti et al. ( ) but with a 40-cm column packed with 3-μm Magic C18 AQ particles (Bruker, MA).

    Concentration Assay:

    Article Title: Novel Core Promoter Elements and a Cognate Transcription Factor in the Divergent Unicellular Eukaryote Trichomonas vaginalis ▿
    Article Snippet: Affinity-purified samples were precipitated by the addition of trichloroacetic acid to a final concentration of 10%, washed with ice-cold acetone, and then resuspended in digestion buffer (100 mM Tris-HCl, pH 8.5, 8 M urea). .. Proteolytic digestion was performed by the sequential addition of Lys-C and trypsin proteases and analyzed using shotgun proteomic methods on an LTQ-OrbitrapXL mass spectrometer (Thermofisher) as previously described ( , , ).

    Generated:

    Article Title: Arhodomonas sp. Strain Seminole and Its Genetic Potential To Degrade Aromatic Compounds under High-Salinity Conditions
    Article Snippet: After overnight digestion at 37°C, the trypsinolytic peptide products were extracted with 0.5% trifluoroacetic acid and analyzed by LC-MS/MS using an LTQ-OrbitrapXL mass spectrometer (ThermoFisher Scientific, MA) basically as described by Voruganti et al. ( ) but with a 40-cm column packed with 3-μm Magic C18 AQ particles (Bruker, MA). .. Proteins were identified by using Mascot (v2.2.2 from Matrix Science, Boston, MA) and a database generated by in silico digestion of the strain Seminole proteome predicted from the genome, as well as sequences for 114 common adventitious laboratory contaminants ( ).

    Affinity Chromatography:

    Article Title: Lack of Tyrosylprotein Sulfotransferase-2 Activity Results in Altered Sperm-Egg Interactions and Loss of ADAM3 and ADAM6 in Epididymal Sperm *
    Article Snippet: Paragraph title: Affinity Chromatography on the PSG2 Anti-sulfotyrosine Antibody ... An LTQ-ORBITRAPXL mass spectrometer (ThermoFisher Scientific, San Jose, CA) equipped with a nano-HPLC(C18 ) system was used to collect the spectra.

    In Silico:

    Article Title: Arhodomonas sp. Strain Seminole and Its Genetic Potential To Degrade Aromatic Compounds under High-Salinity Conditions
    Article Snippet: After overnight digestion at 37°C, the trypsinolytic peptide products were extracted with 0.5% trifluoroacetic acid and analyzed by LC-MS/MS using an LTQ-OrbitrapXL mass spectrometer (ThermoFisher Scientific, MA) basically as described by Voruganti et al. ( ) but with a 40-cm column packed with 3-μm Magic C18 AQ particles (Bruker, MA). .. Proteins were identified by using Mascot (v2.2.2 from Matrix Science, Boston, MA) and a database generated by in silico digestion of the strain Seminole proteome predicted from the genome, as well as sequences for 114 common adventitious laboratory contaminants ( ).

    Mass Spectrometry:

    Article Title: NINL and DZANK1 Co-function in Vesicle Transport and Are Essential for Photoreceptor Development in Zebrafish
    Article Snippet: .. Mass spectrometry and data analysis LC-MS/MS analysis was performed on an Ultimate3000 nano RSLC system (Thermo Scientific) coupled to a LTQ Orbitrap Velos or to an LTQ OrbitrapXL mass spectrometer (Thermo Scientific) by a nano spray ion source. .. Tryptic peptide mixtures were automatically injected and loaded at a flow rate of 6 μl/min in 0.1% trifluoroacetic acid in HPLC-grade water onto a nano trap column (75 μm internal diameter (i.d.)

    Article Title: Novel Mass Spectrometric Method for Phosphorylation Quantification Using Cerium Oxide Nanoparticles and Tandem Mass Tags
    Article Snippet: .. Nano-RPLC tandem mass spectrometric analyses were performed on a LTQ-OrbitrapXL mass spectrometer (ThermoFisher, San Jose, CA) equipped with an ADVANCE nanospray ion source (Bruker-Michrom, Auburn, CA). .. MALDI-TOF/TOF measurements were acquired on a 4700 Proteomics Analyzer (ABSciex, Foster City, CA) with parameters described in .

    Article Title: Direct Detection of Biotinylated Proteins by Mass Spectrometry
    Article Snippet: .. As peptides eluted from the microcapillary column, they were electrosprayed directly into an LTQ-OrbitrapXL mass spectrometer (ThermoFinnigan, Palo Alto, CA) with the application of a distal 2.4 kV spray voltage. .. A cycle of one full-scan FT mass spectrum (300–1600 m /z ) at 60,000 resolution followed by 10 data-dependent IT MS/MS spectra at a 35% normalized collision energy was repeated continuously throughout each step of the multidimensional separation.

    Article Title: Probing the orientation of inhibitor and epoxy-eicosatrienoic acid binding in the active site of soluble epoxide hydrolase
    Article Snippet: .. Nano-RPLC tandem mass spectrometric analysis was performed on an LTQ-OrbitrapXL mass spectrometer (ThermoFisher Scientific, San Jose, CA) equipped with an ADVANCE nanospray ion source (Michrom Bioresources, Auburn, CA), a Surveyor MS pump (ThermoFisher), and a microautosampler (ThermoFisher). .. The lyophillized tryptic peptide mixture was resuspended in 2% acetonitrile with 0.5% TFA and separated on a 100-μm-ID IntegraFrit” column (New Objective, Woburn, MA) packed in house with reversed-phase Magic C18 AQ material (15 cm length, 5 μm particle size, 200-Å pore size, Michrom Bioresources).

    Article Title: Deciphering Membrane-Associated Molecular Processes in Target Tissue of Autoimmune Uveitis by Label-Free Quantitative Mass Spectrometry *
    Article Snippet: .. LC-MSMS analysis was performed on an Ultimate3000 nano HPLC system (Dionex, Sunnyvale, CA) online coupled to a LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific) by a nano spray ion source. .. Samples originating from in-solution digests of membrane fractions of retinal tissue were acidified with TFA and automatically loaded to the HPLC system equipped with a nano trap column (100 μm i.d.

    Article Title: A Horizontally Acquired Transcription Factor Coordinates Salmonella Adaptations to Host Microenvironments
    Article Snippet: .. Approximately 2 µg of tryptic peptides per isoelectric focusing fraction were analyzed on an LTQ-OrbitrapXL mass spectrometer (Thermo, Fisher Scientific) coupled to an Agilent 1100 series high-performance liquid chromatograph with a nanospray electrospray ionization source (Proxeon Biosystems), as previously described ( ). ..

    Article Title: microRNA regulatory circuits in a mouse model of inherited retinal degeneration
    Article Snippet: .. LC-MS/MS analysis was carried out on an Ultimate3000 nano HPLC system (Dionex, Sunnyvale, CA, USA) coupled to a LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific Inc., Waltham, MA, USA) . .. MS spectra were acquired in OrbitrapXL and up to 10 of the most abundant peptide ions selected for fragmentation in the linear ion trap.

    Article Title: Type IV Pilus Assembly Proficiency and Dynamics Influence Pilin Subunit Phospho-Form Macro- and Microheterogeneity in Neisseria gonorrhoeae
    Article Snippet: .. Mass spectra were acquired in the positive-ion mode applying a data-dependent automatic switch between survey scan and MS2 acquisition on the Thermo Scientific LTQ OrbitrapXL mass spectrometer operated by Xcalibur 2.0. ..

    Article Title: Lack of Tyrosylprotein Sulfotransferase-2 Activity Results in Altered Sperm-Egg Interactions and Loss of ADAM3 and ADAM6 in Epididymal Sperm *
    Article Snippet: .. An LTQ-ORBITRAPXL mass spectrometer (ThermoFisher Scientific, San Jose, CA) equipped with a nano-HPLC(C18 ) system was used to collect the spectra. ..

    Article Title: Novel Protein Substrates of the Phospho-Form Modification System in Neisseria gonorrhoeae and Their Connection to O-Linked Protein Glycosylation
    Article Snippet: .. The LC system was connected to a nanoelectrospray source of a Thermo Scientific LTQ OrbitrapXL mass spectrometer operated by Xcalibur 2.0. .. Nanospray ionization was achieved by applying 1.2 kV between an 8-μm-diameter emitter (PicoTip Emitter, New Objective, Woburn, MA) and the capillary entrance of the Orbitrap.

    Article Title: Arhodomonas sp. Strain Seminole and Its Genetic Potential To Degrade Aromatic Compounds under High-Salinity Conditions
    Article Snippet: .. After overnight digestion at 37°C, the trypsinolytic peptide products were extracted with 0.5% trifluoroacetic acid and analyzed by LC-MS/MS using an LTQ-OrbitrapXL mass spectrometer (ThermoFisher Scientific, MA) basically as described by Voruganti et al. ( ) but with a 40-cm column packed with 3-μm Magic C18 AQ particles (Bruker, MA). .. Proteins were identified by using Mascot (v2.2.2 from Matrix Science, Boston, MA) and a database generated by in silico digestion of the strain Seminole proteome predicted from the genome, as well as sequences for 114 common adventitious laboratory contaminants ( ).

    Article Title: Hydra Mesoglea Proteome Identifies Thrombospondin as a Conserved Component Active in Head Organizer Restriction
    Article Snippet: .. The nano UPLC system was coupled online to an LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific). .. Data dependent acquisition with Xcalibur 2.0.6 (Thermo Fisher Scientific) was performed by one FTMS scan with a resolution of 60000 and a range from 370 to 2000 m/z in parallel with six MS/MS scans of the most intense precursor ions in the ion trap.

    Article Title: A QUICK Screen for Lrrk2 Interaction Partners - Leucine-rich Repeat Kinase 2 is Involved in Actin Cytoskeleton Dynamics *
    Article Snippet: .. LC-MS/MS analysis was performed on an Ultimate3000 nano high-pressure liquid chromatography (HPLC) system (Dionex, Sunnyvale, CA) coupled to a LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific) by a nano spray ion source. .. Tryptic peptide mixtures were automatically injected and loaded at a flow rate of 30 μl/min in 95% buffer C (2% acetonitrile, 0.1% trifluoroacetic acid in HPLC grade water) and 5% buffer B (98% acetonitrile, 0.1% formic acid in HPLC grade water) onto a nano trap column (100 μm i.d.

    Article Title: Novel Core Promoter Elements and a Cognate Transcription Factor in the Divergent Unicellular Eukaryote Trichomonas vaginalis ▿
    Article Snippet: .. Proteolytic digestion was performed by the sequential addition of Lys-C and trypsin proteases and analyzed using shotgun proteomic methods on an LTQ-OrbitrapXL mass spectrometer (Thermofisher) as previously described ( , , ). ..

    Article Title: HLA ligandomics identifies histone deacetylase 1 as target for ovarian cancer immunotherapy
    Article Snippet: .. Nanoflow LC-MS/MS Mass spectrometry was performed on an LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific, Bremen, Germany) equipped with a nanoelectron spray ion source and coupled to an Ultimate 3000 RSLC Nano UHPLC System (Dionex, Sunnyvale, CA, USA). .. After injection, samples were loaded with 3% of solvent B (20% H2 O, 80% acetonitrile and 0.04% formic acid) on a 2 cm PepMap 100 C18 Nanotrap column (Dionex) at a flowrate of 4 μL/min for 10 min. Peptides were then separated on a 50 cm PepMap C18 column with a particle size of 2 μm (Dionex) running at 45°C with a gradient ranging from 3 to 30% solvent B within 140 min and a flow rate of 300 nL/min.

    Tandem Mass Spectroscopy:

    Article Title: Direct Detection of Biotinylated Proteins by Mass Spectrometry
    Article Snippet: As peptides eluted from the microcapillary column, they were electrosprayed directly into an LTQ-OrbitrapXL mass spectrometer (ThermoFinnigan, Palo Alto, CA) with the application of a distal 2.4 kV spray voltage. .. A cycle of one full-scan FT mass spectrum (300–1600 m /z ) at 60,000 resolution followed by 10 data-dependent IT MS/MS spectra at a 35% normalized collision energy was repeated continuously throughout each step of the multidimensional separation.

    Article Title: Hydra Mesoglea Proteome Identifies Thrombospondin as a Conserved Component Active in Head Organizer Restriction
    Article Snippet: The nano UPLC system was coupled online to an LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific). .. Data dependent acquisition with Xcalibur 2.0.6 (Thermo Fisher Scientific) was performed by one FTMS scan with a resolution of 60000 and a range from 370 to 2000 m/z in parallel with six MS/MS scans of the most intense precursor ions in the ion trap.

    Staining:

    Article Title: Arhodomonas sp. Strain Seminole and Its Genetic Potential To Degrade Aromatic Compounds under High-Salinity Conditions
    Article Snippet: Protein extracts were prefractionated on 12% SDS-PAGE gels and stained with Coomassie blue, and each lane was excised into 3 slices. .. After overnight digestion at 37°C, the trypsinolytic peptide products were extracted with 0.5% trifluoroacetic acid and analyzed by LC-MS/MS using an LTQ-OrbitrapXL mass spectrometer (ThermoFisher Scientific, MA) basically as described by Voruganti et al. ( ) but with a 40-cm column packed with 3-μm Magic C18 AQ particles (Bruker, MA).

    Article Title: Hydra Mesoglea Proteome Identifies Thrombospondin as a Conserved Component Active in Head Organizer Restriction
    Article Snippet: The gel was stained with colloidal Coomassie (Bio-Rad) and the lane was cut into 27 pieces. .. The nano UPLC system was coupled online to an LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific).

    Affinity Purification:

    Article Title: Novel Core Promoter Elements and a Cognate Transcription Factor in the Divergent Unicellular Eukaryote Trichomonas vaginalis ▿
    Article Snippet: Affinity-purified samples were precipitated by the addition of trichloroacetic acid to a final concentration of 10%, washed with ice-cold acetone, and then resuspended in digestion buffer (100 mM Tris-HCl, pH 8.5, 8 M urea). .. Proteolytic digestion was performed by the sequential addition of Lys-C and trypsin proteases and analyzed using shotgun proteomic methods on an LTQ-OrbitrapXL mass spectrometer (Thermofisher) as previously described ( , , ).

    Liquid Chromatography:

    Article Title: A QUICK Screen for Lrrk2 Interaction Partners - Leucine-rich Repeat Kinase 2 is Involved in Actin Cytoskeleton Dynamics *
    Article Snippet: Paragraph title: Liquid Chromatography (LC)-MS/MS and Data Analysis ... LC-MS/MS analysis was performed on an Ultimate3000 nano high-pressure liquid chromatography (HPLC) system (Dionex, Sunnyvale, CA) coupled to a LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific) by a nano spray ion source.

    Software:

    Article Title: microRNA regulatory circuits in a mouse model of inherited retinal degeneration
    Article Snippet: LC-MS/MS analysis was carried out on an Ultimate3000 nano HPLC system (Dionex, Sunnyvale, CA, USA) coupled to a LTQ OrbitrapXL mass spectrometer (Thermo Fisher Scientific Inc., Waltham, MA, USA) . .. Peptides were quantified using Progenesis QI (Waters, Milford, MA, USA) and identified with Mascot (version 2.5; Matrix Science, Boston, MA, USA) software.

    Article Title: Arhodomonas sp. Strain Seminole and Its Genetic Potential To Degrade Aromatic Compounds under High-Salinity Conditions
    Article Snippet: After overnight digestion at 37°C, the trypsinolytic peptide products were extracted with 0.5% trifluoroacetic acid and analyzed by LC-MS/MS using an LTQ-OrbitrapXL mass spectrometer (ThermoFisher Scientific, MA) basically as described by Voruganti et al. ( ) but with a 40-cm column packed with 3-μm Magic C18 AQ particles (Bruker, MA). .. Peptide and protein identifications were validated using Scaffold (v4.0.6.1; Proteome Software, Inc., Portland, OR) using the ProteinProphet and PeptideProphet algorithms , with a minimum of 2 peptides identified and at protein thresholds that yielded a 1% false discovery rate for the data set.

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  • 99
    Thermo Fisher ltq orbitrap mass spectrometer
    Human Parkin Ser 65 is a substrate of human PINK1 upon CCCP stimulation. ( a ) Confirmation by mass spectrometry that Ser 65 of human Parkin is phosphorylated by CCCP-induced activation of human wild-type PINK1-FLAG. Flp-In T-Rex HEK293 cells expressing FLAG-empty, wild-type PINK1-FLAG, and kinase-inactive PINK1-FLAG (D384A) were co-transfected with HA-Parkin, induced with doxycycline and stimulated with 10 μM of CCCP for 3 h. Whole-cell extracts were obtained following lysis with 1% Triton and approximately 30 mg of whole-cell extract were subjected to immunoprecipitation with anti-HA-agarose and run on 10% SDS-PAGE and stained with colloidal Coomassie blue. Coomassie-stained bands migrating with the expected molecular mass of HA-Parkin were excised from the gel, digested with trypsin, and subjected to high performance liquid chromatography with tandem mass spectrometry (LC-MS-MS) on an <t>LTQ-Orbitrap</t> mass spectrometer. Extracted ion chromatogram analysis of Ser 131 and Ser 65 phosphopeptide (3 + R.NDWTVQNCDLDQQ S IVHIVQRPWR.K+P). The total signal intensity of the phosphopeptide is plotted on the y -axis and retention time is plotted on the x -axis. The m / z value corresponding to the Ser 131 phosphopeptide was detected in all conditions whilst that of the Ser 65 phosphopeptide was only detected in samples from wild-type PINK1-FLAG-expressing cells following CCCP treatment. ( b ) Characterization of Parkin phospho-Ser 65 antibody. Flp-In T-Rex HEK293 cells expressing FLAG-empty, wild-type PINK1-FLAG, and kinase-inactive PINK1-FLAG were co-transfected with untagged wild-type (WT) or Ser 65 Ala (S65A) mutant Parkin, induced with doxycycline and stimulated with 10 μM of CCCP for 3 h. 0.25 mg of 1% Triton whole-cell lysate were subjected to immunoprecipitation with anti-Parkin antibody (S966C) covalently coupled to protein G Sepharose and then immunoblotted with anti-phospho-Ser 65 antibody in the presence of dephosphorylated peptide. Ten per cent of the immunoprecipitate (IP) was immunoblotted with total anti-Parkin antibody. Twenty five micrograms of whole cell lysate was immunoblotted with total PINK1 antibody.
    Ltq Orbitrap Mass Spectrometer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 436 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ltq orbitrap mass spectrometer/product/Thermo Fisher
    Average 99 stars, based on 436 article reviews
    Price from $9.99 to $1999.99
    ltq orbitrap mass spectrometer - by Bioz Stars, 2020-02
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    99
    Thermo Fisher ltq orbitrap velos mass spectrometer
    A schematic workflow illustrating the steps involved in the differential analysis of RA and OA synovial fluid proteome. Proteins from RA and OA synovial fluid were extracted and depleted to remove the 14 most abundant proteins using multiple affinity removal system, Human-14. The depleted protein from RA and OA were then digested with trypsin and labeled with iTRAQ reagents, 117 and 116 respectively. The labeled samples were pooled and subjected to fractionation using strong cation exchange chromatography. The fractions were then analyzed on a <t>LTQ-Orbitrap</t> <t>Velos</t> mass spectrometer. The MS/MS data obtained was searched against Human RefSeq 50 database using Sequest and Mascot search algorithms. Validation of the iTRAQ quantitation data was carried out using multiple reaction monitoring and Western blot.
    Ltq Orbitrap Velos Mass Spectrometer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 81 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ltq orbitrap velos mass spectrometer/product/Thermo Fisher
    Average 99 stars, based on 81 article reviews
    Price from $9.99 to $1999.99
    ltq orbitrap velos mass spectrometer - by Bioz Stars, 2020-02
    99/100 stars
      Buy from Supplier

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    Human Parkin Ser 65 is a substrate of human PINK1 upon CCCP stimulation. ( a ) Confirmation by mass spectrometry that Ser 65 of human Parkin is phosphorylated by CCCP-induced activation of human wild-type PINK1-FLAG. Flp-In T-Rex HEK293 cells expressing FLAG-empty, wild-type PINK1-FLAG, and kinase-inactive PINK1-FLAG (D384A) were co-transfected with HA-Parkin, induced with doxycycline and stimulated with 10 μM of CCCP for 3 h. Whole-cell extracts were obtained following lysis with 1% Triton and approximately 30 mg of whole-cell extract were subjected to immunoprecipitation with anti-HA-agarose and run on 10% SDS-PAGE and stained with colloidal Coomassie blue. Coomassie-stained bands migrating with the expected molecular mass of HA-Parkin were excised from the gel, digested with trypsin, and subjected to high performance liquid chromatography with tandem mass spectrometry (LC-MS-MS) on an LTQ-Orbitrap mass spectrometer. Extracted ion chromatogram analysis of Ser 131 and Ser 65 phosphopeptide (3 + R.NDWTVQNCDLDQQ S IVHIVQRPWR.K+P). The total signal intensity of the phosphopeptide is plotted on the y -axis and retention time is plotted on the x -axis. The m / z value corresponding to the Ser 131 phosphopeptide was detected in all conditions whilst that of the Ser 65 phosphopeptide was only detected in samples from wild-type PINK1-FLAG-expressing cells following CCCP treatment. ( b ) Characterization of Parkin phospho-Ser 65 antibody. Flp-In T-Rex HEK293 cells expressing FLAG-empty, wild-type PINK1-FLAG, and kinase-inactive PINK1-FLAG were co-transfected with untagged wild-type (WT) or Ser 65 Ala (S65A) mutant Parkin, induced with doxycycline and stimulated with 10 μM of CCCP for 3 h. 0.25 mg of 1% Triton whole-cell lysate were subjected to immunoprecipitation with anti-Parkin antibody (S966C) covalently coupled to protein G Sepharose and then immunoblotted with anti-phospho-Ser 65 antibody in the presence of dephosphorylated peptide. Ten per cent of the immunoprecipitate (IP) was immunoblotted with total anti-Parkin antibody. Twenty five micrograms of whole cell lysate was immunoblotted with total PINK1 antibody.

    Journal: Open Biology

    Article Title: PINK1 is activated by mitochondrial membrane potential depolarization and stimulates Parkin E3 ligase activity by phosphorylating Serine 65

    doi: 10.1098/rsob.120080

    Figure Lengend Snippet: Human Parkin Ser 65 is a substrate of human PINK1 upon CCCP stimulation. ( a ) Confirmation by mass spectrometry that Ser 65 of human Parkin is phosphorylated by CCCP-induced activation of human wild-type PINK1-FLAG. Flp-In T-Rex HEK293 cells expressing FLAG-empty, wild-type PINK1-FLAG, and kinase-inactive PINK1-FLAG (D384A) were co-transfected with HA-Parkin, induced with doxycycline and stimulated with 10 μM of CCCP for 3 h. Whole-cell extracts were obtained following lysis with 1% Triton and approximately 30 mg of whole-cell extract were subjected to immunoprecipitation with anti-HA-agarose and run on 10% SDS-PAGE and stained with colloidal Coomassie blue. Coomassie-stained bands migrating with the expected molecular mass of HA-Parkin were excised from the gel, digested with trypsin, and subjected to high performance liquid chromatography with tandem mass spectrometry (LC-MS-MS) on an LTQ-Orbitrap mass spectrometer. Extracted ion chromatogram analysis of Ser 131 and Ser 65 phosphopeptide (3 + R.NDWTVQNCDLDQQ S IVHIVQRPWR.K+P). The total signal intensity of the phosphopeptide is plotted on the y -axis and retention time is plotted on the x -axis. The m / z value corresponding to the Ser 131 phosphopeptide was detected in all conditions whilst that of the Ser 65 phosphopeptide was only detected in samples from wild-type PINK1-FLAG-expressing cells following CCCP treatment. ( b ) Characterization of Parkin phospho-Ser 65 antibody. Flp-In T-Rex HEK293 cells expressing FLAG-empty, wild-type PINK1-FLAG, and kinase-inactive PINK1-FLAG were co-transfected with untagged wild-type (WT) or Ser 65 Ala (S65A) mutant Parkin, induced with doxycycline and stimulated with 10 μM of CCCP for 3 h. 0.25 mg of 1% Triton whole-cell lysate were subjected to immunoprecipitation with anti-Parkin antibody (S966C) covalently coupled to protein G Sepharose and then immunoblotted with anti-phospho-Ser 65 antibody in the presence of dephosphorylated peptide. Ten per cent of the immunoprecipitate (IP) was immunoblotted with total anti-Parkin antibody. Twenty five micrograms of whole cell lysate was immunoblotted with total PINK1 antibody.

    Article Snippet: Isolated phosphopeptides were analysed by LC-MS-MS on a proxeon Easy-nLC nano liquid chromatography system coupled to a Thermo LTQ-orbitrap mass spectrometer.

    Techniques: Mass Spectrometry, Activation Assay, Expressing, Transfection, Lysis, Immunoprecipitation, SDS Page, Staining, High Performance Liquid Chromatography, Liquid Chromatography with Mass Spectroscopy, Mutagenesis

    Schematic diagram of proteomic analysis of the mouse organ of Corti (OC) sample. ( 1 ) The OCs were reconstituted in 100 μL lysis buffer (100 mM ammonium bicarbonate, pH 8.4); ( 2 ) The lysates were reduced by 5 mM DL-Dithiothreitol (DTT) and digested by trypsin overnight; ( 3 ) The digests were desalted and dried in a vacuum centrifuge immediately after digestion; ( 4 ) Dried peptides were subjected to the in-house assembled reverse phase metal-free multiple-column nanoLC system coupled with ( 5 ) LTQ-Orbitrap XL mass spectrometer for MS analysis.

    Journal: International Journal of Molecular Sciences

    Article Title: Proteomic Analysis of the Organ of Corti Using Nanoscale Liquid Chromatography Coupled with Tandem Mass Spectrometry

    doi: 10.3390/ijms13078171

    Figure Lengend Snippet: Schematic diagram of proteomic analysis of the mouse organ of Corti (OC) sample. ( 1 ) The OCs were reconstituted in 100 μL lysis buffer (100 mM ammonium bicarbonate, pH 8.4); ( 2 ) The lysates were reduced by 5 mM DL-Dithiothreitol (DTT) and digested by trypsin overnight; ( 3 ) The digests were desalted and dried in a vacuum centrifuge immediately after digestion; ( 4 ) Dried peptides were subjected to the in-house assembled reverse phase metal-free multiple-column nanoLC system coupled with ( 5 ) LTQ-Orbitrap XL mass spectrometer for MS analysis.

    Article Snippet: The nanoLC system was interfaced with a LTQ-Orbitrap XL mass spectrometer equipped with a nanoelectrospray ion source (Thermo Scientific, Waltham, MA, USA).

    Techniques: Lysis, Mass Spectrometry

    Quantification of calcium-dependent regulation of phosphorylation sites of Toxoplasma invasion motor complex components. A ) Work flow to identify individual phosphorylation sites and quantitatively assess their responsiveness to calcium signals using a SILAC-based proteomics approach. A 1∶1 mixture of Triton X-100 lysates from “Heavy” (H; Arg4/Lys8)-labeled ethanol-stimulated tachyzoites or

    Journal: PLoS Pathogens

    Article Title: Quantitative in vivo Analyses Reveal Calcium-dependent Phosphorylation Sites and Identifies a Novel Component of the Toxoplasma Invasion Motor Complex

    doi: 10.1371/journal.ppat.1002222

    Figure Lengend Snippet: Quantification of calcium-dependent regulation of phosphorylation sites of Toxoplasma invasion motor complex components. A ) Work flow to identify individual phosphorylation sites and quantitatively assess their responsiveness to calcium signals using a SILAC-based proteomics approach. A 1∶1 mixture of Triton X-100 lysates from “Heavy” (H; Arg4/Lys8)-labeled ethanol-stimulated tachyzoites or "Light" (L; Arg0/Lys0)-labeled non-stimulated parasites was generated, and a TiO 2 -enriched phosphopeptide sample of H/L-labeled Toxoplasma invasion motor complexes was prepared and analysed by LC-MS/MS on an LTQ-Orbitrap instrument. Mascot and MaxQuant search engines facilitated subsequent manual identification, phosphosite localization and quantification of proteins or peptides as detailed in materials and mathods. B ) Sypro Ruby-stained SDS-PAGE separation of the relative amounts of light (lane 1) or heavy (lane 2) Triton X-100 whole protein extracts are shown. Intact tachyzoite invasion motor complexes comprising the five major components MyoA, GAP50, GAP45 and MLC1 were precipitated from a 1∶1 H/L mixture by GAP45-specific immuno-affinity chromatography (lane 3).

    Article Snippet: The nano HPLC was coupled on-line to an LTQ-Orbitrap mass spectrometer equipped with a nanoelectrospray ion source (Thermo Fisher Scientific) for automated MS/MS.

    Techniques: Flow Cytometry, Labeling, Generated, Liquid Chromatography with Mass Spectroscopy, Mass Spectrometry, Staining, SDS Page, Affinity Chromatography

    A schematic workflow illustrating the steps involved in the differential analysis of RA and OA synovial fluid proteome. Proteins from RA and OA synovial fluid were extracted and depleted to remove the 14 most abundant proteins using multiple affinity removal system, Human-14. The depleted protein from RA and OA were then digested with trypsin and labeled with iTRAQ reagents, 117 and 116 respectively. The labeled samples were pooled and subjected to fractionation using strong cation exchange chromatography. The fractions were then analyzed on a LTQ-Orbitrap Velos mass spectrometer. The MS/MS data obtained was searched against Human RefSeq 50 database using Sequest and Mascot search algorithms. Validation of the iTRAQ quantitation data was carried out using multiple reaction monitoring and Western blot.

    Journal: Clinical proteomics

    Article Title: Differential proteomic analysis of synovial fluid from rheumatoid arthritis and osteoarthritis patients

    doi: 10.1186/1559-0275-11-1

    Figure Lengend Snippet: A schematic workflow illustrating the steps involved in the differential analysis of RA and OA synovial fluid proteome. Proteins from RA and OA synovial fluid were extracted and depleted to remove the 14 most abundant proteins using multiple affinity removal system, Human-14. The depleted protein from RA and OA were then digested with trypsin and labeled with iTRAQ reagents, 117 and 116 respectively. The labeled samples were pooled and subjected to fractionation using strong cation exchange chromatography. The fractions were then analyzed on a LTQ-Orbitrap Velos mass spectrometer. The MS/MS data obtained was searched against Human RefSeq 50 database using Sequest and Mascot search algorithms. Validation of the iTRAQ quantitation data was carried out using multiple reaction monitoring and Western blot.

    Article Snippet: LC-MS/MS analysis Tandem mass spectrometric analysis of the iTRAQ labeled peptides were carried out using LTQ-Orbitrap Velos mass spectrometer (Thermo Scientific, Bremen, Germany) interfaced with Easy nanoLC II (previously Proxeon, Thermo Scientific, Bremen, Germany).

    Techniques: Labeling, Fractionation, Chromatography, Mass Spectrometry, Quantitation Assay, Western Blot