high performance liquid chromatography tandem mass spectrometry Search Results


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  • 99
    Thermo Fisher ultimate 3000 hplc system
    Ultimate 3000 Hplc System, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 5055 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Agilent technologies 1100 hplc system
    ( A ) Coomassie blue staining of the purified region-1 and region-2 of mPCSK9 proteins. Region-1 (residues 152–351) and region-2 (residues 249–452) of mouse catalytic domain were cloned and expressed by pET-3a plasmids. The proteins were reduced and purified by a reverse-phase high-performance liquid chromatography <t>(HPLC)</t> on an Agilent <t>1100</t> HPLC system. The purified proteins were analyzed by SDS/PAGE and stained with Coomassie blue. The positions of expressed proteins are shown; ( B ) MALDI mass spectrometry on Region-1. The molecular mass of purified PCSK9 region-1 in fully reduced form (PCSK9-1R). The molecular mass of PCSK9 region-1 as determined by MALDI was 21,201 Da; ( C ) MALDI mass spectrometry on Region-2. The molecular mass of purified PCSK9 region-2 in fully reduced form (PCSK9-2R). The molecular mass of PCSK9 region-2 as determined by MALDI was 21,355 Da.
    1100 Hplc System, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 94/100, based on 8933 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Agilent technologies 1200 hplc system
    The catumaxomab sample was analyzed using <t>RP-HPLC</t> mass spectrometry using a PLRP-S column (Varian) with an acetonitrile gradient from 27% to 42% over 40 minutes on a <t>1200</t> HPLC system (Agilent) for separation, coupled with an Agilent 6220 ESI-TOF mass
    1200 Hplc System, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 94/100, based on 8192 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Agilent technologies hplc system
    Trans -acylation reaction assays of various AT4 FkbB mutants after substitution of Val187 with Asp, or Cys, Ile, Lys, Trp, and Phe with allmal- and ethmal-CoA as substrates and the Production of <t>FK506</t> and FK520 in WT, YN06-01, and YN06-02. (A) The change of transferring allmal/ethmal unit to ACP10 FkbA in trans -acylation reactions by AT4 FkbB and its mutants V187D, V187C, V187I, V187K, V187W, and V187F. <t>HPLC</t> analyses of transferring allmal-CoA : ethmal-CoA = 1:1 to holo-ACP10 FkbA in the presence of V187K (B) and in the absence of AT4 FkbB (D) . MS analyses of transferring allmal-CoA : ethmal-CoA = 1:1 to holo-ACP10 FkbA in the presence of V187K (C) and in the absence of AT4 FkbB (E) . The peaks were assigned as follows: b 0 , holo-ACP10 FkbA ; b 1 , allmal-ACP10 FkbA ; b 2 , ethmal-ACP10 FkbA . (F) Production of FK506 and FK520 in WT, YN06-01 and YN06-02. Analyses of variance were conducted to determine the difference of WT and mutants using SPSS 20. The LSD multiple range tests were evaluated for significant differences among WT and mutants ( P
    Hplc System, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 95/100, based on 8784 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Shimadzu Corporation shimadzu hplc system
    Representative chromatograms of LASSBio-1773 (7) (20 µM) and its metabolite formed by incubation with rat plasma. Notes: ( A ) Incubation in rat plasma at 0 minute; ( B ) incubation in rat plasma at 1 hour, and appearance of metabolite (M) formed at 4.57 minutes; ( C ) <t>HPLC/MS</t> ion scan of M [M-H] − at m/z 363.09 (retention time at 4.57); ( D ) UV chromatogram of M at 4.57 minutes; ( E ) synthesized M standard (10); ( F ) UV chromatogram of standard (10) at 4.57 minutes; ( G ) co-injection of M standard (10) in 1 hour rat plasma experiment; ( H ) compound and internal standard (IS) in acetonitrile. IS: internal standard (eg, biphenyl-4-carboxylate methyl, C =20 µM). Apparatus: <t>Shimadzu</t> – LC20AD, column: Kromasil 100-5 C18 250 to 4.6 mm. Mobile phase: 50% acetonitrile, 50% water-gradient, 0.1% formic acid, flow: 1 mL/min. Detector: SPD-M20A (Diode Array); wavelength: 254 nm. The mass spectrometer analyses were performed by the mass spectrometer model Esquire 6000 – ESI Ion Trap MSn System Bruker Daltonics (LASSBio ® -UFRJ). ‘M’ refers to the metabolite of the compound. Abbreviations: HPLC/MS, high performance liquid chromatography/mass spectrometry; min, minutes; Intens, Intensity.
    Shimadzu Hplc System, supplied by Shimadzu Corporation, used in various techniques. Bioz Stars score: 95/100, based on 6431 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher q exactive mass spectrometer
    Representative chromatograms of LASSBio-1773 (7) (20 µM) and its metabolite formed by incubation with rat plasma. Notes: ( A ) Incubation in rat plasma at 0 minute; ( B ) incubation in rat plasma at 1 hour, and appearance of metabolite (M) formed at 4.57 minutes; ( C ) <t>HPLC/MS</t> ion scan of M [M-H] − at m/z 363.09 (retention time at 4.57); ( D ) UV chromatogram of M at 4.57 minutes; ( E ) synthesized M standard (10); ( F ) UV chromatogram of standard (10) at 4.57 minutes; ( G ) co-injection of M standard (10) in 1 hour rat plasma experiment; ( H ) compound and internal standard (IS) in acetonitrile. IS: internal standard (eg, biphenyl-4-carboxylate methyl, C =20 µM). Apparatus: <t>Shimadzu</t> – LC20AD, column: Kromasil 100-5 C18 250 to 4.6 mm. Mobile phase: 50% acetonitrile, 50% water-gradient, 0.1% formic acid, flow: 1 mL/min. Detector: SPD-M20A (Diode Array); wavelength: 254 nm. The mass spectrometer analyses were performed by the mass spectrometer model Esquire 6000 – ESI Ion Trap MSn System Bruker Daltonics (LASSBio ® -UFRJ). ‘M’ refers to the metabolite of the compound. Abbreviations: HPLC/MS, high performance liquid chromatography/mass spectrometry; min, minutes; Intens, Intensity.
    Q Exactive Mass Spectrometer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 11445 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher surveyor hplc system
    Representative chromatograms of LASSBio-1773 (7) (20 µM) and its metabolite formed by incubation with rat plasma. Notes: ( A ) Incubation in rat plasma at 0 minute; ( B ) incubation in rat plasma at 1 hour, and appearance of metabolite (M) formed at 4.57 minutes; ( C ) <t>HPLC/MS</t> ion scan of M [M-H] − at m/z 363.09 (retention time at 4.57); ( D ) UV chromatogram of M at 4.57 minutes; ( E ) synthesized M standard (10); ( F ) UV chromatogram of standard (10) at 4.57 minutes; ( G ) co-injection of M standard (10) in 1 hour rat plasma experiment; ( H ) compound and internal standard (IS) in acetonitrile. IS: internal standard (eg, biphenyl-4-carboxylate methyl, C =20 µM). Apparatus: <t>Shimadzu</t> – LC20AD, column: Kromasil 100-5 C18 250 to 4.6 mm. Mobile phase: 50% acetonitrile, 50% water-gradient, 0.1% formic acid, flow: 1 mL/min. Detector: SPD-M20A (Diode Array); wavelength: 254 nm. The mass spectrometer analyses were performed by the mass spectrometer model Esquire 6000 – ESI Ion Trap MSn System Bruker Daltonics (LASSBio ® -UFRJ). ‘M’ refers to the metabolite of the compound. Abbreviations: HPLC/MS, high performance liquid chromatography/mass spectrometry; min, minutes; Intens, Intensity.
    Surveyor Hplc System, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1380 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher hplc
    Representative chromatograms of LASSBio-1773 (7) (20 µM) and its metabolite formed by incubation with rat plasma. Notes: ( A ) Incubation in rat plasma at 0 minute; ( B ) incubation in rat plasma at 1 hour, and appearance of metabolite (M) formed at 4.57 minutes; ( C ) <t>HPLC/MS</t> ion scan of M [M-H] − at m/z 363.09 (retention time at 4.57); ( D ) UV chromatogram of M at 4.57 minutes; ( E ) synthesized M standard (10); ( F ) UV chromatogram of standard (10) at 4.57 minutes; ( G ) co-injection of M standard (10) in 1 hour rat plasma experiment; ( H ) compound and internal standard (IS) in acetonitrile. IS: internal standard (eg, biphenyl-4-carboxylate methyl, C =20 µM). Apparatus: <t>Shimadzu</t> – LC20AD, column: Kromasil 100-5 C18 250 to 4.6 mm. Mobile phase: 50% acetonitrile, 50% water-gradient, 0.1% formic acid, flow: 1 mL/min. Detector: SPD-M20A (Diode Array); wavelength: 254 nm. The mass spectrometer analyses were performed by the mass spectrometer model Esquire 6000 – ESI Ion Trap MSn System Bruker Daltonics (LASSBio ® -UFRJ). ‘M’ refers to the metabolite of the compound. Abbreviations: HPLC/MS, high performance liquid chromatography/mass spectrometry; min, minutes; Intens, Intensity.
    Hplc, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 5329 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Agilent technologies 1260 hplc system
    Representative chromatograms of LASSBio-1773 (7) (20 µM) and its metabolite formed by incubation with rat plasma. Notes: ( A ) Incubation in rat plasma at 0 minute; ( B ) incubation in rat plasma at 1 hour, and appearance of metabolite (M) formed at 4.57 minutes; ( C ) <t>HPLC/MS</t> ion scan of M [M-H] − at m/z 363.09 (retention time at 4.57); ( D ) UV chromatogram of M at 4.57 minutes; ( E ) synthesized M standard (10); ( F ) UV chromatogram of standard (10) at 4.57 minutes; ( G ) co-injection of M standard (10) in 1 hour rat plasma experiment; ( H ) compound and internal standard (IS) in acetonitrile. IS: internal standard (eg, biphenyl-4-carboxylate methyl, C =20 µM). Apparatus: <t>Shimadzu</t> – LC20AD, column: Kromasil 100-5 C18 250 to 4.6 mm. Mobile phase: 50% acetonitrile, 50% water-gradient, 0.1% formic acid, flow: 1 mL/min. Detector: SPD-M20A (Diode Array); wavelength: 254 nm. The mass spectrometer analyses were performed by the mass spectrometer model Esquire 6000 – ESI Ion Trap MSn System Bruker Daltonics (LASSBio ® -UFRJ). ‘M’ refers to the metabolite of the compound. Abbreviations: HPLC/MS, high performance liquid chromatography/mass spectrometry; min, minutes; Intens, Intensity.
    1260 Hplc System, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 94/100, based on 1925 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher ultimate 3000 nano hplc system
    Representative chromatograms of LASSBio-1773 (7) (20 µM) and its metabolite formed by incubation with rat plasma. Notes: ( A ) Incubation in rat plasma at 0 minute; ( B ) incubation in rat plasma at 1 hour, and appearance of metabolite (M) formed at 4.57 minutes; ( C ) <t>HPLC/MS</t> ion scan of M [M-H] − at m/z 363.09 (retention time at 4.57); ( D ) UV chromatogram of M at 4.57 minutes; ( E ) synthesized M standard (10); ( F ) UV chromatogram of standard (10) at 4.57 minutes; ( G ) co-injection of M standard (10) in 1 hour rat plasma experiment; ( H ) compound and internal standard (IS) in acetonitrile. IS: internal standard (eg, biphenyl-4-carboxylate methyl, C =20 µM). Apparatus: <t>Shimadzu</t> – LC20AD, column: Kromasil 100-5 C18 250 to 4.6 mm. Mobile phase: 50% acetonitrile, 50% water-gradient, 0.1% formic acid, flow: 1 mL/min. Detector: SPD-M20A (Diode Array); wavelength: 254 nm. The mass spectrometer analyses were performed by the mass spectrometer model Esquire 6000 – ESI Ion Trap MSn System Bruker Daltonics (LASSBio ® -UFRJ). ‘M’ refers to the metabolite of the compound. Abbreviations: HPLC/MS, high performance liquid chromatography/mass spectrometry; min, minutes; Intens, Intensity.
    Ultimate 3000 Nano Hplc System, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1317 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    pcpe2  (Abcam)
    90
    Abcam pcpe2
    Mass spectrometric analysis and the ratio of mature to pro-apoA-I in HDL from <t>PCPE2</t> −/− mouse plasma. A and B , molecular masses of HDL apoA-I after maximum entropy transformation of the electrospray spectrum of FPLC purified HDL apoA-I from chow-fed C57BL/6 mice ( A ) and PCPE2 −/− mice ( B ). The most abundant mass in each spectrum was identical within the ±1.4 Da S.D. to the mass of mature mouse apoA-I at 27,950 Da. The minor spectrum peak is consistent with the mass of mouse pro-apoA-I at 28,815 Da. C and D , the relative intensity of apoA-I tryptic peptides from LC-MS/MS analysis. ApoA-I from whole mouse plasma was separated by 12% SDS-PAGE followed by in-gel trypsin digest and then analyzed for the N-terminal tryptic pro-apoA-I peptide and mature apoA-I peptide: C57BL/6 mouse plasma ( C ) and PCPE2 −/− mouse plasma ( D ) as described under “Experimental Procedures.” Peptides were separated by reverse-phase HPLC and analyzed for tryptic peptide of the N-terminal T1 for mature peptide ( m / z 1132.48) and the T1 propeptide ( m / z 1995.88), shown in selective ion chromatograms. The mature murine T1 tryptic peptide has the amino acid sequence DEPQSQWDK, and the unprocessed pro-form has the sequence WHVWQQDEPQSQWDK.
    Pcpe2, supplied by Abcam, used in various techniques. Bioz Stars score: 90/100, based on 168 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    JCR Pharmaceuticals atb100
    Coformulation with AT1001 results in greater tissue levels and biodistribution of <t>ATB100</t> in Gla KO (knockout) mice. ( a ) Twelve-week-old male Gla KO mice were given a single 30-minute intravenous infusion of 1 or 3 mg/kg ATB100, either alone or coformulated with 1.22, 3.66, or 12.2 mg/kg AT1001 (equivalent to 1, 3, and 10 mg/kg free base, respectively). Skin, heart, and kidney were collected 7 days postinfusion, and α-galactosidase A (α-Gal A) activities were determined. Each bar represents the mean ± SEM of five to six mice per group. * P
    Atb100, supplied by JCR Pharmaceuticals, used in various techniques. Bioz Stars score: 90/100, based on 142 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Agilent technologies lc ms ms analysis
    Coformulation with AT1001 results in greater tissue levels and biodistribution of <t>ATB100</t> in Gla KO (knockout) mice. ( a ) Twelve-week-old male Gla KO mice were given a single 30-minute intravenous infusion of 1 or 3 mg/kg ATB100, either alone or coformulated with 1.22, 3.66, or 12.2 mg/kg AT1001 (equivalent to 1, 3, and 10 mg/kg free base, respectively). Skin, heart, and kidney were collected 7 days postinfusion, and α-galactosidase A (α-Gal A) activities were determined. Each bar represents the mean ± SEM of five to six mice per group. * P
    Lc Ms Ms Analysis, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 94/100, based on 2045 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Agilent technologies agilent 1260 hplc
    Coformulation with AT1001 results in greater tissue levels and biodistribution of <t>ATB100</t> in Gla KO (knockout) mice. ( a ) Twelve-week-old male Gla KO mice were given a single 30-minute intravenous infusion of 1 or 3 mg/kg ATB100, either alone or coformulated with 1.22, 3.66, or 12.2 mg/kg AT1001 (equivalent to 1, 3, and 10 mg/kg free base, respectively). Skin, heart, and kidney were collected 7 days postinfusion, and α-galactosidase A (α-Gal A) activities were determined. Each bar represents the mean ± SEM of five to six mice per group. * P
    Agilent 1260 Hplc, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 99/100, based on 1268 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    ( A ) Coomassie blue staining of the purified region-1 and region-2 of mPCSK9 proteins. Region-1 (residues 152–351) and region-2 (residues 249–452) of mouse catalytic domain were cloned and expressed by pET-3a plasmids. The proteins were reduced and purified by a reverse-phase high-performance liquid chromatography (HPLC) on an Agilent 1100 HPLC system. The purified proteins were analyzed by SDS/PAGE and stained with Coomassie blue. The positions of expressed proteins are shown; ( B ) MALDI mass spectrometry on Region-1. The molecular mass of purified PCSK9 region-1 in fully reduced form (PCSK9-1R). The molecular mass of PCSK9 region-1 as determined by MALDI was 21,201 Da; ( C ) MALDI mass spectrometry on Region-2. The molecular mass of purified PCSK9 region-2 in fully reduced form (PCSK9-2R). The molecular mass of PCSK9 region-2 as determined by MALDI was 21,355 Da.

    Journal: International Journal of Molecular Sciences

    Article Title: Non-Native Conformational Isomers of the Catalytic Domain of PCSK9 Induce an Immune Response, Reduce Lipids and Increase LDL Receptor Levels

    doi: 10.3390/ijms19020640

    Figure Lengend Snippet: ( A ) Coomassie blue staining of the purified region-1 and region-2 of mPCSK9 proteins. Region-1 (residues 152–351) and region-2 (residues 249–452) of mouse catalytic domain were cloned and expressed by pET-3a plasmids. The proteins were reduced and purified by a reverse-phase high-performance liquid chromatography (HPLC) on an Agilent 1100 HPLC system. The purified proteins were analyzed by SDS/PAGE and stained with Coomassie blue. The positions of expressed proteins are shown; ( B ) MALDI mass spectrometry on Region-1. The molecular mass of purified PCSK9 region-1 in fully reduced form (PCSK9-1R). The molecular mass of PCSK9 region-1 as determined by MALDI was 21,201 Da; ( C ) MALDI mass spectrometry on Region-2. The molecular mass of purified PCSK9 region-2 in fully reduced form (PCSK9-2R). The molecular mass of PCSK9 region-2 as determined by MALDI was 21,355 Da.

    Article Snippet: The authors used a reverse-phase high-performance liquid chromatography (HPLC) on an Agilent 1100 HPLC system (Column ZORBAX 3000 SB-C18, 9.4 mm × 25 cm) to purify the proteins.

    Techniques: Staining, Purification, Clone Assay, Positron Emission Tomography, High Performance Liquid Chromatography, SDS Page, Mass Spectrometry

    aps1 and pgm leaves accumulate WT ADPG content. (A) HPLC-MS/MS detection of ADPG in WT, aps1 and pgm leaves. Upper panels: Total ion chromatograms (TIC) of extracts from the indicated plants in which the selected fragmentation parent ion was 587.8 m/z. Middle panels: Extracted ion chromatograms (EIC) in which the selected ion for fragmentation of the parent ion was 346.1 m/z. Lower panels: Mass spectra (MS2) obtained from fragmentation of parent ion. ADPG was measured using an Agilent 1100 HPLC fitted with a Xbridge C18 column (100×3.0 mm I.D. particle size 3.5 µm) coupled to a MSD-Trap spectrometer (Agilent) (see Materials and Methods for further details). (B) ADPG content in WT, aps1, pgm and aps1/pgm leaves. Plants were simultaneously grown either in soil or solid MS. Leaves from 4-weeks old WT, aps1, pgm and aps1/pgm plants were simultaneously harvested after 10 h of illumination. ADPG was simultaneouly extracted from leaves of WT, aps1, pgm and aps1/pgm plants and content was simultaneously measured by HPLC-MS/MS as described in Materials and Methods . Note that, consistent with [15] , leaves of aps1, pgm and aps1/pgm plants accumulated WT ADPG content. Values represent the mean ±SD of determinations on three independent samples.

    Journal: PLoS ONE

    Article Title: HPLC-MS/MS Analyses Show That the Near-Starchless aps1 and pgm Leaves Accumulate Wild Type Levels of ADPglucose: Further Evidence for the Occurrence of Important ADPglucose Biosynthetic Pathway(s) Alternative to the pPGI-pPGM-AGP Pathway

    doi: 10.1371/journal.pone.0104997

    Figure Lengend Snippet: aps1 and pgm leaves accumulate WT ADPG content. (A) HPLC-MS/MS detection of ADPG in WT, aps1 and pgm leaves. Upper panels: Total ion chromatograms (TIC) of extracts from the indicated plants in which the selected fragmentation parent ion was 587.8 m/z. Middle panels: Extracted ion chromatograms (EIC) in which the selected ion for fragmentation of the parent ion was 346.1 m/z. Lower panels: Mass spectra (MS2) obtained from fragmentation of parent ion. ADPG was measured using an Agilent 1100 HPLC fitted with a Xbridge C18 column (100×3.0 mm I.D. particle size 3.5 µm) coupled to a MSD-Trap spectrometer (Agilent) (see Materials and Methods for further details). (B) ADPG content in WT, aps1, pgm and aps1/pgm leaves. Plants were simultaneously grown either in soil or solid MS. Leaves from 4-weeks old WT, aps1, pgm and aps1/pgm plants were simultaneously harvested after 10 h of illumination. ADPG was simultaneouly extracted from leaves of WT, aps1, pgm and aps1/pgm plants and content was simultaneously measured by HPLC-MS/MS as described in Materials and Methods . Note that, consistent with [15] , leaves of aps1, pgm and aps1/pgm plants accumulated WT ADPG content. Values represent the mean ±SD of determinations on three independent samples.

    Article Snippet: ADPG content in leaves of plants cultured on soil was measured in the Research Support Service at the Public University of Navarra using an Agilent 1100 HPLC fitted with a Xbridge C18 column (100×3.0 mm I.D. particle size 3.5 µm) coupled to a MSD-Trap spectrometer (Agilent).

    Techniques: High Performance Liquid Chromatography, Mass Spectrometry

    The catumaxomab sample was analyzed using RP-HPLC mass spectrometry using a PLRP-S column (Varian) with an acetonitrile gradient from 27% to 42% over 40 minutes on a 1200 HPLC system (Agilent) for separation, coupled with an Agilent 6220 ESI-TOF mass

    Journal: mAbs

    Article Title: Structural and functional characterization of the trifunctional antibody catumaxomab

    doi:

    Figure Lengend Snippet: The catumaxomab sample was analyzed using RP-HPLC mass spectrometry using a PLRP-S column (Varian) with an acetonitrile gradient from 27% to 42% over 40 minutes on a 1200 HPLC system (Agilent) for separation, coupled with an Agilent 6220 ESI-TOF mass

    Article Snippet: The peptides were separated on a reversed-phase HPLC using an Agilent 1200 HPLC system equipped with a diode-array detector, an auto-sampler, and a temperature controlled column compartment (Agilent, Waldbronn, Germany).

    Techniques: High Performance Liquid Chromatography, Mass Spectrometry

    ( a ) Representative HPLC-MS chromatogram (HILIC-ESI-MS) from site U1357, 7.15 mbsf, showing the presence of major intact polar lipid isoprenoidal glycerol dialkyl glycerol tetraethers (IPL-GDGTs) and bacterial lipids in the mass range m/z 1200–1800. The HPLC-MS chromatogram is depicted as a density map showing retention time on the x axis, m/z on the y axis and the relative peak intensity by grey shading. ( b ) Structural variety in the head groups and biphytanyl moieties of the detected IPL-GDGTs. Note that during the ether cleavage reaction the hydroxyl group in the biphytane (bp) side chain becomes dehydrated and forms biphytene (bp0:1). Position of the hydroxyl group and double bond were not determined. 1G, monohexose; 2G, dihexose; PG, phosphatidyl glycerol; HPH, hexose-phoshohexose; cren, crenarchaeol.

    Journal: The ISME Journal

    Article Title: Acetoclastic Methanosaeta are dominant methanogens in organic-rich Antarctic marine sediments

    doi: 10.1038/ismej.2017.150

    Figure Lengend Snippet: ( a ) Representative HPLC-MS chromatogram (HILIC-ESI-MS) from site U1357, 7.15 mbsf, showing the presence of major intact polar lipid isoprenoidal glycerol dialkyl glycerol tetraethers (IPL-GDGTs) and bacterial lipids in the mass range m/z 1200–1800. The HPLC-MS chromatogram is depicted as a density map showing retention time on the x axis, m/z on the y axis and the relative peak intensity by grey shading. ( b ) Structural variety in the head groups and biphytanyl moieties of the detected IPL-GDGTs. Note that during the ether cleavage reaction the hydroxyl group in the biphytane (bp) side chain becomes dehydrated and forms biphytene (bp0:1). Position of the hydroxyl group and double bond were not determined. 1G, monohexose; 2G, dihexose; PG, phosphatidyl glycerol; HPH, hexose-phoshohexose; cren, crenarchaeol.

    Article Snippet: Identification and quantification of IPLs and core GDGTs was achieved on a Agilent 1200 series HPLC system coupled to an Agilent 6520 accurate-mass quadrupole time-of-flight mass spectrometer (Summons laboratory, Agilent, Santa Clara, CA, USA) and Dionex Ultimate 3000 UHPLC (Thermo Fisher Scientific, Waltham, MA, USA) coupled to a Bruker maXis ultra-high-resolution orthogonal acceleration quadrupole time-of-flight tandem MS2 instrument (Hinrichs laboratory, Bruker Daltonics Inc., Billerica, MA, USA) following previously described protocols ( ; ; ; ).

    Techniques: High Performance Liquid Chromatography, Mass Spectrometry, Hydrophilic Interaction Liquid Chromatography

    A C-type lectin was identified by TripleTOF 5600 MS. The mass spectrometer was fitted with a nanospray III ion source and coupled to an Agilent 1200 HPLC. A typical spectrum of a single peptide [(K)IFNELKAWK(D), m/z, 574.83] is shown (A). The mass and the amino acid sequence of the C-type lectin are shown and the sequence obtained from spectrometry is underlined (B). This protein was identified as Q7T228 in Uniprot database with coverage of 94%.

    Journal: PLoS ONE

    Article Title: A C-Type Lectin from Bothrops jararacussu Venom Disrupts Staphylococcal Biofilms

    doi: 10.1371/journal.pone.0120514

    Figure Lengend Snippet: A C-type lectin was identified by TripleTOF 5600 MS. The mass spectrometer was fitted with a nanospray III ion source and coupled to an Agilent 1200 HPLC. A typical spectrum of a single peptide [(K)IFNELKAWK(D), m/z, 574.83] is shown (A). The mass and the amino acid sequence of the C-type lectin are shown and the sequence obtained from spectrometry is underlined (B). This protein was identified as Q7T228 in Uniprot database with coverage of 94%.

    Article Snippet: Using a TripleTOF 5600 mass spectrometer coupled to an Agilent 1200 HPLC, we identified a C-type lectin from Bothrops jararacussu with 94% coverage.

    Techniques: Mass Spectrometry, High Performance Liquid Chromatography, Sequencing

    Trans -acylation reaction assays of various AT4 FkbB mutants after substitution of Val187 with Asp, or Cys, Ile, Lys, Trp, and Phe with allmal- and ethmal-CoA as substrates and the Production of FK506 and FK520 in WT, YN06-01, and YN06-02. (A) The change of transferring allmal/ethmal unit to ACP10 FkbA in trans -acylation reactions by AT4 FkbB and its mutants V187D, V187C, V187I, V187K, V187W, and V187F. HPLC analyses of transferring allmal-CoA : ethmal-CoA = 1:1 to holo-ACP10 FkbA in the presence of V187K (B) and in the absence of AT4 FkbB (D) . MS analyses of transferring allmal-CoA : ethmal-CoA = 1:1 to holo-ACP10 FkbA in the presence of V187K (C) and in the absence of AT4 FkbB (E) . The peaks were assigned as follows: b 0 , holo-ACP10 FkbA ; b 1 , allmal-ACP10 FkbA ; b 2 , ethmal-ACP10 FkbA . (F) Production of FK506 and FK520 in WT, YN06-01 and YN06-02. Analyses of variance were conducted to determine the difference of WT and mutants using SPSS 20. The LSD multiple range tests were evaluated for significant differences among WT and mutants ( P

    Journal: Frontiers in Microbiology

    Article Title: Substrate Specificity of Acyltransferase Domains for Efficient Transfer of Acyl Groups

    doi: 10.3389/fmicb.2018.01840

    Figure Lengend Snippet: Trans -acylation reaction assays of various AT4 FkbB mutants after substitution of Val187 with Asp, or Cys, Ile, Lys, Trp, and Phe with allmal- and ethmal-CoA as substrates and the Production of FK506 and FK520 in WT, YN06-01, and YN06-02. (A) The change of transferring allmal/ethmal unit to ACP10 FkbA in trans -acylation reactions by AT4 FkbB and its mutants V187D, V187C, V187I, V187K, V187W, and V187F. HPLC analyses of transferring allmal-CoA : ethmal-CoA = 1:1 to holo-ACP10 FkbA in the presence of V187K (B) and in the absence of AT4 FkbB (D) . MS analyses of transferring allmal-CoA : ethmal-CoA = 1:1 to holo-ACP10 FkbA in the presence of V187K (C) and in the absence of AT4 FkbB (E) . The peaks were assigned as follows: b 0 , holo-ACP10 FkbA ; b 1 , allmal-ACP10 FkbA ; b 2 , ethmal-ACP10 FkbA . (F) Production of FK506 and FK520 in WT, YN06-01 and YN06-02. Analyses of variance were conducted to determine the difference of WT and mutants using SPSS 20. The LSD multiple range tests were evaluated for significant differences among WT and mutants ( P

    Article Snippet: The concentration of FK506 and FK520 was determined using an HPLC system (Agilent Series 1100, Agilent) equipped with a SB-C18 column (150 mm × 2.1 mm, Agilent).

    Techniques: Transferring, High Performance Liquid Chromatography, Mass Spectrometry

    Representative chromatograms of LASSBio-1773 (7) (20 µM) and its metabolite formed by incubation with rat plasma. Notes: ( A ) Incubation in rat plasma at 0 minute; ( B ) incubation in rat plasma at 1 hour, and appearance of metabolite (M) formed at 4.57 minutes; ( C ) HPLC/MS ion scan of M [M-H] − at m/z 363.09 (retention time at 4.57); ( D ) UV chromatogram of M at 4.57 minutes; ( E ) synthesized M standard (10); ( F ) UV chromatogram of standard (10) at 4.57 minutes; ( G ) co-injection of M standard (10) in 1 hour rat plasma experiment; ( H ) compound and internal standard (IS) in acetonitrile. IS: internal standard (eg, biphenyl-4-carboxylate methyl, C =20 µM). Apparatus: Shimadzu – LC20AD, column: Kromasil 100-5 C18 250 to 4.6 mm. Mobile phase: 50% acetonitrile, 50% water-gradient, 0.1% formic acid, flow: 1 mL/min. Detector: SPD-M20A (Diode Array); wavelength: 254 nm. The mass spectrometer analyses were performed by the mass spectrometer model Esquire 6000 – ESI Ion Trap MSn System Bruker Daltonics (LASSBio ® -UFRJ). ‘M’ refers to the metabolite of the compound. Abbreviations: HPLC/MS, high performance liquid chromatography/mass spectrometry; min, minutes; Intens, Intensity.

    Journal: Drug Design, Development and Therapy

    Article Title: Synthesis, solubility, plasma stability, and pharmacological evaluation of novel sulfonylhydrazones designed as anti-diabetic agents

    doi: 10.2147/DDDT.S108327

    Figure Lengend Snippet: Representative chromatograms of LASSBio-1773 (7) (20 µM) and its metabolite formed by incubation with rat plasma. Notes: ( A ) Incubation in rat plasma at 0 minute; ( B ) incubation in rat plasma at 1 hour, and appearance of metabolite (M) formed at 4.57 minutes; ( C ) HPLC/MS ion scan of M [M-H] − at m/z 363.09 (retention time at 4.57); ( D ) UV chromatogram of M at 4.57 minutes; ( E ) synthesized M standard (10); ( F ) UV chromatogram of standard (10) at 4.57 minutes; ( G ) co-injection of M standard (10) in 1 hour rat plasma experiment; ( H ) compound and internal standard (IS) in acetonitrile. IS: internal standard (eg, biphenyl-4-carboxylate methyl, C =20 µM). Apparatus: Shimadzu – LC20AD, column: Kromasil 100-5 C18 250 to 4.6 mm. Mobile phase: 50% acetonitrile, 50% water-gradient, 0.1% formic acid, flow: 1 mL/min. Detector: SPD-M20A (Diode Array); wavelength: 254 nm. The mass spectrometer analyses were performed by the mass spectrometer model Esquire 6000 – ESI Ion Trap MSn System Bruker Daltonics (LASSBio ® -UFRJ). ‘M’ refers to the metabolite of the compound. Abbreviations: HPLC/MS, high performance liquid chromatography/mass spectrometry; min, minutes; Intens, Intensity.

    Article Snippet: HPLC-UV analysis The organic fraction was analyzed with the Shimadzu Prominence HPLC system (Shimadzu, Tokyo, Japan) consisting of a vacuum degasser (DGU-20A5), a binary pump (LC-20AD), an autosampler (SIL-20A), UV/VIS Photodiode Array Detector (SPD-M20A), and fitted with a guard column (CLC G-ODS) and Kromasil 100-5C18 (4.6×250 mm) analytical column running at room temperature.

    Techniques: Incubation, High Performance Liquid Chromatography, Mass Spectrometry, Synthesized, Injection, Flow Cytometry

    Representative chromatograms of LASSBio-1771 (8) (20 µM) and its metabolite formed by incubation with rat plasma. Notes: ( A ) Incubation in rat plasma at 0 minute; ( B ) incubation in rat plasma at 1 hour, and appearance of metabolite (M) formed at 4.57 minutes; ( C ) HPLC/MS ion scan of M [M-H] − at m/z 363.09 (retention time at 4.57); ( D ) UV chromatogram of M at 4.57 minutes; ( E ) co-injection of synthesized standard M (10) in 1 hour rat plasma experiment (M); ( F ) UV chromatogram of standard (10) at 4.57 minutes. IS: internal standard (eg, methyl biphenyl-4-carboxylate, C =20 µM). Apparatus: Shimadzu – LC20AD, column: Kromasil 100-5 C18 250 to 4.6 mm. Mobile phase: 50% acetonitrile, 50% water-gradient, 0.1% formic acid, flow: 1 mL/min. Detector: SPD-M20A (Diode Array); wavelength: 254 nm. The mass spectrometer analyses were performed by the mass spectrometer model Esquire 6000 – ESI Ion Trap MSn System Bruker Daltonics (LASSBio ® -UFRJ). Abbreviations: HPLC/MS, high performance liquid chromatography/mass spectrometry; min, minutes; UV, ultraviolet spectroscopy; Intens, Intensity.

    Journal: Drug Design, Development and Therapy

    Article Title: Synthesis, solubility, plasma stability, and pharmacological evaluation of novel sulfonylhydrazones designed as anti-diabetic agents

    doi: 10.2147/DDDT.S108327

    Figure Lengend Snippet: Representative chromatograms of LASSBio-1771 (8) (20 µM) and its metabolite formed by incubation with rat plasma. Notes: ( A ) Incubation in rat plasma at 0 minute; ( B ) incubation in rat plasma at 1 hour, and appearance of metabolite (M) formed at 4.57 minutes; ( C ) HPLC/MS ion scan of M [M-H] − at m/z 363.09 (retention time at 4.57); ( D ) UV chromatogram of M at 4.57 minutes; ( E ) co-injection of synthesized standard M (10) in 1 hour rat plasma experiment (M); ( F ) UV chromatogram of standard (10) at 4.57 minutes. IS: internal standard (eg, methyl biphenyl-4-carboxylate, C =20 µM). Apparatus: Shimadzu – LC20AD, column: Kromasil 100-5 C18 250 to 4.6 mm. Mobile phase: 50% acetonitrile, 50% water-gradient, 0.1% formic acid, flow: 1 mL/min. Detector: SPD-M20A (Diode Array); wavelength: 254 nm. The mass spectrometer analyses were performed by the mass spectrometer model Esquire 6000 – ESI Ion Trap MSn System Bruker Daltonics (LASSBio ® -UFRJ). Abbreviations: HPLC/MS, high performance liquid chromatography/mass spectrometry; min, minutes; UV, ultraviolet spectroscopy; Intens, Intensity.

    Article Snippet: HPLC-UV analysis The organic fraction was analyzed with the Shimadzu Prominence HPLC system (Shimadzu, Tokyo, Japan) consisting of a vacuum degasser (DGU-20A5), a binary pump (LC-20AD), an autosampler (SIL-20A), UV/VIS Photodiode Array Detector (SPD-M20A), and fitted with a guard column (CLC G-ODS) and Kromasil 100-5C18 (4.6×250 mm) analytical column running at room temperature.

    Techniques: Incubation, High Performance Liquid Chromatography, Mass Spectrometry, Injection, Synthesized, Flow Cytometry, Spectroscopy

    Mass spectrometric analysis and the ratio of mature to pro-apoA-I in HDL from PCPE2 −/− mouse plasma. A and B , molecular masses of HDL apoA-I after maximum entropy transformation of the electrospray spectrum of FPLC purified HDL apoA-I from chow-fed C57BL/6 mice ( A ) and PCPE2 −/− mice ( B ). The most abundant mass in each spectrum was identical within the ±1.4 Da S.D. to the mass of mature mouse apoA-I at 27,950 Da. The minor spectrum peak is consistent with the mass of mouse pro-apoA-I at 28,815 Da. C and D , the relative intensity of apoA-I tryptic peptides from LC-MS/MS analysis. ApoA-I from whole mouse plasma was separated by 12% SDS-PAGE followed by in-gel trypsin digest and then analyzed for the N-terminal tryptic pro-apoA-I peptide and mature apoA-I peptide: C57BL/6 mouse plasma ( C ) and PCPE2 −/− mouse plasma ( D ) as described under “Experimental Procedures.” Peptides were separated by reverse-phase HPLC and analyzed for tryptic peptide of the N-terminal T1 for mature peptide ( m / z 1132.48) and the T1 propeptide ( m / z 1995.88), shown in selective ion chromatograms. The mature murine T1 tryptic peptide has the amino acid sequence DEPQSQWDK, and the unprocessed pro-form has the sequence WHVWQQDEPQSQWDK.

    Journal: The Journal of Biological Chemistry

    Article Title: Procollagen C-endopeptidase Enhancer Protein 2 (PCPE2) Reduces Atherosclerosis in Mice by Enhancing Scavenger Receptor Class B1 (SR-BI)-mediated High-density Lipoprotein (HDL)-Cholesteryl Ester Uptake *

    doi: 10.1074/jbc.M115.646240

    Figure Lengend Snippet: Mass spectrometric analysis and the ratio of mature to pro-apoA-I in HDL from PCPE2 −/− mouse plasma. A and B , molecular masses of HDL apoA-I after maximum entropy transformation of the electrospray spectrum of FPLC purified HDL apoA-I from chow-fed C57BL/6 mice ( A ) and PCPE2 −/− mice ( B ). The most abundant mass in each spectrum was identical within the ±1.4 Da S.D. to the mass of mature mouse apoA-I at 27,950 Da. The minor spectrum peak is consistent with the mass of mouse pro-apoA-I at 28,815 Da. C and D , the relative intensity of apoA-I tryptic peptides from LC-MS/MS analysis. ApoA-I from whole mouse plasma was separated by 12% SDS-PAGE followed by in-gel trypsin digest and then analyzed for the N-terminal tryptic pro-apoA-I peptide and mature apoA-I peptide: C57BL/6 mouse plasma ( C ) and PCPE2 −/− mouse plasma ( D ) as described under “Experimental Procedures.” Peptides were separated by reverse-phase HPLC and analyzed for tryptic peptide of the N-terminal T1 for mature peptide ( m / z 1132.48) and the T1 propeptide ( m / z 1995.88), shown in selective ion chromatograms. The mature murine T1 tryptic peptide has the amino acid sequence DEPQSQWDK, and the unprocessed pro-form has the sequence WHVWQQDEPQSQWDK.

    Article Snippet: As anticipated, HDL from LDLr−/− , PCPE2−/− mice contained larger sized particles, indicated by elution in earlier eluting fractions as seen in PCPE2−/− mouse plasma ( A ).

    Techniques: Transformation Assay, Fast Protein Liquid Chromatography, Purification, Mouse Assay, Liquid Chromatography with Mass Spectroscopy, Mass Spectrometry, SDS Page, High Performance Liquid Chromatography, Sequencing

    PCPE2 deficiency exacerbates atherosclerosis despite elevated HDL cholesterol levels. A , representative aortic root sections stained with Oil Red O from 12 weeks atherogenic diet-fed LDLr −/− , LDLr −/− PCPE2 −/− , and LDLr −/− , apoA-I −/− mice. B and C , quantification of the atherosclerotic lesion area as a percentage of the total aortic area ( B ) and the total lesion area in μm 2 ( C ). D , representative aortic root sections stained with fluorescently labeled antibodies to CD68, a macrophage marker, from 12 weeks atherogenic diet-fed LDLr −/− mice, LDLr −/− ,PCPE2 −/− mice, and LDLr −/− , apoA-I −/− mice. E and F , quantification of the macrophage content was performed by measuring CD68+ staining over background as a percentage of the total lesion area ( E ) and of the total lesion area in μm 2 ( F ). Data represent the mean ± S.D. of n = 10 male mice/group. Different letters indicate statistical significance at p

    Journal: The Journal of Biological Chemistry

    Article Title: Procollagen C-endopeptidase Enhancer Protein 2 (PCPE2) Reduces Atherosclerosis in Mice by Enhancing Scavenger Receptor Class B1 (SR-BI)-mediated High-density Lipoprotein (HDL)-Cholesteryl Ester Uptake *

    doi: 10.1074/jbc.M115.646240

    Figure Lengend Snippet: PCPE2 deficiency exacerbates atherosclerosis despite elevated HDL cholesterol levels. A , representative aortic root sections stained with Oil Red O from 12 weeks atherogenic diet-fed LDLr −/− , LDLr −/− PCPE2 −/− , and LDLr −/− , apoA-I −/− mice. B and C , quantification of the atherosclerotic lesion area as a percentage of the total aortic area ( B ) and the total lesion area in μm 2 ( C ). D , representative aortic root sections stained with fluorescently labeled antibodies to CD68, a macrophage marker, from 12 weeks atherogenic diet-fed LDLr −/− mice, LDLr −/− ,PCPE2 −/− mice, and LDLr −/− , apoA-I −/− mice. E and F , quantification of the macrophage content was performed by measuring CD68+ staining over background as a percentage of the total lesion area ( E ) and of the total lesion area in μm 2 ( F ). Data represent the mean ± S.D. of n = 10 male mice/group. Different letters indicate statistical significance at p

    Article Snippet: As anticipated, HDL from LDLr−/− , PCPE2−/− mice contained larger sized particles, indicated by elution in earlier eluting fractions as seen in PCPE2−/− mouse plasma ( A ).

    Techniques: Staining, Mouse Assay, Labeling, Marker

    Absence of liver PCPE2 affects HDL-mediated reverse cholesterol transport and SR-BI expression. A , HDL from diet-fed LDLr −/− and LDLr −/− , PCPE2 −/− mice was purified and then radiolabeled with 125 I as described under “Experimental Procedures.” Approximately 10 × 10 6 cpm of 125 I-labeled HDL was retro-orbitally injected into atherogenic diet-fed recipient mice of the indicated genotype. Blood was collected from the contralateral retro-orbital sinus at the indicated times. B , the FCR of 125 I-labeled HDL were determined from plasma decay curves assuming a one-pool model as described under ”Experimental Procedures.” Data represent the mean ± S.D. of n = 3 mice/group. C , [ 3 H]cholesterol levels in plasma during macrophage reverse cholesterol transport study in LDLr −/− ( gray circles ) and LDLr −/− , PCPE2 −/− ( black squares ) mice fed an atherogenic diet. Mice were injected intraperitoneally with [ 3 H]cholesterol-labeled J774 foam cells as described under “Experimental Procedures.” D , [ 3 H]cholesterol in feces after 0–48 h collection. The numerical data shown is the mean ± S.D. of n = 5–6 male mice for each genotype. The asterisk indicates statistical significance at p

    Journal: The Journal of Biological Chemistry

    Article Title: Procollagen C-endopeptidase Enhancer Protein 2 (PCPE2) Reduces Atherosclerosis in Mice by Enhancing Scavenger Receptor Class B1 (SR-BI)-mediated High-density Lipoprotein (HDL)-Cholesteryl Ester Uptake *

    doi: 10.1074/jbc.M115.646240

    Figure Lengend Snippet: Absence of liver PCPE2 affects HDL-mediated reverse cholesterol transport and SR-BI expression. A , HDL from diet-fed LDLr −/− and LDLr −/− , PCPE2 −/− mice was purified and then radiolabeled with 125 I as described under “Experimental Procedures.” Approximately 10 × 10 6 cpm of 125 I-labeled HDL was retro-orbitally injected into atherogenic diet-fed recipient mice of the indicated genotype. Blood was collected from the contralateral retro-orbital sinus at the indicated times. B , the FCR of 125 I-labeled HDL were determined from plasma decay curves assuming a one-pool model as described under ”Experimental Procedures.” Data represent the mean ± S.D. of n = 3 mice/group. C , [ 3 H]cholesterol levels in plasma during macrophage reverse cholesterol transport study in LDLr −/− ( gray circles ) and LDLr −/− , PCPE2 −/− ( black squares ) mice fed an atherogenic diet. Mice were injected intraperitoneally with [ 3 H]cholesterol-labeled J774 foam cells as described under “Experimental Procedures.” D , [ 3 H]cholesterol in feces after 0–48 h collection. The numerical data shown is the mean ± S.D. of n = 5–6 male mice for each genotype. The asterisk indicates statistical significance at p

    Article Snippet: As anticipated, HDL from LDLr−/− , PCPE2−/− mice contained larger sized particles, indicated by elution in earlier eluting fractions as seen in PCPE2−/− mouse plasma ( A ).

    Techniques: Expressing, Mouse Assay, Purification, Labeling, Injection

    PCPE2 deficiency decreases collagen content in aorta and liver. A , representative aortic root sections following Masson's trichrome staining. Sections were obtained from LDLr −/− and LDLr −/− , PCPE2 −/− mice fed an atherogenic diet for 12 weeks. Quantification of the absolute connective tissue area and the percent of total collagen content per aortic area is shown adjacent to representative sections from each genotype. Data represent the mean ± S.D. of n = 10 male mice/group. Different letters indicate statistical significance at p

    Journal: The Journal of Biological Chemistry

    Article Title: Procollagen C-endopeptidase Enhancer Protein 2 (PCPE2) Reduces Atherosclerosis in Mice by Enhancing Scavenger Receptor Class B1 (SR-BI)-mediated High-density Lipoprotein (HDL)-Cholesteryl Ester Uptake *

    doi: 10.1074/jbc.M115.646240

    Figure Lengend Snippet: PCPE2 deficiency decreases collagen content in aorta and liver. A , representative aortic root sections following Masson's trichrome staining. Sections were obtained from LDLr −/− and LDLr −/− , PCPE2 −/− mice fed an atherogenic diet for 12 weeks. Quantification of the absolute connective tissue area and the percent of total collagen content per aortic area is shown adjacent to representative sections from each genotype. Data represent the mean ± S.D. of n = 10 male mice/group. Different letters indicate statistical significance at p

    Article Snippet: As anticipated, HDL from LDLr−/− , PCPE2−/− mice contained larger sized particles, indicated by elution in earlier eluting fractions as seen in PCPE2−/− mouse plasma ( A ).

    Techniques: Staining, Mouse Assay

    Coformulation with AT1001 results in greater tissue levels and biodistribution of ATB100 in Gla KO (knockout) mice. ( a ) Twelve-week-old male Gla KO mice were given a single 30-minute intravenous infusion of 1 or 3 mg/kg ATB100, either alone or coformulated with 1.22, 3.66, or 12.2 mg/kg AT1001 (equivalent to 1, 3, and 10 mg/kg free base, respectively). Skin, heart, and kidney were collected 7 days postinfusion, and α-galactosidase A (α-Gal A) activities were determined. Each bar represents the mean ± SEM of five to six mice per group. * P

    Journal: Molecular Therapy

    Article Title: Coformulation of a Novel Human α-Galactosidase A With the Pharmacological Chaperone AT1001 Leads to Improved Substrate Reduction in Fabry Mice

    doi: 10.1038/mt.2015.87

    Figure Lengend Snippet: Coformulation with AT1001 results in greater tissue levels and biodistribution of ATB100 in Gla KO (knockout) mice. ( a ) Twelve-week-old male Gla KO mice were given a single 30-minute intravenous infusion of 1 or 3 mg/kg ATB100, either alone or coformulated with 1.22, 3.66, or 12.2 mg/kg AT1001 (equivalent to 1, 3, and 10 mg/kg free base, respectively). Skin, heart, and kidney were collected 7 days postinfusion, and α-galactosidase A (α-Gal A) activities were determined. Each bar represents the mean ± SEM of five to six mice per group. * P

    Article Snippet: In vitro , coincubation of Fabry patient fibroblasts with ATB100 and AT1001 resulted in ~2.6-fold leftward shift in the EC50 value of ATB100 for GL-3 reduction, indicating that AT1001 increases the potency of ATB100.

    Techniques: Knock-Out, Mouse Assay

    Coformulation of ATB100 with AT1001 leads to greater substrate reduction in Gla KO (knockout) mice . Fourteen-week-old male Gla KO mice were administered ATB100 (0.5, 1, or 3 mg/kg), either alone or coformulated with 1.22, 3.66, or 12.2 mg/kg AT1001 (equivalent to 1, 3, and 10 mg/kg free base, respectively) via intravenous bolus injection (four biweekly administrations). Plasma and tissues were collected 7 days after the final administration, and globotriaosylceramide (GL-3) levels were measured by liquid chromatography in combination with tandem mass spectrometry. GL-3 levels were normalized to those in untreated Gla KO (100%) and wild-type C57BL/6 (0%) mouse tissues (GL-3 levels in Gla KO mouse skin, heart, and kidney were 2265, 559, and 778 µg/g tissue, respectively; GL-3 levels in wild-type mouse skin, heart, and kidney were 7, 5, and 75 µg/g tissue, respectively). Each bar represents the mean ± SEM of seven mice per group. * P

    Journal: Molecular Therapy

    Article Title: Coformulation of a Novel Human α-Galactosidase A With the Pharmacological Chaperone AT1001 Leads to Improved Substrate Reduction in Fabry Mice

    doi: 10.1038/mt.2015.87

    Figure Lengend Snippet: Coformulation of ATB100 with AT1001 leads to greater substrate reduction in Gla KO (knockout) mice . Fourteen-week-old male Gla KO mice were administered ATB100 (0.5, 1, or 3 mg/kg), either alone or coformulated with 1.22, 3.66, or 12.2 mg/kg AT1001 (equivalent to 1, 3, and 10 mg/kg free base, respectively) via intravenous bolus injection (four biweekly administrations). Plasma and tissues were collected 7 days after the final administration, and globotriaosylceramide (GL-3) levels were measured by liquid chromatography in combination with tandem mass spectrometry. GL-3 levels were normalized to those in untreated Gla KO (100%) and wild-type C57BL/6 (0%) mouse tissues (GL-3 levels in Gla KO mouse skin, heart, and kidney were 2265, 559, and 778 µg/g tissue, respectively; GL-3 levels in wild-type mouse skin, heart, and kidney were 7, 5, and 75 µg/g tissue, respectively). Each bar represents the mean ± SEM of seven mice per group. * P

    Article Snippet: In vitro , coincubation of Fabry patient fibroblasts with ATB100 and AT1001 resulted in ~2.6-fold leftward shift in the EC50 value of ATB100 for GL-3 reduction, indicating that AT1001 increases the potency of ATB100.

    Techniques: Knock-Out, Mouse Assay, Injection, Liquid Chromatography, Mass Spectrometry

    Coincubation of ATB100 with AT1001 enhances cellular α-galactosidase A (α-Gal A) activity and globotriaosylceramide (GL-3) reduction in male Fabry fibroblasts. ( a ) C52S Fabry fibroblasts were incubated with ATB100 (0.5 nmol/l) alone or in the presence of increasing concentrations of AT1001 (0.1, 1, 10, or 100 μmol/l) for 5 hours. α-Gal A activity (blue bars) and GL-3 levels (red bars) in cell lysates were then measured 2 and 10 days later, respectively. The data points shown are the mean ± SEM of four independent experiments. * P

    Journal: Molecular Therapy

    Article Title: Coformulation of a Novel Human α-Galactosidase A With the Pharmacological Chaperone AT1001 Leads to Improved Substrate Reduction in Fabry Mice

    doi: 10.1038/mt.2015.87

    Figure Lengend Snippet: Coincubation of ATB100 with AT1001 enhances cellular α-galactosidase A (α-Gal A) activity and globotriaosylceramide (GL-3) reduction in male Fabry fibroblasts. ( a ) C52S Fabry fibroblasts were incubated with ATB100 (0.5 nmol/l) alone or in the presence of increasing concentrations of AT1001 (0.1, 1, 10, or 100 μmol/l) for 5 hours. α-Gal A activity (blue bars) and GL-3 levels (red bars) in cell lysates were then measured 2 and 10 days later, respectively. The data points shown are the mean ± SEM of four independent experiments. * P

    Article Snippet: In vitro , coincubation of Fabry patient fibroblasts with ATB100 and AT1001 resulted in ~2.6-fold leftward shift in the EC50 value of ATB100 for GL-3 reduction, indicating that AT1001 increases the potency of ATB100.

    Techniques: Activity Assay, Incubation

    Coformulation of ATB100 with AT1001 leads to greater plasma globotriaosylceramide (GL-3) and globotriaosylsphingosine (lyso-Gb 3 ) reductions in Gla KO (knockout) mice. Fourteen-week-old male Gla KO mice were administered ATB100 (0.5, 1, or 3 mg/kg), either alone or coformulated with 1.22, 3.66, or 12.2 mg/kg AT1001 (equivalent to 1, 3, and 10 mg/kg free base, respectively) via intravenous bolus injection (four biweekly administrations). Plasma samples were collected 7 days after the final administration, and substrate levels: GL-3 ( a ) and lyso-Gb 3 ( b ) were measured by liquid chromatography in combination with tandem mass spectrometry. Substrate levels were normalized to those in Gla KO mouse plasma (5 μg/ml and 246 ng/ml, respectively). Each bar represents the mean ± SEM of seven mice per group. * P

    Journal: Molecular Therapy

    Article Title: Coformulation of a Novel Human α-Galactosidase A With the Pharmacological Chaperone AT1001 Leads to Improved Substrate Reduction in Fabry Mice

    doi: 10.1038/mt.2015.87

    Figure Lengend Snippet: Coformulation of ATB100 with AT1001 leads to greater plasma globotriaosylceramide (GL-3) and globotriaosylsphingosine (lyso-Gb 3 ) reductions in Gla KO (knockout) mice. Fourteen-week-old male Gla KO mice were administered ATB100 (0.5, 1, or 3 mg/kg), either alone or coformulated with 1.22, 3.66, or 12.2 mg/kg AT1001 (equivalent to 1, 3, and 10 mg/kg free base, respectively) via intravenous bolus injection (four biweekly administrations). Plasma samples were collected 7 days after the final administration, and substrate levels: GL-3 ( a ) and lyso-Gb 3 ( b ) were measured by liquid chromatography in combination with tandem mass spectrometry. Substrate levels were normalized to those in Gla KO mouse plasma (5 μg/ml and 246 ng/ml, respectively). Each bar represents the mean ± SEM of seven mice per group. * P

    Article Snippet: In vitro , coincubation of Fabry patient fibroblasts with ATB100 and AT1001 resulted in ~2.6-fold leftward shift in the EC50 value of ATB100 for GL-3 reduction, indicating that AT1001 increases the potency of ATB100.

    Techniques: Knock-Out, Mouse Assay, Injection, Liquid Chromatography, Mass Spectrometry

    Coformulation of ATB100 with AT1001 increases the total exposure of active ATB100 in mice . ( a ) Eight-week-old male wild-type C57BL/6 mice were given a single 30-minute intravenous infusion of 1 mg/kg of ATB100, either alone or coformulated with 3.66 or 12.2 mg/kg AT1001 (equivalent to 3 and 10 mg/kg free base, respectively). Plasma samples were collected over the 24-hour period from the start of infusion, and α-galactosidase A (α-Gal A) activity (upper panel) and protein levels (Western blot, lower panel) were measured. For the graph, each time point represents the mean ± SEM of the activity measured from five mice. * P

    Journal: Molecular Therapy

    Article Title: Coformulation of a Novel Human α-Galactosidase A With the Pharmacological Chaperone AT1001 Leads to Improved Substrate Reduction in Fabry Mice

    doi: 10.1038/mt.2015.87

    Figure Lengend Snippet: Coformulation of ATB100 with AT1001 increases the total exposure of active ATB100 in mice . ( a ) Eight-week-old male wild-type C57BL/6 mice were given a single 30-minute intravenous infusion of 1 mg/kg of ATB100, either alone or coformulated with 3.66 or 12.2 mg/kg AT1001 (equivalent to 3 and 10 mg/kg free base, respectively). Plasma samples were collected over the 24-hour period from the start of infusion, and α-galactosidase A (α-Gal A) activity (upper panel) and protein levels (Western blot, lower panel) were measured. For the graph, each time point represents the mean ± SEM of the activity measured from five mice. * P

    Article Snippet: In vitro , coincubation of Fabry patient fibroblasts with ATB100 and AT1001 resulted in ~2.6-fold leftward shift in the EC50 value of ATB100 for GL-3 reduction, indicating that AT1001 increases the potency of ATB100.

    Techniques: Mouse Assay, Activity Assay, Western Blot

    AT1001 increases the physical stability of ATB100 in vitro . ( a ) Thermal stability scans of ATB100 in the absence and presence of AT1001. Unfolding and denaturation of ATB100 were monitored by changes in the fluorescence of SYPRO Orange as a function of temperature. The thermal stability scans were performed at pH 7.4 in the absence (blue line) and presence of 10 µmol/l (purple line) and 100 µmol/l (green line) AT1001, and at pH 5.2 in the absence of AT1001 (red line). ( b ) Aggregation of ATB100 in the absence and presence of AT1001 at neutral pH for 4 weeks at 40 °C. Size exclusion-high performance liquid chromatography chromatograms showed a single homodimeric enzyme peak with 5 mg/ml ATB100 coincubated with 10 mg/ml AT1001 (black line), but a high molecular weight peak (indicative of aggregate) along with enzyme peak with ATB100 alone (blue line). Inset : Aggregation of ATB100 as monitored visually at neutral pH for 4 weeks at 40 °C. The vial of 50 mg/ml ATB100 alone showed clear aggregation as depicted by white turbid solution, whereas the vial coincubated with 10 mg/ml AT1001 resulted in a clear solution. AT, Amicus Therapeutics; ATB, Amicus Therapeutics Biologics.

    Journal: Molecular Therapy

    Article Title: Coformulation of a Novel Human α-Galactosidase A With the Pharmacological Chaperone AT1001 Leads to Improved Substrate Reduction in Fabry Mice

    doi: 10.1038/mt.2015.87

    Figure Lengend Snippet: AT1001 increases the physical stability of ATB100 in vitro . ( a ) Thermal stability scans of ATB100 in the absence and presence of AT1001. Unfolding and denaturation of ATB100 were monitored by changes in the fluorescence of SYPRO Orange as a function of temperature. The thermal stability scans were performed at pH 7.4 in the absence (blue line) and presence of 10 µmol/l (purple line) and 100 µmol/l (green line) AT1001, and at pH 5.2 in the absence of AT1001 (red line). ( b ) Aggregation of ATB100 in the absence and presence of AT1001 at neutral pH for 4 weeks at 40 °C. Size exclusion-high performance liquid chromatography chromatograms showed a single homodimeric enzyme peak with 5 mg/ml ATB100 coincubated with 10 mg/ml AT1001 (black line), but a high molecular weight peak (indicative of aggregate) along with enzyme peak with ATB100 alone (blue line). Inset : Aggregation of ATB100 as monitored visually at neutral pH for 4 weeks at 40 °C. The vial of 50 mg/ml ATB100 alone showed clear aggregation as depicted by white turbid solution, whereas the vial coincubated with 10 mg/ml AT1001 resulted in a clear solution. AT, Amicus Therapeutics; ATB, Amicus Therapeutics Biologics.

    Article Snippet: In vitro , coincubation of Fabry patient fibroblasts with ATB100 and AT1001 resulted in ~2.6-fold leftward shift in the EC50 value of ATB100 for GL-3 reduction, indicating that AT1001 increases the potency of ATB100.

    Techniques: In Vitro, Fluorescence, High Performance Liquid Chromatography, Molecular Weight

    Coformulation of ATB100 with AT1001 enhances cell type-specific globotriaosylceramide (GL-3) reduction in Gla KO (knockout) mouse tissues. Fourteen-week-old male Gla KO mice were administered ATB1001 (0.5, 1, or 3 mg/kg), either alone or coformulated with 1.22 or 3.66 mg/kg AT1001 (equivalent to 1 and 3 mg/kg free base, respectively) via intravenous bolus injection (four biweekly administrations). Tissues were collected 7 days after the final administration, and GL-3 levels were examined by IHC using an anti-GL-3 antibody. GL-3 signals are shown as brown spots in dermal fibroblasts (arrows) and vascular smooth muscle cells of blood vessels (asterisks), cardiac vascular smooth muscle cells (arrows, with lumens marked by asterisks), and renal distal tubular epithelial cells (arrows). Images were taken with a ×40 objective lens for heart and skin, and a ×20 objective lens for kidney.

    Journal: Molecular Therapy

    Article Title: Coformulation of a Novel Human α-Galactosidase A With the Pharmacological Chaperone AT1001 Leads to Improved Substrate Reduction in Fabry Mice

    doi: 10.1038/mt.2015.87

    Figure Lengend Snippet: Coformulation of ATB100 with AT1001 enhances cell type-specific globotriaosylceramide (GL-3) reduction in Gla KO (knockout) mouse tissues. Fourteen-week-old male Gla KO mice were administered ATB1001 (0.5, 1, or 3 mg/kg), either alone or coformulated with 1.22 or 3.66 mg/kg AT1001 (equivalent to 1 and 3 mg/kg free base, respectively) via intravenous bolus injection (four biweekly administrations). Tissues were collected 7 days after the final administration, and GL-3 levels were examined by IHC using an anti-GL-3 antibody. GL-3 signals are shown as brown spots in dermal fibroblasts (arrows) and vascular smooth muscle cells of blood vessels (asterisks), cardiac vascular smooth muscle cells (arrows, with lumens marked by asterisks), and renal distal tubular epithelial cells (arrows). Images were taken with a ×40 objective lens for heart and skin, and a ×20 objective lens for kidney.

    Article Snippet: In vitro , coincubation of Fabry patient fibroblasts with ATB100 and AT1001 resulted in ~2.6-fold leftward shift in the EC50 value of ATB100 for GL-3 reduction, indicating that AT1001 increases the potency of ATB100.

    Techniques: Knock-Out, Mouse Assay, Injection, Immunohistochemistry