liquid chromatography tandem mass spectrometry  (SCIEX)

 
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    Structured Review

    SCIEX liquid chromatography tandem mass spectrometry
    Proteins that interact with RRAD. Notes: ( A ) Proteins that predicted to interact with RRAD on STRING (Szklarczyk D, Franceschini A, Wyder S, et al. STRING v10: protein-protein interaction networks, integrated over the tree of life. Nucleic Acids Res . 2015;43(Database issue):D447–D452). 35 ( B ) Expression of P53 after overexpression of RRAD and expression of RRAD after overexpression of P53 by Western blotting. ( C ) The Venn diagram of the number of proteins after Co-IP assays and LC-MS/MS analysis. ( D ) GST (glutathione-S-transferase) pull-down assay for ACTG1/EF1A1 with RRAD. Abbreviations: RRAD, Ras-related associated with diabetes; Co-IP, co-immunoprecipitation; LC-MS/MS, <t>liquid</t> <t>chromatography–tandem</t> <t>mass</t> <t>spectrometry;</t> ACTG1, actin gamma 1; NC, negative control.
    Liquid Chromatography Tandem Mass Spectrometry, supplied by SCIEX, used in various techniques. Bioz Stars score: 93/100, based on 54 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 93 stars, based on 54 article reviews
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    liquid chromatography tandem mass spectrometry - by Bioz Stars, 2020-07
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    Images

    1) Product Images from "RRAD suppresses the Warburg effect by downregulating ACTG1 in hepatocellular carcinoma"

    Article Title: RRAD suppresses the Warburg effect by downregulating ACTG1 in hepatocellular carcinoma

    Journal: OncoTargets and therapy

    doi: 10.2147/OTT.S197844

    Proteins that interact with RRAD. Notes: ( A ) Proteins that predicted to interact with RRAD on STRING (Szklarczyk D, Franceschini A, Wyder S, et al. STRING v10: protein-protein interaction networks, integrated over the tree of life. Nucleic Acids Res . 2015;43(Database issue):D447–D452). 35 ( B ) Expression of P53 after overexpression of RRAD and expression of RRAD after overexpression of P53 by Western blotting. ( C ) The Venn diagram of the number of proteins after Co-IP assays and LC-MS/MS analysis. ( D ) GST (glutathione-S-transferase) pull-down assay for ACTG1/EF1A1 with RRAD. Abbreviations: RRAD, Ras-related associated with diabetes; Co-IP, co-immunoprecipitation; LC-MS/MS, liquid chromatography–tandem mass spectrometry; ACTG1, actin gamma 1; NC, negative control.
    Figure Legend Snippet: Proteins that interact with RRAD. Notes: ( A ) Proteins that predicted to interact with RRAD on STRING (Szklarczyk D, Franceschini A, Wyder S, et al. STRING v10: protein-protein interaction networks, integrated over the tree of life. Nucleic Acids Res . 2015;43(Database issue):D447–D452). 35 ( B ) Expression of P53 after overexpression of RRAD and expression of RRAD after overexpression of P53 by Western blotting. ( C ) The Venn diagram of the number of proteins after Co-IP assays and LC-MS/MS analysis. ( D ) GST (glutathione-S-transferase) pull-down assay for ACTG1/EF1A1 with RRAD. Abbreviations: RRAD, Ras-related associated with diabetes; Co-IP, co-immunoprecipitation; LC-MS/MS, liquid chromatography–tandem mass spectrometry; ACTG1, actin gamma 1; NC, negative control.

    Techniques Used: Expressing, Over Expression, Western Blot, Co-Immunoprecipitation Assay, Liquid Chromatography with Mass Spectroscopy, Mass Spectrometry, Pull Down Assay, Immunoprecipitation, Liquid Chromatography, Negative Control

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    High Performance Liquid Chromatography:

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    Article Snippet: .. Analysis with Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) High Performance Liquid Chromatography (HPLC) SystemsLC-MS/MS analysis was performed with Absciex 3200 Q trap MS/MS detector. .. Experiments were carried out using Shimadzu 20A HPLC system coupled to an Applied Biosystems 3200 Q-Trap LC-MS/MS instrument equipped with an electrospray ionization (ESI) source operating in negative ion mode.

    Chromatography:

    Article Title: Cardiorespiratory and anesthetic effects of combined alfaxalone, butorphanol, and medetomidine in Thoroughbred horses
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    Article Title: Potential Antioxidant and Enzyme Inhibitory Effects of Nanoliposomal Formulation Prepared from Salvia aramiensis Rech. f. Extract
    Article Snippet: .. Analysis with Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) High Performance Liquid Chromatography (HPLC) SystemsLC-MS/MS analysis was performed with Absciex 3200 Q trap MS/MS detector. .. Experiments were carried out using Shimadzu 20A HPLC system coupled to an Applied Biosystems 3200 Q-Trap LC-MS/MS instrument equipped with an electrospray ionization (ESI) source operating in negative ion mode.

    Article Title: RRAD suppresses the Warburg effect by downregulating ACTG1 in hepatocellular carcinoma
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    Methylation:

    Article Title: Higher Number of Night Shifts Associates with Good Perception of Work Capacity and Optimal Lung Function but Correlates with Increased Oxidative Damage and Telomere Attrition
    Article Snippet: .. Oxidized and Methylated Nucleic Acids Analysis Biomarkers of nucleic acids oxidation, as 8-oxo-7,8-dihydro 2'deoxyguanosine (8-oxodGuo), 8-oxo-7,8-dihydro-guanosine (8-oxoGuo), and 8-oxo-7,8-dihydroguanine (8-oxoGua), and methylation, such as 5-methyl-cytosine (5-MeCyto), 5-hydroxymethyl-cytosine (5-OHMeCyto), 5-methylcytidine (5-MeCyt), 5-methyl-2'deoxycytidine (5-MedCyt), 5-hydroxymethyl-2'deoxycytidine (5-OHMedCyt), 1-methyl-guanine (1-MeGua), 7-methyl-guanine (7-MeGua), and 7-methyl-guanosine (7-MeGuo), were determined by isotopic dilution liquid chromatography tandem mass spectrometry (LC-MS/MS) using a API 4000 triple quadrupole mass spectrometer (SCIEX, Framingham, MA USA) equipped with a TurboIonSpray interface for pneumatically assisted electrospray according to the method by Andreoli et al. [ ], with some modifications to determine urinary methylated biomarkers in the same chromatographic run. .. Briefly, after centrifugation urinary samples (50 μ L) were diluted with 150 μ L of internal standards (including [13 C1 , 15 N2 ] 8-oxo-7,8-dihydroguanine, [15 N5 ] 8-oxo-7,8-dihydro-guanosine, [15 N5 ] 8-oxo-7,8-dihydro 2'deoxyguanosine, and 5-hydroxymethyl-2'deoxycytidine-d3 ) aqueous mixture and injected (5 μ l).

    Mass Spectrometry:

    Article Title: Cardiorespiratory and anesthetic effects of combined alfaxalone, butorphanol, and medetomidine in Thoroughbred horses
    Article Snippet: .. The extracted substances were analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS; AB Sciex., Framingham, MA, U.S.A.) by electrospray ionization in positive ion mode. ..

    Article Title: Heterogeneous nuclear ribonucleoprotein A2/B1 is a negative regulator of human breast cancer metastasis by maintaining the balance of multiple genes and pathways
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    Article Title: Potential Antioxidant and Enzyme Inhibitory Effects of Nanoliposomal Formulation Prepared from Salvia aramiensis Rech. f. Extract
    Article Snippet: .. Analysis with Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) High Performance Liquid Chromatography (HPLC) SystemsLC-MS/MS analysis was performed with Absciex 3200 Q trap MS/MS detector. .. Experiments were carried out using Shimadzu 20A HPLC system coupled to an Applied Biosystems 3200 Q-Trap LC-MS/MS instrument equipped with an electrospray ionization (ESI) source operating in negative ion mode.

    Article Title: RRAD suppresses the Warburg effect by downregulating ACTG1 in hepatocellular carcinoma
    Article Snippet: .. Analysis was performed using liquid chromatography–tandem mass spectrometry (LC-MS/MS; Ekspert NanoLC and TripleTOF 5600+; Sciex, Framingham, MA, USA). .. Cell proliferation and cell cycle analysis Cell Counting Kit-8 (Dojindo Laboratories, Kumamoto, Japan) and EdU assay (Cell-Light™ EdU Apollo567 In Vitro Imaging Kit; Ribobio, Guangzhou, China) were used for cell proliferation analysis.

    Article Title: A Mollusk Retinoic Acid Receptor (RAR) Ortholog Sheds Light on the Evolution of Ligand Binding
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    Article Title: Effects of Estrogen on Bone mRNA Levels of Sclerostin and Other Genes Relevant to Bone Metabolism in Postmenopausal Women
    Article Snippet: .. 25-Hydroxyvitamin D [25(OH)D] (within assay CV of 2.4% and between assay CV of 6.8%) was measured using liquid chromatography-tandem mass spectrometry (API 5000; Applied Biosystems-MDS Sciex). ..

    Article Title: Higher Number of Night Shifts Associates with Good Perception of Work Capacity and Optimal Lung Function but Correlates with Increased Oxidative Damage and Telomere Attrition
    Article Snippet: .. Oxidized and Methylated Nucleic Acids Analysis Biomarkers of nucleic acids oxidation, as 8-oxo-7,8-dihydro 2'deoxyguanosine (8-oxodGuo), 8-oxo-7,8-dihydro-guanosine (8-oxoGuo), and 8-oxo-7,8-dihydroguanine (8-oxoGua), and methylation, such as 5-methyl-cytosine (5-MeCyto), 5-hydroxymethyl-cytosine (5-OHMeCyto), 5-methylcytidine (5-MeCyt), 5-methyl-2'deoxycytidine (5-MedCyt), 5-hydroxymethyl-2'deoxycytidine (5-OHMedCyt), 1-methyl-guanine (1-MeGua), 7-methyl-guanine (7-MeGua), and 7-methyl-guanosine (7-MeGuo), were determined by isotopic dilution liquid chromatography tandem mass spectrometry (LC-MS/MS) using a API 4000 triple quadrupole mass spectrometer (SCIEX, Framingham, MA USA) equipped with a TurboIonSpray interface for pneumatically assisted electrospray according to the method by Andreoli et al. [ ], with some modifications to determine urinary methylated biomarkers in the same chromatographic run. .. Briefly, after centrifugation urinary samples (50 μ L) were diluted with 150 μ L of internal standards (including [13 C1 , 15 N2 ] 8-oxo-7,8-dihydroguanine, [15 N5 ] 8-oxo-7,8-dihydro-guanosine, [15 N5 ] 8-oxo-7,8-dihydro 2'deoxyguanosine, and 5-hydroxymethyl-2'deoxycytidine-d3 ) aqueous mixture and injected (5 μ l).

    Article Title: DNA methylation of the cancer-related genes F2RL3 and AHRR is associated with occupational exposure to polycyclic aromatic hydrocarbons
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    Tandem Mass Spectroscopy:

    Article Title: Potential Antioxidant and Enzyme Inhibitory Effects of Nanoliposomal Formulation Prepared from Salvia aramiensis Rech. f. Extract
    Article Snippet: .. Analysis with Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) High Performance Liquid Chromatography (HPLC) SystemsLC-MS/MS analysis was performed with Absciex 3200 Q trap MS/MS detector. .. Experiments were carried out using Shimadzu 20A HPLC system coupled to an Applied Biosystems 3200 Q-Trap LC-MS/MS instrument equipped with an electrospray ionization (ESI) source operating in negative ion mode.

    Liquid Chromatography:

    Article Title: Heterogeneous nuclear ribonucleoprotein A2/B1 is a negative regulator of human breast cancer metastasis by maintaining the balance of multiple genes and pathways
    Article Snippet: .. Liquid chromatography tandem mass spectrometry (LC–MS/MS) analysis was performed (AB Sciex Triple TOF 5600). .. 2.6 Tissue microarray Tissue microarray (US Biomax, BR10010d) of 47 cases of infiltrative ductal carcinoma and lymph node metastasis and three cases of infiltrative lobular carcinoma was evaluated through immunohistochemical staining according to the protocol provided by US Biomax Inc.

    Article Title: Higher Number of Night Shifts Associates with Good Perception of Work Capacity and Optimal Lung Function but Correlates with Increased Oxidative Damage and Telomere Attrition
    Article Snippet: .. Oxidized and Methylated Nucleic Acids Analysis Biomarkers of nucleic acids oxidation, as 8-oxo-7,8-dihydro 2'deoxyguanosine (8-oxodGuo), 8-oxo-7,8-dihydro-guanosine (8-oxoGuo), and 8-oxo-7,8-dihydroguanine (8-oxoGua), and methylation, such as 5-methyl-cytosine (5-MeCyto), 5-hydroxymethyl-cytosine (5-OHMeCyto), 5-methylcytidine (5-MeCyt), 5-methyl-2'deoxycytidine (5-MedCyt), 5-hydroxymethyl-2'deoxycytidine (5-OHMedCyt), 1-methyl-guanine (1-MeGua), 7-methyl-guanine (7-MeGua), and 7-methyl-guanosine (7-MeGuo), were determined by isotopic dilution liquid chromatography tandem mass spectrometry (LC-MS/MS) using a API 4000 triple quadrupole mass spectrometer (SCIEX, Framingham, MA USA) equipped with a TurboIonSpray interface for pneumatically assisted electrospray according to the method by Andreoli et al. [ ], with some modifications to determine urinary methylated biomarkers in the same chromatographic run. .. Briefly, after centrifugation urinary samples (50 μ L) were diluted with 150 μ L of internal standards (including [13 C1 , 15 N2 ] 8-oxo-7,8-dihydroguanine, [15 N5 ] 8-oxo-7,8-dihydro-guanosine, [15 N5 ] 8-oxo-7,8-dihydro 2'deoxyguanosine, and 5-hydroxymethyl-2'deoxycytidine-d3 ) aqueous mixture and injected (5 μ l).

    Article Title: DNA methylation of the cancer-related genes F2RL3 and AHRR is associated with occupational exposure to polycyclic aromatic hydrocarbons
    Article Snippet: .. Liquid chromatography tandem mass spectrometry (LC-MS/MS; QTRAP 5500, AB Sciex, Foster City, CA) was employed for measurements. .. Urine samples were hydrolysed with glucuronidase b and 5 µl of sample was injected onto a C18 column for analysis of 1-OH-PYR (four aromatic rings) and 2-OH-PH (three aromatic rings).

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    SCIEX quantitative lc ms ms results
    Correlation of <t>results</t> obtained by both MNP-bsELISA and <t>LC-MS/MS</t> for zearalenone detection in natural cereal and feed samples.
    Quantitative Lc Ms Ms Results, supplied by SCIEX, used in various techniques. Bioz Stars score: 88/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    SCIEX rv2607
    PLP formation catalyzed by PNPOx and associated vitamin B6 metabolic pathways. The role of <t>Rv2607</t> is shown in red. In E. coli , PNPOx catalyzes the last step in the DXP-dependent PLP biosynthetic pathway [2] . Most organisms capable of PLP biosynthesis produce PLP via PLP synthase, a macromolecular complex consisting of Pdx1 and Pdx2 [2] . Organisms with genes that encode both PLP synthase and PNPOx likely use PNPOx to salavage PLP from PNP and PMP, which are produced by enzymes that use PLP as a cofactor.
    Rv2607, supplied by SCIEX, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    SCIEX maldi ms ms
    Calibration curve of PPD (A) on the <t>MALDI-MS/MS</t> system (50–1000 <t>µmol/L)</t> and (B) on the HPLC-UV system (150–1000 µmol/L). Graphs shows the mean values of ( A ) 5 and ( B ) 3 independent measurements and the corresponding r 2 value. Mean +/− SEM.
    Maldi Ms Ms, supplied by SCIEX, used in various techniques. Bioz Stars score: 90/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 90 stars, based on 7 article reviews
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    maldi ms ms - by Bioz Stars, 2020-07
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    Image Search Results


    Correlation of results obtained by both MNP-bsELISA and LC-MS/MS for zearalenone detection in natural cereal and feed samples.

    Journal: Toxins

    Article Title: A Magnetic Nanoparticle Based Enzyme-Linked Immunosorbent Assay for Sensitive Quantification of Zearalenone in Cereal and Feed Samples

    doi: 10.3390/toxins7104216

    Figure Lengend Snippet: Correlation of results obtained by both MNP-bsELISA and LC-MS/MS for zearalenone detection in natural cereal and feed samples.

    Article Snippet: Quantitative LC-MS/MS results were analyzed using Analyst software (AB SCIEX, Framingham, MA, USA).

    Techniques: Liquid Chromatography with Mass Spectroscopy, Mass Spectrometry

    PLP formation catalyzed by PNPOx and associated vitamin B6 metabolic pathways. The role of Rv2607 is shown in red. In E. coli , PNPOx catalyzes the last step in the DXP-dependent PLP biosynthetic pathway [2] . Most organisms capable of PLP biosynthesis produce PLP via PLP synthase, a macromolecular complex consisting of Pdx1 and Pdx2 [2] . Organisms with genes that encode both PLP synthase and PNPOx likely use PNPOx to salavage PLP from PNP and PMP, which are produced by enzymes that use PLP as a cofactor.

    Journal: PLoS ONE

    Article Title: Rv2607 from Mycobacterium tuberculosis Is a Pyridoxine 5?-Phosphate Oxidase with Unusual Substrate Specificity

    doi: 10.1371/journal.pone.0027643

    Figure Lengend Snippet: PLP formation catalyzed by PNPOx and associated vitamin B6 metabolic pathways. The role of Rv2607 is shown in red. In E. coli , PNPOx catalyzes the last step in the DXP-dependent PLP biosynthetic pathway [2] . Most organisms capable of PLP biosynthesis produce PLP via PLP synthase, a macromolecular complex consisting of Pdx1 and Pdx2 [2] . Organisms with genes that encode both PLP synthase and PNPOx likely use PNPOx to salavage PLP from PNP and PMP, which are produced by enzymes that use PLP as a cofactor.

    Article Snippet: Extracted cofactor from Rv2607 and authentic FMN were analyzed with a Q Star Elite Mass Spectrometer (Applied Biosystems/MDS Sciex) by electrospray ionization positive ion mode with a gold-plated nanospray tip ( ).

    Techniques: Plasmid Purification, Produced

    Nano-ESI mass spectrum of intact Rv2607 with co-purified FMN. The major species, C , has peaks with charges ranging from 15+ to 12+ in the 3500-4500 m/z range that correspond to a molecular weight of 55376±12. This experimental mass is consistent with the calculated mass for the complex of dimeric Rv2607 with one molecule of FMN bound (55442 Da), which is derived from the amino acid sequence of the 6x-histidine tagged Rv2607 (54986 Da; 27493 Da per protomer) and the molecular weight of FMN (456 Da). Monomeric tagged Rv2607 and truncated Rv2607 monomer (missing 10-11 residues), are present in solution and correspond to molecular weights of A (27450±52 Da) and B (26215±54 Da) respectively. A minor dimeric species comprised of full length and truncated monomer is represented as species D . Higher molecular weight oligomers (trimers E and tetramers F with molecular weights of 110.6 and 82.7 kDa, respectively) are electrospray-induced non-specific association of monomers and dimers.

    Journal: PLoS ONE

    Article Title: Rv2607 from Mycobacterium tuberculosis Is a Pyridoxine 5?-Phosphate Oxidase with Unusual Substrate Specificity

    doi: 10.1371/journal.pone.0027643

    Figure Lengend Snippet: Nano-ESI mass spectrum of intact Rv2607 with co-purified FMN. The major species, C , has peaks with charges ranging from 15+ to 12+ in the 3500-4500 m/z range that correspond to a molecular weight of 55376±12. This experimental mass is consistent with the calculated mass for the complex of dimeric Rv2607 with one molecule of FMN bound (55442 Da), which is derived from the amino acid sequence of the 6x-histidine tagged Rv2607 (54986 Da; 27493 Da per protomer) and the molecular weight of FMN (456 Da). Monomeric tagged Rv2607 and truncated Rv2607 monomer (missing 10-11 residues), are present in solution and correspond to molecular weights of A (27450±52 Da) and B (26215±54 Da) respectively. A minor dimeric species comprised of full length and truncated monomer is represented as species D . Higher molecular weight oligomers (trimers E and tetramers F with molecular weights of 110.6 and 82.7 kDa, respectively) are electrospray-induced non-specific association of monomers and dimers.

    Article Snippet: Extracted cofactor from Rv2607 and authentic FMN were analyzed with a Q Star Elite Mass Spectrometer (Applied Biosystems/MDS Sciex) by electrospray ionization positive ion mode with a gold-plated nanospray tip ( ).

    Techniques: Purification, Molecular Weight, Derivative Assay, Sequencing

    Reverse-phase HPLC, NMR, and spectrophotometric analysis of the PNPOx activity of Rv2607. (A) HPLC chromatograms (270 nm) of reaction mixtures containing PNP (i-iv) or PMP (v-viii), FMN, and Rv2607, and the associated control reactions. All reactions were carried out in 25 mM potassium phosphate buffer (pH 7.8), incubated at 25°C for 3 h, and quenched with DNPH (0.7 mM final concentration). Where present, the reaction components were at the following concentrations: 1 mM PNP or PMP, 10 µM FMN, and 10 µM Rv2607. Reaction mixtures contained: (i) PNP, FMN, and Rv2607, (ii) PNP and FMN (enzyme negative control), (iii) PNP and Rv2607 (no added FMN), (iv) FMN and Rv2607 (substrate negative control), (v) PMP, FMN, and Rv2607, (vi) PMP and FMN (enzyme negative control), (vii) PMP and Rv2607 (no added FMN), (viii) FMN and Rv2607 (substrate negative control). The peak to the right of PLP-DNPH is DNPH, which has a retention time of 8.6 min. (B) 1 H NMR analysis of the conversion of PNP into PLP with time. The enzymatic reaction (1 mM PNP, 28 µM enzyme, 10% D 2 O in 25 mM potassium phosphate buffer, pH 7.8) was incubated for 18 hours at 298 K in the spectrometer. Left; stacked 1 H NMR spectra recorded at various time points and after the addition of authentic PLP. The starred peak corresponds to PLP hydrate. Right; a plot of percent substrate conversion versus time. Substrate conversion was determined by comparing integrals of the C2- 1 H signals associated with PLP (7.80 ppm) to that of PNP (7.75 ppm). (C) Michaelis-Menten plot for Rv2607 with PNP as a substrate. The rate of PLP formation was monitored spectrophotometrically (λ max = 388 nm, ε = 4900 cm −1 M −1 ) for various concentrations of PNP. All solutions were made in 100 mM potassium phosphate buffer (pH 7.8).

    Journal: PLoS ONE

    Article Title: Rv2607 from Mycobacterium tuberculosis Is a Pyridoxine 5?-Phosphate Oxidase with Unusual Substrate Specificity

    doi: 10.1371/journal.pone.0027643

    Figure Lengend Snippet: Reverse-phase HPLC, NMR, and spectrophotometric analysis of the PNPOx activity of Rv2607. (A) HPLC chromatograms (270 nm) of reaction mixtures containing PNP (i-iv) or PMP (v-viii), FMN, and Rv2607, and the associated control reactions. All reactions were carried out in 25 mM potassium phosphate buffer (pH 7.8), incubated at 25°C for 3 h, and quenched with DNPH (0.7 mM final concentration). Where present, the reaction components were at the following concentrations: 1 mM PNP or PMP, 10 µM FMN, and 10 µM Rv2607. Reaction mixtures contained: (i) PNP, FMN, and Rv2607, (ii) PNP and FMN (enzyme negative control), (iii) PNP and Rv2607 (no added FMN), (iv) FMN and Rv2607 (substrate negative control), (v) PMP, FMN, and Rv2607, (vi) PMP and FMN (enzyme negative control), (vii) PMP and Rv2607 (no added FMN), (viii) FMN and Rv2607 (substrate negative control). The peak to the right of PLP-DNPH is DNPH, which has a retention time of 8.6 min. (B) 1 H NMR analysis of the conversion of PNP into PLP with time. The enzymatic reaction (1 mM PNP, 28 µM enzyme, 10% D 2 O in 25 mM potassium phosphate buffer, pH 7.8) was incubated for 18 hours at 298 K in the spectrometer. Left; stacked 1 H NMR spectra recorded at various time points and after the addition of authentic PLP. The starred peak corresponds to PLP hydrate. Right; a plot of percent substrate conversion versus time. Substrate conversion was determined by comparing integrals of the C2- 1 H signals associated with PLP (7.80 ppm) to that of PNP (7.75 ppm). (C) Michaelis-Menten plot for Rv2607 with PNP as a substrate. The rate of PLP formation was monitored spectrophotometrically (λ max = 388 nm, ε = 4900 cm −1 M −1 ) for various concentrations of PNP. All solutions were made in 100 mM potassium phosphate buffer (pH 7.8).

    Article Snippet: Extracted cofactor from Rv2607 and authentic FMN were analyzed with a Q Star Elite Mass Spectrometer (Applied Biosystems/MDS Sciex) by electrospray ionization positive ion mode with a gold-plated nanospray tip ( ).

    Techniques: High Performance Liquid Chromatography, Nuclear Magnetic Resonance, Activity Assay, Incubation, Concentration Assay, Negative Control, Plasmid Purification

    Calibration curve of PPD (A) on the MALDI-MS/MS system (50–1000 µmol/L) and (B) on the HPLC-UV system (150–1000 µmol/L). Graphs shows the mean values of ( A ) 5 and ( B ) 3 independent measurements and the corresponding r 2 value. Mean +/− SEM.

    Journal: PLoS ONE

    Article Title: Analytical Investigations of Toxic p-Phenylenediamine (PPD) Levels in Clinical Urine Samples with Special Focus on MALDI-MS/MS

    doi: 10.1371/journal.pone.0022191

    Figure Lengend Snippet: Calibration curve of PPD (A) on the MALDI-MS/MS system (50–1000 µmol/L) and (B) on the HPLC-UV system (150–1000 µmol/L). Graphs shows the mean values of ( A ) 5 and ( B ) 3 independent measurements and the corresponding r 2 value. Mean +/− SEM.

    Article Snippet: Compared to the MALDI-MS/MS higher concentrations ( > 1000 µmol/L) were still in the linear range (data not shown).

    Techniques: Mass Spectrometry, High Performance Liquid Chromatography

    MALDI-MS/MS measurements of urine samples from patients with suspected PPD intoxication and controls. ( A ) Measured PPD concentrations in clinical urine samples ( n = 15 ); dotted lines show the LLOQ of the MALDI-MS/MS and the HPLC-UV application, resp. ( B ) Box-and-whisker blot with the 5–95 percentiles for the comparison of PPD, MAPPD and DAPPD peak areas in drug free control urine ( n = 5 ) and clinical samples of intoxication ( n = 15 ). ( C ) Data correlation ( r 2 = 0.7618) of PPD and DAPPD in patient urine ( n = 15 ). Respective p-values are given in the graphs.

    Journal: PLoS ONE

    Article Title: Analytical Investigations of Toxic p-Phenylenediamine (PPD) Levels in Clinical Urine Samples with Special Focus on MALDI-MS/MS

    doi: 10.1371/journal.pone.0022191

    Figure Lengend Snippet: MALDI-MS/MS measurements of urine samples from patients with suspected PPD intoxication and controls. ( A ) Measured PPD concentrations in clinical urine samples ( n = 15 ); dotted lines show the LLOQ of the MALDI-MS/MS and the HPLC-UV application, resp. ( B ) Box-and-whisker blot with the 5–95 percentiles for the comparison of PPD, MAPPD and DAPPD peak areas in drug free control urine ( n = 5 ) and clinical samples of intoxication ( n = 15 ). ( C ) Data correlation ( r 2 = 0.7618) of PPD and DAPPD in patient urine ( n = 15 ). Respective p-values are given in the graphs.

    Article Snippet: For reasons of speed, sensitivity, selectivity and the possibility to simultaneously scan for the PPD metabolites, patient measurements were conducted using MALDI-MS/MS.

    Techniques: Mass Spectrometry, High Performance Liquid Chromatography, Whisker Assay