flight maldi tof mass spectrometry  (Thermo Fisher)


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    Structured Review

    Thermo Fisher flight maldi tof mass spectrometry
    Amino acid sequence alignment of H. volcanii PanA and PanB. Identical residues are shaded in black. Functionally conserved and semiconserved amino acid residues are shaded in grey. Dashes indicate gaps introduced in the protein sequence alignment. Boxed residues were predicted with a > 90% probability to form a coiled-coil conformation (see Materials and Methods). Consensus sequences of the Walker A and B boxes of the P-loop nucleotidase core are indicated below the alignment as GX 2 GXGKT and DEXD, respectively (where X is any amino acid residue). The AAA ATPase second region of homology or SHR motif [(T/S)-(N/S)-X 5 -DXA-X 2 -R-X 2 -RX-(D/E)] is also indicated. The N-terminal sequences of the PanA-His and PanA Δ1-40 -His antigens were identical to residues 2 to 14 and 41 to 51 of the deduced primary sequence, respectively. <t>MALDI-TOF</t> Q-STAR detected the mass spectra of 11 tryptic fragments of the PanB-His antigen, which encompassed 36% of the primary amino acid sequence. The masses (in daltons) and corresponding residue numbers of PanB-His were as follows: 716.3148, 30 to 34; 749.3711, 35 to 40; 1,004.5522, 330 to 337; 1,264.6408, 205 to 261; 1,307.7720, 317 to 328; 1,490.6807, 379 to 392; 1,548.7975, 301 to 313; 1,836.7691, 275 to 289; 2,180.0845, 118 to 137; 2,408.1564, 4 to 24; 2,739.3637, 138 to 162.
    Flight Maldi Tof Mass Spectrometry, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 89/100, based on 42 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "Differential Regulation of the PanA and PanB Proteasome-Activating Nucleotidase and 20S Proteasomal Proteins of the Haloarchaeon Haloferax volcanii †"

    Article Title: Differential Regulation of the PanA and PanB Proteasome-Activating Nucleotidase and 20S Proteasomal Proteins of the Haloarchaeon Haloferax volcanii †

    Journal: Journal of Bacteriology

    doi: 10.1128/JB.186.22.7763-7772.2004

    Amino acid sequence alignment of H. volcanii PanA and PanB. Identical residues are shaded in black. Functionally conserved and semiconserved amino acid residues are shaded in grey. Dashes indicate gaps introduced in the protein sequence alignment. Boxed residues were predicted with a > 90% probability to form a coiled-coil conformation (see Materials and Methods). Consensus sequences of the Walker A and B boxes of the P-loop nucleotidase core are indicated below the alignment as GX 2 GXGKT and DEXD, respectively (where X is any amino acid residue). The AAA ATPase second region of homology or SHR motif [(T/S)-(N/S)-X 5 -DXA-X 2 -R-X 2 -RX-(D/E)] is also indicated. The N-terminal sequences of the PanA-His and PanA Δ1-40 -His antigens were identical to residues 2 to 14 and 41 to 51 of the deduced primary sequence, respectively. MALDI-TOF Q-STAR detected the mass spectra of 11 tryptic fragments of the PanB-His antigen, which encompassed 36% of the primary amino acid sequence. The masses (in daltons) and corresponding residue numbers of PanB-His were as follows: 716.3148, 30 to 34; 749.3711, 35 to 40; 1,004.5522, 330 to 337; 1,264.6408, 205 to 261; 1,307.7720, 317 to 328; 1,490.6807, 379 to 392; 1,548.7975, 301 to 313; 1,836.7691, 275 to 289; 2,180.0845, 118 to 137; 2,408.1564, 4 to 24; 2,739.3637, 138 to 162.
    Figure Legend Snippet: Amino acid sequence alignment of H. volcanii PanA and PanB. Identical residues are shaded in black. Functionally conserved and semiconserved amino acid residues are shaded in grey. Dashes indicate gaps introduced in the protein sequence alignment. Boxed residues were predicted with a > 90% probability to form a coiled-coil conformation (see Materials and Methods). Consensus sequences of the Walker A and B boxes of the P-loop nucleotidase core are indicated below the alignment as GX 2 GXGKT and DEXD, respectively (where X is any amino acid residue). The AAA ATPase second region of homology or SHR motif [(T/S)-(N/S)-X 5 -DXA-X 2 -R-X 2 -RX-(D/E)] is also indicated. The N-terminal sequences of the PanA-His and PanA Δ1-40 -His antigens were identical to residues 2 to 14 and 41 to 51 of the deduced primary sequence, respectively. MALDI-TOF Q-STAR detected the mass spectra of 11 tryptic fragments of the PanB-His antigen, which encompassed 36% of the primary amino acid sequence. The masses (in daltons) and corresponding residue numbers of PanB-His were as follows: 716.3148, 30 to 34; 749.3711, 35 to 40; 1,004.5522, 330 to 337; 1,264.6408, 205 to 261; 1,307.7720, 317 to 328; 1,490.6807, 379 to 392; 1,548.7975, 301 to 313; 1,836.7691, 275 to 289; 2,180.0845, 118 to 137; 2,408.1564, 4 to 24; 2,739.3637, 138 to 162.

    Techniques Used: Sequencing

    2) Product Images from "Characterization of Amylolysin, a Novel Lantibiotic from Bacillus amyloliquefaciens GA1"

    Article Title: Characterization of Amylolysin, a Novel Lantibiotic from Bacillus amyloliquefaciens GA1

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0083037

    MALDI-TOF MS and LC-MS analysis of amylolysin. Mass spectrum of purified amylolysin sample was recorded in negative mode. Insert: LC-MS chromatograms of commercial lanthionine standard and amylolysin hydrolysate. Intensity (%, Y-scale) was recorded by setting the SQD mass analyzer on the specific mass of lanthionine (315 Da).
    Figure Legend Snippet: MALDI-TOF MS and LC-MS analysis of amylolysin. Mass spectrum of purified amylolysin sample was recorded in negative mode. Insert: LC-MS chromatograms of commercial lanthionine standard and amylolysin hydrolysate. Intensity (%, Y-scale) was recorded by setting the SQD mass analyzer on the specific mass of lanthionine (315 Da).

    Techniques Used: Mass Spectrometry, Liquid Chromatography with Mass Spectroscopy, Purification

    3) Product Images from "Physical and Functional Interaction between DNA Ligase III? and Poly(ADP-Ribose) Polymerase 1 in DNA Single-Strand Break Repair"

    Article Title: Physical and Functional Interaction between DNA Ligase III? and Poly(ADP-Ribose) Polymerase 1 in DNA Single-Strand Break Repair

    Journal: Molecular and Cellular Biology

    doi: 10.1128/MCB.23.16.5919-5927.2003

    Identification of DNA ligase III-associated proteins by affinity chromatography. A HeLa nuclear extract was fractionated by using a DNA ligase III affinity chromatography column as described in Materials and Methods. (A) Proteins in comparable fractions eluted with 0.3 M NaCl from the beads liganded by GST and by GST-DNA ligase III (Lig III) were detected by silver staining after separation by SDS-PAGE. The polypeptides indicated were identified by MALDI-TOF mass spectrometry. The identities of PARP-1, Ku70, and Ku80 were verified by immunoblotting with PARP-1 (B) and Ku70 and Ku80 (C) monoclonal antibodies.
    Figure Legend Snippet: Identification of DNA ligase III-associated proteins by affinity chromatography. A HeLa nuclear extract was fractionated by using a DNA ligase III affinity chromatography column as described in Materials and Methods. (A) Proteins in comparable fractions eluted with 0.3 M NaCl from the beads liganded by GST and by GST-DNA ligase III (Lig III) were detected by silver staining after separation by SDS-PAGE. The polypeptides indicated were identified by MALDI-TOF mass spectrometry. The identities of PARP-1, Ku70, and Ku80 were verified by immunoblotting with PARP-1 (B) and Ku70 and Ku80 (C) monoclonal antibodies.

    Techniques Used: Affinity Chromatography, Affinity Column, Silver Staining, SDS Page, Mass Spectrometry

    4) Product Images from "An Engineered Saccharomyces cerevisiae Strain Binds the Broadly Neutralizing Human Immunodeficiency Virus Type 1 Antibody 2G12 and Elicits Mannose-Specific gp120-Binding Antibodies "

    Article Title: An Engineered Saccharomyces cerevisiae Strain Binds the Broadly Neutralizing Human Immunodeficiency Virus Type 1 Antibody 2G12 and Elicits Mannose-Specific gp120-Binding Antibodies

    Journal: Journal of Virology

    doi: 10.1128/JVI.00412-08

    Cross-reactivity of 2G12 with TM yeast expressing mostly Man8 glycans. (A) MALDI-TOF MS was performed on permethylated glycans, released from homogenized TM yeast with PNGase F. The major mass peak at a mass-to-charge ratio ( m/z ) of 2191.73 is representative
    Figure Legend Snippet: Cross-reactivity of 2G12 with TM yeast expressing mostly Man8 glycans. (A) MALDI-TOF MS was performed on permethylated glycans, released from homogenized TM yeast with PNGase F. The major mass peak at a mass-to-charge ratio ( m/z ) of 2191.73 is representative

    Techniques Used: Expressing, Mass Spectrometry

    5) Product Images from "Members of the Large Maf Transcription Family Regulate Insulin Gene Transcription in Islet ? Cells"

    Article Title: Members of the Large Maf Transcription Family Regulate Insulin Gene Transcription in Islet ? Cells

    Journal: Molecular and Cellular Biology

    doi: 10.1128/MCB.23.17.6049-6062.2003

    Biochemical isolation of the InsC1/RIPE3b1 DNA-binding subunit. (A) InsC1 affinity chromatography was performed on βTC-3 nuclear extract (NE). The protein composition of each step was determined by SDS-PAGE and silver staining. First affinity, InsC1-(AC)5 Sepharose chromatography; (AC)5, Sepharose chromatography alone; second affinity, InsC1-(AC)5 Sepharose chromatography. (B) The pooled InsC1/RIPE3b1 binding fractions from the second affinity step were subjected to 2-D electrophoresis. The Coomassie blue-stained spots at pH 7.0 and 4.5 (arrow) of approximately 46-kDa were subjected to MALDI-TOF mass spectrometry. (C) βTC-3 nuclear extract (NE) was resolved by 2-D electrophoresis and probed by Western analysis with αMafA.
    Figure Legend Snippet: Biochemical isolation of the InsC1/RIPE3b1 DNA-binding subunit. (A) InsC1 affinity chromatography was performed on βTC-3 nuclear extract (NE). The protein composition of each step was determined by SDS-PAGE and silver staining. First affinity, InsC1-(AC)5 Sepharose chromatography; (AC)5, Sepharose chromatography alone; second affinity, InsC1-(AC)5 Sepharose chromatography. (B) The pooled InsC1/RIPE3b1 binding fractions from the second affinity step were subjected to 2-D electrophoresis. The Coomassie blue-stained spots at pH 7.0 and 4.5 (arrow) of approximately 46-kDa were subjected to MALDI-TOF mass spectrometry. (C) βTC-3 nuclear extract (NE) was resolved by 2-D electrophoresis and probed by Western analysis with αMafA.

    Techniques Used: Isolation, Binding Assay, Affinity Chromatography, SDS Page, Silver Staining, Chromatography, Electrophoresis, Staining, Mass Spectrometry, Western Blot

    6) Product Images from "Synthesis of Maleimide-End Functionalized Star Polymers and Multimeric Protein-Polymer Conjugates"

    Article Title: Synthesis of Maleimide-End Functionalized Star Polymers and Multimeric Protein-Polymer Conjugates

    Journal: Macromolecules

    doi: 10.1021/ma901540p

    GPC and mass spectra of four-arm poly(NIPAAm) ( 2 ). GPC trace of the 4-armed poly(NIPAAm) (left); MALDI-TOF spectra before (middle) and after (right) aminolysis.
    Figure Legend Snippet: GPC and mass spectra of four-arm poly(NIPAAm) ( 2 ). GPC trace of the 4-armed poly(NIPAAm) (left); MALDI-TOF spectra before (middle) and after (right) aminolysis.

    Techniques Used: Gel Permeation Chromatography

    7) Product Images from "Chemical validation of molecular mimicry: interaction of cholera toxin with Campylobacter lipooligosaccharides"

    Article Title: Chemical validation of molecular mimicry: interaction of cholera toxin with Campylobacter lipooligosaccharides

    Journal:

    doi: 10.1007/s10719-006-9025-9

    MALDI-TOF mass spectra of LS and LF. a ) obtained from mild acid hydrolysis of LS. b : The DMB-labeled oligosaccharides (see peak B in ) obtained from mild acid hydrolysis of LS. c : LF
    Figure Legend Snippet: MALDI-TOF mass spectra of LS and LF. a ) obtained from mild acid hydrolysis of LS. b : The DMB-labeled oligosaccharides (see peak B in ) obtained from mild acid hydrolysis of LS. c : LF

    Techniques Used: Labeling

    Related Articles

    Clone Assay:

    Article Title: Novel Anti-inflammatory Activity of Epoxyazadiradione against Macrophage Migration Inhibitory Factor
    Article Snippet: MIFs were cloned, overexpressed, and purified using Ni-NTA affinity column chromatography. .. The molecular weights of the purified huMIF, Pf MIF, and Py MIF were confirmed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Foster City, CA), which showed exact molecular masses ( m / z ) of 13.430, 13.266, and 13.251 kDa, respectively ( , A , B , and C ).

    Centrifugation:

    Article Title: Members of the Large Maf Transcription Family Regulate Insulin Gene Transcription in Islet ? Cells
    Article Snippet: Peptides were extracted with 60% acetonitrile-0.1% trifluoroacetic acid, dried by vacuum centrifugation, and reconstituted in 10 μl of 0.1% trifluoroacetic acid. .. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry and TOF/TOF tandem mass spectrometry was carried out by using a Voyager 4700 mass spectrometer (Applied Biosystems, Foster City, Calif.) operated in reflectron mode.

    Article Title: Differential Regulation of the PanA and PanB Proteasome-Activating Nucleotidase and 20S Proteasomal Proteins of the Haloarchaeon Haloferax volcanii †
    Article Snippet: Prior to antibody preparation, the N-terminal sequences of PanAΔ1-40 -His and PanA-His were determined by Edman degradation, and the mass spectra of the tryptic fragments of PanB-His were determined by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (Applied Biosystems QSTAR XL hybrid LC/MS/MS; University of Florida ICBR Protein Chemistry Core). .. For immunoblotting, cells were harvested by centrifugation (5,000 × g ; 10 min at 4°C) and stored at −70°C.

    Filtration:

    Article Title: Novel Anti-inflammatory Activity of Epoxyazadiradione against Macrophage Migration Inhibitory Factor
    Article Snippet: Because Ni-NTA affinity chromatography purification co-purifies some contaminating proteins along with the desired MIFs, we further subjected Ni-NTA-purified MIFs to Sephadex 75 gel filtration (Amersham Biosciences), contaminating protein-free fractions of MIFs were collected, and purity was confirmed by silver staining ( , A , B , and C , insets ). .. The molecular weights of the purified huMIF, Pf MIF, and Py MIF were confirmed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Foster City, CA), which showed exact molecular masses ( m / z ) of 13.430, 13.266, and 13.251 kDa, respectively ( , A , B , and C ).

    Article Title: Cooperative Hybridization of γPNA Miniprobes to a Repeating Sequence Motif and Application to Telomere Analysis
    Article Snippet: DNA oligonucleotides were purchased from Integrated DNA Technologies and used after standard desalting by gel filtration chromatography. .. DNAs contained different numbers of telomeric repeats Telo-n (5’-TTAGGG-3’)n , where n = 1-4 or 6. γ-minipeg-t-Boc-protected PNA monomers and oligomers were synthesized according to published procedures, , purified using reverse-phase high performance liquid chromatography (HPLC), and characterized by matrix-assisted laser desorption/ionization time of flight (MALDI-ToF) mass spectrometry (Applied Biosystems, Voyager DE sSTR) using α-cyano-4-hydroxycinnamic acid as the matrix.

    Mass Spectrometry:

    Article Title: Engineering an antibody with picomolar affinity to DOTA chelates of multiple radionuclides for pretargeted radioimmunotherapy and imaging
    Article Snippet: .. Fractions containing DOTA-Bn-biotin were confirmed using matrix assisted laser desorption instrument time of flight (MALDI-TOF) mass spectrometry (Applied Biosystems Model Voyager DE-STR). .. Chemical purity was assessed by analytical HPLC (Agilent Model 1100 HPLC, 2.1 × 150 mm, buffer A = 0.05% TFA, buffer B = 0.0425% TFA in 80% acetonitrile, 2-100% B gradient for 45 min).

    Article Title: Chemical validation of molecular mimicry: interaction of cholera toxin with Campylobacter lipooligosaccharides
    Article Snippet: .. The ABOE-labeled oligosaccharides and the DMB-labeled oligosaccharides obtained from the Sep-Pak C18 cartridge were analyzed by Matrix-Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) mass spectrometry using a Model 4700 Proteomics Analyzer (Applied Biosystems, Foster City, CA, USA) equipped with a nitrogen UV laser (λ=337 nm) with a matrix of 2,5-dihydroxybenzoic acid at a concentration of 10 mg/ml in 10% aqueous ethanol. ..

    Article Title: Physical and Functional Interaction between DNA Ligase III? and Poly(ADP-Ribose) Polymerase 1 in DNA Single-Strand Break Repair
    Article Snippet: .. After concentration to 5 μl, an equal volume of 50% acetonitrile-5% formic acid was added, and an aliquot (1 μl) was analyzed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (Voyager-DE Pro; Applied Biosystems). ..

    Article Title: An Engineered Saccharomyces cerevisiae Strain Binds the Broadly Neutralizing Human Immunodeficiency Virus Type 1 Antibody 2G12 and Elicits Mannose-Specific gp120-Binding Antibodies
    Article Snippet: .. Glycans were permethylated for analysis by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) using a QSTAR hybrid mass spectrometer (Applied Biosystems, Foster City, CA) and derivatized with 2-aminobenzamide (2AB) for normal-phase high-performance liquid chromatography (NP-HPLC) analysis on a Dionex DX-600 HPLC system (Sunnyvale, CA). .. 2AB-derivatized N-linked glycans released from RNase B were used as a standard for HPLC.

    Article Title: The Level of Ets-1 Protein Is Regulated by Poly(ADP-Ribose) Polymerase-1 (PARP-1) in Cancer Cells to Prevent DNA Damage
    Article Snippet: .. The trypsin digests were then analysed by matrix-assisted laser desorption/ionisation-time of flight (MALDI-TOF) mass spectrometry on a Voyager-DE STR (Applied Biosystems) as described in reference . ..

    Article Title: The Peptide ERα17p Is a GPER Inverse Agonist that Exerts Antiproliferative Effects in Breast Cancer Cells
    Article Snippet: .. The purified peptides were then characterized by matrix assisted laser desorption ionization time of flight (MALDI-TOF) mass spectrometry using a 4700 Proteomic Analyzer (Applied Biosystems, Foster City, CA, USA). α cyano-4-hydroxycinnamic acid (HCCA) was used as a matrix. ..

    Article Title: Novel Anti-inflammatory Activity of Epoxyazadiradione against Macrophage Migration Inhibitory Factor
    Article Snippet: .. The molecular weights of the purified huMIF, Pf MIF, and Py MIF were confirmed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Foster City, CA), which showed exact molecular masses ( m / z ) of 13.430, 13.266, and 13.251 kDa, respectively ( , A , B , and C ). .. These purified MIFs were then used for further studies.

    Article Title: Active, soluble recombinant melittin purified by extracting insoluble lysate of Escherichia coli without denaturation
    Article Snippet: .. Protein molecular weights were confirmed with matrix-assisted laser desorption-ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Grand Island, NY). .. The surfactant-containing insoluble fraction extracts were analyzed by SDS-PAGE.

    Article Title: Members of the Large Maf Transcription Family Regulate Insulin Gene Transcription in Islet ? Cells
    Article Snippet: .. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry and TOF/TOF tandem mass spectrometry was carried out by using a Voyager 4700 mass spectrometer (Applied Biosystems, Foster City, Calif.) operated in reflectron mode. ..

    Article Title: Cooperative Hybridization of γPNA Miniprobes to a Repeating Sequence Motif and Application to Telomere Analysis
    Article Snippet: .. DNAs contained different numbers of telomeric repeats Telo-n (5’-TTAGGG-3’)n , where n = 1-4 or 6. γ-minipeg-t-Boc-protected PNA monomers and oligomers were synthesized according to published procedures, , purified using reverse-phase high performance liquid chromatography (HPLC), and characterized by matrix-assisted laser desorption/ionization time of flight (MALDI-ToF) mass spectrometry (Applied Biosystems, Voyager DE sSTR) using α-cyano-4-hydroxycinnamic acid as the matrix. .. Cy3 was a generous gift from B. Schmidt of the Molecular Biosensor and Imaging Center at Carnegie Mellon University.

    Article Title: Cationized liposomal keto-mycolic acids isolated from Mycobacterium bovis bacillus Calmette-Guérin induce antitumor immunity in a syngeneic murine bladder cancer model
    Article Snippet: .. The molecular weight of each MA methylester was determined by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry on a Voyager DE-STR workstation (Applied Biosystems, Foster City, CA) using 2.4-dihydroxybenzoic acid (2.5-DHB) as a matrix, as reported previously [ ]. ..

    Article Title: Characterization of Amylolysin, a Novel Lantibiotic from Bacillus amyloliquefaciens GA1
    Article Snippet: .. Mass spectrometry The molecular mass of the purified amylolysin was determined by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry using a 4700 Proteomics analyzer (Applied Biosystem). .. Purified amylolysin was mixed with an equal volume of α-cyanohydroxycinnamic acid solution (10 mg/ml) and spotted onto the MALDI plate.

    Article Title: Synthesis of Maleimide-End Functionalized Star Polymers and Multimeric Protein-Polymer Conjugates
    Article Snippet: .. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry was performed on an Applied Biosystems Voyager-DE STR and operated in linear mode with an external calibration. ..

    Article Title: Differential Regulation of the PanA and PanB Proteasome-Activating Nucleotidase and 20S Proteasomal Proteins of the Haloarchaeon Haloferax volcanii †
    Article Snippet: .. Prior to antibody preparation, the N-terminal sequences of PanAΔ1-40 -His and PanA-His were determined by Edman degradation, and the mass spectra of the tryptic fragments of PanB-His were determined by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (Applied Biosystems QSTAR XL hybrid LC/MS/MS; University of Florida ICBR Protein Chemistry Core). .. PanAΔ1-40 -His and PanB-His proteins purified from recombinant E. coli were separated by SDS-7.5% PAGE, excised from the gel, and used as antigen to raise polyclonal antibodies in rabbit (Cocalico Biologicals, Reamstown, Pa.).

    Article Title: Mycoplasma genitalium Rapidly Disseminates to the Upper Reproductive Tracts and Knees of Female Mice following Vaginal Inoculation ▿
    Article Snippet: .. Each of the immunogenic M. genitalium proteins that were recognized by infected mouse sera was carefully excised from separate Coomassie-stained gels, digested with trypsin, and then identified using matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (UTMB Biomolecular Resource Facility) and a 4700 proteomics analyzer (Applied Biosystems). .. To evaluate antibody responses from individual mice, we tested serum samples collected upon euthanasia from the 12 estradiol-treated animals 10 weeks p.i. in the hydrosalpinx study described above.

    Synthesized:

    Article Title: Engineering an antibody with picomolar affinity to DOTA chelates of multiple radionuclides for pretargeted radioimmunotherapy and imaging
    Article Snippet: DOTA-Bn-biotin ( ) was synthesized by dissolving Amine-PEG3 -Biotin purchased from Thermo Scientific (Rockford, IL) and S-2-(4-Isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane-tetraacetic acid (p-SCN-Bn-DOTA) purchased from Macrocyclics in dimethyl sulfoxide (DMSO) with a 10 fold molar excess of triethylamine. .. Fractions containing DOTA-Bn-biotin were confirmed using matrix assisted laser desorption instrument time of flight (MALDI-TOF) mass spectrometry (Applied Biosystems Model Voyager DE-STR).

    Article Title: The Peptide ERα17p Is a GPER Inverse Agonist that Exerts Antiproliferative Effects in Breast Cancer Cells
    Article Snippet: The peptides were manually synthesized via Fmoc solid phase peptide synthesis (SPPS) using a preloaded Fmoc-Thr-Novasyn TGA resin (Merck, Fontenay sous Bois, France), as previously described [ , ]. .. The purified peptides were then characterized by matrix assisted laser desorption ionization time of flight (MALDI-TOF) mass spectrometry using a 4700 Proteomic Analyzer (Applied Biosystems, Foster City, CA, USA). α cyano-4-hydroxycinnamic acid (HCCA) was used as a matrix.

    Article Title: Cooperative Hybridization of γPNA Miniprobes to a Repeating Sequence Motif and Application to Telomere Analysis
    Article Snippet: .. DNAs contained different numbers of telomeric repeats Telo-n (5’-TTAGGG-3’)n , where n = 1-4 or 6. γ-minipeg-t-Boc-protected PNA monomers and oligomers were synthesized according to published procedures, , purified using reverse-phase high performance liquid chromatography (HPLC), and characterized by matrix-assisted laser desorption/ionization time of flight (MALDI-ToF) mass spectrometry (Applied Biosystems, Voyager DE sSTR) using α-cyano-4-hydroxycinnamic acid as the matrix. .. Cy3 was a generous gift from B. Schmidt of the Molecular Biosensor and Imaging Center at Carnegie Mellon University.

    Incubation:

    Article Title: Physical and Functional Interaction between DNA Ligase III? and Poly(ADP-Ribose) Polymerase 1 in DNA Single-Strand Break Repair
    Article Snippet: The supernatant was discarded and 20 μl of 25 mM ammonium bicarbonate was added to the gel pieces, which were then incubated at 37°C for 16 h. After the addition of 100 μl of H2 O, samples were vortexed for 10 min. .. After concentration to 5 μl, an equal volume of 50% acetonitrile-5% formic acid was added, and an aliquot (1 μl) was analyzed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (Voyager-DE Pro; Applied Biosystems).

    Expressing:

    Article Title: Active, soluble recombinant melittin purified by extracting insoluble lysate of Escherichia coli without denaturation
    Article Snippet: Paragraph title: Expression and purification of Melittin ... Protein molecular weights were confirmed with matrix-assisted laser desorption-ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Grand Island, NY).

    Bradford Assay:

    Article Title: Active, soluble recombinant melittin purified by extracting insoluble lysate of Escherichia coli without denaturation
    Article Snippet: Total protein was determined by the Bradford Assay (Thermo Scientific, Hanover Park, IL) used according to the manufacturer recommendations and measured with a Beckman model DU640 spectrophotometer (Brea, CA). .. Protein molecular weights were confirmed with matrix-assisted laser desorption-ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Grand Island, NY).

    Modification:

    Article Title: Members of the Large Maf Transcription Family Regulate Insulin Gene Transcription in Islet ? Cells
    Article Snippet: The gel slice was then dehydrated with acetonitrile and rehydrated with 15 μl of 25 mM NH4 HCO3 containing 0.01 μg of modified trypsin (Promega)/μl, and trypsin digestion was carried out for 2 h at 37°C. .. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry and TOF/TOF tandem mass spectrometry was carried out by using a Voyager 4700 mass spectrometer (Applied Biosystems, Foster City, Calif.) operated in reflectron mode.

    High Performance Liquid Chromatography:

    Article Title: Engineering an antibody with picomolar affinity to DOTA chelates of multiple radionuclides for pretargeted radioimmunotherapy and imaging
    Article Snippet: HPLC purification was performed on a C-18 reverse-phase column (Agilent Model 1100 HPLC, 1 × 25 cm, buffer A = 0.05% trifluoroacetic acid (TFA), buffer B = 0.0425% TFA in 80% acetonitrile, 2 – 100% B gradient for 98 min). .. Fractions containing DOTA-Bn-biotin were confirmed using matrix assisted laser desorption instrument time of flight (MALDI-TOF) mass spectrometry (Applied Biosystems Model Voyager DE-STR).

    Article Title: Chemical validation of molecular mimicry: interaction of cholera toxin with Campylobacter lipooligosaccharides
    Article Snippet: The fluorescent peak at the retention time of 26.1 min, corresponding to that of the DMB-labeled oligosaccharide, was recovered from HPLC and desalted by a Sep-Pak C18 cartridge. .. The ABOE-labeled oligosaccharides and the DMB-labeled oligosaccharides obtained from the Sep-Pak C18 cartridge were analyzed by Matrix-Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) mass spectrometry using a Model 4700 Proteomics Analyzer (Applied Biosystems, Foster City, CA, USA) equipped with a nitrogen UV laser (λ=337 nm) with a matrix of 2,5-dihydroxybenzoic acid at a concentration of 10 mg/ml in 10% aqueous ethanol.

    Article Title: An Engineered Saccharomyces cerevisiae Strain Binds the Broadly Neutralizing Human Immunodeficiency Virus Type 1 Antibody 2G12 and Elicits Mannose-Specific gp120-Binding Antibodies
    Article Snippet: .. Glycans were permethylated for analysis by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) using a QSTAR hybrid mass spectrometer (Applied Biosystems, Foster City, CA) and derivatized with 2-aminobenzamide (2AB) for normal-phase high-performance liquid chromatography (NP-HPLC) analysis on a Dionex DX-600 HPLC system (Sunnyvale, CA). .. 2AB-derivatized N-linked glycans released from RNase B were used as a standard for HPLC.

    Article Title: The Peptide ERα17p Is a GPER Inverse Agonist that Exerts Antiproliferative Effects in Breast Cancer Cells
    Article Snippet: The crude products were purified by semi-preparative reverse-phase high performance liquid chromatography RP-HPLC (Waters, Saint-Quentin en Yveline, France) using a Waters 600 pump and controller and a 2487 UV–Vis detector (λ = 220 nm, flow rate = 5 mL/min). .. The purified peptides were then characterized by matrix assisted laser desorption ionization time of flight (MALDI-TOF) mass spectrometry using a 4700 Proteomic Analyzer (Applied Biosystems, Foster City, CA, USA). α cyano-4-hydroxycinnamic acid (HCCA) was used as a matrix.

    Article Title: Cooperative Hybridization of γPNA Miniprobes to a Repeating Sequence Motif and Application to Telomere Analysis
    Article Snippet: .. DNAs contained different numbers of telomeric repeats Telo-n (5’-TTAGGG-3’)n , where n = 1-4 or 6. γ-minipeg-t-Boc-protected PNA monomers and oligomers were synthesized according to published procedures, , purified using reverse-phase high performance liquid chromatography (HPLC), and characterized by matrix-assisted laser desorption/ionization time of flight (MALDI-ToF) mass spectrometry (Applied Biosystems, Voyager DE sSTR) using α-cyano-4-hydroxycinnamic acid as the matrix. .. Cy3 was a generous gift from B. Schmidt of the Molecular Biosensor and Imaging Center at Carnegie Mellon University.

    Flow Cytometry:

    Article Title: Engineering an antibody with picomolar affinity to DOTA chelates of multiple radionuclides for pretargeted radioimmunotherapy and imaging
    Article Snippet: Flow through was monitored by absorbance detection at 280 nanometers. .. Fractions containing DOTA-Bn-biotin were confirmed using matrix assisted laser desorption instrument time of flight (MALDI-TOF) mass spectrometry (Applied Biosystems Model Voyager DE-STR).

    Article Title: The Peptide ERα17p Is a GPER Inverse Agonist that Exerts Antiproliferative Effects in Breast Cancer Cells
    Article Snippet: Analytical RP-HPLC conditions: λ = 220 nm, flow rate = 1 mL/min. .. The purified peptides were then characterized by matrix assisted laser desorption ionization time of flight (MALDI-TOF) mass spectrometry using a 4700 Proteomic Analyzer (Applied Biosystems, Foster City, CA, USA). α cyano-4-hydroxycinnamic acid (HCCA) was used as a matrix.

    Silver Staining:

    Article Title: Novel Anti-inflammatory Activity of Epoxyazadiradione against Macrophage Migration Inhibitory Factor
    Article Snippet: Because Ni-NTA affinity chromatography purification co-purifies some contaminating proteins along with the desired MIFs, we further subjected Ni-NTA-purified MIFs to Sephadex 75 gel filtration (Amersham Biosciences), contaminating protein-free fractions of MIFs were collected, and purity was confirmed by silver staining ( , A , B , and C , insets ). .. The molecular weights of the purified huMIF, Pf MIF, and Py MIF were confirmed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Foster City, CA), which showed exact molecular masses ( m / z ) of 13.430, 13.266, and 13.251 kDa, respectively ( , A , B , and C ).

    Chromatography:

    Article Title: Chemical validation of molecular mimicry: interaction of cholera toxin with Campylobacter lipooligosaccharides
    Article Snippet: The DMB-labeled oligosaccharides were analyzed by HPLC using an ODS-column Axxiom chromatography with a fluorescence detector at an excitation wavelength of 373 nm and an emission wavelength of 448 nm. .. The ABOE-labeled oligosaccharides and the DMB-labeled oligosaccharides obtained from the Sep-Pak C18 cartridge were analyzed by Matrix-Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) mass spectrometry using a Model 4700 Proteomics Analyzer (Applied Biosystems, Foster City, CA, USA) equipped with a nitrogen UV laser (λ=337 nm) with a matrix of 2,5-dihydroxybenzoic acid at a concentration of 10 mg/ml in 10% aqueous ethanol.

    Article Title: Novel Anti-inflammatory Activity of Epoxyazadiradione against Macrophage Migration Inhibitory Factor
    Article Snippet: MIFs were cloned, overexpressed, and purified using Ni-NTA affinity column chromatography. .. The molecular weights of the purified huMIF, Pf MIF, and Py MIF were confirmed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Foster City, CA), which showed exact molecular masses ( m / z ) of 13.430, 13.266, and 13.251 kDa, respectively ( , A , B , and C ).

    Article Title: Cooperative Hybridization of γPNA Miniprobes to a Repeating Sequence Motif and Application to Telomere Analysis
    Article Snippet: DNA oligonucleotides were purchased from Integrated DNA Technologies and used after standard desalting by gel filtration chromatography. .. DNAs contained different numbers of telomeric repeats Telo-n (5’-TTAGGG-3’)n , where n = 1-4 or 6. γ-minipeg-t-Boc-protected PNA monomers and oligomers were synthesized according to published procedures, , purified using reverse-phase high performance liquid chromatography (HPLC), and characterized by matrix-assisted laser desorption/ionization time of flight (MALDI-ToF) mass spectrometry (Applied Biosystems, Voyager DE sSTR) using α-cyano-4-hydroxycinnamic acid as the matrix.

    Article Title: Synthesis of Maleimide-End Functionalized Star Polymers and Multimeric Protein-Polymer Conjugates
    Article Snippet: Chromatograms were processed with the EZStart 7.2 chromatography software. .. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry was performed on an Applied Biosystems Voyager-DE STR and operated in linear mode with an external calibration.

    Transferring:

    Article Title: Members of the Large Maf Transcription Family Regulate Insulin Gene Transcription in Islet ? Cells
    Article Snippet: The peptides were then desalted and concentrated into 2 μl of 60% acetonitrile-0.1% trifluoroacetic acid by using ZipTipC18 pipette tips (Millipore), and then 0.4 μl was applied to a target plate and overlaid with 0.4 μl of α-cyano-4-hydroxycinnamic acid matrix. .. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry and TOF/TOF tandem mass spectrometry was carried out by using a Voyager 4700 mass spectrometer (Applied Biosystems, Foster City, Calif.) operated in reflectron mode.

    Infection:

    Article Title: Mycoplasma genitalium Rapidly Disseminates to the Upper Reproductive Tracts and Knees of Female Mice following Vaginal Inoculation ▿
    Article Snippet: .. Each of the immunogenic M. genitalium proteins that were recognized by infected mouse sera was carefully excised from separate Coomassie-stained gels, digested with trypsin, and then identified using matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (UTMB Biomolecular Resource Facility) and a 4700 proteomics analyzer (Applied Biosystems). .. To evaluate antibody responses from individual mice, we tested serum samples collected upon euthanasia from the 12 estradiol-treated animals 10 weeks p.i. in the hydrosalpinx study described above.

    Imaging:

    Article Title: Cooperative Hybridization of γPNA Miniprobes to a Repeating Sequence Motif and Application to Telomere Analysis
    Article Snippet: DNAs contained different numbers of telomeric repeats Telo-n (5’-TTAGGG-3’)n , where n = 1-4 or 6. γ-minipeg-t-Boc-protected PNA monomers and oligomers were synthesized according to published procedures, , purified using reverse-phase high performance liquid chromatography (HPLC), and characterized by matrix-assisted laser desorption/ionization time of flight (MALDI-ToF) mass spectrometry (Applied Biosystems, Voyager DE sSTR) using α-cyano-4-hydroxycinnamic acid as the matrix. .. Cy3 was a generous gift from B. Schmidt of the Molecular Biosensor and Imaging Center at Carnegie Mellon University.

    Sequencing:

    Article Title: Physical and Functional Interaction between DNA Ligase III? and Poly(ADP-Ribose) Polymerase 1 in DNA Single-Strand Break Repair
    Article Snippet: The dried gel pieces were resuspended in 20 μl of 25 mM ammonium bicarbonate containing 5 μg of sequencing-grade trypsin (Promega)/ml, vortexed for 10 min, and then incubated at 4°C for 30 min. .. After concentration to 5 μl, an equal volume of 50% acetonitrile-5% formic acid was added, and an aliquot (1 μl) was analyzed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (Voyager-DE Pro; Applied Biosystems).

    Article Title: Members of the Large Maf Transcription Family Regulate Insulin Gene Transcription in Islet ? Cells
    Article Snippet: Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry and TOF/TOF tandem mass spectrometry was carried out by using a Voyager 4700 mass spectrometer (Applied Biosystems, Foster City, Calif.) operated in reflectron mode. .. Ions ([M+H]) corresponding to peptide masses were entered into the mouse expressed sequence tag (EST) database, which was searched by using the MS-FIT algorithm (prospector.ucsf.edu).

    Quantitation Assay:

    Article Title: Engineering an antibody with picomolar affinity to DOTA chelates of multiple radionuclides for pretargeted radioimmunotherapy and imaging
    Article Snippet: Fractions containing DOTA-Bn-biotin were confirmed using matrix assisted laser desorption instrument time of flight (MALDI-TOF) mass spectrometry (Applied Biosystems Model Voyager DE-STR). .. DOTA-Bn-biotin concentration was determined using a biotin quantitation kit (Thermo Scientific) following the manufacturer’s instructions. (+)-(2S)-2-(4-Aminobutyl)-1,4,7,10-tetra-azacyclododecane-1,4,7,10-tetrayltetra-acetic acid (DOTA-alkyl) has been synthesized previously [ ].

    Recombinant:

    Article Title: Active, soluble recombinant melittin purified by extracting insoluble lysate of Escherichia coli without denaturation
    Article Snippet: Recombinant melittin was purified as described, then digested with 2 units (U) of thrombin for 2 hours at 37°C. .. Protein molecular weights were confirmed with matrix-assisted laser desorption-ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Grand Island, NY).

    Article Title: Differential Regulation of the PanA and PanB Proteasome-Activating Nucleotidase and 20S Proteasomal Proteins of the Haloarchaeon Haloferax volcanii †
    Article Snippet: Prior to antibody preparation, the N-terminal sequences of PanAΔ1-40 -His and PanA-His were determined by Edman degradation, and the mass spectra of the tryptic fragments of PanB-His were determined by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (Applied Biosystems QSTAR XL hybrid LC/MS/MS; University of Florida ICBR Protein Chemistry Core). .. PanAΔ1-40 -His and PanB-His proteins purified from recombinant E. coli were separated by SDS-7.5% PAGE, excised from the gel, and used as antigen to raise polyclonal antibodies in rabbit (Cocalico Biologicals, Reamstown, Pa.).

    Molecular Weight:

    Article Title: Cationized liposomal keto-mycolic acids isolated from Mycobacterium bovis bacillus Calmette-Guérin induce antitumor immunity in a syngeneic murine bladder cancer model
    Article Snippet: .. The molecular weight of each MA methylester was determined by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry on a Voyager DE-STR workstation (Applied Biosystems, Foster City, CA) using 2.4-dihydroxybenzoic acid (2.5-DHB) as a matrix, as reported previously [ ]. ..

    Article Title: Synthesis of Maleimide-End Functionalized Star Polymers and Multimeric Protein-Polymer Conjugates
    Article Snippet: The number-average molecular weight (Mn ) by dynamic light scattering (DLS) was collected from a Viscotek GPCmax VE2001 GPC solvent/sample module with 270 Dual detector and VE 3580 RI detector. .. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry was performed on an Applied Biosystems Voyager-DE STR and operated in linear mode with an external calibration.

    Nucleic Acid Electrophoresis:

    Article Title: Members of the Large Maf Transcription Family Regulate Insulin Gene Transcription in Islet ? Cells
    Article Snippet: Proteins were separated by 2-D gel electrophoresis and visualized by using the Colloidal Blue staining kit (Invitrogen, Carlsbad, Calif.). .. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry and TOF/TOF tandem mass spectrometry was carried out by using a Voyager 4700 mass spectrometer (Applied Biosystems, Foster City, Calif.) operated in reflectron mode.

    Article Title: Mycoplasma genitalium Rapidly Disseminates to the Upper Reproductive Tracts and Knees of Female Mice following Vaginal Inoculation ▿
    Article Snippet: To investigate the host response to M. genitalium infection and determine the most immunodominant proteins, serum was collected from M. genitalium G37-infected or mock-inoculated mice (at the termination date [77 d.p.i.]) for immunoblot experiments using M. genitalium G37 lysates, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels, and standard methods. .. Each of the immunogenic M. genitalium proteins that were recognized by infected mouse sera was carefully excised from separate Coomassie-stained gels, digested with trypsin, and then identified using matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (UTMB Biomolecular Resource Facility) and a 4700 proteomics analyzer (Applied Biosystems).

    Fluorescence:

    Article Title: Chemical validation of molecular mimicry: interaction of cholera toxin with Campylobacter lipooligosaccharides
    Article Snippet: The DMB-labeled oligosaccharides were analyzed by HPLC using an ODS-column Axxiom chromatography with a fluorescence detector at an excitation wavelength of 373 nm and an emission wavelength of 448 nm. .. The ABOE-labeled oligosaccharides and the DMB-labeled oligosaccharides obtained from the Sep-Pak C18 cartridge were analyzed by Matrix-Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) mass spectrometry using a Model 4700 Proteomics Analyzer (Applied Biosystems, Foster City, CA, USA) equipped with a nitrogen UV laser (λ=337 nm) with a matrix of 2,5-dihydroxybenzoic acid at a concentration of 10 mg/ml in 10% aqueous ethanol.

    Labeling:

    Article Title: The Peptide ERα17p Is a GPER Inverse Agonist that Exerts Antiproliferative Effects in Breast Cancer Cells
    Article Snippet: The purified peptides were then characterized by matrix assisted laser desorption ionization time of flight (MALDI-TOF) mass spectrometry using a 4700 Proteomic Analyzer (Applied Biosystems, Foster City, CA, USA). α cyano-4-hydroxycinnamic acid (HCCA) was used as a matrix. .. N-terminal ERα17p labeling was carried out by using 5(6)-carboxyfluorescein (fluorescein-Ahx-ERα17p-COOH, where Ahx corresponds to aminohexanoic acid), following the standard Fmoc peptide synthesis protocol [ , ].

    Article Title: Cooperative Hybridization of γPNA Miniprobes to a Repeating Sequence Motif and Application to Telomere Analysis
    Article Snippet: DNAs contained different numbers of telomeric repeats Telo-n (5’-TTAGGG-3’)n , where n = 1-4 or 6. γ-minipeg-t-Boc-protected PNA monomers and oligomers were synthesized according to published procedures, , purified using reverse-phase high performance liquid chromatography (HPLC), and characterized by matrix-assisted laser desorption/ionization time of flight (MALDI-ToF) mass spectrometry (Applied Biosystems, Voyager DE sSTR) using α-cyano-4-hydroxycinnamic acid as the matrix. .. For Cy3 labeling, a carboxylic acid derivative of Cy3 was coupled to the γPNA prior to cleavage using similar procedures to all of the other monomers.

    Mouse Assay:

    Article Title: Mycoplasma genitalium Rapidly Disseminates to the Upper Reproductive Tracts and Knees of Female Mice following Vaginal Inoculation ▿
    Article Snippet: To investigate the host response to M. genitalium infection and determine the most immunodominant proteins, serum was collected from M. genitalium G37-infected or mock-inoculated mice (at the termination date [77 d.p.i.]) for immunoblot experiments using M. genitalium G37 lysates, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels, and standard methods. .. Each of the immunogenic M. genitalium proteins that were recognized by infected mouse sera was carefully excised from separate Coomassie-stained gels, digested with trypsin, and then identified using matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (UTMB Biomolecular Resource Facility) and a 4700 proteomics analyzer (Applied Biosystems).

    Affinity Column:

    Article Title: Novel Anti-inflammatory Activity of Epoxyazadiradione against Macrophage Migration Inhibitory Factor
    Article Snippet: MIFs were cloned, overexpressed, and purified using Ni-NTA affinity column chromatography. .. The molecular weights of the purified huMIF, Pf MIF, and Py MIF were confirmed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Foster City, CA), which showed exact molecular masses ( m / z ) of 13.430, 13.266, and 13.251 kDa, respectively ( , A , B , and C ).

    Polyacrylamide Gel Electrophoresis:

    Article Title: Synthesis of Maleimide-End Functionalized Star Polymers and Multimeric Protein-Polymer Conjugates
    Article Snippet: Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was carried out with 4–20% Tris-Glycine gels (Invitrogen, 1.0 mm × 12 well). .. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry was performed on an Applied Biosystems Voyager-DE STR and operated in linear mode with an external calibration.

    Article Title: Differential Regulation of the PanA and PanB Proteasome-Activating Nucleotidase and 20S Proteasomal Proteins of the Haloarchaeon Haloferax volcanii †
    Article Snippet: Prior to antibody preparation, the N-terminal sequences of PanAΔ1-40 -His and PanA-His were determined by Edman degradation, and the mass spectra of the tryptic fragments of PanB-His were determined by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (Applied Biosystems QSTAR XL hybrid LC/MS/MS; University of Florida ICBR Protein Chemistry Core). .. PanAΔ1-40 -His and PanB-His proteins purified from recombinant E. coli were separated by SDS-7.5% PAGE, excised from the gel, and used as antigen to raise polyclonal antibodies in rabbit (Cocalico Biologicals, Reamstown, Pa.).

    Purification:

    Article Title: Engineering an antibody with picomolar affinity to DOTA chelates of multiple radionuclides for pretargeted radioimmunotherapy and imaging
    Article Snippet: HPLC purification was performed on a C-18 reverse-phase column (Agilent Model 1100 HPLC, 1 × 25 cm, buffer A = 0.05% trifluoroacetic acid (TFA), buffer B = 0.0425% TFA in 80% acetonitrile, 2 – 100% B gradient for 98 min). .. Fractions containing DOTA-Bn-biotin were confirmed using matrix assisted laser desorption instrument time of flight (MALDI-TOF) mass spectrometry (Applied Biosystems Model Voyager DE-STR).

    Article Title: An Engineered Saccharomyces cerevisiae Strain Binds the Broadly Neutralizing Human Immunodeficiency Virus Type 1 Antibody 2G12 and Elicits Mannose-Specific gp120-Binding Antibodies
    Article Snippet: N-linked glycans on log-phase TM yeast cells or purified Gp38 glycoprotein were analyzed by the Glycotechnology Core Resource, University of California, San Diego. .. Glycans were permethylated for analysis by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) using a QSTAR hybrid mass spectrometer (Applied Biosystems, Foster City, CA) and derivatized with 2-aminobenzamide (2AB) for normal-phase high-performance liquid chromatography (NP-HPLC) analysis on a Dionex DX-600 HPLC system (Sunnyvale, CA).

    Article Title: The Peptide ERα17p Is a GPER Inverse Agonist that Exerts Antiproliferative Effects in Breast Cancer Cells
    Article Snippet: .. The purified peptides were then characterized by matrix assisted laser desorption ionization time of flight (MALDI-TOF) mass spectrometry using a 4700 Proteomic Analyzer (Applied Biosystems, Foster City, CA, USA). α cyano-4-hydroxycinnamic acid (HCCA) was used as a matrix. ..

    Article Title: Novel Anti-inflammatory Activity of Epoxyazadiradione against Macrophage Migration Inhibitory Factor
    Article Snippet: .. The molecular weights of the purified huMIF, Pf MIF, and Py MIF were confirmed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Foster City, CA), which showed exact molecular masses ( m / z ) of 13.430, 13.266, and 13.251 kDa, respectively ( , A , B , and C ). .. These purified MIFs were then used for further studies.

    Article Title: Active, soluble recombinant melittin purified by extracting insoluble lysate of Escherichia coli without denaturation
    Article Snippet: Paragraph title: Expression and purification of Melittin ... Protein molecular weights were confirmed with matrix-assisted laser desorption-ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Grand Island, NY).

    Article Title: Cooperative Hybridization of γPNA Miniprobes to a Repeating Sequence Motif and Application to Telomere Analysis
    Article Snippet: .. DNAs contained different numbers of telomeric repeats Telo-n (5’-TTAGGG-3’)n , where n = 1-4 or 6. γ-minipeg-t-Boc-protected PNA monomers and oligomers were synthesized according to published procedures, , purified using reverse-phase high performance liquid chromatography (HPLC), and characterized by matrix-assisted laser desorption/ionization time of flight (MALDI-ToF) mass spectrometry (Applied Biosystems, Voyager DE sSTR) using α-cyano-4-hydroxycinnamic acid as the matrix. .. Cy3 was a generous gift from B. Schmidt of the Molecular Biosensor and Imaging Center at Carnegie Mellon University.

    Article Title: Characterization of Amylolysin, a Novel Lantibiotic from Bacillus amyloliquefaciens GA1
    Article Snippet: .. Mass spectrometry The molecular mass of the purified amylolysin was determined by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry using a 4700 Proteomics analyzer (Applied Biosystem). .. Purified amylolysin was mixed with an equal volume of α-cyanohydroxycinnamic acid solution (10 mg/ml) and spotted onto the MALDI plate.

    Article Title: Differential Regulation of the PanA and PanB Proteasome-Activating Nucleotidase and 20S Proteasomal Proteins of the Haloarchaeon Haloferax volcanii †
    Article Snippet: Prior to antibody preparation, the N-terminal sequences of PanAΔ1-40 -His and PanA-His were determined by Edman degradation, and the mass spectra of the tryptic fragments of PanB-His were determined by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (Applied Biosystems QSTAR XL hybrid LC/MS/MS; University of Florida ICBR Protein Chemistry Core). .. PanAΔ1-40 -His and PanB-His proteins purified from recombinant E. coli were separated by SDS-7.5% PAGE, excised from the gel, and used as antigen to raise polyclonal antibodies in rabbit (Cocalico Biologicals, Reamstown, Pa.).

    SDS Page:

    Article Title: Active, soluble recombinant melittin purified by extracting insoluble lysate of Escherichia coli without denaturation
    Article Snippet: Protein molecular weights were confirmed with matrix-assisted laser desorption-ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Grand Island, NY). .. The surfactant-containing insoluble fraction extracts were analyzed by SDS-PAGE.

    Article Title: Synthesis of Maleimide-End Functionalized Star Polymers and Multimeric Protein-Polymer Conjugates
    Article Snippet: Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was carried out with 4–20% Tris-Glycine gels (Invitrogen, 1.0 mm × 12 well). .. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry was performed on an Applied Biosystems Voyager-DE STR and operated in linear mode with an external calibration.

    Article Title: Mycoplasma genitalium Rapidly Disseminates to the Upper Reproductive Tracts and Knees of Female Mice following Vaginal Inoculation ▿
    Article Snippet: To investigate the host response to M. genitalium infection and determine the most immunodominant proteins, serum was collected from M. genitalium G37-infected or mock-inoculated mice (at the termination date [77 d.p.i.]) for immunoblot experiments using M. genitalium G37 lysates, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels, and standard methods. .. Each of the immunogenic M. genitalium proteins that were recognized by infected mouse sera was carefully excised from separate Coomassie-stained gels, digested with trypsin, and then identified using matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (UTMB Biomolecular Resource Facility) and a 4700 proteomics analyzer (Applied Biosystems).

    Software:

    Article Title: Synthesis of Maleimide-End Functionalized Star Polymers and Multimeric Protein-Polymer Conjugates
    Article Snippet: Chromatograms were processed with the EZStart 7.2 chromatography software. .. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry was performed on an Applied Biosystems Voyager-DE STR and operated in linear mode with an external calibration.

    Affinity Chromatography:

    Article Title: Novel Anti-inflammatory Activity of Epoxyazadiradione against Macrophage Migration Inhibitory Factor
    Article Snippet: Because Ni-NTA affinity chromatography purification co-purifies some contaminating proteins along with the desired MIFs, we further subjected Ni-NTA-purified MIFs to Sephadex 75 gel filtration (Amersham Biosciences), contaminating protein-free fractions of MIFs were collected, and purity was confirmed by silver staining ( , A , B , and C , insets ). .. The molecular weights of the purified huMIF, Pf MIF, and Py MIF were confirmed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Foster City, CA), which showed exact molecular masses ( m / z ) of 13.430, 13.266, and 13.251 kDa, respectively ( , A , B , and C ).

    Gel Permeation Chromatography:

    Article Title: Synthesis of Maleimide-End Functionalized Star Polymers and Multimeric Protein-Polymer Conjugates
    Article Snippet: The number-average molecular weight (Mn ) by dynamic light scattering (DLS) was collected from a Viscotek GPCmax VE2001 GPC solvent/sample module with 270 Dual detector and VE 3580 RI detector. .. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry was performed on an Applied Biosystems Voyager-DE STR and operated in linear mode with an external calibration.

    Nuclear Magnetic Resonance:

    Article Title: Synthesis of Maleimide-End Functionalized Star Polymers and Multimeric Protein-Polymer Conjugates
    Article Snippet: Monomer conversions were calculated by 1 H NMR by monitoring the disappearance of the peaks corresponding to the vinylic protons with the CH of the poly(NIPAAm) as internal standard. .. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry was performed on an Applied Biosystems Voyager-DE STR and operated in linear mode with an external calibration.

    Spectrophotometry:

    Article Title: Chemical validation of molecular mimicry: interaction of cholera toxin with Campylobacter lipooligosaccharides
    Article Snippet: Fractions of 1 ml each were collected, and the fluorescence intensity in each fraction was measured using a fluorescence spectrophotometer (Model F-2500, Hitachi, Tokyo, Japan). .. The ABOE-labeled oligosaccharides and the DMB-labeled oligosaccharides obtained from the Sep-Pak C18 cartridge were analyzed by Matrix-Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) mass spectrometry using a Model 4700 Proteomics Analyzer (Applied Biosystems, Foster City, CA, USA) equipped with a nitrogen UV laser (λ=337 nm) with a matrix of 2,5-dihydroxybenzoic acid at a concentration of 10 mg/ml in 10% aqueous ethanol.

    Article Title: Active, soluble recombinant melittin purified by extracting insoluble lysate of Escherichia coli without denaturation
    Article Snippet: Total protein was determined by the Bradford Assay (Thermo Scientific, Hanover Park, IL) used according to the manufacturer recommendations and measured with a Beckman model DU640 spectrophotometer (Brea, CA). .. Protein molecular weights were confirmed with matrix-assisted laser desorption-ionization-time of flight (MALDI-TOF) mass spectroscopy (Applied Biosystems, Grand Island, NY).

    Concentration Assay:

    Article Title: Engineering an antibody with picomolar affinity to DOTA chelates of multiple radionuclides for pretargeted radioimmunotherapy and imaging
    Article Snippet: Fractions containing DOTA-Bn-biotin were confirmed using matrix assisted laser desorption instrument time of flight (MALDI-TOF) mass spectrometry (Applied Biosystems Model Voyager DE-STR). .. DOTA-Bn-biotin concentration was determined using a biotin quantitation kit (Thermo Scientific) following the manufacturer’s instructions. (+)-(2S)-2-(4-Aminobutyl)-1,4,7,10-tetra-azacyclododecane-1,4,7,10-tetrayltetra-acetic acid (DOTA-alkyl) has been synthesized previously [ ].

    Article Title: Chemical validation of molecular mimicry: interaction of cholera toxin with Campylobacter lipooligosaccharides
    Article Snippet: .. The ABOE-labeled oligosaccharides and the DMB-labeled oligosaccharides obtained from the Sep-Pak C18 cartridge were analyzed by Matrix-Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) mass spectrometry using a Model 4700 Proteomics Analyzer (Applied Biosystems, Foster City, CA, USA) equipped with a nitrogen UV laser (λ=337 nm) with a matrix of 2,5-dihydroxybenzoic acid at a concentration of 10 mg/ml in 10% aqueous ethanol. ..

    Article Title: Physical and Functional Interaction between DNA Ligase III? and Poly(ADP-Ribose) Polymerase 1 in DNA Single-Strand Break Repair
    Article Snippet: .. After concentration to 5 μl, an equal volume of 50% acetonitrile-5% formic acid was added, and an aliquot (1 μl) was analyzed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (Voyager-DE Pro; Applied Biosystems). ..

    Article Title: Cationized liposomal keto-mycolic acids isolated from Mycobacterium bovis bacillus Calmette-Guérin induce antitumor immunity in a syngeneic murine bladder cancer model
    Article Snippet: The molecular weight of each MA methylester was determined by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry on a Voyager DE-STR workstation (Applied Biosystems, Foster City, CA) using 2.4-dihydroxybenzoic acid (2.5-DHB) as a matrix, as reported previously [ ]. .. Next, 2.5-DHB at the concentration of 10 mg/ml was mixed with 1.0 ml of the matrix solution.

    Staining:

    Article Title: Members of the Large Maf Transcription Family Regulate Insulin Gene Transcription in Islet ? Cells
    Article Snippet: Proteins were separated by 2-D gel electrophoresis and visualized by using the Colloidal Blue staining kit (Invitrogen, Carlsbad, Calif.). .. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry and TOF/TOF tandem mass spectrometry was carried out by using a Voyager 4700 mass spectrometer (Applied Biosystems, Foster City, Calif.) operated in reflectron mode.

    Normal Phase Liquid Chromatography:

    Article Title: An Engineered Saccharomyces cerevisiae Strain Binds the Broadly Neutralizing Human Immunodeficiency Virus Type 1 Antibody 2G12 and Elicits Mannose-Specific gp120-Binding Antibodies
    Article Snippet: .. Glycans were permethylated for analysis by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) using a QSTAR hybrid mass spectrometer (Applied Biosystems, Foster City, CA) and derivatized with 2-aminobenzamide (2AB) for normal-phase high-performance liquid chromatography (NP-HPLC) analysis on a Dionex DX-600 HPLC system (Sunnyvale, CA). .. 2AB-derivatized N-linked glycans released from RNase B were used as a standard for HPLC.

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  • 76
    Thermo Fisher flight maldi tof mass spectrometry uv irradiated
    Representative <t>MALDI-TOF</t> spectra of trypsin-digested samples of γS-WT and γS-G18V after UV-A irradiation. Mass fragments were identified by manual comparison of mass-to-charge ratio peaks to the theoretical peaks and isotopic mass distributions from MS-Digest ( http://prospector.ucsf.edu , in the public domain). Additional mass fragments corresponding to identifiable molecular weight changes are labeled with the specific mass difference, that is, m + 16. Top : The γS-WT spectrum is shown with selected fragments annotated. Insets A and B display the observed cases of m + 4 peaks in matched fragments containing a Trp residue. An m + 4 mass difference matches a Trp-to-Kyn PTM. The masses for each peak are labeled as m Trp and m Kyn . Bottom : The γS-G18V distribution is shown with annotations indicating each of the observed peaks matching an MS-Digest fragment. Kyn, kynurenine.
    Flight Maldi Tof Mass Spectrometry Uv Irradiated, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 76/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Thermo Fisher matrix assisted laser desorption ionization time of flight maldi tof mass spectrometric analysis
    Production of hirudin and a Fab fragment containing synthetic amino acids in BL21(DE3) and B-95.ΔA. (a) <t>MALDI-TOF</t> analyses of the recombinant HV1 products from the gene with TAG at position 63. The products were from the growth medium (left) and the cytoplasm (right) of BL21(DE3) and B-95.ΔAΔ fabR . The calculated masses (m/z) of the full-length HV1 with and without O -sulfation are 7050.5 and 6970.5, respectively, for [M+H] + . Note that sulfotyrosine can easily be deacylated in this matrix. The calculated mass (m/z) of the truncated HV1 with translation terminated at UAG is 6566.1 for [M+H] + . (b) Fab fragments produced in BL21(DE3) and B-95.ΔA. The purified products and those subsequently labeled with a fluorescent probe were fractionated on NuPAGE gels (in the upper and lower gels, respectively). Fluorescence was detected using an LAS4010 image analyzer. The labeled products correspond to the heavy chain. “WT” and “Am” indicate the products expressed from genes with and without an in-frame TAG, respectively. The figures indicate the yields of the products (mg/l). (c) The Fab fragment from B-95.ΔA, labeled and then purified by protein-G column chromatography, was analyzed by electrophoresis on a NuPAGE gel. (d) Fab fragments containing three 4-azidophenylalanines at the positions marked with “Am” were synthesized in BL21(DE3) (“RF-1+”) and B-95.ΔA (“RF-1−”), and then were analyzed by electrophoresis on a NuPAGE gel. The figures at the bottom of the gel indicate relative yields for the variants. (e) The Fab fragments containing three 4-azidophenylalanines at the positions marked with “Am” were synthesized in B-95.ΔA and labeled, followed by a NuPAGE analysis (upper) and fluorescence detection (lower). The positions corresponding to the Fab fragments labeled at one, two, and three sites are indicated. The size markers in the right-most or left-most lanes in panels b–e indicate 20 and 30 kDa.
    Matrix Assisted Laser Desorption Ionization Time Of Flight Maldi Tof Mass Spectrometric Analysis, supplied by Thermo Fisher, 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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    79
    Thermo Fisher matrix assisted laser desorption ionization time of flight maldi tof mass spectrometer
    Production of hirudin and a Fab fragment containing synthetic amino acids in BL21(DE3) and B-95.ΔA. (a) <t>MALDI-TOF</t> analyses of the recombinant HV1 products from the gene with TAG at position 63. The products were from the growth medium (left) and the cytoplasm (right) of BL21(DE3) and B-95.ΔAΔ fabR . The calculated masses (m/z) of the full-length HV1 with and without O -sulfation are 7050.5 and 6970.5, respectively, for [M+H] + . Note that sulfotyrosine can easily be deacylated in this matrix. The calculated mass (m/z) of the truncated HV1 with translation terminated at UAG is 6566.1 for [M+H] + . (b) Fab fragments produced in BL21(DE3) and B-95.ΔA. The purified products and those subsequently labeled with a fluorescent probe were fractionated on NuPAGE gels (in the upper and lower gels, respectively). Fluorescence was detected using an LAS4010 image analyzer. The labeled products correspond to the heavy chain. “WT” and “Am” indicate the products expressed from genes with and without an in-frame TAG, respectively. The figures indicate the yields of the products (mg/l). (c) The Fab fragment from B-95.ΔA, labeled and then purified by protein-G column chromatography, was analyzed by electrophoresis on a NuPAGE gel. (d) Fab fragments containing three 4-azidophenylalanines at the positions marked with “Am” were synthesized in BL21(DE3) (“RF-1+”) and B-95.ΔA (“RF-1−”), and then were analyzed by electrophoresis on a NuPAGE gel. The figures at the bottom of the gel indicate relative yields for the variants. (e) The Fab fragments containing three 4-azidophenylalanines at the positions marked with “Am” were synthesized in B-95.ΔA and labeled, followed by a NuPAGE analysis (upper) and fluorescence detection (lower). The positions corresponding to the Fab fragments labeled at one, two, and three sites are indicated. The size markers in the right-most or left-most lanes in panels b–e indicate 20 and 30 kDa.
    Matrix Assisted Laser Desorption Ionization Time Of Flight Maldi Tof Mass Spectrometer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 79/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Thermo Fisher maldi tof ms
    Production of hirudin and a Fab fragment containing synthetic amino acids in BL21(DE3) and B-95.ΔA. (a) <t>MALDI-TOF</t> analyses of the recombinant HV1 products from the gene with TAG at position 63. The products were from the growth medium (left) and the cytoplasm (right) of BL21(DE3) and B-95.ΔAΔ fabR . The calculated masses (m/z) of the full-length HV1 with and without O -sulfation are 7050.5 and 6970.5, respectively, for [M+H] + . Note that sulfotyrosine can easily be deacylated in this matrix. The calculated mass (m/z) of the truncated HV1 with translation terminated at UAG is 6566.1 for [M+H] + . (b) Fab fragments produced in BL21(DE3) and B-95.ΔA. The purified products and those subsequently labeled with a fluorescent probe were fractionated on NuPAGE gels (in the upper and lower gels, respectively). Fluorescence was detected using an LAS4010 image analyzer. The labeled products correspond to the heavy chain. “WT” and “Am” indicate the products expressed from genes with and without an in-frame TAG, respectively. The figures indicate the yields of the products (mg/l). (c) The Fab fragment from B-95.ΔA, labeled and then purified by protein-G column chromatography, was analyzed by electrophoresis on a NuPAGE gel. (d) Fab fragments containing three 4-azidophenylalanines at the positions marked with “Am” were synthesized in BL21(DE3) (“RF-1+”) and B-95.ΔA (“RF-1−”), and then were analyzed by electrophoresis on a NuPAGE gel. The figures at the bottom of the gel indicate relative yields for the variants. (e) The Fab fragments containing three 4-azidophenylalanines at the positions marked with “Am” were synthesized in B-95.ΔA and labeled, followed by a NuPAGE analysis (upper) and fluorescence detection (lower). The positions corresponding to the Fab fragments labeled at one, two, and three sites are indicated. The size markers in the right-most or left-most lanes in panels b–e indicate 20 and 30 kDa.
    Maldi Tof Ms, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 95/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Representative MALDI-TOF spectra of trypsin-digested samples of γS-WT and γS-G18V after UV-A irradiation. Mass fragments were identified by manual comparison of mass-to-charge ratio peaks to the theoretical peaks and isotopic mass distributions from MS-Digest ( http://prospector.ucsf.edu , in the public domain). Additional mass fragments corresponding to identifiable molecular weight changes are labeled with the specific mass difference, that is, m + 16. Top : The γS-WT spectrum is shown with selected fragments annotated. Insets A and B display the observed cases of m + 4 peaks in matched fragments containing a Trp residue. An m + 4 mass difference matches a Trp-to-Kyn PTM. The masses for each peak are labeled as m Trp and m Kyn . Bottom : The γS-G18V distribution is shown with annotations indicating each of the observed peaks matching an MS-Digest fragment. Kyn, kynurenine.

    Journal: Investigative Ophthalmology & Visual Science

    Article Title: Multiple Aggregation Pathways in Human γS-Crystallin and Its Aggregation-Prone G18V Variant

    doi: 10.1167/iovs.16-20621

    Figure Lengend Snippet: Representative MALDI-TOF spectra of trypsin-digested samples of γS-WT and γS-G18V after UV-A irradiation. Mass fragments were identified by manual comparison of mass-to-charge ratio peaks to the theoretical peaks and isotopic mass distributions from MS-Digest ( http://prospector.ucsf.edu , in the public domain). Additional mass fragments corresponding to identifiable molecular weight changes are labeled with the specific mass difference, that is, m + 16. Top : The γS-WT spectrum is shown with selected fragments annotated. Insets A and B display the observed cases of m + 4 peaks in matched fragments containing a Trp residue. An m + 4 mass difference matches a Trp-to-Kyn PTM. The masses for each peak are labeled as m Trp and m Kyn . Bottom : The γS-G18V distribution is shown with annotations indicating each of the observed peaks matching an MS-Digest fragment. Kyn, kynurenine.

    Article Snippet: Matrix Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) Mass Spectrometry UV-irradiated and unexposed proteins were digested using MS Grade Pierce Trypsin Protease (ThermoFischer Scientific, Rockford, IL, USA) at 37°C.

    Techniques: Irradiation, Mass Spectrometry, Molecular Weight, Labeling

    Production of hirudin and a Fab fragment containing synthetic amino acids in BL21(DE3) and B-95.ΔA. (a) MALDI-TOF analyses of the recombinant HV1 products from the gene with TAG at position 63. The products were from the growth medium (left) and the cytoplasm (right) of BL21(DE3) and B-95.ΔAΔ fabR . The calculated masses (m/z) of the full-length HV1 with and without O -sulfation are 7050.5 and 6970.5, respectively, for [M+H] + . Note that sulfotyrosine can easily be deacylated in this matrix. The calculated mass (m/z) of the truncated HV1 with translation terminated at UAG is 6566.1 for [M+H] + . (b) Fab fragments produced in BL21(DE3) and B-95.ΔA. The purified products and those subsequently labeled with a fluorescent probe were fractionated on NuPAGE gels (in the upper and lower gels, respectively). Fluorescence was detected using an LAS4010 image analyzer. The labeled products correspond to the heavy chain. “WT” and “Am” indicate the products expressed from genes with and without an in-frame TAG, respectively. The figures indicate the yields of the products (mg/l). (c) The Fab fragment from B-95.ΔA, labeled and then purified by protein-G column chromatography, was analyzed by electrophoresis on a NuPAGE gel. (d) Fab fragments containing three 4-azidophenylalanines at the positions marked with “Am” were synthesized in BL21(DE3) (“RF-1+”) and B-95.ΔA (“RF-1−”), and then were analyzed by electrophoresis on a NuPAGE gel. The figures at the bottom of the gel indicate relative yields for the variants. (e) The Fab fragments containing three 4-azidophenylalanines at the positions marked with “Am” were synthesized in B-95.ΔA and labeled, followed by a NuPAGE analysis (upper) and fluorescence detection (lower). The positions corresponding to the Fab fragments labeled at one, two, and three sites are indicated. The size markers in the right-most or left-most lanes in panels b–e indicate 20 and 30 kDa.

    Journal: Scientific Reports

    Article Title: Highly reproductive Escherichia coli cells with no specific assignment to the UAG codon

    doi: 10.1038/srep09699

    Figure Lengend Snippet: Production of hirudin and a Fab fragment containing synthetic amino acids in BL21(DE3) and B-95.ΔA. (a) MALDI-TOF analyses of the recombinant HV1 products from the gene with TAG at position 63. The products were from the growth medium (left) and the cytoplasm (right) of BL21(DE3) and B-95.ΔAΔ fabR . The calculated masses (m/z) of the full-length HV1 with and without O -sulfation are 7050.5 and 6970.5, respectively, for [M+H] + . Note that sulfotyrosine can easily be deacylated in this matrix. The calculated mass (m/z) of the truncated HV1 with translation terminated at UAG is 6566.1 for [M+H] + . (b) Fab fragments produced in BL21(DE3) and B-95.ΔA. The purified products and those subsequently labeled with a fluorescent probe were fractionated on NuPAGE gels (in the upper and lower gels, respectively). Fluorescence was detected using an LAS4010 image analyzer. The labeled products correspond to the heavy chain. “WT” and “Am” indicate the products expressed from genes with and without an in-frame TAG, respectively. The figures indicate the yields of the products (mg/l). (c) The Fab fragment from B-95.ΔA, labeled and then purified by protein-G column chromatography, was analyzed by electrophoresis on a NuPAGE gel. (d) Fab fragments containing three 4-azidophenylalanines at the positions marked with “Am” were synthesized in BL21(DE3) (“RF-1+”) and B-95.ΔA (“RF-1−”), and then were analyzed by electrophoresis on a NuPAGE gel. The figures at the bottom of the gel indicate relative yields for the variants. (e) The Fab fragments containing three 4-azidophenylalanines at the positions marked with “Am” were synthesized in B-95.ΔA and labeled, followed by a NuPAGE analysis (upper) and fluorescence detection (lower). The positions corresponding to the Fab fragments labeled at one, two, and three sites are indicated. The size markers in the right-most or left-most lanes in panels b–e indicate 20 and 30 kDa.

    Article Snippet: Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometric analysis of recombinant hirudin HV1 was reduced by an incubation in Bond-Breaker TCEP solution (neutral pH) (Thermo Scientific) with TECP at a final concentration of 50 mM at 70°C for 10 min, and then desalted by using a SPE C-TIP(C18) column (Nikkyo Technos, Co., Ltd., Japan), followed by elution in 80% acetonitrile and 0.5% TFA.

    Techniques: Recombinant, Produced, Purification, Labeling, Fluorescence, Column Chromatography, Electrophoresis, Synthesized