reversed-phase hplc Search Results


  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    Thermo Fisher reverse phase hplc
    Reverse Phase Hplc, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 736 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/reverse phase hplc/product/Thermo Fisher
    Average 99 stars, based on 736 article reviews
    Price from $9.99 to $1999.99
    reverse phase hplc - by Bioz Stars, 2020-10
    99/100 stars
      Buy from Supplier

    99
    Millipore rp hplc
    Rp Hplc, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 371 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rp hplc/product/Millipore
    Average 99 stars, based on 371 article reviews
    Price from $9.99 to $1999.99
    rp hplc - by Bioz Stars, 2020-10
    99/100 stars
      Buy from Supplier

    94
    Waters Corporation rp hplc
    Figure 1. <t>RP-HPLC</t> chromatogram of ( A ) Cell culture harvest and ( B ) <t>mAb1</t> Protein A intermediate.
    Rp Hplc, supplied by Waters Corporation, used in various techniques. Bioz Stars score: 94/100, based on 2110 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rp hplc/product/Waters Corporation
    Average 94 stars, based on 2110 article reviews
    Price from $9.99 to $1999.99
    rp hplc - by Bioz Stars, 2020-10
    94/100 stars
      Buy from Supplier

    94
    Phenomenex rp hplc
    ( a ) Reverse phase high performance liquid chromatography <t>(HPLC)</t> chromatogram of skin secretion of Phyllomedusa camba monitored at 214 nm. The arrow indicated the retention time of <t>PSN-PC;</t> ( b ) Tandem mass (MS/MS) fragmentation spectrum of PSN-PC; ( c ) Predicted singly-charged b ions and y ions arising from MS/MS fragmentation. The observed b- and y-ions were indicated in blue and red typefaces.
    Rp Hplc, supplied by Phenomenex, used in various techniques. Bioz Stars score: 94/100, based on 2382 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rp hplc/product/Phenomenex
    Average 94 stars, based on 2382 article reviews
    Price from $9.99 to $1999.99
    rp hplc - by Bioz Stars, 2020-10
    94/100 stars
      Buy from Supplier

    94
    Agilent technologies rp hplc
    Solid-phase purification of the phosphorothioate DNA sequence 10b . <t>RP-HPLC</t> analysis of solid-phase-purified 12b that was released from the support 11b . Inset: Purity analysis of the solid-phase-purified 12b by <t>PAGE.</t> Abbreviation: BP, bromophenol blue dye.
    Rp Hplc, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 94/100, based on 2100 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rp hplc/product/Agilent technologies
    Average 94 stars, based on 2100 article reviews
    Price from $9.99 to $1999.99
    rp hplc - by Bioz Stars, 2020-10
    94/100 stars
      Buy from Supplier

    99
    Agilent technologies reverse phase hplc
    Solid-phase purification of the phosphorothioate DNA sequence 10b . <t>RP-HPLC</t> analysis of solid-phase-purified 12b that was released from the support 11b . Inset: Purity analysis of the solid-phase-purified 12b by <t>PAGE.</t> Abbreviation: BP, bromophenol blue dye.
    Reverse Phase Hplc, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 99/100, based on 2244 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/reverse phase hplc/product/Agilent technologies
    Average 99 stars, based on 2244 article reviews
    Price from $9.99 to $1999.99
    reverse phase hplc - by Bioz Stars, 2020-10
    99/100 stars
      Buy from Supplier

    93
    Shimadzu Corporation rp hplc
    Analytical <t>HPLC</t> of purified <t>mSAM.</t> Gradient of 50–100% acetonitrile. RT: 10.7 min (lower panel), 11.5 min (upper panel). Absorbances at 450 nm (coumarin 343), 540 nm (TAMRA), 254 nm, and 280 nm are shown. Both 5-(6-)TAMRA isomers are shown separately (upper and lower panel).
    Rp Hplc, supplied by Shimadzu Corporation, used in various techniques. Bioz Stars score: 93/100, based on 823 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rp hplc/product/Shimadzu Corporation
    Average 93 stars, based on 823 article reviews
    Price from $9.99 to $1999.99
    rp hplc - by Bioz Stars, 2020-10
    93/100 stars
      Buy from Supplier

    93
    Waters Corporation reverse phase hplc
    Enzymatic assay of recombinant TcLAR. A , <t>HPLC</t> analysis of products from incubation of leucocyanidin with boiled protein as control, monitored by UV spectroscopy. B , as above, for incubation of purified recombinant TcLAR with 3 H-leucocyanidin. C , As in (A) , but monitoring radioactivity of fractions; D , As in (B) , but monitoring radioactivity of fractions; ( E ), UV spectrum of catechin product from (B) ; F , HPLC analysis of an authentic standard of (−)-catechin. mAU, milliabsorbance units; CPM, counts per minute.
    Reverse Phase Hplc, supplied by Waters Corporation, used in various techniques. Bioz Stars score: 93/100, based on 1851 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/reverse phase hplc/product/Waters Corporation
    Average 93 stars, based on 1851 article reviews
    Price from $9.99 to $1999.99
    reverse phase hplc - by Bioz Stars, 2020-10
    93/100 stars
      Buy from Supplier

    93
    Phenomenex reverse phase hplc
    The synthesis of sugar nucleotides from 2-chloro-4-nitrophenyl glucosides. ( a ) General reaction scheme. ( b ) Structures of 2-chloro-4-nitrophenyl glycoside donors evaluated for D-sugars within this series, the differences between each member and the native OleD sugar substrate (β-D-glucose) are highlighted in red. ( c ) Maximum observed percent conversion of (U/T)DP to (U/T)DP-glucose within a 21 hour time course assay for each donor (n ≥ 2, standard deviation ≤ 5%). Standard reactions contained 7 μM TDP-16, 1 mM (U/T)DP, and 1 mM of 2-chloro-4-nitrophenyl glycoside donor ( 9 , 34 – 47 ) in Tris-HCl buffer (50 mM, pH 8.5) with a final volume of 300 μl. Over 21 hours at 25°C, aliquots taken at various times were flash frozen and analyzed by <t>HPLC</t> ( Supplementary Methods ). For reactions with <t>UDP</t> yielding
    Reverse Phase Hplc, supplied by Phenomenex, used in various techniques. Bioz Stars score: 93/100, based on 1307 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/reverse phase hplc/product/Phenomenex
    Average 93 stars, based on 1307 article reviews
    Price from $9.99 to $1999.99
    reverse phase hplc - by Bioz Stars, 2020-10
    93/100 stars
      Buy from Supplier

    93
    Thermo Fisher reversed phase hplc
    The synthesis of sugar nucleotides from 2-chloro-4-nitrophenyl glucosides. ( a ) General reaction scheme. ( b ) Structures of 2-chloro-4-nitrophenyl glycoside donors evaluated for D-sugars within this series, the differences between each member and the native OleD sugar substrate (β-D-glucose) are highlighted in red. ( c ) Maximum observed percent conversion of (U/T)DP to (U/T)DP-glucose within a 21 hour time course assay for each donor (n ≥ 2, standard deviation ≤ 5%). Standard reactions contained 7 μM TDP-16, 1 mM (U/T)DP, and 1 mM of 2-chloro-4-nitrophenyl glycoside donor ( 9 , 34 – 47 ) in Tris-HCl buffer (50 mM, pH 8.5) with a final volume of 300 μl. Over 21 hours at 25°C, aliquots taken at various times were flash frozen and analyzed by <t>HPLC</t> ( Supplementary Methods ). For reactions with <t>UDP</t> yielding
    Reversed Phase Hplc, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 539 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/reversed phase hplc/product/Thermo Fisher
    Average 93 stars, based on 539 article reviews
    Price from $9.99 to $1999.99
    reversed phase hplc - by Bioz Stars, 2020-10
    93/100 stars
      Buy from Supplier

    92
    Shimadzu Corporation reverse phase hplc
    The radioactive signal from <t>oligopeptides</t> being modified with radioactive acetyl CoA . The oligopeptide 1 MLGTE-RRR 24 (200 μM) was incubated with [1- 14 C] acetyl CoA (final 300 μM with specific activity 11.2 mCi/mmol) and purified MBP-hNaa30p (80 nM) in acetylation buffer for 60 minutes at 37°C. Samples were collected at indicated time points and analysed with reverse phase <t>HPLC</t> that had been connected to a radioactivity flow detector after the absorbance detector. A clear time dependent increase in the radioactivity signal is observed, verifying that the eluted oligopeptides are labelled with radioactive acetyl groups.
    Reverse Phase Hplc, supplied by Shimadzu Corporation, used in various techniques. Bioz Stars score: 92/100, based on 628 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/reverse phase hplc/product/Shimadzu Corporation
    Average 92 stars, based on 628 article reviews
    Price from $9.99 to $1999.99
    reverse phase hplc - by Bioz Stars, 2020-10
    92/100 stars
      Buy from Supplier

    92
    Waters Corporation preparative rp hplc
    Purification of stingray ghrelin-like peptide (GRLN-LP) from stomach extracts . Black bars indicate the measured fluorescence changes in intracellular calcium ion concentrations in CHO cells expressing rat GHS-R1a (CHO-GHSR62). (A) Carboxymethyl (CM)-cation ion-exchange <t>HPLC</t> (pH 4.8) of the SP-III fraction of stomach extracts. The GRLN-like activity was divided into four groups (A-D). (B) Preparative reverse-phase (RP)-HPLC (Symmetry <t>C18,</t> 3.9 × 150 mm) of group B after purification with an anti-rat GRLN1-11 immuno-affinity column. (C) Final purification of the active fraction indicated in (B) by another RP-HPLC (Vydac diphenyl, 219TP5215, 2.1 × 150 mm).
    Preparative Rp Hplc, supplied by Waters Corporation, used in various techniques. Bioz Stars score: 92/100, based on 315 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/preparative rp hplc/product/Waters Corporation
    Average 92 stars, based on 315 article reviews
    Price from $9.99 to $1999.99
    preparative rp hplc - by Bioz Stars, 2020-10
    92/100 stars
      Buy from Supplier

    93
    Hitachi Ltd rp hplc
    <t>RP-HPLC</t> separation of TPIs from N. attenuata of the wild-type genotype. Numbers refer to <t>TPI</t> fractions containing inhibitory activity against trypsin. RP-HPLC profile of TPIs from uninduced control leaves (dashed line, right y axis) and leaves elicited
    Rp Hplc, supplied by Hitachi Ltd, used in various techniques. Bioz Stars score: 93/100, based on 261 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rp hplc/product/Hitachi Ltd
    Average 93 stars, based on 261 article reviews
    Price from $9.99 to $1999.99
    rp hplc - by Bioz Stars, 2020-10
    93/100 stars
      Buy from Supplier

    91
    Phenomenex preparative reversed phase hplc
    <t>RP-HPLC</t> separation of TPIs from N. attenuata of the wild-type genotype. Numbers refer to <t>TPI</t> fractions containing inhibitory activity against trypsin. RP-HPLC profile of TPIs from uninduced control leaves (dashed line, right y axis) and leaves elicited
    Preparative Reversed Phase Hplc, supplied by Phenomenex, used in various techniques. Bioz Stars score: 91/100, based on 121 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/preparative reversed phase hplc/product/Phenomenex
    Average 91 stars, based on 121 article reviews
    Price from $9.99 to $1999.99
    preparative reversed phase hplc - by Bioz Stars, 2020-10
    91/100 stars
      Buy from Supplier

    92
    JASCO Inc rp hplc
    <t>RP-HPLC</t> separation of TPIs from N. attenuata of the wild-type genotype. Numbers refer to <t>TPI</t> fractions containing inhibitory activity against trypsin. RP-HPLC profile of TPIs from uninduced control leaves (dashed line, right y axis) and leaves elicited
    Rp Hplc, supplied by JASCO Inc, used in various techniques. Bioz Stars score: 92/100, based on 165 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rp hplc/product/JASCO Inc
    Average 92 stars, based on 165 article reviews
    Price from $9.99 to $1999.99
    rp hplc - by Bioz Stars, 2020-10
    92/100 stars
      Buy from Supplier

    94
    Merck & Co reversed phase hplc
    Translocation. A. Colicin E3-cleaved (closed circles) or control (open circles) pre-translocation complexes (0.1 μM) were mixed in a quench-flow apparatus with EF-G·GTP (3 μM) and Pmn (10 mM) and translocation analysed by formation of <t>fMetPhe-Pmn.</t> The amount of pre-translocation complex (PTC) was determined by <t>HPLC</t> analysis of fMetPhe. B. Translocation was monitored by the fluorescence change of fMetPhe-tRNA Phe (Prf16/17). Curve 1 – control ribosomes; curve 2 – colicin E3-cleaved ribosomes; curve 3 – control ribosomes in the presence of viomycin (200 μM); curve 4 – colicin E3-cleaved ribosomes with viomycin (200 μM). Normalized ΔF, fluorescence change normalized by the maximum extent of translocation determined by the Pmn reaction. C. Pi release upon GTP hydrolysis by EF-G (curves 1 and 2) or buffer control (curves 3 and 4) with control ribosomes (curves 1 and 3) or colicin E3-cleaved (curves 2 and 4). Pi release was measured by the fluorescence change of MDCC-PBP upon Pi binding ( Experimental procedures ). D. Multiple-turnover GTP hydrolysis by EF-G. Velocity of GTP hydrolysis was measured with catalytic concentrations of EF-G (10 nM) and increasing concentrations of colicin E3-cleaved ribosomes (closed circles) or control ribosomes (open circles) in the presence of excess [γ- 32 P]-GTP (200 μM).
    Reversed Phase Hplc, supplied by Merck & Co, used in various techniques. Bioz Stars score: 94/100, based on 220 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/reversed phase hplc/product/Merck & Co
    Average 94 stars, based on 220 article reviews
    Price from $9.99 to $1999.99
    reversed phase hplc - by Bioz Stars, 2020-10
    94/100 stars
      Buy from Supplier

    91
    Phenomenex reverse phase hplc column
    Translocation. A. Colicin E3-cleaved (closed circles) or control (open circles) pre-translocation complexes (0.1 μM) were mixed in a quench-flow apparatus with EF-G·GTP (3 μM) and Pmn (10 mM) and translocation analysed by formation of <t>fMetPhe-Pmn.</t> The amount of pre-translocation complex (PTC) was determined by <t>HPLC</t> analysis of fMetPhe. B. Translocation was monitored by the fluorescence change of fMetPhe-tRNA Phe (Prf16/17). Curve 1 – control ribosomes; curve 2 – colicin E3-cleaved ribosomes; curve 3 – control ribosomes in the presence of viomycin (200 μM); curve 4 – colicin E3-cleaved ribosomes with viomycin (200 μM). Normalized ΔF, fluorescence change normalized by the maximum extent of translocation determined by the Pmn reaction. C. Pi release upon GTP hydrolysis by EF-G (curves 1 and 2) or buffer control (curves 3 and 4) with control ribosomes (curves 1 and 3) or colicin E3-cleaved (curves 2 and 4). Pi release was measured by the fluorescence change of MDCC-PBP upon Pi binding ( Experimental procedures ). D. Multiple-turnover GTP hydrolysis by EF-G. Velocity of GTP hydrolysis was measured with catalytic concentrations of EF-G (10 nM) and increasing concentrations of colicin E3-cleaved ribosomes (closed circles) or control ribosomes (open circles) in the presence of excess [γ- 32 P]-GTP (200 μM).
    Reverse Phase Hplc Column, supplied by Phenomenex, used in various techniques. Bioz Stars score: 91/100, based on 320 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/reverse phase hplc column/product/Phenomenex
    Average 91 stars, based on 320 article reviews
    Price from $9.99 to $1999.99
    reverse phase hplc column - by Bioz Stars, 2020-10
    91/100 stars
      Buy from Supplier

    90
    Phenomenex semi preparative rp hplc
    Recombinant production of <t>Mb1a.</t> Semi-preparative <t>RP-HPLC</t> chromatogram of recombinant Mb1a released by TEV protease cleavage of the MBP-Mb1a fusion protein (see Materials and Methods for more details). The dotted line indicates the gradient of solvent B (90% acetonitrile/0.043% TFA). Top inset: SDS-PAGE gel showing pre-cleaved MBP-Mb1a fusion protein (lane 1) and remaining MBP after cleavage (lane 2). Lane M contains molecular markers (masses in kDa). Bottom inset: MALDI-TOF mass spectrum of pure recombinant Mb1a.
    Semi Preparative Rp Hplc, supplied by Phenomenex, used in various techniques. Bioz Stars score: 90/100, based on 78 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/semi preparative rp hplc/product/Phenomenex
    Average 90 stars, based on 78 article reviews
    Price from $9.99 to $1999.99
    semi preparative rp hplc - by Bioz Stars, 2020-10
    90/100 stars
      Buy from Supplier

    91
    Phenomenex rp hplc column
    Distinct HRNR-derived CIDAMPs show similar ribosomal protein binding patterns. ( a ) SulfoLink®–column-bound proteins of an E . coli -extract were separated on a Jupiter® <t>C18</t> <t>RP-HPLC</t> column with a Prp-gradient. HPLC fractions containing UV-absorbing peaks (C1–D10) were divided into five aliquots and adjusted in parallel to five PAGE-gels and separated. ( b ) Silver-stained proteins. ( c ) HRNR-Far-Western blot for probing with biotinylated HR1-18 (HRNR 2556–2677 ) using Strep - Tactin ®, ( d ) HRNR-Far-Western blot for probing with biotinylated rSumo3-HRNR 2591–2684 using Strep - Tactin ®, ( e ) HRNR-Far-Western blot for probing with rHRNR 2591–2684 using anti-HRNR 2591–2684 antibodies, ( f ) HRNR-Far-Western blot for probing with rHRNR 1075–1172 using anti-HRNR 1075–1172 antibodies. Note similarities of the staining patterns, irrespective the CIDAMP AA-sequence or biotin-labeling and irrespective whether a Strep - Tactin ®- or antibody-detectable CIDAMP was used to probe and detect the target protein on the membrane. Note the presence of 70 kDa bands upon HRNR-Far-Western blot analyses in most of the investigated HPLC fractions with highest intensity for rHRNR 1075–1172 binding ( f ). The most intensive band, corresponding to a 37 kDa protein in fraction number C13, was identified as E . coli ribosomal protein L2.
    Rp Hplc Column, supplied by Phenomenex, used in various techniques. Bioz Stars score: 91/100, based on 95 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rp hplc column/product/Phenomenex
    Average 91 stars, based on 95 article reviews
    Price from $9.99 to $1999.99
    rp hplc column - by Bioz Stars, 2020-10
    91/100 stars
      Buy from Supplier

    92
    Supelco reversed phase hplc
    Isolation of vurtoxin and Vur-S49 by reverse-phase <t>HPLC.</t> Separation was done on a Discovery <t>BIO</t> Wide Pore C18 column (10×250 mm, Supelco) in a gradient of 25–40% (v/v) acetonitrile in 60 min in the presence of 0.1% (v/v) trifluoroacetic acid, at a flow rate of 2.0 ml/min. Fraction containing Vur-S49 (4) and vurtoxin (11) are indicated by horizontal bars.
    Reversed Phase Hplc, supplied by Supelco, used in various techniques. Bioz Stars score: 92/100, based on 136 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/reversed phase hplc/product/Supelco
    Average 92 stars, based on 136 article reviews
    Price from $9.99 to $1999.99
    reversed phase hplc - by Bioz Stars, 2020-10
    92/100 stars
      Buy from Supplier

    93
    Gilson Inc rp hplc
    Isolation of vurtoxin and Vur-S49 by reverse-phase <t>HPLC.</t> Separation was done on a Discovery <t>BIO</t> Wide Pore C18 column (10×250 mm, Supelco) in a gradient of 25–40% (v/v) acetonitrile in 60 min in the presence of 0.1% (v/v) trifluoroacetic acid, at a flow rate of 2.0 ml/min. Fraction containing Vur-S49 (4) and vurtoxin (11) are indicated by horizontal bars.
    Rp Hplc, supplied by Gilson Inc, used in various techniques. Bioz Stars score: 93/100, based on 207 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rp hplc/product/Gilson Inc
    Average 93 stars, based on 207 article reviews
    Price from $9.99 to $1999.99
    rp hplc - by Bioz Stars, 2020-10
    93/100 stars
      Buy from Supplier

    Image Search Results


    Figure 1. RP-HPLC chromatogram of ( A ) Cell culture harvest and ( B ) mAb1 Protein A intermediate.

    Journal: mAbs

    Article Title: Characterization of antibody variants during process development

    doi: 10.4161/mabs.21614

    Figure Lengend Snippet: Figure 1. RP-HPLC chromatogram of ( A ) Cell culture harvest and ( B ) mAb1 Protein A intermediate.

    Article Snippet: RP-HPLC of mAb1 was performed on a Waters nanoAcquity UPLC system consisting of a temperature-controlled autosampler.

    Techniques: High Performance Liquid Chromatography, Cell Culture

    Figure 6. LC/ESI-MS of the leader peptide. ( A ) The molecular mass of the peptide is detected at 3262.8153 Da. ( B ) LC/ESI-MS/MS of the leader peptide ( m/z 1088.26, 3+) of mAb1 enriched in RP-HPLC post peak.

    Journal: mAbs

    Article Title: Characterization of antibody variants during process development

    doi: 10.4161/mabs.21614

    Figure Lengend Snippet: Figure 6. LC/ESI-MS of the leader peptide. ( A ) The molecular mass of the peptide is detected at 3262.8153 Da. ( B ) LC/ESI-MS/MS of the leader peptide ( m/z 1088.26, 3+) of mAb1 enriched in RP-HPLC post peak.

    Article Snippet: RP-HPLC of mAb1 was performed on a Waters nanoAcquity UPLC system consisting of a temperature-controlled autosampler.

    Techniques: Mass Spectrometry, High Performance Liquid Chromatography

    Figure 5. LC/ESI-MS/MS in DDA mode of trypsin-digested and PNGase F-deglycosylated mAb1 enriched in RP-HPLC post peak. The leader peptide (2–31 HC) eluted at 127.04 min in the TIC.

    Journal: mAbs

    Article Title: Characterization of antibody variants during process development

    doi: 10.4161/mabs.21614

    Figure Lengend Snippet: Figure 5. LC/ESI-MS/MS in DDA mode of trypsin-digested and PNGase F-deglycosylated mAb1 enriched in RP-HPLC post peak. The leader peptide (2–31 HC) eluted at 127.04 min in the TIC.

    Article Snippet: RP-HPLC of mAb1 was performed on a Waters nanoAcquity UPLC system consisting of a temperature-controlled autosampler.

    Techniques: Mass Spectrometry, High Performance Liquid Chromatography

    Figure 4. LC/ESI-MS of DTT treated heavy chains of mAb1 ( A ) control and ( B ) enriched in RP-HPLC post peak.

    Journal: mAbs

    Article Title: Characterization of antibody variants during process development

    doi: 10.4161/mabs.21614

    Figure Lengend Snippet: Figure 4. LC/ESI-MS of DTT treated heavy chains of mAb1 ( A ) control and ( B ) enriched in RP-HPLC post peak.

    Article Snippet: RP-HPLC of mAb1 was performed on a Waters nanoAcquity UPLC system consisting of a temperature-controlled autosampler.

    Techniques: Mass Spectrometry, High Performance Liquid Chromatography

    Figure 2. Overlays of ( A ) RP-HPLC; ( B ) SEC-HPLC; and ( C ) IEX-HPLC chromatograms of mAb1 control and mAb1 enriched in RP-HPLC post peak.

    Journal: mAbs

    Article Title: Characterization of antibody variants during process development

    doi: 10.4161/mabs.21614

    Figure Lengend Snippet: Figure 2. Overlays of ( A ) RP-HPLC; ( B ) SEC-HPLC; and ( C ) IEX-HPLC chromatograms of mAb1 control and mAb1 enriched in RP-HPLC post peak.

    Article Snippet: RP-HPLC of mAb1 was performed on a Waters nanoAcquity UPLC system consisting of a temperature-controlled autosampler.

    Techniques: High Performance Liquid Chromatography, Size-exclusion Chromatography

    Figure 3. LC/ESI-MS of PNGase F deglycosylated mAb1 ( A ) control and ( B ) enriched in RP-HPLC post peak.

    Journal: mAbs

    Article Title: Characterization of antibody variants during process development

    doi: 10.4161/mabs.21614

    Figure Lengend Snippet: Figure 3. LC/ESI-MS of PNGase F deglycosylated mAb1 ( A ) control and ( B ) enriched in RP-HPLC post peak.

    Article Snippet: RP-HPLC of mAb1 was performed on a Waters nanoAcquity UPLC system consisting of a temperature-controlled autosampler.

    Techniques: Mass Spectrometry, High Performance Liquid Chromatography

    RP-HPLC post peak is due to the presence of unprocessed secretion leader on the N-terminus of mAb1 heavy chain

    Journal: mAbs

    Article Title: Characterization of antibody variants during process development

    doi: 10.4161/mabs.21614

    Figure Lengend Snippet: RP-HPLC post peak is due to the presence of unprocessed secretion leader on the N-terminus of mAb1 heavy chain

    Article Snippet: RP-HPLC of mAb1 was performed on a Waters nanoAcquity UPLC system consisting of a temperature-controlled autosampler.

    Techniques: High Performance Liquid Chromatography

    Purification of active fraction F2 using reverse phase high performance liquid chromatography (RP-HPLC) and the activities of sub-fractions assay: ( A ) percentage inhibition of hydrophilic and hydrophobic extracts of F2; ( B ) chromatogram of active fraction F2 by RP-HPLC, measured at 280 nm; ( C ) percentage inhibition of F2-1 and F2-2; and (D) H 2 O 2 production capacity of F2-1 and F2-2. Spots in ( A , C , D ) represent the raw data. The results are expressed as the mean ± standard deviation ( n = 3). The symbol of “**” and “*” in ( A , C , D ) represent significant differences of p

    Journal: Marine Drugs

    Article Title: Novel Antibacterial Peptides Isolated from the Maillard Reaction Products of Half-Fin Anchovy (Setipinna taty) Hydrolysates/Glucose and Their Mode of Action in Escherichia Coli

    doi: 10.3390/md17010047

    Figure Lengend Snippet: Purification of active fraction F2 using reverse phase high performance liquid chromatography (RP-HPLC) and the activities of sub-fractions assay: ( A ) percentage inhibition of hydrophilic and hydrophobic extracts of F2; ( B ) chromatogram of active fraction F2 by RP-HPLC, measured at 280 nm; ( C ) percentage inhibition of F2-1 and F2-2; and (D) H 2 O 2 production capacity of F2-1 and F2-2. Spots in ( A , C , D ) represent the raw data. The results are expressed as the mean ± standard deviation ( n = 3). The symbol of “**” and “*” in ( A , C , D ) represent significant differences of p

    Article Snippet: The hydrophobic extract of F2, which showed the largest percentage inhibition, was loaded onto a RP-HPLC system equipped with a C18 column (4.6 × 250 mm, 5 μm, Sunfire™, Waters, MA, USA) for further separation.

    Techniques: Purification, High Performance Liquid Chromatography, Inhibition, Standard Deviation

    ( a ) Reverse phase high performance liquid chromatography (HPLC) chromatogram of skin secretion of Phyllomedusa camba monitored at 214 nm. The arrow indicated the retention time of PSN-PC; ( b ) Tandem mass (MS/MS) fragmentation spectrum of PSN-PC; ( c ) Predicted singly-charged b ions and y ions arising from MS/MS fragmentation. The observed b- and y-ions were indicated in blue and red typefaces.

    Journal: Molecules : A Journal of Synthetic Chemistry and Natural Product Chemistry

    Article Title: PSN-PC: A Novel Antimicrobial and Anti-Biofilm Peptide from the Skin Secretion of Phyllomedusa-camba with Cytotoxicity on Human Lung Cancer Cell

    doi: 10.3390/molecules22111896

    Figure Lengend Snippet: ( a ) Reverse phase high performance liquid chromatography (HPLC) chromatogram of skin secretion of Phyllomedusa camba monitored at 214 nm. The arrow indicated the retention time of PSN-PC; ( b ) Tandem mass (MS/MS) fragmentation spectrum of PSN-PC; ( c ) Predicted singly-charged b ions and y ions arising from MS/MS fragmentation. The observed b- and y-ions were indicated in blue and red typefaces.

    Article Snippet: The authenticity of the purified synthetic PSN-PC was verified by RP-HPLC with an analytical Jupiter C5 column (250 mm × 4.6 mm, Phenomenex, UK).

    Techniques: High Performance Liquid Chromatography, Mass Spectrometry

    ( a ) The RP-HPLC chromatogram of the purified synthetic phylloseptin-PC (PSN-PC); ( b ) MALDI-TOF mass spectrum of synthetic PSN-PC; ( c ) Predicted secondary structure of PSN-PC using I-TASSER; ( d ) Predicted 3D model of PSN-PC using I-TASSER; ( e ) Z-score plot using ProSA-web; ( f ) Helical wheel plot of PSN-PC; ( g ) Circular dichroism (CD) spectra recorded for PSN-PC (100 μM) in 10 mM ammonium acetate buffer and 50% TFE ammonium acetate buffer.

    Journal: Molecules : A Journal of Synthetic Chemistry and Natural Product Chemistry

    Article Title: PSN-PC: A Novel Antimicrobial and Anti-Biofilm Peptide from the Skin Secretion of Phyllomedusa-camba with Cytotoxicity on Human Lung Cancer Cell

    doi: 10.3390/molecules22111896

    Figure Lengend Snippet: ( a ) The RP-HPLC chromatogram of the purified synthetic phylloseptin-PC (PSN-PC); ( b ) MALDI-TOF mass spectrum of synthetic PSN-PC; ( c ) Predicted secondary structure of PSN-PC using I-TASSER; ( d ) Predicted 3D model of PSN-PC using I-TASSER; ( e ) Z-score plot using ProSA-web; ( f ) Helical wheel plot of PSN-PC; ( g ) Circular dichroism (CD) spectra recorded for PSN-PC (100 μM) in 10 mM ammonium acetate buffer and 50% TFE ammonium acetate buffer.

    Article Snippet: The authenticity of the purified synthetic PSN-PC was verified by RP-HPLC with an analytical Jupiter C5 column (250 mm × 4.6 mm, Phenomenex, UK).

    Techniques: High Performance Liquid Chromatography, Purification

    Solid-phase purification of the phosphorothioate DNA sequence 10b . RP-HPLC analysis of solid-phase-purified 12b that was released from the support 11b . Inset: Purity analysis of the solid-phase-purified 12b by PAGE. Abbreviation: BP, bromophenol blue dye.

    Journal: Current protocols in nucleic acid chemistry

    Article Title: A High-Throughput Process for the Solid-Phase Purification of Synthetic DNA Sequences

    doi: 10.1002/cpnc.31

    Figure Lengend Snippet: Solid-phase purification of the phosphorothioate DNA sequence 10b . RP-HPLC analysis of solid-phase-purified 12b that was released from the support 11b . Inset: Purity analysis of the solid-phase-purified 12b by PAGE. Abbreviation: BP, bromophenol blue dye.

    Article Snippet: The purity of these sequences has been evaluated by RP-HPLC and by PAGE under denaturing conditions ( , , , and ).

    Techniques: Purification, Sequencing, High Performance Liquid Chromatography, Polyacrylamide Gel Electrophoresis

    Solid-phase purification of the native DNA sequence 10f . RP-HPLC analysis of solid-phase-purified 12f that was released from the support 11f . Inset: Purity analysis of the solid-phase-purified 12f by PAGE. Abbreviation: BP, bromophenol blue dye.

    Journal: Current protocols in nucleic acid chemistry

    Article Title: A High-Throughput Process for the Solid-Phase Purification of Synthetic DNA Sequences

    doi: 10.1002/cpnc.31

    Figure Lengend Snippet: Solid-phase purification of the native DNA sequence 10f . RP-HPLC analysis of solid-phase-purified 12f that was released from the support 11f . Inset: Purity analysis of the solid-phase-purified 12f by PAGE. Abbreviation: BP, bromophenol blue dye.

    Article Snippet: The purity of these sequences has been evaluated by RP-HPLC and by PAGE under denaturing conditions ( , , , and ).

    Techniques: Purification, Sequencing, High Performance Liquid Chromatography, Polyacrylamide Gel Electrophoresis

    Solid-phase purification of the phosphorothioate DNA sequence 10a . RP-HPLC analysis of solid-phase-purified 12a that was released from the support 11a . Inset: Purity analysis of the solid-phase-purified 12a by PAGE. Chromatographic conditions are reported in the annotation of step 3 of Basic Protocol 4 . Abbreviation: BP, bromophenol blue dye.

    Journal: Current protocols in nucleic acid chemistry

    Article Title: A High-Throughput Process for the Solid-Phase Purification of Synthetic DNA Sequences

    doi: 10.1002/cpnc.31

    Figure Lengend Snippet: Solid-phase purification of the phosphorothioate DNA sequence 10a . RP-HPLC analysis of solid-phase-purified 12a that was released from the support 11a . Inset: Purity analysis of the solid-phase-purified 12a by PAGE. Chromatographic conditions are reported in the annotation of step 3 of Basic Protocol 4 . Abbreviation: BP, bromophenol blue dye.

    Article Snippet: The purity of these sequences has been evaluated by RP-HPLC and by PAGE under denaturing conditions ( , , , and ).

    Techniques: Purification, Sequencing, High Performance Liquid Chromatography, Polyacrylamide Gel Electrophoresis

    Solid-phase purification of the phosphorothioate DNA sequence 10c . RP-HPLC analysis of solid-phase-purified 12c that was released from the support 11c . Inset: Purity analysis of the solid-phase-purified 12c by PAGE. Abbreviation: BP, bromophenol blue dye.

    Journal: Current protocols in nucleic acid chemistry

    Article Title: A High-Throughput Process for the Solid-Phase Purification of Synthetic DNA Sequences

    doi: 10.1002/cpnc.31

    Figure Lengend Snippet: Solid-phase purification of the phosphorothioate DNA sequence 10c . RP-HPLC analysis of solid-phase-purified 12c that was released from the support 11c . Inset: Purity analysis of the solid-phase-purified 12c by PAGE. Abbreviation: BP, bromophenol blue dye.

    Article Snippet: The purity of these sequences has been evaluated by RP-HPLC and by PAGE under denaturing conditions ( , , , and ).

    Techniques: Purification, Sequencing, High Performance Liquid Chromatography, Polyacrylamide Gel Electrophoresis

    10-Fold scale up solid-phase purification of the phosphorothioate DNA sequence 12a. (A) RP-HPLC profile of unpurified 10a . (B) RP-HPLC profile of unpurified 10a after capture by the support 3 . (C) RP-HPLC analysis of solid-phase purified 12a that has been released from the support 11a . (D) Photograph of precipitated 12a accounting for 995 OD 260 of the DNA sequence. Inset: Purity analysis of the solid-phase purified 12a by PAGE. Abbreviation: BP, bromophenol blue dye.

    Journal: Current protocols in nucleic acid chemistry

    Article Title: A High-Throughput Process for the Solid-Phase Purification of Synthetic DNA Sequences

    doi: 10.1002/cpnc.31

    Figure Lengend Snippet: 10-Fold scale up solid-phase purification of the phosphorothioate DNA sequence 12a. (A) RP-HPLC profile of unpurified 10a . (B) RP-HPLC profile of unpurified 10a after capture by the support 3 . (C) RP-HPLC analysis of solid-phase purified 12a that has been released from the support 11a . (D) Photograph of precipitated 12a accounting for 995 OD 260 of the DNA sequence. Inset: Purity analysis of the solid-phase purified 12a by PAGE. Abbreviation: BP, bromophenol blue dye.

    Article Snippet: The purity of these sequences has been evaluated by RP-HPLC and by PAGE under denaturing conditions ( , , , and ).

    Techniques: Purification, Sequencing, High Performance Liquid Chromatography, Polyacrylamide Gel Electrophoresis

    Solid-phase purification of the phosphorothioate DNA sequence 10d . RP-HPLC analysis of solid-phase-purified 12d that was released from the support 11d . Inset: Purity analysis of the solid-phase-purified 12d by PAGE. Abbreviation: BP, bromophenol blue dye.

    Journal: Current protocols in nucleic acid chemistry

    Article Title: A High-Throughput Process for the Solid-Phase Purification of Synthetic DNA Sequences

    doi: 10.1002/cpnc.31

    Figure Lengend Snippet: Solid-phase purification of the phosphorothioate DNA sequence 10d . RP-HPLC analysis of solid-phase-purified 12d that was released from the support 11d . Inset: Purity analysis of the solid-phase-purified 12d by PAGE. Abbreviation: BP, bromophenol blue dye.

    Article Snippet: The purity of these sequences has been evaluated by RP-HPLC and by PAGE under denaturing conditions ( , , , and ).

    Techniques: Purification, Sequencing, High Performance Liquid Chromatography, Polyacrylamide Gel Electrophoresis

    Size exclusion and reversed-phase HPLC analysis of purified R0.6C. a Size exclusion chromatography was performed under native conditions in a phosphate buffer pH 6.7, to determine the amount of monomer in the sample. The peak corresponding to the monomer is indicated with the integrated area of the peak written above. b Reversed-phase HPLC–UV chromatograms recorded following analysis of purified protein batches. The peak at 17 min corresponds to monomeric R0.6C antigen

    Journal: Microbial Cell Factories

    Article Title: Construct design, production, and characterization of Plasmodium falciparum 48/45 R0.6C subunit protein produced in Lactococcus lactis as candidate vaccine

    doi: 10.1186/s12934-017-0710-0

    Figure Lengend Snippet: Size exclusion and reversed-phase HPLC analysis of purified R0.6C. a Size exclusion chromatography was performed under native conditions in a phosphate buffer pH 6.7, to determine the amount of monomer in the sample. The peak corresponding to the monomer is indicated with the integrated area of the peak written above. b Reversed-phase HPLC–UV chromatograms recorded following analysis of purified protein batches. The peak at 17 min corresponds to monomeric R0.6C antigen

    Article Snippet: SEC–HPLC and RP-HPLC analysis of R0.6C Native size exclusion high-performance liquid chromatography (SEC–HPLC) or reversed-phase HPLC (RP-HPLC) of intact R0.6C were performed using an Agilent 1100 Series HPLC System (Agilent Technologies, USA) equipped with a TSKgel G3000SWXL SEC column, 5 µm, 7.8 × 300 mm (Tosoh Bioscience, Japan) or equipped with a Vydac 214TP C4 reversed-phase column, 5 µm, 4.6 × 250 mm (The Separations Group, CA, US), respectively.

    Techniques: High Performance Liquid Chromatography, Purification, Size-exclusion Chromatography

    Analytical HPLC of purified mSAM. Gradient of 50–100% acetonitrile. RT: 10.7 min (lower panel), 11.5 min (upper panel). Absorbances at 450 nm (coumarin 343), 540 nm (TAMRA), 254 nm, and 280 nm are shown. Both 5-(6-)TAMRA isomers are shown separately (upper and lower panel).

    Journal: ACS Central Science

    Article Title: Cathepsin G Activity as a New Marker for Detecting Airway Inflammation by Microscopy and Flow Cytometry

    doi: 10.1021/acscentsci.8b00933

    Figure Lengend Snippet: Analytical HPLC of purified mSAM. Gradient of 50–100% acetonitrile. RT: 10.7 min (lower panel), 11.5 min (upper panel). Absorbances at 450 nm (coumarin 343), 540 nm (TAMRA), 254 nm, and 280 nm are shown. Both 5-(6-)TAMRA isomers are shown separately (upper and lower panel).

    Article Snippet: Reporter Purification sSAM and mSAM were analyzed and purified by RP-HPLC on a Shimadzu system equipped with a photodiode array detector (Duisburg, Germany).

    Techniques: High Performance Liquid Chromatography, Purification

    Mass spectra of sSAM. Expected masses: [M+1] + , 1902; [M+2] 2+ , 951; [M+3] 3+ , 634. The two spectra correspond to the two isomer peaks observed by HPLC ( Figure 6 ).

    Journal: ACS Central Science

    Article Title: Cathepsin G Activity as a New Marker for Detecting Airway Inflammation by Microscopy and Flow Cytometry

    doi: 10.1021/acscentsci.8b00933

    Figure Lengend Snippet: Mass spectra of sSAM. Expected masses: [M+1] + , 1902; [M+2] 2+ , 951; [M+3] 3+ , 634. The two spectra correspond to the two isomer peaks observed by HPLC ( Figure 6 ).

    Article Snippet: Reporter Purification sSAM and mSAM were analyzed and purified by RP-HPLC on a Shimadzu system equipped with a photodiode array detector (Duisburg, Germany).

    Techniques: High Performance Liquid Chromatography

    Analytical HPLC of purified sSAM. Gradient of 10–100% acetonitrile, RT: 9 and 9.5 min. Absorbance at 450 nm (coumarin 343), 540 nm (TAMRA), 254 nm, and 280 nm are shown. The two peaks represent the two 5-(6-)TAMRA isomers.

    Journal: ACS Central Science

    Article Title: Cathepsin G Activity as a New Marker for Detecting Airway Inflammation by Microscopy and Flow Cytometry

    doi: 10.1021/acscentsci.8b00933

    Figure Lengend Snippet: Analytical HPLC of purified sSAM. Gradient of 10–100% acetonitrile, RT: 9 and 9.5 min. Absorbance at 450 nm (coumarin 343), 540 nm (TAMRA), 254 nm, and 280 nm are shown. The two peaks represent the two 5-(6-)TAMRA isomers.

    Article Snippet: Reporter Purification sSAM and mSAM were analyzed and purified by RP-HPLC on a Shimadzu system equipped with a photodiode array detector (Duisburg, Germany).

    Techniques: High Performance Liquid Chromatography, Purification

    Enzymatic assay of recombinant TcLAR. A , HPLC analysis of products from incubation of leucocyanidin with boiled protein as control, monitored by UV spectroscopy. B , as above, for incubation of purified recombinant TcLAR with 3 H-leucocyanidin. C , As in (A) , but monitoring radioactivity of fractions; D , As in (B) , but monitoring radioactivity of fractions; ( E ), UV spectrum of catechin product from (B) ; F , HPLC analysis of an authentic standard of (−)-catechin. mAU, milliabsorbance units; CPM, counts per minute.

    Journal: BMC Plant Biology

    Article Title: Proanthocyanidin synthesis in Theobroma cacao: genes encoding anthocyanidin synthase, anthocyanidin reductase, and leucoanthocyanidin reductase

    doi: 10.1186/1471-2229-13-202

    Figure Lengend Snippet: Enzymatic assay of recombinant TcLAR. A , HPLC analysis of products from incubation of leucocyanidin with boiled protein as control, monitored by UV spectroscopy. B , as above, for incubation of purified recombinant TcLAR with 3 H-leucocyanidin. C , As in (A) , but monitoring radioactivity of fractions; D , As in (B) , but monitoring radioactivity of fractions; ( E ), UV spectrum of catechin product from (B) ; F , HPLC analysis of an authentic standard of (−)-catechin. mAU, milliabsorbance units; CPM, counts per minute.

    Article Snippet: Catechin and epicatechin content was determined by reverse-phase HPLC using an Alliance separations module (Model 2695; Waters, Milford, MA, USA) equipped with a multi λ fluorescence detector (Model 2475; Waters, Milford, MA, USA).

    Techniques: Enzymatic Assay, Recombinant, High Performance Liquid Chromatography, Incubation, Spectroscopy, Purification, Radioactivity

    Complementation of the Arabidopsis PA-deficient ldox mutant by constitutively expressing TcLAR . A , Catechin and epicatechin standards analyzed by HPLC. B , HPLC analysis of Col-0 young siliques. C , HPLC analysis of ldox 35S:TcLAR Line14 young siliques. D , HPLC analysis of ldox mutant young siliques. E . TcLAR and AtUbiquitin transcripts in total RNA from leaves of ldox 35S:TcLAR transgenic plants, (line 9, line 14 and line 36), Col-O and ldox muant plants by RT-PCR. PCR products from the TcLAR-PGEM plasmid were loaded on the last lane as a positive control for the TcLAR primer set and as a negative control for the AtUbiquitin primer set. F , Catechin and epicatechin levels in young siliques of plants that were the source of the total RNA used in (E) . Catechin and epicatechin levels were determined by extraction and HPLC analysis. The data are presented as means ± SE, n = 3. *P

    Journal: BMC Plant Biology

    Article Title: Proanthocyanidin synthesis in Theobroma cacao: genes encoding anthocyanidin synthase, anthocyanidin reductase, and leucoanthocyanidin reductase

    doi: 10.1186/1471-2229-13-202

    Figure Lengend Snippet: Complementation of the Arabidopsis PA-deficient ldox mutant by constitutively expressing TcLAR . A , Catechin and epicatechin standards analyzed by HPLC. B , HPLC analysis of Col-0 young siliques. C , HPLC analysis of ldox 35S:TcLAR Line14 young siliques. D , HPLC analysis of ldox mutant young siliques. E . TcLAR and AtUbiquitin transcripts in total RNA from leaves of ldox 35S:TcLAR transgenic plants, (line 9, line 14 and line 36), Col-O and ldox muant plants by RT-PCR. PCR products from the TcLAR-PGEM plasmid were loaded on the last lane as a positive control for the TcLAR primer set and as a negative control for the AtUbiquitin primer set. F , Catechin and epicatechin levels in young siliques of plants that were the source of the total RNA used in (E) . Catechin and epicatechin levels were determined by extraction and HPLC analysis. The data are presented as means ± SE, n = 3. *P

    Article Snippet: Catechin and epicatechin content was determined by reverse-phase HPLC using an Alliance separations module (Model 2695; Waters, Milford, MA, USA) equipped with a multi λ fluorescence detector (Model 2475; Waters, Milford, MA, USA).

    Techniques: Mutagenesis, Expressing, High Performance Liquid Chromatography, Transgenic Assay, Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Plasmid Preparation, Positive Control, Negative Control

    Expression analysis of key PA biosynthesis genes, and PA composition in various tissues of Theobroma cacao . A , Gene expression levels of TcANR , TcLAR1 and TcLAR1 . Expression was determined by semi-quantitative RT-PCR normalized relative to the expression of TcActin in each sample. B , Levels of soluble PAs expressed as mg PA per g of dry weight. C , Levels of insoluble PAs expressed as mg PA per g of dry weight. D , Catechin and epicatechin standards analyzed by HPLC. E , Representative HPLC analysis of flavan-3-ols extracted from cacao leaves. F , Flavan-3-ol accumulation and composition analyzed by HPLC. All data are presented as means ± SE, for gene expression data, n ≥ 3, for PA level data, n ≥ 5. DW, dry weight.

    Journal: BMC Plant Biology

    Article Title: Proanthocyanidin synthesis in Theobroma cacao: genes encoding anthocyanidin synthase, anthocyanidin reductase, and leucoanthocyanidin reductase

    doi: 10.1186/1471-2229-13-202

    Figure Lengend Snippet: Expression analysis of key PA biosynthesis genes, and PA composition in various tissues of Theobroma cacao . A , Gene expression levels of TcANR , TcLAR1 and TcLAR1 . Expression was determined by semi-quantitative RT-PCR normalized relative to the expression of TcActin in each sample. B , Levels of soluble PAs expressed as mg PA per g of dry weight. C , Levels of insoluble PAs expressed as mg PA per g of dry weight. D , Catechin and epicatechin standards analyzed by HPLC. E , Representative HPLC analysis of flavan-3-ols extracted from cacao leaves. F , Flavan-3-ol accumulation and composition analyzed by HPLC. All data are presented as means ± SE, for gene expression data, n ≥ 3, for PA level data, n ≥ 5. DW, dry weight.

    Article Snippet: Catechin and epicatechin content was determined by reverse-phase HPLC using an Alliance separations module (Model 2695; Waters, Milford, MA, USA) equipped with a multi λ fluorescence detector (Model 2475; Waters, Milford, MA, USA).

    Techniques: Expressing, Quantitative RT-PCR, High Performance Liquid Chromatography

    The synthesis of sugar nucleotides from 2-chloro-4-nitrophenyl glucosides. ( a ) General reaction scheme. ( b ) Structures of 2-chloro-4-nitrophenyl glycoside donors evaluated for D-sugars within this series, the differences between each member and the native OleD sugar substrate (β-D-glucose) are highlighted in red. ( c ) Maximum observed percent conversion of (U/T)DP to (U/T)DP-glucose within a 21 hour time course assay for each donor (n ≥ 2, standard deviation ≤ 5%). Standard reactions contained 7 μM TDP-16, 1 mM (U/T)DP, and 1 mM of 2-chloro-4-nitrophenyl glycoside donor ( 9 , 34 – 47 ) in Tris-HCl buffer (50 mM, pH 8.5) with a final volume of 300 μl. Over 21 hours at 25°C, aliquots taken at various times were flash frozen and analyzed by HPLC ( Supplementary Methods ). For reactions with UDP yielding

    Journal: Nature chemical biology

    Article Title: Using Simple Donors to Drive the Equilibria of Glycosyltransferase-Catalyzed Reactions

    doi: 10.1038/nchembio.638

    Figure Lengend Snippet: The synthesis of sugar nucleotides from 2-chloro-4-nitrophenyl glucosides. ( a ) General reaction scheme. ( b ) Structures of 2-chloro-4-nitrophenyl glycoside donors evaluated for D-sugars within this series, the differences between each member and the native OleD sugar substrate (β-D-glucose) are highlighted in red. ( c ) Maximum observed percent conversion of (U/T)DP to (U/T)DP-glucose within a 21 hour time course assay for each donor (n ≥ 2, standard deviation ≤ 5%). Standard reactions contained 7 μM TDP-16, 1 mM (U/T)DP, and 1 mM of 2-chloro-4-nitrophenyl glycoside donor ( 9 , 34 – 47 ) in Tris-HCl buffer (50 mM, pH 8.5) with a final volume of 300 μl. Over 21 hours at 25°C, aliquots taken at various times were flash frozen and analyzed by HPLC ( Supplementary Methods ). For reactions with UDP yielding

    Article Snippet: Following, samples were thawed at 4°C and filtered through a MultiScreen Filter Plate (Millipore, Billerica, MA, USA) according to manufacturer’s instructions and evaluated for formation of UDP- ( 33a ) or TDP-α-D-glucose ( 33b ) by analytical reverse-phase HPLC with a 250 mm × 4.6 mm Gemini-NX 5μ C18 column (Phenomenex, Torrance, CA, USA) using a linear gradient of 0% to 15% CH3 CN (solvent B) over 15 minutes (solvent A = aqueous 50 mM triethylammonium acetate buffer [Sigma-Aldrich, St. Louis, MO, USA], flow rate = 1 ml min−1 , with detection monitored at 254 nm).

    Techniques: Standard Deviation, High Performance Liquid Chromatography

    Evaluation of putative donors for sugar nucleotide synthesis. ( a ) General reaction scheme. ( b ) Structures of the β-D-glucopyranoside donors which led to (U/T)DP-glucose formation. ( c ) Percent conversion of (U/T)DP to (U/T)DP-glucose with various donors (n ≥ 2, standard deviation ≤ 5%). Reactions contained 2.1 μM (10 μg) OleD variant, 1 mM of (U/T)DP, and 1 mM of aromatic donor ( 1–9 ) in Tris-HCl buffer (50 mM, pH 8.5) with a final volume of 100 μl. After one hour at 25°C, reactions were flash frozen and analyzed by HPLC ( Supplementary Methods ). The pK a for each corresponding donor aglycon is highlighted in parentheses. ( d ) Plot depicting the relative Gibbs free energy of selected donors/acceptors in relation to 33a . Small glycoside donors display large shifts in relative free energy, transforming formation of UDP-Glc ( 33a ) from an endo- to an exothermic process. The ΔG° pH8.5 for 1 , 2, 4, 7, and 9 with UDP in Tris-HCl buffer (50 mM, pH 8.5) at 298K relative to 33a were determined in this study ( Supplementary Methods ). The ΔG° for 61a was previously determined (at pH 9.0 and 310K) ( 5 ) .

    Journal: Nature chemical biology

    Article Title: Using Simple Donors to Drive the Equilibria of Glycosyltransferase-Catalyzed Reactions

    doi: 10.1038/nchembio.638

    Figure Lengend Snippet: Evaluation of putative donors for sugar nucleotide synthesis. ( a ) General reaction scheme. ( b ) Structures of the β-D-glucopyranoside donors which led to (U/T)DP-glucose formation. ( c ) Percent conversion of (U/T)DP to (U/T)DP-glucose with various donors (n ≥ 2, standard deviation ≤ 5%). Reactions contained 2.1 μM (10 μg) OleD variant, 1 mM of (U/T)DP, and 1 mM of aromatic donor ( 1–9 ) in Tris-HCl buffer (50 mM, pH 8.5) with a final volume of 100 μl. After one hour at 25°C, reactions were flash frozen and analyzed by HPLC ( Supplementary Methods ). The pK a for each corresponding donor aglycon is highlighted in parentheses. ( d ) Plot depicting the relative Gibbs free energy of selected donors/acceptors in relation to 33a . Small glycoside donors display large shifts in relative free energy, transforming formation of UDP-Glc ( 33a ) from an endo- to an exothermic process. The ΔG° pH8.5 for 1 , 2, 4, 7, and 9 with UDP in Tris-HCl buffer (50 mM, pH 8.5) at 298K relative to 33a were determined in this study ( Supplementary Methods ). The ΔG° for 61a was previously determined (at pH 9.0 and 310K) ( 5 ) .

    Article Snippet: Following, samples were thawed at 4°C and filtered through a MultiScreen Filter Plate (Millipore, Billerica, MA, USA) according to manufacturer’s instructions and evaluated for formation of UDP- ( 33a ) or TDP-α-D-glucose ( 33b ) by analytical reverse-phase HPLC with a 250 mm × 4.6 mm Gemini-NX 5μ C18 column (Phenomenex, Torrance, CA, USA) using a linear gradient of 0% to 15% CH3 CN (solvent B) over 15 minutes (solvent A = aqueous 50 mM triethylammonium acetate buffer [Sigma-Aldrich, St. Louis, MO, USA], flow rate = 1 ml min−1 , with detection monitored at 254 nm).

    Techniques: Standard Deviation, Variant Assay, High Performance Liquid Chromatography, Gas Chromatography

    Evaluation of 2-chloro-4-nitrophenyl glycosides as sugar donors in coupled GT-catalyzed transglycosylation reactions. ( a ) The scheme for a single enzyme (TDP-16) coupled system with 4-methylumbelliferone ( 58) as the final acceptor (left) and a representative HPLC analysis (right) using the donor for 6-azido-6-deoxy-D-glucose ( 37 ). Reactions contained 1 mM glycoside donor, 1 mM 58 , 1 mM UDP, and 11 μM TDP-16 in a total volume of 100 μl with Tris-HCl buffer (50 mM, pH 8.5) at 25°C for 24 hour and were subsequently analyzed by HPLC ( Supplementary Methods ). For the representative reaction: (i) control reaction lacking TDP-16; (ii) control reaction lacking UDP; (iii) full reaction where 37 is donor, 58 is acceptor, 59d is desired product and ⋄ represents 2-chloro-4-nitrophenolate. ( b ) The scheme for a double enzyme (TDP-16 and GtfE) coupled system with vancomycin aglycon ( 60 ) as the final acceptor (left) and a representative HPLC analysis (right) using the donor for 6-azido-6-deoxy-D-glucose ( 37 ). Reactions contained 1 mM glycoside donor, 0.1 mM 60 , 1 mM UDP, 11 μM TDP-16, and 11 μM GtfE in a total volume of 100 μl with Tris-HCl buffer (50 mM, pH 8.5) at 25°C for 24 hour and were subsequently analyzed by HPLC ( Supplementary Methods ). For the representative reaction: (i) control reaction lacking TDP-16; (ii) control reaction lacking GtfE; (iii) full reaction where 37 is donor, 60 is acceptor, 61e is desired product and ⋄ represents 2-chloro-4-nitrophenolate. Sample preparation and HPLC parameters, along with chromatograms ( Supplementary Fig. 14 and 17 ), conversion rates, and mass characterization ( Supplementary Table 4 and 5 ) for all products are presented in supporting online material .

    Journal: Nature chemical biology

    Article Title: Using Simple Donors to Drive the Equilibria of Glycosyltransferase-Catalyzed Reactions

    doi: 10.1038/nchembio.638

    Figure Lengend Snippet: Evaluation of 2-chloro-4-nitrophenyl glycosides as sugar donors in coupled GT-catalyzed transglycosylation reactions. ( a ) The scheme for a single enzyme (TDP-16) coupled system with 4-methylumbelliferone ( 58) as the final acceptor (left) and a representative HPLC analysis (right) using the donor for 6-azido-6-deoxy-D-glucose ( 37 ). Reactions contained 1 mM glycoside donor, 1 mM 58 , 1 mM UDP, and 11 μM TDP-16 in a total volume of 100 μl with Tris-HCl buffer (50 mM, pH 8.5) at 25°C for 24 hour and were subsequently analyzed by HPLC ( Supplementary Methods ). For the representative reaction: (i) control reaction lacking TDP-16; (ii) control reaction lacking UDP; (iii) full reaction where 37 is donor, 58 is acceptor, 59d is desired product and ⋄ represents 2-chloro-4-nitrophenolate. ( b ) The scheme for a double enzyme (TDP-16 and GtfE) coupled system with vancomycin aglycon ( 60 ) as the final acceptor (left) and a representative HPLC analysis (right) using the donor for 6-azido-6-deoxy-D-glucose ( 37 ). Reactions contained 1 mM glycoside donor, 0.1 mM 60 , 1 mM UDP, 11 μM TDP-16, and 11 μM GtfE in a total volume of 100 μl with Tris-HCl buffer (50 mM, pH 8.5) at 25°C for 24 hour and were subsequently analyzed by HPLC ( Supplementary Methods ). For the representative reaction: (i) control reaction lacking TDP-16; (ii) control reaction lacking GtfE; (iii) full reaction where 37 is donor, 60 is acceptor, 61e is desired product and ⋄ represents 2-chloro-4-nitrophenolate. Sample preparation and HPLC parameters, along with chromatograms ( Supplementary Fig. 14 and 17 ), conversion rates, and mass characterization ( Supplementary Table 4 and 5 ) for all products are presented in supporting online material .

    Article Snippet: Following, samples were thawed at 4°C and filtered through a MultiScreen Filter Plate (Millipore, Billerica, MA, USA) according to manufacturer’s instructions and evaluated for formation of UDP- ( 33a ) or TDP-α-D-glucose ( 33b ) by analytical reverse-phase HPLC with a 250 mm × 4.6 mm Gemini-NX 5μ C18 column (Phenomenex, Torrance, CA, USA) using a linear gradient of 0% to 15% CH3 CN (solvent B) over 15 minutes (solvent A = aqueous 50 mM triethylammonium acetate buffer [Sigma-Aldrich, St. Louis, MO, USA], flow rate = 1 ml min−1 , with detection monitored at 254 nm).

    Techniques: High Performance Liquid Chromatography, Sample Prep

    Utilizing a colorimetric screen for glycosyl transfer. ( a ) Scheme for colorimetric screen using the single enzyme (TDP-16) coupled format. ( b ) Evaluation of the colorimetric assay with 58 as the final acceptor. The reactions contained 0.5 mM 9 as donor, 0.5 mM 58 as acceptor, 5 μM UDP, and 11μM TDP-16 in a final total volume of 100 μl with Tris-HCl buffer (50 mM, pH 8.5) in a 96-well plate incubated at 25°C for one hour. ( i ) Qualitative color change after one hour for the full reaction (yellow square), a control lacking the final acceptor 58 (white circle), and a control lacking UDP (red triangle). ( ii ) Δ410 nm over one hour for the full reaction (yellow squares), a control lacking the final acceptor 58 (white circles), and a control reaction lacking UDP (red triangles). ( iii ) HPLC chromatograms of full reaction at 1, 5, and 60 min where 1 is desired product, 9 is the donor, 58 is the target aglycon and ⋄ represents 2-chloro-4-nitrophenolate. (c) The absorbance data and HPLC chromatograms of three representative hits [( i ) 62 (genistein), ( ii ) 79 (tyrphostin), or ( iii ) 92 (ciprofloxacin)] from the broad 50 compound panel screen using the single enzyme (TDP-16) coupled format. In HPLC chromatograms 9 indicates donor; 62 , 79 or 92 represent target aglycon; ⋄ indicates 2-chloro-4-nitrophenolate; and ● depicts glucosylated product(s). For the overall results of the 50 compound screen, additional representative absorbance plots and chromatograms, and combined HPLC and LC/MS characterization, see Supplementary Fig. 19–21 and Supplementary Table 6 .

    Journal: Nature chemical biology

    Article Title: Using Simple Donors to Drive the Equilibria of Glycosyltransferase-Catalyzed Reactions

    doi: 10.1038/nchembio.638

    Figure Lengend Snippet: Utilizing a colorimetric screen for glycosyl transfer. ( a ) Scheme for colorimetric screen using the single enzyme (TDP-16) coupled format. ( b ) Evaluation of the colorimetric assay with 58 as the final acceptor. The reactions contained 0.5 mM 9 as donor, 0.5 mM 58 as acceptor, 5 μM UDP, and 11μM TDP-16 in a final total volume of 100 μl with Tris-HCl buffer (50 mM, pH 8.5) in a 96-well plate incubated at 25°C for one hour. ( i ) Qualitative color change after one hour for the full reaction (yellow square), a control lacking the final acceptor 58 (white circle), and a control lacking UDP (red triangle). ( ii ) Δ410 nm over one hour for the full reaction (yellow squares), a control lacking the final acceptor 58 (white circles), and a control reaction lacking UDP (red triangles). ( iii ) HPLC chromatograms of full reaction at 1, 5, and 60 min where 1 is desired product, 9 is the donor, 58 is the target aglycon and ⋄ represents 2-chloro-4-nitrophenolate. (c) The absorbance data and HPLC chromatograms of three representative hits [( i ) 62 (genistein), ( ii ) 79 (tyrphostin), or ( iii ) 92 (ciprofloxacin)] from the broad 50 compound panel screen using the single enzyme (TDP-16) coupled format. In HPLC chromatograms 9 indicates donor; 62 , 79 or 92 represent target aglycon; ⋄ indicates 2-chloro-4-nitrophenolate; and ● depicts glucosylated product(s). For the overall results of the 50 compound screen, additional representative absorbance plots and chromatograms, and combined HPLC and LC/MS characterization, see Supplementary Fig. 19–21 and Supplementary Table 6 .

    Article Snippet: Following, samples were thawed at 4°C and filtered through a MultiScreen Filter Plate (Millipore, Billerica, MA, USA) according to manufacturer’s instructions and evaluated for formation of UDP- ( 33a ) or TDP-α-D-glucose ( 33b ) by analytical reverse-phase HPLC with a 250 mm × 4.6 mm Gemini-NX 5μ C18 column (Phenomenex, Torrance, CA, USA) using a linear gradient of 0% to 15% CH3 CN (solvent B) over 15 minutes (solvent A = aqueous 50 mM triethylammonium acetate buffer [Sigma-Aldrich, St. Louis, MO, USA], flow rate = 1 ml min−1 , with detection monitored at 254 nm).

    Techniques: Colorimetric Assay, Incubation, High Performance Liquid Chromatography, Liquid Chromatography with Mass Spectroscopy

    The radioactive signal from oligopeptides being modified with radioactive acetyl CoA . The oligopeptide 1 MLGTE-RRR 24 (200 μM) was incubated with [1- 14 C] acetyl CoA (final 300 μM with specific activity 11.2 mCi/mmol) and purified MBP-hNaa30p (80 nM) in acetylation buffer for 60 minutes at 37°C. Samples were collected at indicated time points and analysed with reverse phase HPLC that had been connected to a radioactivity flow detector after the absorbance detector. A clear time dependent increase in the radioactivity signal is observed, verifying that the eluted oligopeptides are labelled with radioactive acetyl groups.

    Journal: BMC Proceedings

    Article Title: Application of reverse-phase HPLC to quantify oligopeptide acetylation eliminates interference from unspecific acetyl CoA hydrolysis

    doi: 10.1186/1753-6561-3-S6-S5

    Figure Lengend Snippet: The radioactive signal from oligopeptides being modified with radioactive acetyl CoA . The oligopeptide 1 MLGTE-RRR 24 (200 μM) was incubated with [1- 14 C] acetyl CoA (final 300 μM with specific activity 11.2 mCi/mmol) and purified MBP-hNaa30p (80 nM) in acetylation buffer for 60 minutes at 37°C. Samples were collected at indicated time points and analysed with reverse phase HPLC that had been connected to a radioactivity flow detector after the absorbance detector. A clear time dependent increase in the radioactivity signal is observed, verifying that the eluted oligopeptides are labelled with radioactive acetyl groups.

    Article Snippet: Separation of non acetylated and acetylated oligopeptides using reverse phase HPLC The acetylation activity was analysed by a reverse phase HPLC system, consisting of a LC-20AB solvent delivery module, an SPD-M20A photodiode array detector and a SIL-20AC autosampler (Shimadzu Prominence), and a 250 mm × 3 mm Nucleosil C18 HD column (Macherey-Nagel) reverse phase HPLC column.

    Techniques: Modification, Incubation, Activity Assay, Purification, High Performance Liquid Chromatography, Radioactivity, Flow Cytometry

    MBP-hNaa30p specificity constants (V/K) for selected oligopeptides based on detection of acetylated oligopeptides (215 nm) . Purified MBP-hNaa30p (80 nM) was incubated with selected oligopeptides and acetyl CoA (300 μM) in acetylation buffer for 30 minutes at 37°C. The acetylation kinetics was analysed by reverse phase HPLC. V/K is the V max /K m (oligopeptides) . Error bars indicate S.D. Experiments are performed in triplicates.

    Journal: BMC Proceedings

    Article Title: Application of reverse-phase HPLC to quantify oligopeptide acetylation eliminates interference from unspecific acetyl CoA hydrolysis

    doi: 10.1186/1753-6561-3-S6-S5

    Figure Lengend Snippet: MBP-hNaa30p specificity constants (V/K) for selected oligopeptides based on detection of acetylated oligopeptides (215 nm) . Purified MBP-hNaa30p (80 nM) was incubated with selected oligopeptides and acetyl CoA (300 μM) in acetylation buffer for 30 minutes at 37°C. The acetylation kinetics was analysed by reverse phase HPLC. V/K is the V max /K m (oligopeptides) . Error bars indicate S.D. Experiments are performed in triplicates.

    Article Snippet: Separation of non acetylated and acetylated oligopeptides using reverse phase HPLC The acetylation activity was analysed by a reverse phase HPLC system, consisting of a LC-20AB solvent delivery module, an SPD-M20A photodiode array detector and a SIL-20AC autosampler (Shimadzu Prominence), and a 250 mm × 3 mm Nucleosil C18 HD column (Macherey-Nagel) reverse phase HPLC column.

    Techniques: Purification, Incubation, High Performance Liquid Chromatography

    Reverse phase HPLC absorbance profile at 215 nm for the separation of acetylated and non-acetylated peptides . A ; The oligopeptide 1 MLGTE-RRR 24 (200 μM) was incubated with acetyl CoA (300 μM) and purified MBP-hNaa30p (80 nM) in acetylation buffer for 60 minutes at 37°C. Samples were collected at indicated time points and analysed with reverse phase HPLC. The resulting absorbance profile at 215 nm indicate good separation of unacetylated ('a') and acetylated oligopeptides ('b'). B ; An expanded version of the absorbance profile for the formation of acetylated oligopeptide. A clear time dependent increase in the absorption signal is observed.

    Journal: BMC Proceedings

    Article Title: Application of reverse-phase HPLC to quantify oligopeptide acetylation eliminates interference from unspecific acetyl CoA hydrolysis

    doi: 10.1186/1753-6561-3-S6-S5

    Figure Lengend Snippet: Reverse phase HPLC absorbance profile at 215 nm for the separation of acetylated and non-acetylated peptides . A ; The oligopeptide 1 MLGTE-RRR 24 (200 μM) was incubated with acetyl CoA (300 μM) and purified MBP-hNaa30p (80 nM) in acetylation buffer for 60 minutes at 37°C. Samples were collected at indicated time points and analysed with reverse phase HPLC. The resulting absorbance profile at 215 nm indicate good separation of unacetylated ('a') and acetylated oligopeptides ('b'). B ; An expanded version of the absorbance profile for the formation of acetylated oligopeptide. A clear time dependent increase in the absorption signal is observed.

    Article Snippet: Separation of non acetylated and acetylated oligopeptides using reverse phase HPLC The acetylation activity was analysed by a reverse phase HPLC system, consisting of a LC-20AB solvent delivery module, an SPD-M20A photodiode array detector and a SIL-20AC autosampler (Shimadzu Prominence), and a 250 mm × 3 mm Nucleosil C18 HD column (Macherey-Nagel) reverse phase HPLC column.

    Techniques: High Performance Liquid Chromatography, Incubation, Purification

    Purification of stingray ghrelin-like peptide (GRLN-LP) from stomach extracts . Black bars indicate the measured fluorescence changes in intracellular calcium ion concentrations in CHO cells expressing rat GHS-R1a (CHO-GHSR62). (A) Carboxymethyl (CM)-cation ion-exchange HPLC (pH 4.8) of the SP-III fraction of stomach extracts. The GRLN-like activity was divided into four groups (A-D). (B) Preparative reverse-phase (RP)-HPLC (Symmetry C18, 3.9 × 150 mm) of group B after purification with an anti-rat GRLN1-11 immuno-affinity column. (C) Final purification of the active fraction indicated in (B) by another RP-HPLC (Vydac diphenyl, 219TP5215, 2.1 × 150 mm).

    Journal: BMC Biochemistry

    Article Title: Ghrelin-like peptide with fatty acid modification and O-glycosylation in the red stingray, Dasyatis akajei

    doi: 10.1186/1471-2091-10-30

    Figure Lengend Snippet: Purification of stingray ghrelin-like peptide (GRLN-LP) from stomach extracts . Black bars indicate the measured fluorescence changes in intracellular calcium ion concentrations in CHO cells expressing rat GHS-R1a (CHO-GHSR62). (A) Carboxymethyl (CM)-cation ion-exchange HPLC (pH 4.8) of the SP-III fraction of stomach extracts. The GRLN-like activity was divided into four groups (A-D). (B) Preparative reverse-phase (RP)-HPLC (Symmetry C18, 3.9 × 150 mm) of group B after purification with an anti-rat GRLN1-11 immuno-affinity column. (C) Final purification of the active fraction indicated in (B) by another RP-HPLC (Vydac diphenyl, 219TP5215, 2.1 × 150 mm).

    Article Snippet: The samples were first applied to a preparative RP-HPLC with a Symmetry C18 column (3.9 × 150 mm, Waters) at a flow rate of 1 ml/min under a linear gradient from 10% to 60% acetonitlile containing 0.1% TFA for 40 min.

    Techniques: Purification, Fluorescence, Expressing, High Performance Liquid Chromatography, Activity Assay, Affinity Column

    RP-HPLC separation of TPIs from N. attenuata of the wild-type genotype. Numbers refer to TPI fractions containing inhibitory activity against trypsin. RP-HPLC profile of TPIs from uninduced control leaves (dashed line, right y axis) and leaves elicited

    Journal: Plant Physiology

    Article Title: Differential Elicitation of Two Processing Proteases Controls the Processing Pattern of the Trypsin Proteinase Inhibitor Precursor in Nicotiana attenuata 1

    doi: 10.1104/pp.105.064006

    Figure Lengend Snippet: RP-HPLC separation of TPIs from N. attenuata of the wild-type genotype. Numbers refer to TPI fractions containing inhibitory activity against trypsin. RP-HPLC profile of TPIs from uninduced control leaves (dashed line, right y axis) and leaves elicited

    Article Snippet: The total TPI pool was chromatographed on RP-HPLC performed on a Hitachi LaChrom L7100.

    Techniques: High Performance Liquid Chromatography, Activity Assay

    Chain composition of the two-chain TPI from N. attenuata . A, Separation of the chains. The purified two-chain TPI (5 μ g) was reductively carboxymethylated, and the liberated chains were purified by RP-HPLC (solid line). The position of the original

    Journal: Plant Physiology

    Article Title: Differential Elicitation of Two Processing Proteases Controls the Processing Pattern of the Trypsin Proteinase Inhibitor Precursor in Nicotiana attenuata 1

    doi: 10.1104/pp.105.064006

    Figure Lengend Snippet: Chain composition of the two-chain TPI from N. attenuata . A, Separation of the chains. The purified two-chain TPI (5 μ g) was reductively carboxymethylated, and the liberated chains were purified by RP-HPLC (solid line). The position of the original

    Article Snippet: The total TPI pool was chromatographed on RP-HPLC performed on a Hitachi LaChrom L7100.

    Techniques: Purification, High Performance Liquid Chromatography

    RP-HPLC separation of TPIs from the Arizona genotype S++, which was transformed with the TPI gene from Utah under control of a constitutive promoter to restore TPI production. Numbers refer to TPI fractions containing inhibitory activity

    Journal: Plant Physiology

    Article Title: Differential Elicitation of Two Processing Proteases Controls the Processing Pattern of the Trypsin Proteinase Inhibitor Precursor in Nicotiana attenuata 1

    doi: 10.1104/pp.105.064006

    Figure Lengend Snippet: RP-HPLC separation of TPIs from the Arizona genotype S++, which was transformed with the TPI gene from Utah under control of a constitutive promoter to restore TPI production. Numbers refer to TPI fractions containing inhibitory activity

    Article Snippet: The total TPI pool was chromatographed on RP-HPLC performed on a Hitachi LaChrom L7100.

    Techniques: High Performance Liquid Chromatography, Transformation Assay, Activity Assay

    Translocation. A. Colicin E3-cleaved (closed circles) or control (open circles) pre-translocation complexes (0.1 μM) were mixed in a quench-flow apparatus with EF-G·GTP (3 μM) and Pmn (10 mM) and translocation analysed by formation of fMetPhe-Pmn. The amount of pre-translocation complex (PTC) was determined by HPLC analysis of fMetPhe. B. Translocation was monitored by the fluorescence change of fMetPhe-tRNA Phe (Prf16/17). Curve 1 – control ribosomes; curve 2 – colicin E3-cleaved ribosomes; curve 3 – control ribosomes in the presence of viomycin (200 μM); curve 4 – colicin E3-cleaved ribosomes with viomycin (200 μM). Normalized ΔF, fluorescence change normalized by the maximum extent of translocation determined by the Pmn reaction. C. Pi release upon GTP hydrolysis by EF-G (curves 1 and 2) or buffer control (curves 3 and 4) with control ribosomes (curves 1 and 3) or colicin E3-cleaved (curves 2 and 4). Pi release was measured by the fluorescence change of MDCC-PBP upon Pi binding ( Experimental procedures ). D. Multiple-turnover GTP hydrolysis by EF-G. Velocity of GTP hydrolysis was measured with catalytic concentrations of EF-G (10 nM) and increasing concentrations of colicin E3-cleaved ribosomes (closed circles) or control ribosomes (open circles) in the presence of excess [γ- 32 P]-GTP (200 μM).

    Journal: Molecular Microbiology

    Article Title: Colicin E3 cleavage of 16S rRNA impairs decoding and accelerates tRNA translocation on Escherichia coli ribosomes

    doi: 10.1111/j.1365-2958.2008.06283.x

    Figure Lengend Snippet: Translocation. A. Colicin E3-cleaved (closed circles) or control (open circles) pre-translocation complexes (0.1 μM) were mixed in a quench-flow apparatus with EF-G·GTP (3 μM) and Pmn (10 mM) and translocation analysed by formation of fMetPhe-Pmn. The amount of pre-translocation complex (PTC) was determined by HPLC analysis of fMetPhe. B. Translocation was monitored by the fluorescence change of fMetPhe-tRNA Phe (Prf16/17). Curve 1 – control ribosomes; curve 2 – colicin E3-cleaved ribosomes; curve 3 – control ribosomes in the presence of viomycin (200 μM); curve 4 – colicin E3-cleaved ribosomes with viomycin (200 μM). Normalized ΔF, fluorescence change normalized by the maximum extent of translocation determined by the Pmn reaction. C. Pi release upon GTP hydrolysis by EF-G (curves 1 and 2) or buffer control (curves 3 and 4) with control ribosomes (curves 1 and 3) or colicin E3-cleaved (curves 2 and 4). Pi release was measured by the fluorescence change of MDCC-PBP upon Pi binding ( Experimental procedures ). D. Multiple-turnover GTP hydrolysis by EF-G. Velocity of GTP hydrolysis was measured with catalytic concentrations of EF-G (10 nM) and increasing concentrations of colicin E3-cleaved ribosomes (closed circles) or control ribosomes (open circles) in the presence of excess [γ- 32 P]-GTP (200 μM).

    Article Snippet: The amount of fMetPhe formed was determined by reversed-phase HPLC (Lichrospher 100 RP-8, Merck).

    Techniques: Translocation Assay, Flow Cytometry, High Performance Liquid Chromatography, Fluorescence, Binding Assay

    Recombinant production of Mb1a. Semi-preparative RP-HPLC chromatogram of recombinant Mb1a released by TEV protease cleavage of the MBP-Mb1a fusion protein (see Materials and Methods for more details). The dotted line indicates the gradient of solvent B (90% acetonitrile/0.043% TFA). Top inset: SDS-PAGE gel showing pre-cleaved MBP-Mb1a fusion protein (lane 1) and remaining MBP after cleavage (lane 2). Lane M contains molecular markers (masses in kDa). Bottom inset: MALDI-TOF mass spectrum of pure recombinant Mb1a.

    Journal: Toxins

    Article Title: Insect-Active Toxins with Promiscuous Pharmacology from the African Theraphosid Spider Monocentropus balfouri

    doi: 10.3390/toxins9050155

    Figure Lengend Snippet: Recombinant production of Mb1a. Semi-preparative RP-HPLC chromatogram of recombinant Mb1a released by TEV protease cleavage of the MBP-Mb1a fusion protein (see Materials and Methods for more details). The dotted line indicates the gradient of solvent B (90% acetonitrile/0.043% TFA). Top inset: SDS-PAGE gel showing pre-cleaved MBP-Mb1a fusion protein (lane 1) and remaining MBP after cleavage (lane 2). Lane M contains molecular markers (masses in kDa). Bottom inset: MALDI-TOF mass spectrum of pure recombinant Mb1a.

    Article Snippet: The supernatant was filtered with a 0.45 μm syringe filter (EMD Millipore, Billerica, MA, USA) before purification of recombinant Mb1a or Mb1b using semi-preparative RP-HPLC (Phenomenex Jupiter C4 column; 250 × 10 mm, 10 μm; flow rate 5 mL/min).

    Techniques: Recombinant, High Performance Liquid Chromatography, SDS Page

    ( a ) Photo of a female M. balfouri ; ( b ) Chromatogram resulting from RP-HPLC fractionation of M. balfouri venom. The peak highlighted in red contains the µ/ω-TRTX-Mb1a/b peptide. The dotted line indicates the gradient of solvent B (90% acetonitrile/0.1% formic acid). Inset is a MALDI-TOF mass spectrum of the isolated µ/ω-TRTX-Mb1a/b peptide.

    Journal: Toxins

    Article Title: Insect-Active Toxins with Promiscuous Pharmacology from the African Theraphosid Spider Monocentropus balfouri

    doi: 10.3390/toxins9050155

    Figure Lengend Snippet: ( a ) Photo of a female M. balfouri ; ( b ) Chromatogram resulting from RP-HPLC fractionation of M. balfouri venom. The peak highlighted in red contains the µ/ω-TRTX-Mb1a/b peptide. The dotted line indicates the gradient of solvent B (90% acetonitrile/0.1% formic acid). Inset is a MALDI-TOF mass spectrum of the isolated µ/ω-TRTX-Mb1a/b peptide.

    Article Snippet: The supernatant was filtered with a 0.45 μm syringe filter (EMD Millipore, Billerica, MA, USA) before purification of recombinant Mb1a or Mb1b using semi-preparative RP-HPLC (Phenomenex Jupiter C4 column; 250 × 10 mm, 10 μm; flow rate 5 mL/min).

    Techniques: High Performance Liquid Chromatography, Fractionation, Isolation

    Distinct HRNR-derived CIDAMPs show similar ribosomal protein binding patterns. ( a ) SulfoLink®–column-bound proteins of an E . coli -extract were separated on a Jupiter® C18 RP-HPLC column with a Prp-gradient. HPLC fractions containing UV-absorbing peaks (C1–D10) were divided into five aliquots and adjusted in parallel to five PAGE-gels and separated. ( b ) Silver-stained proteins. ( c ) HRNR-Far-Western blot for probing with biotinylated HR1-18 (HRNR 2556–2677 ) using Strep - Tactin ®, ( d ) HRNR-Far-Western blot for probing with biotinylated rSumo3-HRNR 2591–2684 using Strep - Tactin ®, ( e ) HRNR-Far-Western blot for probing with rHRNR 2591–2684 using anti-HRNR 2591–2684 antibodies, ( f ) HRNR-Far-Western blot for probing with rHRNR 1075–1172 using anti-HRNR 1075–1172 antibodies. Note similarities of the staining patterns, irrespective the CIDAMP AA-sequence or biotin-labeling and irrespective whether a Strep - Tactin ®- or antibody-detectable CIDAMP was used to probe and detect the target protein on the membrane. Note the presence of 70 kDa bands upon HRNR-Far-Western blot analyses in most of the investigated HPLC fractions with highest intensity for rHRNR 1075–1172 binding ( f ). The most intensive band, corresponding to a 37 kDa protein in fraction number C13, was identified as E . coli ribosomal protein L2.

    Journal: Scientific Reports

    Article Title: Hornerin contains a Linked Series of Ribosome-Targeting Peptide Antibiotics

    doi: 10.1038/s41598-018-34467-8

    Figure Lengend Snippet: Distinct HRNR-derived CIDAMPs show similar ribosomal protein binding patterns. ( a ) SulfoLink®–column-bound proteins of an E . coli -extract were separated on a Jupiter® C18 RP-HPLC column with a Prp-gradient. HPLC fractions containing UV-absorbing peaks (C1–D10) were divided into five aliquots and adjusted in parallel to five PAGE-gels and separated. ( b ) Silver-stained proteins. ( c ) HRNR-Far-Western blot for probing with biotinylated HR1-18 (HRNR 2556–2677 ) using Strep - Tactin ®, ( d ) HRNR-Far-Western blot for probing with biotinylated rSumo3-HRNR 2591–2684 using Strep - Tactin ®, ( e ) HRNR-Far-Western blot for probing with rHRNR 2591–2684 using anti-HRNR 2591–2684 antibodies, ( f ) HRNR-Far-Western blot for probing with rHRNR 1075–1172 using anti-HRNR 1075–1172 antibodies. Note similarities of the staining patterns, irrespective the CIDAMP AA-sequence or biotin-labeling and irrespective whether a Strep - Tactin ®- or antibody-detectable CIDAMP was used to probe and detect the target protein on the membrane. Note the presence of 70 kDa bands upon HRNR-Far-Western blot analyses in most of the investigated HPLC fractions with highest intensity for rHRNR 1075–1172 binding ( f ). The most intensive band, corresponding to a 37 kDa protein in fraction number C13, was identified as E . coli ribosomal protein L2.

    Article Snippet: We either used a Jupiter® 300 Å, 250 × 12.6 mm, C18-RP-HPLC column (Phenomenex) or an Aeris® widepore XB C18, 250 × 12.6 mm, RP-HPLC column (Phenomenex) and either a gradient of acetonitrile (ACN) in aqueous 0.1% TFA, or a gradient of 2-propanol (Prp) in aqueous 0.1% TFA – as indicated for separation of ribosomal proteins.

    Techniques: Derivative Assay, Protein Binding, High Performance Liquid Chromatography, Polyacrylamide Gel Electrophoresis, Staining, Far Western Blot, Sequencing, Labeling, Binding Assay

    Isolation of vurtoxin and Vur-S49 by reverse-phase HPLC. Separation was done on a Discovery BIO Wide Pore C18 column (10×250 mm, Supelco) in a gradient of 25–40% (v/v) acetonitrile in 60 min in the presence of 0.1% (v/v) trifluoroacetic acid, at a flow rate of 2.0 ml/min. Fraction containing Vur-S49 (4) and vurtoxin (11) are indicated by horizontal bars.

    Journal: PLoS ONE

    Article Title: Inhibition of Nicotinic Acetylcholine Receptors, a Novel Facet in the Pleiotropic Activities of Snake Venom Phospholipases A2

    doi: 10.1371/journal.pone.0115428

    Figure Lengend Snippet: Isolation of vurtoxin and Vur-S49 by reverse-phase HPLC. Separation was done on a Discovery BIO Wide Pore C18 column (10×250 mm, Supelco) in a gradient of 25–40% (v/v) acetonitrile in 60 min in the presence of 0.1% (v/v) trifluoroacetic acid, at a flow rate of 2.0 ml/min. Fraction containing Vur-S49 (4) and vurtoxin (11) are indicated by horizontal bars.

    Article Snippet: Fraction 3+4 ( in ) was further purified by reversed phase HPLC on a Discovery BIO Wide Pore C18 column (10×250 mm, Supelco) in a gradient of 25–40% (v/v) acetonitrile in 60 min in the presence of 0.1% (v/v) trifluoroacetic acid, at a flow rate of 2.0 ml/min ( ).

    Techniques: Isolation, High Performance Liquid Chromatography, Flow Cytometry