vhs  (New England Biolabs)


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  • 99
    Name:
    BamHI HF
    Description:
    BamHI HF 50 000 units
    Catalog Number:
    r3136l
    Price:
    249
    Size:
    50 000 units
    Category:
    Restriction Enzymes
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    Structured Review

    New England Biolabs vhs
    BamHI HF
    BamHI HF 50 000 units
    https://www.bioz.com/result/vhs/product/New England Biolabs
    Average 99 stars, based on 4 article reviews
    Price from $9.99 to $1999.99
    vhs - by Bioz Stars, 2020-07
    99/100 stars

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    Related Articles

    RNA Expression:

    Article Title: A genome-scale CRISPR interference guide library enables comprehensive phenotypic profiling in yeast
    Article Snippet: .. Guide library construction The guide RNA expression vector pNTI661 was digested by taking 3.0 μ g plasmid in a 75. μ l reaction with 1x final concentration CutSmart buffer (NEB B7204S) with 60 U BamHI-HF (NEB R3136L) and 60 U HindIII-HF (NEB R3104S), incubated for 1 hour at 37 °C, and then purified with a DNA Clean & Concentrator (Zymo D4013). .. The guide RNA oligonucleotide library was amplified using Q5 polymerase (NEB M0491S) according to the manufacturers instructions, using 100 pg guide oligonucleotide pool (CustomArray, Inc.) as a template and oligonucleotides NM636 and NM637 ( ) for amplification, with 15 cycles of amplification using 10 s denaturation, 15 s annealing at 58 °C, and 15 s extension.

    Agarose Gel Electrophoresis:

    Article Title: Efficient Dual-Negative Selection for Bacterial Genome Editing
    Article Snippet: .. Vectors were digested using EcoRI-HF and BamHI-HF (New England BioLabs) for 1h at 37°C, or PCR-amplified, and purified on agarose gel. ..

    Spectrophotometry:

    Article Title: The SMC5/6 Complex Interacts with the Papillomavirus E2 Protein and Influences Maintenance of Viral Episomal DNA
    Article Snippet: .. DNA was measured by use of the NanoDrop spectrophotometer (Thermo Fisher Scientific) and digested overnight at 37°C with BamHI-HF and XbaI (New England BioLabs). ..

    Purification:

    Article Title: A genome-scale CRISPR interference guide library enables comprehensive phenotypic profiling in yeast
    Article Snippet: .. Guide library construction The guide RNA expression vector pNTI661 was digested by taking 3.0 μ g plasmid in a 75. μ l reaction with 1x final concentration CutSmart buffer (NEB B7204S) with 60 U BamHI-HF (NEB R3136L) and 60 U HindIII-HF (NEB R3104S), incubated for 1 hour at 37 °C, and then purified with a DNA Clean & Concentrator (Zymo D4013). .. The guide RNA oligonucleotide library was amplified using Q5 polymerase (NEB M0491S) according to the manufacturers instructions, using 100 pg guide oligonucleotide pool (CustomArray, Inc.) as a template and oligonucleotides NM636 and NM637 ( ) for amplification, with 15 cycles of amplification using 10 s denaturation, 15 s annealing at 58 °C, and 15 s extension.

    Article Title: Human herpesvirus–encoded kinase induces B cell lymphomas in vivo
    Article Snippet: .. For the probe, plasmid vPK (Ub.vPK.hgH) was double digested with BamHI-HF and HindIII-HF and gel purified. .. One hundred nanograms of this DNA was used as a template to make a radiolabeled deoxycytidine triphosphate [α-32 P]-dCTP (PerkinElmer, BLU513Z250UC) probe per the manufacturer’s instructions (Thermo Scientific DecaLabel DNA Labeling Kit, K0622).

    Article Title: Efficient Dual-Negative Selection for Bacterial Genome Editing
    Article Snippet: .. Vectors were digested using EcoRI-HF and BamHI-HF (New England BioLabs) for 1h at 37°C, or PCR-amplified, and purified on agarose gel. ..

    Concentration Assay:

    Article Title: A genome-scale CRISPR interference guide library enables comprehensive phenotypic profiling in yeast
    Article Snippet: .. Guide library construction The guide RNA expression vector pNTI661 was digested by taking 3.0 μ g plasmid in a 75. μ l reaction with 1x final concentration CutSmart buffer (NEB B7204S) with 60 U BamHI-HF (NEB R3136L) and 60 U HindIII-HF (NEB R3104S), incubated for 1 hour at 37 °C, and then purified with a DNA Clean & Concentrator (Zymo D4013). .. The guide RNA oligonucleotide library was amplified using Q5 polymerase (NEB M0491S) according to the manufacturers instructions, using 100 pg guide oligonucleotide pool (CustomArray, Inc.) as a template and oligonucleotides NM636 and NM637 ( ) for amplification, with 15 cycles of amplification using 10 s denaturation, 15 s annealing at 58 °C, and 15 s extension.

    Incubation:

    Article Title: A genome-scale CRISPR interference guide library enables comprehensive phenotypic profiling in yeast
    Article Snippet: .. Guide library construction The guide RNA expression vector pNTI661 was digested by taking 3.0 μ g plasmid in a 75. μ l reaction with 1x final concentration CutSmart buffer (NEB B7204S) with 60 U BamHI-HF (NEB R3136L) and 60 U HindIII-HF (NEB R3104S), incubated for 1 hour at 37 °C, and then purified with a DNA Clean & Concentrator (Zymo D4013). .. The guide RNA oligonucleotide library was amplified using Q5 polymerase (NEB M0491S) according to the manufacturers instructions, using 100 pg guide oligonucleotide pool (CustomArray, Inc.) as a template and oligonucleotides NM636 and NM637 ( ) for amplification, with 15 cycles of amplification using 10 s denaturation, 15 s annealing at 58 °C, and 15 s extension.

    Article Title: A bacterial DNA repair pathway specific to a natural antibiotic
    Article Snippet: .. To generate linear or nicked substrate, pUC19 was first incubated with BamHI-HF (NEB) or Nt.BSPQ1 (NEB), respectively, for 30 minutes at 37°C. .. To test metal dependency of MrfB, the linearized pUC19 was purified using a silica spin-column.

    Polymerase Chain Reaction:

    Article Title: Efficient Dual-Negative Selection for Bacterial Genome Editing
    Article Snippet: .. Vectors were digested using EcoRI-HF and BamHI-HF (New England BioLabs) for 1h at 37°C, or PCR-amplified, and purified on agarose gel. ..

    Plasmid Preparation:

    Article Title: A genome-scale CRISPR interference guide library enables comprehensive phenotypic profiling in yeast
    Article Snippet: .. Guide library construction The guide RNA expression vector pNTI661 was digested by taking 3.0 μ g plasmid in a 75. μ l reaction with 1x final concentration CutSmart buffer (NEB B7204S) with 60 U BamHI-HF (NEB R3136L) and 60 U HindIII-HF (NEB R3104S), incubated for 1 hour at 37 °C, and then purified with a DNA Clean & Concentrator (Zymo D4013). .. The guide RNA oligonucleotide library was amplified using Q5 polymerase (NEB M0491S) according to the manufacturers instructions, using 100 pg guide oligonucleotide pool (CustomArray, Inc.) as a template and oligonucleotides NM636 and NM637 ( ) for amplification, with 15 cycles of amplification using 10 s denaturation, 15 s annealing at 58 °C, and 15 s extension.

    Article Title: Human herpesvirus–encoded kinase induces B cell lymphomas in vivo
    Article Snippet: .. For the probe, plasmid vPK (Ub.vPK.hgH) was double digested with BamHI-HF and HindIII-HF and gel purified. .. One hundred nanograms of this DNA was used as a template to make a radiolabeled deoxycytidine triphosphate [α-32 P]-dCTP (PerkinElmer, BLU513Z250UC) probe per the manufacturer’s instructions (Thermo Scientific DecaLabel DNA Labeling Kit, K0622).

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  • 88
    New England Biolabs vhs fragment
    An in vitro assay of RNase activity mediated by PrV <t>vhs</t> using various substrates. ( A ) Different types of RNA, including single stranded RNA without cap (ssRNA), with cap and polyA tail (mRNA) and cellular total rRNA, were incubated with assay buffer (mock), recombinant vhs and the appropriate tag proteins, either NUS or Thioredoxin (THX) (negative control), for various times (as indicated above the gel). The degradation pattern was resolved by agarose electrophoresis. In addition to RNA, various <t>DNA</t> substrates ( B ) and a DNA-RNA hybrid ( C ) were also tested.
    Vhs Fragment, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 88/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/vhs fragment/product/New England Biolabs
    Average 88 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    vhs fragment - by Bioz Stars, 2020-07
    88/100 stars
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    85
    New England Biolabs a5 1 vh hs
    V H Hs bind TcdA with high affinity. SPR sensorgrams of TcdA-specific V H Hs A4.2, <t>A5.1,</t> A19.2, A20.1, A24.1, and A26.8 binding to immobilized TcdA ( A–F ) and TcdB-RBD-f1 binding to immobilized TcdB-specific V H Hs B5.2, B13.6, and B15.5 ( G–I
    A5 1 Vh Hs, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/a5 1 vh hs/product/New England Biolabs
    Average 85 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    a5 1 vh hs - by Bioz Stars, 2020-07
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    85
    New England Biolabs hc 1 vh mutant
    Binding and neutralization of <t>HC-1</t> against HCVpp 1a and 1b isolates. A , dose-dependent binding to two panels of HCV 1a and 1b isolates by ELISA. Error bars indicate range in replicate studies. B , correlating neutralization and binding affinity against
    Hc 1 Vh Mutant, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hc 1 vh mutant/product/New England Biolabs
    Average 85 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    hc 1 vh mutant - by Bioz Stars, 2020-07
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    85
    New England Biolabs hc 1 vh gene fragments
    Binding and neutralization of <t>HC-1</t> against HCVpp 1a and 1b isolates. A , dose-dependent binding to two panels of HCV 1a and 1b isolates by ELISA. Error bars indicate range in replicate studies. B , correlating neutralization and binding affinity against
    Hc 1 Vh Gene Fragments, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hc 1 vh gene fragments/product/New England Biolabs
    Average 85 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    hc 1 vh gene fragments - by Bioz Stars, 2020-07
    85/100 stars
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    Image Search Results


    An in vitro assay of RNase activity mediated by PrV vhs using various substrates. ( A ) Different types of RNA, including single stranded RNA without cap (ssRNA), with cap and polyA tail (mRNA) and cellular total rRNA, were incubated with assay buffer (mock), recombinant vhs and the appropriate tag proteins, either NUS or Thioredoxin (THX) (negative control), for various times (as indicated above the gel). The degradation pattern was resolved by agarose electrophoresis. In addition to RNA, various DNA substrates ( B ) and a DNA-RNA hybrid ( C ) were also tested.

    Journal: Veterinary Research

    Article Title: Roles of nucleic acid substrates and cofactors in the vhs protein activity of pseudorabies virus

    doi: 10.1186/s13567-015-0284-y

    Figure Lengend Snippet: An in vitro assay of RNase activity mediated by PrV vhs using various substrates. ( A ) Different types of RNA, including single stranded RNA without cap (ssRNA), with cap and polyA tail (mRNA) and cellular total rRNA, were incubated with assay buffer (mock), recombinant vhs and the appropriate tag proteins, either NUS or Thioredoxin (THX) (negative control), for various times (as indicated above the gel). The degradation pattern was resolved by agarose electrophoresis. In addition to RNA, various DNA substrates ( B ) and a DNA-RNA hybrid ( C ) were also tested.

    Article Snippet: ∆Box I of vhs fragment was amplified by PCR using Pfu DNA polymerase (NEB, NEW ENGLAND BioLabs® INC.) and primer sets with built-in sequences of Bam HI/Xho I as shown in underlines (Box 1–F: AA GGATCC GCCATGGAGAAGCACTAC and PrV-R: AA CTCGAG TTATTTTCTCCTGTGGG).

    Techniques: In Vitro, Activity Assay, Incubation, Recombinant, Negative Control, Electrophoresis

    Sequence alignment of the vhs coding regions. The vhs sequences of HSV-1, HSV-2, and PRV were analysed by DNA Star MegaAlign software. Identical residues are denoted as “.” and deleted amino acids are indicate as “-”. Four conserved region, designated as Boxes I–IV (reported in Berthomme et al. [ 15 ]) are marked. In addition, several deduced residues, numbered according to the PrV vhs gene, that have been reported as being responsible for ribonuclease activities are indicated with arrowheads.

    Journal: Veterinary Research

    Article Title: Roles of nucleic acid substrates and cofactors in the vhs protein activity of pseudorabies virus

    doi: 10.1186/s13567-015-0284-y

    Figure Lengend Snippet: Sequence alignment of the vhs coding regions. The vhs sequences of HSV-1, HSV-2, and PRV were analysed by DNA Star MegaAlign software. Identical residues are denoted as “.” and deleted amino acids are indicate as “-”. Four conserved region, designated as Boxes I–IV (reported in Berthomme et al. [ 15 ]) are marked. In addition, several deduced residues, numbered according to the PrV vhs gene, that have been reported as being responsible for ribonuclease activities are indicated with arrowheads.

    Article Snippet: ∆Box I of vhs fragment was amplified by PCR using Pfu DNA polymerase (NEB, NEW ENGLAND BioLabs® INC.) and primer sets with built-in sequences of Bam HI/Xho I as shown in underlines (Box 1–F: AA GGATCC GCCATGGAGAAGCACTAC and PrV-R: AA CTCGAG TTATTTTCTCCTGTGGG).

    Techniques: Sequencing, Software

    V H Hs bind TcdA with high affinity. SPR sensorgrams of TcdA-specific V H Hs A4.2, A5.1, A19.2, A20.1, A24.1, and A26.8 binding to immobilized TcdA ( A–F ) and TcdB-RBD-f1 binding to immobilized TcdB-specific V H Hs B5.2, B13.6, and B15.5 ( G–I

    Journal: The Journal of Biological Chemistry

    Article Title: Neutralization of Clostridium difficile Toxin A with Single-domain Antibodies Targeting the Cell Receptor Binding Domain *

    doi: 10.1074/jbc.M110.198754

    Figure Lengend Snippet: V H Hs bind TcdA with high affinity. SPR sensorgrams of TcdA-specific V H Hs A4.2, A5.1, A19.2, A20.1, A24.1, and A26.8 binding to immobilized TcdA ( A–F ) and TcdB-RBD-f1 binding to immobilized TcdB-specific V H Hs B5.2, B13.6, and B15.5 ( G–I

    Article Snippet: Furthermore, we were unsuccessful at identifying peptide binders, i.e. linear epitopes, to A4.2 and A5.1 VH Hs through panning of the 12-mer peptide library (New England Biolabs) (data not shown).

    Techniques: SPR Assay, Binding Assay

    Antitoxin V H Hs recognize conformational (A4.2, A5.1, A20.1, and A26.8) and linear (A19.2) epitopes. ELISA demonstrates that purified anti-TcdA V H Hs ( A ) and anti-TcdB V H Hs ( B ) recognize both native toxins and recombinant RBD fragments. Wells were coated

    Journal: The Journal of Biological Chemistry

    Article Title: Neutralization of Clostridium difficile Toxin A with Single-domain Antibodies Targeting the Cell Receptor Binding Domain *

    doi: 10.1074/jbc.M110.198754

    Figure Lengend Snippet: Antitoxin V H Hs recognize conformational (A4.2, A5.1, A20.1, and A26.8) and linear (A19.2) epitopes. ELISA demonstrates that purified anti-TcdA V H Hs ( A ) and anti-TcdB V H Hs ( B ) recognize both native toxins and recombinant RBD fragments. Wells were coated

    Article Snippet: Furthermore, we were unsuccessful at identifying peptide binders, i.e. linear epitopes, to A4.2 and A5.1 VH Hs through panning of the 12-mer peptide library (New England Biolabs) (data not shown).

    Techniques: Enzyme-linked Immunosorbent Assay, Purification, Recombinant

    Mixtures of A4.2, A5.1, and A26.8 V H Hs show binding at overlapping TcdA epitopes. The three V H Hs suspected of sharing an overlapping epitope () were further analyzed by SPR by injecting the V H Hs alone (A4.2, A5.1, or A26.8) or as a triplet mixture

    Journal: The Journal of Biological Chemistry

    Article Title: Neutralization of Clostridium difficile Toxin A with Single-domain Antibodies Targeting the Cell Receptor Binding Domain *

    doi: 10.1074/jbc.M110.198754

    Figure Lengend Snippet: Mixtures of A4.2, A5.1, and A26.8 V H Hs show binding at overlapping TcdA epitopes. The three V H Hs suspected of sharing an overlapping epitope () were further analyzed by SPR by injecting the V H Hs alone (A4.2, A5.1, or A26.8) or as a triplet mixture

    Article Snippet: Furthermore, we were unsuccessful at identifying peptide binders, i.e. linear epitopes, to A4.2 and A5.1 VH Hs through panning of the 12-mer peptide library (New England Biolabs) (data not shown).

    Techniques: Binding Assay, SPR Assay

    Anti-TcdA V H Hs recognize overlapping and nonoverlapping epitopes. SPR co-injection experiments were used to determine whether pairs of V H Hs could bind TcdA simultaneously. The sensorgrams of all of the possible paired combinations of A4.2, A5.1, A20.1,

    Journal: The Journal of Biological Chemistry

    Article Title: Neutralization of Clostridium difficile Toxin A with Single-domain Antibodies Targeting the Cell Receptor Binding Domain *

    doi: 10.1074/jbc.M110.198754

    Figure Lengend Snippet: Anti-TcdA V H Hs recognize overlapping and nonoverlapping epitopes. SPR co-injection experiments were used to determine whether pairs of V H Hs could bind TcdA simultaneously. The sensorgrams of all of the possible paired combinations of A4.2, A5.1, A20.1,

    Article Snippet: Furthermore, we were unsuccessful at identifying peptide binders, i.e. linear epitopes, to A4.2 and A5.1 VH Hs through panning of the 12-mer peptide library (New England Biolabs) (data not shown).

    Techniques: SPR Assay, Injection

    Binding and neutralization of HC-1 against HCVpp 1a and 1b isolates. A , dose-dependent binding to two panels of HCV 1a and 1b isolates by ELISA. Error bars indicate range in replicate studies. B , correlating neutralization and binding affinity against

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: Binding and neutralization of HC-1 against HCVpp 1a and 1b isolates. A , dose-dependent binding to two panels of HCV 1a and 1b isolates by ELISA. Error bars indicate range in replicate studies. B , correlating neutralization and binding affinity against

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: Binding Assay, Neutralization, Enzyme-linked Immunosorbent Assay

    Measurement of WT HC-1 and affinity-matured mutants IgG affinity to HCV 1b E2 on SPR. A , overlay plot of association and dissociation curves obtained for HCV1b 34E2 antigen at 4.49 μ m concentration against immobilized HC-1 IgG. The HC-1 affinity-matured

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: Measurement of WT HC-1 and affinity-matured mutants IgG affinity to HCV 1b E2 on SPR. A , overlay plot of association and dissociation curves obtained for HCV1b 34E2 antigen at 4.49 μ m concentration against immobilized HC-1 IgG. The HC-1 affinity-matured

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: SPR Assay, Concentration Assay

    Neutralization improvement of HC-1 affinity-matured mutants against HCVpp 1a and 1b isolates and 2a HCVcc. A , dose-dependent binding by WT HC-1 and affinity-matured IgG to HCV 1A20.8 by ELISA. B , all of the HC-1 affinity-matured clones except for Vk1.9

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: Neutralization improvement of HC-1 affinity-matured mutants against HCVpp 1a and 1b isolates and 2a HCVcc. A , dose-dependent binding by WT HC-1 and affinity-matured IgG to HCV 1A20.8 by ELISA. B , all of the HC-1 affinity-matured clones except for Vk1.9

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: Neutralization, Binding Assay, Enzyme-linked Immunosorbent Assay, Clone Assay

    HC-1 random mutagenesis yeast library construction and affinity maturation. A , two separate yeast surface display libraries: the VH-Mut library contains the random mutated HC-1 VH gene and unchanged Vk, and the VH-Mut library with random mutated HC-1

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: HC-1 random mutagenesis yeast library construction and affinity maturation. A , two separate yeast surface display libraries: the VH-Mut library contains the random mutated HC-1 VH gene and unchanged Vk, and the VH-Mut library with random mutated HC-1

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: Mutagenesis

    Propagating 2a HCVcc in the presence of WT HC-1 and its affinity-matured mutants. A and B , dual antibody immunofluorescence staining of Huh7.5 cells infected with 2a HCVcc after multiple rounds of neutralization by the respective antibodies. R04, a human

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: Propagating 2a HCVcc in the presence of WT HC-1 and its affinity-matured mutants. A and B , dual antibody immunofluorescence staining of Huh7.5 cells infected with 2a HCVcc after multiple rounds of neutralization by the respective antibodies. R04, a human

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: Immunofluorescence, Staining, Infection, Neutralization

    Reactivity of HC-1 affinity-matured mutants against different HCV genotypes. A , dose-dependent binding by WT HC-1 and affinity-matured IgG to HCV UKN4.11.1 E1E2 lysate by ELISA. B , dose-dependent neutralization of HCVpp UKN4.11.1 by WT HC-1 and its affinity-matured

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: Reactivity of HC-1 affinity-matured mutants against different HCV genotypes. A , dose-dependent binding by WT HC-1 and affinity-matured IgG to HCV UKN4.11.1 E1E2 lysate by ELISA. B , dose-dependent neutralization of HCVpp UKN4.11.1 by WT HC-1 and its affinity-matured

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: Binding Assay, Enzyme-linked Immunosorbent Assay, Neutralization

    Sequence Characterization Analysis of WT HC-1 and Affinity-matured Mutants

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: Sequence Characterization Analysis of WT HC-1 and Affinity-matured Mutants

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: Sequencing

    Binding and neutralization of HC-1 against HCVpp 1a and 1b isolates. A , dose-dependent binding to two panels of HCV 1a and 1b isolates by ELISA. Error bars indicate range in replicate studies. B , correlating neutralization and binding affinity against

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: Binding and neutralization of HC-1 against HCVpp 1a and 1b isolates. A , dose-dependent binding to two panels of HCV 1a and 1b isolates by ELISA. Error bars indicate range in replicate studies. B , correlating neutralization and binding affinity against

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: Binding Assay, Neutralization, Enzyme-linked Immunosorbent Assay

    Measurement of WT HC-1 and affinity-matured mutants IgG affinity to HCV 1b E2 on SPR. A , overlay plot of association and dissociation curves obtained for HCV1b 34E2 antigen at 4.49 μ m concentration against immobilized HC-1 IgG. The HC-1 affinity-matured

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: Measurement of WT HC-1 and affinity-matured mutants IgG affinity to HCV 1b E2 on SPR. A , overlay plot of association and dissociation curves obtained for HCV1b 34E2 antigen at 4.49 μ m concentration against immobilized HC-1 IgG. The HC-1 affinity-matured

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: SPR Assay, Concentration Assay

    Neutralization improvement of HC-1 affinity-matured mutants against HCVpp 1a and 1b isolates and 2a HCVcc. A , dose-dependent binding by WT HC-1 and affinity-matured IgG to HCV 1A20.8 by ELISA. B , all of the HC-1 affinity-matured clones except for Vk1.9

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: Neutralization improvement of HC-1 affinity-matured mutants against HCVpp 1a and 1b isolates and 2a HCVcc. A , dose-dependent binding by WT HC-1 and affinity-matured IgG to HCV 1A20.8 by ELISA. B , all of the HC-1 affinity-matured clones except for Vk1.9

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: Neutralization, Binding Assay, Enzyme-linked Immunosorbent Assay, Clone Assay

    HC-1 random mutagenesis yeast library construction and affinity maturation. A , two separate yeast surface display libraries: the VH-Mut library contains the random mutated HC-1 VH gene and unchanged Vk, and the VH-Mut library with random mutated HC-1

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: HC-1 random mutagenesis yeast library construction and affinity maturation. A , two separate yeast surface display libraries: the VH-Mut library contains the random mutated HC-1 VH gene and unchanged Vk, and the VH-Mut library with random mutated HC-1

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: Mutagenesis

    Propagating 2a HCVcc in the presence of WT HC-1 and its affinity-matured mutants. A and B , dual antibody immunofluorescence staining of Huh7.5 cells infected with 2a HCVcc after multiple rounds of neutralization by the respective antibodies. R04, a human

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: Propagating 2a HCVcc in the presence of WT HC-1 and its affinity-matured mutants. A and B , dual antibody immunofluorescence staining of Huh7.5 cells infected with 2a HCVcc after multiple rounds of neutralization by the respective antibodies. R04, a human

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: Immunofluorescence, Staining, Infection, Neutralization

    Reactivity of HC-1 affinity-matured mutants against different HCV genotypes. A , dose-dependent binding by WT HC-1 and affinity-matured IgG to HCV UKN4.11.1 E1E2 lysate by ELISA. B , dose-dependent neutralization of HCVpp UKN4.11.1 by WT HC-1 and its affinity-matured

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: Reactivity of HC-1 affinity-matured mutants against different HCV genotypes. A , dose-dependent binding by WT HC-1 and affinity-matured IgG to HCV UKN4.11.1 E1E2 lysate by ELISA. B , dose-dependent neutralization of HCVpp UKN4.11.1 by WT HC-1 and its affinity-matured

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: Binding Assay, Enzyme-linked Immunosorbent Assay, Neutralization

    Sequence Characterization Analysis of WT HC-1 and Affinity-matured Mutants

    Journal: The Journal of Biological Chemistry

    Article Title: Affinity Maturation to Improve Human Monoclonal Antibody Neutralization Potency and Breadth against Hepatitis C Virus *

    doi: 10.1074/jbc.M111.290783

    Figure Lengend Snippet: Sequence Characterization Analysis of WT HC-1 and Affinity-matured Mutants

    Article Snippet: For the HC-1 VH-mutant (Mut) library, HC-1 VH gene fragments were PCR-reamplified with the primers Gap5 and HuJHR (Gap 5, 5′-TTA AGC TTC TGC AGG CTA GTG-3′; and HuJHR, 5′-ACC TCC GGA GCC ACC TCC GCC TGA ACC GCC TCC ACC TGT CGA CCC TGA-3′) with PhusionTM high fidelity DNA polymerase (New England Biolabs).

    Techniques: Sequencing