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New England Biolabs vhs
Vhs, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 91/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 91 stars, based on 4 article reviews
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Isolation:

Article Title: One-Step Recovery of scFv Clones from High-Throughput Sequencing-Based Screening of Phage Display Libraries Challenged to Cells Expressing Native Claudin-1
Article Snippet: VH Extraction and Purification The double strand DNA plasmids containing the scFvs were isolated from each cycle of selection from a culture of superinfected E. coli TG1 cells using GenElute HP Plasmid Maxiprep Kit (Sigma-Aldrich). .. The VHs were excised by double digestion with restriction enzymes Nco I and Xho I (New England Biolabs) and then purified from a 1.2% agarose gel ( ).

Selection:

Article Title: One-Step Recovery of scFv Clones from High-Throughput Sequencing-Based Screening of Phage Display Libraries Challenged to Cells Expressing Native Claudin-1
Article Snippet: VH Extraction and Purification The double strand DNA plasmids containing the scFvs were isolated from each cycle of selection from a culture of superinfected E. coli TG1 cells using GenElute HP Plasmid Maxiprep Kit (Sigma-Aldrich). .. The VHs were excised by double digestion with restriction enzymes Nco I and Xho I (New England Biolabs) and then purified from a 1.2% agarose gel ( ).

Agarose Gel Electrophoresis:

Article Title: One-Step Recovery of scFv Clones from High-Throughput Sequencing-Based Screening of Phage Display Libraries Challenged to Cells Expressing Native Claudin-1
Article Snippet: .. The VHs were excised by double digestion with restriction enzymes Nco I and Xho I (New England Biolabs) and then purified from a 1.2% agarose gel ( ). .. High-Throughput Sequencing Library preparations of the fragments, sequencing reactions, and preliminary analysis of the data were performed at the Center for Translational Genomics and Bioinformatics, Hospital San Raffaele, Milano, Italy.

Plasmid Preparation:

Article Title: One-Step Recovery of scFv Clones from High-Throughput Sequencing-Based Screening of Phage Display Libraries Challenged to Cells Expressing Native Claudin-1
Article Snippet: VH Extraction and Purification The double strand DNA plasmids containing the scFvs were isolated from each cycle of selection from a culture of superinfected E. coli TG1 cells using GenElute HP Plasmid Maxiprep Kit (Sigma-Aldrich). .. The VHs were excised by double digestion with restriction enzymes Nco I and Xho I (New England Biolabs) and then purified from a 1.2% agarose gel ( ).

Purification:

Article Title: One-Step Recovery of scFv Clones from High-Throughput Sequencing-Based Screening of Phage Display Libraries Challenged to Cells Expressing Native Claudin-1
Article Snippet: .. The VHs were excised by double digestion with restriction enzymes Nco I and Xho I (New England Biolabs) and then purified from a 1.2% agarose gel ( ). .. High-Throughput Sequencing Library preparations of the fragments, sequencing reactions, and preliminary analysis of the data were performed at the Center for Translational Genomics and Bioinformatics, Hospital San Raffaele, Milano, Italy.

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    New England Biolabs vhs
    Vhs, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 90/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/vhs/product/New England Biolabs
    Average 90 stars, based on 4 article reviews
    Price from $9.99 to $1999.99
    vhs - by Bioz Stars, 2020-01
    90/100 stars
      Buy from Supplier

    78
    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: 78/100, 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 78 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    hc 1 vh gene fragments - by Bioz Stars, 2020-01
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    80
    New England Biolabs ganglidiomab vh
    Establishment of a cell line stably producing ganglidiximab. CHO cells were stably co-transfected with two generated expression plasmids (p3-ganglidiximab-HC/ p3-ganglidiximab-LC) and a cell clone “VH-VL-1” stably producing high levels of ganglidiximab was selected for further Ab production. Permanent ganglidiximab expression was confirmed after 15 passages and two freeze-thaw cycles by RT-PCR (A) and standard ELISA (B). ( A ) To amplify coding sequences of ganglidiximab VH (440 bp) and -VL (420 bp), RNA of “VH-VL-1” was used for RT-PCR followed by agarose gel electrophoresis. RNA of <t>ganglidiomab-producing</t> hybridoma cells served as a positive control (+) and RNA of non-transfected CHO cells as a negative control. One representative image is shown. NTC—no-template-control. M—Marker (100-bp). ( B ) Ganglidiximab production by “VH-VL-1” was analyzed in supernatants collected during 15 passages and after two freeze-thaw cycles (P2, 5, 10 and 15). Supernatants of non-transfected CHO cells or cells incubated with control plasmids (mock) or transfection reagent only were utilized as negative controls (white columns). Human/mouse chimeric mAb rituximab and cell culture medium were included as additional negative controls (white-striped columns). Data are shown as mean values ± SEM of three independent experiments performed at least in triplicates. One-way ANOVA on ranks followed by appropriate post hoc comparison test; * P 0.05 vs . negative controls.
    Ganglidiomab Vh, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 80/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 80 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
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    Image Search Results


    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

    Establishment of a cell line stably producing ganglidiximab. CHO cells were stably co-transfected with two generated expression plasmids (p3-ganglidiximab-HC/ p3-ganglidiximab-LC) and a cell clone “VH-VL-1” stably producing high levels of ganglidiximab was selected for further Ab production. Permanent ganglidiximab expression was confirmed after 15 passages and two freeze-thaw cycles by RT-PCR (A) and standard ELISA (B). ( A ) To amplify coding sequences of ganglidiximab VH (440 bp) and -VL (420 bp), RNA of “VH-VL-1” was used for RT-PCR followed by agarose gel electrophoresis. RNA of ganglidiomab-producing hybridoma cells served as a positive control (+) and RNA of non-transfected CHO cells as a negative control. One representative image is shown. NTC—no-template-control. M—Marker (100-bp). ( B ) Ganglidiximab production by “VH-VL-1” was analyzed in supernatants collected during 15 passages and after two freeze-thaw cycles (P2, 5, 10 and 15). Supernatants of non-transfected CHO cells or cells incubated with control plasmids (mock) or transfection reagent only were utilized as negative controls (white columns). Human/mouse chimeric mAb rituximab and cell culture medium were included as additional negative controls (white-striped columns). Data are shown as mean values ± SEM of three independent experiments performed at least in triplicates. One-way ANOVA on ranks followed by appropriate post hoc comparison test; * P 0.05 vs . negative controls.

    Journal: PLoS ONE

    Article Title: Generation and Characterization of a Human/Mouse Chimeric GD2-Mimicking Anti-Idiotype Antibody Ganglidiximab for Active Immunotherapy against Neuroblastoma

    doi: 10.1371/journal.pone.0150479

    Figure Lengend Snippet: Establishment of a cell line stably producing ganglidiximab. CHO cells were stably co-transfected with two generated expression plasmids (p3-ganglidiximab-HC/ p3-ganglidiximab-LC) and a cell clone “VH-VL-1” stably producing high levels of ganglidiximab was selected for further Ab production. Permanent ganglidiximab expression was confirmed after 15 passages and two freeze-thaw cycles by RT-PCR (A) and standard ELISA (B). ( A ) To amplify coding sequences of ganglidiximab VH (440 bp) and -VL (420 bp), RNA of “VH-VL-1” was used for RT-PCR followed by agarose gel electrophoresis. RNA of ganglidiomab-producing hybridoma cells served as a positive control (+) and RNA of non-transfected CHO cells as a negative control. One representative image is shown. NTC—no-template-control. M—Marker (100-bp). ( B ) Ganglidiximab production by “VH-VL-1” was analyzed in supernatants collected during 15 passages and after two freeze-thaw cycles (P2, 5, 10 and 15). Supernatants of non-transfected CHO cells or cells incubated with control plasmids (mock) or transfection reagent only were utilized as negative controls (white columns). Human/mouse chimeric mAb rituximab and cell culture medium were included as additional negative controls (white-striped columns). Data are shown as mean values ± SEM of three independent experiments performed at least in triplicates. One-way ANOVA on ranks followed by appropriate post hoc comparison test; * P 0.05 vs . negative controls.

    Article Snippet: For ganglidiomab VH, plasmid DNA (4 μg) was incubated with Not I-HF (1 unit; New England Biolabs GmbH, Frankfurt/Main, Germany) and Xho I-HF (1 unit; New England Biolabs GmbH, Frankfurt/Main, Germany) in CutSmart® Buffer (1×) for 1 h at +37°C followed by enzyme heat inactivation (20 min, +65°C).

    Techniques: Stable Transfection, Transfection, Generated, Expressing, Reverse Transcription Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Agarose Gel Electrophoresis, Positive Control, Negative Control, Marker, Incubation, Cell Culture

    Binding of ganglidiximab to NK-92tr and ganglidiximab-dependent inhibition of NK-92tr-mediated GD 2 -specific cytotoxicity against NB. ( A ) Binding of ganglidiximab to NK-92tr expressing a GD 2 -specific chimeric antigen receptor was analyzed by flow cytometry. Cells were stained with chimeric ganglidiximab (black solid line), murine anti-Id ganglidiomab (positive control; grey solid line), chimeric rituximab (isotype control; black dashed line) or murine IgG1 (isotype control; grey dashed line) followed by incubation with biotinylated ch14.18/CHO and PE-labeled streptavidin. Staining of the parental NK-92 cell line lacking GD 2 -specific CAR expression was further included as negative control. Results from one representative experiment are shown. ( B ) Inhibition of GD 2 -specific NK-92tr-mediated NB cell lysis (w/o Ab; white-striped column) was analyzed after pre-incubation with excess of ganglidiximab (black column) using a calcein-AM-based cytotoxicity assay. Murine anti-Id ganglidiomab served as a positive control (grey column). Rituximab (white column) and murine IgG1 (white column) were utilized as negative controls. Results are expressed as percentage of lysis inhibition (mean values ± SEM) of two independent experiments performed using six replicates. One-way ANOVA on ranks followed by appropriate post hoc comparison test; * P 0.05 vs . w/o Ab.

    Journal: PLoS ONE

    Article Title: Generation and Characterization of a Human/Mouse Chimeric GD2-Mimicking Anti-Idiotype Antibody Ganglidiximab for Active Immunotherapy against Neuroblastoma

    doi: 10.1371/journal.pone.0150479

    Figure Lengend Snippet: Binding of ganglidiximab to NK-92tr and ganglidiximab-dependent inhibition of NK-92tr-mediated GD 2 -specific cytotoxicity against NB. ( A ) Binding of ganglidiximab to NK-92tr expressing a GD 2 -specific chimeric antigen receptor was analyzed by flow cytometry. Cells were stained with chimeric ganglidiximab (black solid line), murine anti-Id ganglidiomab (positive control; grey solid line), chimeric rituximab (isotype control; black dashed line) or murine IgG1 (isotype control; grey dashed line) followed by incubation with biotinylated ch14.18/CHO and PE-labeled streptavidin. Staining of the parental NK-92 cell line lacking GD 2 -specific CAR expression was further included as negative control. Results from one representative experiment are shown. ( B ) Inhibition of GD 2 -specific NK-92tr-mediated NB cell lysis (w/o Ab; white-striped column) was analyzed after pre-incubation with excess of ganglidiximab (black column) using a calcein-AM-based cytotoxicity assay. Murine anti-Id ganglidiomab served as a positive control (grey column). Rituximab (white column) and murine IgG1 (white column) were utilized as negative controls. Results are expressed as percentage of lysis inhibition (mean values ± SEM) of two independent experiments performed using six replicates. One-way ANOVA on ranks followed by appropriate post hoc comparison test; * P 0.05 vs . w/o Ab.

    Article Snippet: For ganglidiomab VH, plasmid DNA (4 μg) was incubated with Not I-HF (1 unit; New England Biolabs GmbH, Frankfurt/Main, Germany) and Xho I-HF (1 unit; New England Biolabs GmbH, Frankfurt/Main, Germany) in CutSmart® Buffer (1×) for 1 h at +37°C followed by enzyme heat inactivation (20 min, +65°C).

    Techniques: Binding Assay, Inhibition, Expressing, Flow Cytometry, Cytometry, Staining, Positive Control, Incubation, Labeling, Negative Control, Lysis, Cytotoxicity Assay

    Schematic overview of generation of human/mouse chimeric anti-Id Ab ganglidiximab. The human/mouse chimeric anti-Id Ab ganglidiximab is composed of GD 2 mimicking variable regions (VH, VL) of murine anti-Id ganglidiomab and human IgG1 constant regions. Coding sequences of VH and VL were synthesized and inserted into mammalian expression plasmids containing DNA sequences for human IgG1 heavy (p3-IgG1-HC) and light chain (p3-IgG1-LC), respectively. For ganglidiximab production, CHO cells were stably co-transfected with the two generated expression plasmids (p3-ganglidiximab-HC and p3-ganglidiximab-LC).

    Journal: PLoS ONE

    Article Title: Generation and Characterization of a Human/Mouse Chimeric GD2-Mimicking Anti-Idiotype Antibody Ganglidiximab for Active Immunotherapy against Neuroblastoma

    doi: 10.1371/journal.pone.0150479

    Figure Lengend Snippet: Schematic overview of generation of human/mouse chimeric anti-Id Ab ganglidiximab. The human/mouse chimeric anti-Id Ab ganglidiximab is composed of GD 2 mimicking variable regions (VH, VL) of murine anti-Id ganglidiomab and human IgG1 constant regions. Coding sequences of VH and VL were synthesized and inserted into mammalian expression plasmids containing DNA sequences for human IgG1 heavy (p3-IgG1-HC) and light chain (p3-IgG1-LC), respectively. For ganglidiximab production, CHO cells were stably co-transfected with the two generated expression plasmids (p3-ganglidiximab-HC and p3-ganglidiximab-LC).

    Article Snippet: For ganglidiomab VH, plasmid DNA (4 μg) was incubated with Not I-HF (1 unit; New England Biolabs GmbH, Frankfurt/Main, Germany) and Xho I-HF (1 unit; New England Biolabs GmbH, Frankfurt/Main, Germany) in CutSmart® Buffer (1×) for 1 h at +37°C followed by enzyme heat inactivation (20 min, +65°C).

    Techniques: Synthesized, Expressing, Stable Transfection, Transfection, Generated

    Ganglidiximab-dependent inhibition of GD 2 -specific ch14.18/CHO-mediated ADCC and CDC. GD 2 -specific ch14.18/CHO-mediated ADCC ( A ) and CDC ( B ). Ch14.18 induced ADCC and CDC of NB cells LA-N-1 (black column) was compared to rituximab used as negative control (white-striped column). Pre-incubation of ch14.18 with excess of ganglidiximab (grey column) or ganglidiomab (white column) resulted in inhibition of both ADCC and CDC. Results are expressed as percentage of cytotoxicity (mean values ± SEM) of three independent experiments performed at least in triplicates. One-way ANOVA on ranks followed by appropriate post hoc comparison test; * P 0.05 vs . ch14.18/CHO.

    Journal: PLoS ONE

    Article Title: Generation and Characterization of a Human/Mouse Chimeric GD2-Mimicking Anti-Idiotype Antibody Ganglidiximab for Active Immunotherapy against Neuroblastoma

    doi: 10.1371/journal.pone.0150479

    Figure Lengend Snippet: Ganglidiximab-dependent inhibition of GD 2 -specific ch14.18/CHO-mediated ADCC and CDC. GD 2 -specific ch14.18/CHO-mediated ADCC ( A ) and CDC ( B ). Ch14.18 induced ADCC and CDC of NB cells LA-N-1 (black column) was compared to rituximab used as negative control (white-striped column). Pre-incubation of ch14.18 with excess of ganglidiximab (grey column) or ganglidiomab (white column) resulted in inhibition of both ADCC and CDC. Results are expressed as percentage of cytotoxicity (mean values ± SEM) of three independent experiments performed at least in triplicates. One-way ANOVA on ranks followed by appropriate post hoc comparison test; * P 0.05 vs . ch14.18/CHO.

    Article Snippet: For ganglidiomab VH, plasmid DNA (4 μg) was incubated with Not I-HF (1 unit; New England Biolabs GmbH, Frankfurt/Main, Germany) and Xho I-HF (1 unit; New England Biolabs GmbH, Frankfurt/Main, Germany) in CutSmart® Buffer (1×) for 1 h at +37°C followed by enzyme heat inactivation (20 min, +65°C).

    Techniques: Inhibition, Negative Control, Incubation

    Amplification of DNA fragments encoding for GD 2 -mimicking paratopes of ganglidiximab. ( A ) Visualization of coding sequences of GD 2 -mimicking variable heavy (VH; 440 bp) and light chain (VL; 420 bp) amplified by RT-PCR. RNA was isolated from hybridoma cells producing murine anti-Id ganglidiomab. PCR products were analyzed by agarose gel electrophoresis. Representative image is shown. NTC—no-template-control. M—Marker (100-bp). ( B ) PCR products were cloned into pCR ® 2.1-TOPO ® plasmids and analyzed by restriction enzyme digest to excise DNA sequences encoding for VH and VL (product sizes 427 bp and 407 bp, respectively). Resulting DNA fragments were analyzed by agarose gel electrophoresis. Representative image is shown. M—Marker (2-log, 0.1–10.0 kbp).

    Journal: PLoS ONE

    Article Title: Generation and Characterization of a Human/Mouse Chimeric GD2-Mimicking Anti-Idiotype Antibody Ganglidiximab for Active Immunotherapy against Neuroblastoma

    doi: 10.1371/journal.pone.0150479

    Figure Lengend Snippet: Amplification of DNA fragments encoding for GD 2 -mimicking paratopes of ganglidiximab. ( A ) Visualization of coding sequences of GD 2 -mimicking variable heavy (VH; 440 bp) and light chain (VL; 420 bp) amplified by RT-PCR. RNA was isolated from hybridoma cells producing murine anti-Id ganglidiomab. PCR products were analyzed by agarose gel electrophoresis. Representative image is shown. NTC—no-template-control. M—Marker (100-bp). ( B ) PCR products were cloned into pCR ® 2.1-TOPO ® plasmids and analyzed by restriction enzyme digest to excise DNA sequences encoding for VH and VL (product sizes 427 bp and 407 bp, respectively). Resulting DNA fragments were analyzed by agarose gel electrophoresis. Representative image is shown. M—Marker (2-log, 0.1–10.0 kbp).

    Article Snippet: For ganglidiomab VH, plasmid DNA (4 μg) was incubated with Not I-HF (1 unit; New England Biolabs GmbH, Frankfurt/Main, Germany) and Xho I-HF (1 unit; New England Biolabs GmbH, Frankfurt/Main, Germany) in CutSmart® Buffer (1×) for 1 h at +37°C followed by enzyme heat inactivation (20 min, +65°C).

    Techniques: Amplification, Reverse Transcription Polymerase Chain Reaction, Isolation, Polymerase Chain Reaction, Agarose Gel Electrophoresis, Marker, Clone Assay