Recombinant:Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. 1-1) Preparation of Immunogen As an immunogen, a human BMP10 recombinant protein or a human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was used. ..
Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. Specifically, a solution in which the human BMP10 recombinant prepared in Example 1, 1-1) or a human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was prepared at 0.5 μg/mL with a phosphate buffer solution (manufactured by Nacalai Tesque, Inc.) was dispensed in a 96-well ELISA plate (F96 MAXISORP NUNC-IMMNO PLATE, manufactured by Thermo Fisher Scientific, Inc., Cat #442404) at 50 μL/well, and adsorption was carried out by leaving the plate to stand overnight at 4° C. After the immobilization solution was removed, washing was carried out 3 to 5 times with PBS, and 1% BSA-PBS (manufactured by Nacalai Tesque, Inc., Cat #099968-35) was added at 200 μL/well, and blocking was carried out by leaving the plate to stand at room temperature for 1 hour. ..
Binding Assay:Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: The purified antibody was confirmed for its quality by analytical gel filtration chromatography (using an apparatus manufactured by Shimadzu Corporation, and a column TSKgel SuperSW3000 manufactured by Tosoh Corporation) and SDS-PAGE. .. For the purpose of comparing the binding activity to human BMP10 between the ch18C1 antibody obtained in Example 5-3 and the anti-BMP10 humanized antibody obtained in Example 12, the binding activity to a human BMP10 mature dimer (manufactured by R & D Systems, Cat #2926-BP) was measured by a surface plasmon resonance method (SPR method) using Biacore T100 (manufactured by GE Healthcare Bio-Sciences Corporation). ..
Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. 5-5) Evaluation of Binding Activity of Anti-BMP10 Chimeric Antibody to Human BMP10 Protein by Biacore For the purpose of comparing the binding activity to human BMP10 between the anti-BMP10 chimeric antibody obtained in Example 5-4 and MAB2926 (R & D Systems, Inc.) that is a known antibody, the binding activity to a human BMP10 mature dimer (manufactured by R & D Systems, Cat #2926-BP) was measured by a surface plasmon resonance method (SPR method) using Biacore T100 (manufactured by GE Healthcare Bio-Sciences Corporation). ..
Activity Assay:Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: The purified antibody was confirmed for its quality by analytical gel filtration chromatography (using an apparatus manufactured by Shimadzu Corporation, and a column TSKgel SuperSW3000 manufactured by Tosoh Corporation) and SDS-PAGE. .. For the purpose of comparing the binding activity to human BMP10 between the ch18C1 antibody obtained in Example 5-3 and the anti-BMP10 humanized antibody obtained in Example 12, the binding activity to a human BMP10 mature dimer (manufactured by R & D Systems, Cat #2926-BP) was measured by a surface plasmon resonance method (SPR method) using Biacore T100 (manufactured by GE Healthcare Bio-Sciences Corporation). ..
Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. 3-2) Comparison of BMP10 Neutralizing Activity between Newly Obtained Antibody and Known Antibody To a 96-well fluorescence and luminescence plate (manufactured by Corning, Inc., Cat #3916), human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was added at a final concentration of 0.3 ng/mL. .. Subsequently, the 18C1 antibody, the 12H3 antibody, the 11H10 antibody or MAB2926, each of which is an anti-BMP10 antibody, or a control antibody (Purified Rat IgG1λ Isotype control, manufactured by BD, Inc., Cat #553993) was prepared at 6 concentrations by 3-fold serial dilution from the final concentration of 3000 ng/mL, and then added.
Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. 5-5) Evaluation of Binding Activity of Anti-BMP10 Chimeric Antibody to Human BMP10 Protein by Biacore For the purpose of comparing the binding activity to human BMP10 between the anti-BMP10 chimeric antibody obtained in Example 5-4 and MAB2926 (R & D Systems, Inc.) that is a known antibody, the binding activity to a human BMP10 mature dimer (manufactured by R & D Systems, Cat #2926-BP) was measured by a surface plasmon resonance method (SPR method) using Biacore T100 (manufactured by GE Healthcare Bio-Sciences Corporation). ..
SPR Assay:Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: The purified antibody was confirmed for its quality by analytical gel filtration chromatography (using an apparatus manufactured by Shimadzu Corporation, and a column TSKgel SuperSW3000 manufactured by Tosoh Corporation) and SDS-PAGE. .. For the purpose of comparing the binding activity to human BMP10 between the ch18C1 antibody obtained in Example 5-3 and the anti-BMP10 humanized antibody obtained in Example 12, the binding activity to a human BMP10 mature dimer (manufactured by R & D Systems, Cat #2926-BP) was measured by a surface plasmon resonance method (SPR method) using Biacore T100 (manufactured by GE Healthcare Bio-Sciences Corporation). ..
Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. 5-5) Evaluation of Binding Activity of Anti-BMP10 Chimeric Antibody to Human BMP10 Protein by Biacore For the purpose of comparing the binding activity to human BMP10 between the anti-BMP10 chimeric antibody obtained in Example 5-4 and MAB2926 (R & D Systems, Inc.) that is a known antibody, the binding activity to a human BMP10 mature dimer (manufactured by R & D Systems, Cat #2926-BP) was measured by a surface plasmon resonance method (SPR method) using Biacore T100 (manufactured by GE Healthcare Bio-Sciences Corporation). ..
Enzyme-linked Immunosorbent Assay:Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. Specifically, a solution in which the human BMP10 recombinant prepared in Example 1, 1-1) or a human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was prepared at 0.5 μg/mL with a phosphate buffer solution (manufactured by Nacalai Tesque, Inc.) was dispensed in a 96-well ELISA plate (F96 MAXISORP NUNC-IMMNO PLATE, manufactured by Thermo Fisher Scientific, Inc., Cat #442404) at 50 μL/well, and adsorption was carried out by leaving the plate to stand overnight at 4° C. After the immobilization solution was removed, washing was carried out 3 to 5 times with PBS, and 1% BSA-PBS (manufactured by Nacalai Tesque, Inc., Cat #099968-35) was added at 200 μL/well, and blocking was carried out by leaving the plate to stand at room temperature for 1 hour. ..
Adsorption:Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. Specifically, a solution in which the human BMP10 recombinant prepared in Example 1, 1-1) or a human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was prepared at 0.5 μg/mL with a phosphate buffer solution (manufactured by Nacalai Tesque, Inc.) was dispensed in a 96-well ELISA plate (F96 MAXISORP NUNC-IMMNO PLATE, manufactured by Thermo Fisher Scientific, Inc., Cat #442404) at 50 μL/well, and adsorption was carried out by leaving the plate to stand overnight at 4° C. After the immobilization solution was removed, washing was carried out 3 to 5 times with PBS, and 1% BSA-PBS (manufactured by Nacalai Tesque, Inc., Cat #099968-35) was added at 200 μL/well, and blocking was carried out by leaving the plate to stand at room temperature for 1 hour. ..
Blocking Assay:Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. Specifically, a solution in which the human BMP10 recombinant prepared in Example 1, 1-1) or a human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was prepared at 0.5 μg/mL with a phosphate buffer solution (manufactured by Nacalai Tesque, Inc.) was dispensed in a 96-well ELISA plate (F96 MAXISORP NUNC-IMMNO PLATE, manufactured by Thermo Fisher Scientific, Inc., Cat #442404) at 50 μL/well, and adsorption was carried out by leaving the plate to stand overnight at 4° C. After the immobilization solution was removed, washing was carried out 3 to 5 times with PBS, and 1% BSA-PBS (manufactured by Nacalai Tesque, Inc., Cat #099968-35) was added at 200 μL/well, and blocking was carried out by leaving the plate to stand at room temperature for 1 hour. ..
Fluorescence:Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. To a 96-well fluorescence and luminescence plate (manufactured by Corning, Inc., Cat #3916), a human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was added at a final concentration of 3 ng/mL. .. Subsequently, a known antibody MAB2926 (the known antibody refers to MAB2926), and the 18C1 antibody, the 12H3 antibody, and the 11H10 antibody, each of which is an anti-BMP10 antibody purified in Example 1, 1-9), or a control antibody (Purified Rat IgG1λ Isotype control, manufactured by BD, Inc., Cat #553993) were prepared at 6 concentrations by 3-fold serial dilution from the final concentration of 3000 ng/mL, and then added.
Article Title: Anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. To a 96-well fluorescence and luminescence plate (manufactured by Corning, Inc., Cat #3916), a human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was added at a final concentration of 0.3 ng/mL, and subsequently, the anti-BMP10 humanized antibody obtained in Example 12 or the ch18C1 antibody was prepared at 7 concentrations by 3-fold serial dilution from the final concentration of 3000 ng/mL, and then added. .. Thereafter, an ALK1/Id1-Luc/CHO cell liquid in which the cells were suspended in Excell 325 medium [Excell 325 PF CHO (manufactured by SAFC, Inc., Cat #14340C-1000 mL), 4 mM L-glutamine, 1×Penicillin, 1×Streptomycin (manufactured by Nacalai, Inc., Cat #09367-34), 0.5 mg/mL hygromycin] was added at 5×104 cells/well.
Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. 3-2) Comparison of BMP10 Neutralizing Activity between Newly Obtained Antibody and Known Antibody To a 96-well fluorescence and luminescence plate (manufactured by Corning, Inc., Cat #3916), human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was added at a final concentration of 0.3 ng/mL. .. Subsequently, the 18C1 antibody, the 12H3 antibody, the 11H10 antibody or MAB2926, each of which is an anti-BMP10 antibody, or a control antibody (Purified Rat IgG1λ Isotype control, manufactured by BD, Inc., Cat #553993) was prepared at 6 concentrations by 3-fold serial dilution from the final concentration of 3000 ng/mL, and then added.
Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. To a 96-well fluorescence and luminescence plate (manufactured by Corning, Inc., Cat #3916), a human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was added at a final concentration of 0.3 ng/mL, and subsequently, the anti-BMP10 humanized antibody obtained in Example 12 or the ch18C1 antibody was prepared at 7 concentrations by 3-fold serial dilution from the final concentration of 3000 ng/mL, and then added. .. Thereafter, an ALK1/Id1-Luc/CHO cell liquid in which the cells were suspended in Excell 325 medium [Excell 325 PF CHO (manufactured by SAFC, Inc., Cat #14340C-1000 mL), 4 mM L-glutamine, 1× Penicillin, 1× Streptomycin (manufactured by Nacalai, Inc., Cat #09367-34), 0.5 mg/mL hygromycin] was added at 5×104 cells/well.
Concentration Assay:Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. To a 96-well fluorescence and luminescence plate (manufactured by Corning, Inc., Cat #3916), a human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was added at a final concentration of 3 ng/mL. .. Subsequently, a known antibody MAB2926 (the known antibody refers to MAB2926), and the 18C1 antibody, the 12H3 antibody, and the 11H10 antibody, each of which is an anti-BMP10 antibody purified in Example 1, 1-9), or a control antibody (Purified Rat IgG1λ Isotype control, manufactured by BD, Inc., Cat #553993) were prepared at 6 concentrations by 3-fold serial dilution from the final concentration of 3000 ng/mL, and then added.
Article Title: Anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. To a 96-well fluorescence and luminescence plate (manufactured by Corning, Inc., Cat #3916), a human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was added at a final concentration of 0.3 ng/mL, and subsequently, the anti-BMP10 humanized antibody obtained in Example 12 or the ch18C1 antibody was prepared at 7 concentrations by 3-fold serial dilution from the final concentration of 3000 ng/mL, and then added. .. Thereafter, an ALK1/Id1-Luc/CHO cell liquid in which the cells were suspended in Excell 325 medium [Excell 325 PF CHO (manufactured by SAFC, Inc., Cat #14340C-1000 mL), 4 mM L-glutamine, 1×Penicillin, 1×Streptomycin (manufactured by Nacalai, Inc., Cat #09367-34), 0.5 mg/mL hygromycin] was added at 5×104 cells/well.
Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. 3-2) Comparison of BMP10 Neutralizing Activity between Newly Obtained Antibody and Known Antibody To a 96-well fluorescence and luminescence plate (manufactured by Corning, Inc., Cat #3916), human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was added at a final concentration of 0.3 ng/mL. .. Subsequently, the 18C1 antibody, the 12H3 antibody, the 11H10 antibody or MAB2926, each of which is an anti-BMP10 antibody, or a control antibody (Purified Rat IgG1λ Isotype control, manufactured by BD, Inc., Cat #553993) was prepared at 6 concentrations by 3-fold serial dilution from the final concentration of 3000 ng/mL, and then added.
Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. To a 96-well fluorescence and luminescence plate (manufactured by Corning, Inc., Cat #3916), a human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was added at a final concentration of 0.3 ng/mL, and subsequently, the anti-BMP10 humanized antibody obtained in Example 12 or the ch18C1 antibody was prepared at 7 concentrations by 3-fold serial dilution from the final concentration of 3000 ng/mL, and then added. .. Thereafter, an ALK1/Id1-Luc/CHO cell liquid in which the cells were suspended in Excell 325 medium [Excell 325 PF CHO (manufactured by SAFC, Inc., Cat #14340C-1000 mL), 4 mM L-glutamine, 1× Penicillin, 1× Streptomycin (manufactured by Nacalai, Inc., Cat #09367-34), 0.5 mg/mL hygromycin] was added at 5×104 cells/well.
Serial Dilution:Article Title: Anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. To a 96-well fluorescence and luminescence plate (manufactured by Corning, Inc., Cat #3916), a human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was added at a final concentration of 0.3 ng/mL, and subsequently, the anti-BMP10 humanized antibody obtained in Example 12 or the ch18C1 antibody was prepared at 7 concentrations by 3-fold serial dilution from the final concentration of 3000 ng/mL, and then added. .. Thereafter, an ALK1/Id1-Luc/CHO cell liquid in which the cells were suspended in Excell 325 medium [Excell 325 PF CHO (manufactured by SAFC, Inc., Cat #14340C-1000 mL), 4 mM L-glutamine, 1×Penicillin, 1×Streptomycin (manufactured by Nacalai, Inc., Cat #09367-34), 0.5 mg/mL hygromycin] was added at 5×104 cells/well.
Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. To a 96-well fluorescence and luminescence plate (manufactured by Corning, Inc., Cat #3916), a human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was added at a final concentration of 0.3 ng/mL, and subsequently, the anti-BMP10 humanized antibody obtained in Example 12 or the ch18C1 antibody was prepared at 7 concentrations by 3-fold serial dilution from the final concentration of 3000 ng/mL, and then added. .. Thereafter, an ALK1/Id1-Luc/CHO cell liquid in which the cells were suspended in Excell 325 medium [Excell 325 PF CHO (manufactured by SAFC, Inc., Cat #14340C-1000 mL), 4 mM L-glutamine, 1× Penicillin, 1× Streptomycin (manufactured by Nacalai, Inc., Cat #09367-34), 0.5 mg/mL hygromycin] was added at 5×104 cells/well.
Comparison:Article Title: Methods for treating hypertension using an anti-BMP10 monoclonal antibody or fragment thereof
Article Snippet: .. 3-2) Comparison of BMP10 Neutralizing Activity between Newly Obtained Antibody and Known Antibody To a 96-well fluorescence and luminescence plate (manufactured by Corning, Inc., Cat #3916), human BMP10 mature dimer (manufactured by R & D Systems, Inc., Cat #2926-BP) was added at a final concentration of 0.3 ng/mL. .. Subsequently, the 18C1 antibody, the 12H3 antibody, the 11H10 antibody or MAB2926, each of which is an anti-BMP10 antibody, or a control antibody (Purified Rat IgG1λ Isotype control, manufactured by BD, Inc., Cat #553993) was prepared at 6 concentrations by 3-fold serial dilution from the final concentration of 3000 ng/mL, and then added.
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