Variant Assay:Article Title: Method for improving affinity of antibody for antigen and use thereof
Article Snippet: By substituting 3 amino acid residues of heavy chain FR3 of an anti-lysozyme antibody with arginine residues, 68, 70 72 variant, 79, 81, 82A variant, and 82B, 83, 84 variant were prepared as modified antibodies of the anti-lysozyme antibody. (1) Obtainment of Gene of Wild-Type Anti-Lysozyme Antibody A gene of a mouse anti-lysozyme antibody (HyHEL-10) was synthesized by GenScript Japan Inc. on commission, so that a plasmid DNA containing a gene of a wild-type anti-lysozyme antibody was obtained. (2) Preparation of Variant of Anti-Lysozyme Antibody [Reagents] QIAprep Spin Miniprep kit (QIAGEN) PrimeSTAR (registered trademark) Max DNA Polymerase (Takara Bio Inc.) Ligation high ver.2 (TOYOBO CO., LTD.) T4 Polynucleotide Kinase (TOYOBO CO., LTD.) Dpn I (TOYOBO CO., LTD.) Competent high DH5α (TOYOBO CO., LTD.) (2.1) Primer Design and PCR Based on a base sequence of the wild-type anti-lysozyme antibody gene in the plasmid DNA obtained in the above (1), a primer set for obtaining a polynucleotide encoding a light chain and a primer set for obtaining a polynucleotide encoding a heavy chain in which the following 3 amino acid residues in FR3 were substituted with arginine residues were designed.
Modification:Article Title: Method for improving affinity of antibody for antigen and use thereof
Article Snippet: By substituting 3 amino acid residues of heavy chain FR3 of an anti-lysozyme antibody with arginine residues, 68, 70 72 variant, 79, 81, 82A variant, and 82B, 83, 84 variant were prepared as modified antibodies of the anti-lysozyme antibody. (1) Obtainment of Gene of Wild-Type Anti-Lysozyme Antibody A gene of a mouse anti-lysozyme antibody (HyHEL-10) was synthesized by GenScript Japan Inc. on commission, so that a plasmid DNA containing a gene of a wild-type anti-lysozyme antibody was obtained. (2) Preparation of Variant of Anti-Lysozyme Antibody [Reagents] QIAprep Spin Miniprep kit (QIAGEN) PrimeSTAR (registered trademark) Max DNA Polymerase (Takara Bio Inc.) Ligation high ver.2 (TOYOBO CO., LTD.) T4 Polynucleotide Kinase (TOYOBO CO., LTD.) Dpn I (TOYOBO CO., LTD.) Competent high DH5α (TOYOBO CO., LTD.) (2.1) Primer Design and PCR Based on a base sequence of the wild-type anti-lysozyme antibody gene in the plasmid DNA obtained in the above (1), a primer set for obtaining a polynucleotide encoding a light chain and a primer set for obtaining a polynucleotide encoding a heavy chain in which the following 3 amino acid residues in FR3 were substituted with arginine residues were designed.
Synthesized:Article Title: Method for improving affinity of antibody for antigen and use thereof
Article Snippet: By substituting 3 amino acid residues of heavy chain FR3 of an anti-lysozyme antibody with arginine residues, 68, 70 72 variant, 79, 81, 82A variant, and 82B, 83, 84 variant were prepared as modified antibodies of the anti-lysozyme antibody. (1) Obtainment of Gene of Wild-Type Anti-Lysozyme Antibody A gene of a mouse anti-lysozyme antibody (HyHEL-10) was synthesized by GenScript Japan Inc. on commission, so that a plasmid DNA containing a gene of a wild-type anti-lysozyme antibody was obtained. (2) Preparation of Variant of Anti-Lysozyme Antibody [Reagents] QIAprep Spin Miniprep kit (QIAGEN) PrimeSTAR (registered trademark) Max DNA Polymerase (Takara Bio Inc.) Ligation high ver.2 (TOYOBO CO., LTD.) T4 Polynucleotide Kinase (TOYOBO CO., LTD.) Dpn I (TOYOBO CO., LTD.) Competent high DH5α (TOYOBO CO., LTD.) (2.1) Primer Design and PCR Based on a base sequence of the wild-type anti-lysozyme antibody gene in the plasmid DNA obtained in the above (1), a primer set for obtaining a polynucleotide encoding a light chain and a primer set for obtaining a polynucleotide encoding a heavy chain in which the following 3 amino acid residues in FR3 were substituted with arginine residues were designed.
Plasmid Preparation:Article Title: Method for improving affinity of antibody for antigen and use thereof
Article Snippet: By substituting 3 amino acid residues of heavy chain FR3 of an anti-lysozyme antibody with arginine residues, 68, 70 72 variant, 79, 81, 82A variant, and 82B, 83, 84 variant were prepared as modified antibodies of the anti-lysozyme antibody. (1) Obtainment of Gene of Wild-Type Anti-Lysozyme Antibody A gene of a mouse anti-lysozyme antibody (HyHEL-10) was synthesized by GenScript Japan Inc. on commission, so that a plasmid DNA containing a gene of a wild-type anti-lysozyme antibody was obtained. (2) Preparation of Variant of Anti-Lysozyme Antibody [Reagents] QIAprep Spin Miniprep kit (QIAGEN) PrimeSTAR (registered trademark) Max DNA Polymerase (Takara Bio Inc.) Ligation high ver.2 (TOYOBO CO., LTD.) T4 Polynucleotide Kinase (TOYOBO CO., LTD.) Dpn I (TOYOBO CO., LTD.) Competent high DH5α (TOYOBO CO., LTD.) (2.1) Primer Design and PCR Based on a base sequence of the wild-type anti-lysozyme antibody gene in the plasmid DNA obtained in the above (1), a primer set for obtaining a polynucleotide encoding a light chain and a primer set for obtaining a polynucleotide encoding a heavy chain in which the following 3 amino acid residues in FR3 were substituted with arginine residues were designed.
Ligation:Article Title: Method for improving affinity of antibody for antigen and use thereof
Article Snippet: By substituting 3 amino acid residues of heavy chain FR3 of an anti-lysozyme antibody with arginine residues, 68, 70 72 variant, 79, 81, 82A variant, and 82B, 83, 84 variant were prepared as modified antibodies of the anti-lysozyme antibody. (1) Obtainment of Gene of Wild-Type Anti-Lysozyme Antibody A gene of a mouse anti-lysozyme antibody (HyHEL-10) was synthesized by GenScript Japan Inc. on commission, so that a plasmid DNA containing a gene of a wild-type anti-lysozyme antibody was obtained. (2) Preparation of Variant of Anti-Lysozyme Antibody [Reagents] QIAprep Spin Miniprep kit (QIAGEN) PrimeSTAR (registered trademark) Max DNA Polymerase (Takara Bio Inc.) Ligation high ver.2 (TOYOBO CO., LTD.) T4 Polynucleotide Kinase (TOYOBO CO., LTD.) Dpn I (TOYOBO CO., LTD.) Competent high DH5α (TOYOBO CO., LTD.) (2.1) Primer Design and PCR Based on a base sequence of the wild-type anti-lysozyme antibody gene in the plasmid DNA obtained in the above (1), a primer set for obtaining a polynucleotide encoding a light chain and a primer set for obtaining a polynucleotide encoding a heavy chain in which the following 3 amino acid residues in FR3 were substituted with arginine residues were designed.
Polymerase Chain Reaction:Article Title: Method for improving affinity of antibody for antigen and use thereof
Article Snippet: By substituting 3 amino acid residues of heavy chain FR3 of an anti-lysozyme antibody with arginine residues, 68, 70 72 variant, 79, 81, 82A variant, and 82B, 83, 84 variant were prepared as modified antibodies of the anti-lysozyme antibody. (1) Obtainment of Gene of Wild-Type Anti-Lysozyme Antibody A gene of a mouse anti-lysozyme antibody (HyHEL-10) was synthesized by GenScript Japan Inc. on commission, so that a plasmid DNA containing a gene of a wild-type anti-lysozyme antibody was obtained. (2) Preparation of Variant of Anti-Lysozyme Antibody [Reagents] QIAprep Spin Miniprep kit (QIAGEN) PrimeSTAR (registered trademark) Max DNA Polymerase (Takara Bio Inc.) Ligation high ver.2 (TOYOBO CO., LTD.) T4 Polynucleotide Kinase (TOYOBO CO., LTD.) Dpn I (TOYOBO CO., LTD.) Competent high DH5α (TOYOBO CO., LTD.) (2.1) Primer Design and PCR Based on a base sequence of the wild-type anti-lysozyme antibody gene in the plasmid DNA obtained in the above (1), a primer set for obtaining a polynucleotide encoding a light chain and a primer set for obtaining a polynucleotide encoding a heavy chain in which the following 3 amino acid residues in FR3 were substituted with arginine residues were designed.
Sequencing:Article Title: Method for improving affinity of antibody for antigen and use thereof
Article Snippet: By substituting 3 amino acid residues of heavy chain FR3 of an anti-lysozyme antibody with arginine residues, 68, 70 72 variant, 79, 81, 82A variant, and 82B, 83, 84 variant were prepared as modified antibodies of the anti-lysozyme antibody. (1) Obtainment of Gene of Wild-Type Anti-Lysozyme Antibody A gene of a mouse anti-lysozyme antibody (HyHEL-10) was synthesized by GenScript Japan Inc. on commission, so that a plasmid DNA containing a gene of a wild-type anti-lysozyme antibody was obtained. (2) Preparation of Variant of Anti-Lysozyme Antibody [Reagents] QIAprep Spin Miniprep kit (QIAGEN) PrimeSTAR (registered trademark) Max DNA Polymerase (Takara Bio Inc.) Ligation high ver.2 (TOYOBO CO., LTD.) T4 Polynucleotide Kinase (TOYOBO CO., LTD.) Dpn I (TOYOBO CO., LTD.) Competent high DH5α (TOYOBO CO., LTD.) (2.1) Primer Design and PCR Based on a base sequence of the wild-type anti-lysozyme antibody gene in the plasmid DNA obtained in the above (1), a primer set for obtaining a polynucleotide encoding a light chain and a primer set for obtaining a polynucleotide encoding a heavy chain in which the following 3 amino acid residues in FR3 were substituted with arginine residues were designed.
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