High throughput generation and affinity maturation of humanized antibody
First Claim
1. A method for screening a library of humanized antibody in yeast, comprising:
- expressing a library of humanized antibodies in yeast cells;
expressing a specific target protein in the yeast cells expressing the humanized antibodies; and
selecting those yeast cells in which a reporter gene is expressed, the expression of the reporter gene being activated by binding of the humanized antibody to the target protein.
3 Assignments
0 Petitions
Accused Products
Abstract
Compositions, methods, and kits are provided for efficiently generating and screening humanized antibody with high affinity against a specific antigen. The library of humanized antibody is generated by mutagenizing a chimeric antibody template that combines human antibody framework and antigen binding sites of a non-human antibody. Alternatively, the library of humanized antibody is generated by grafting essential antigen-recognition segment(s) such as CDRs of the non-human antibody into the corresponding position(s) of each member of a human antibody library. This library of humanized antibody is then screened for high affinity binding toward a specific antigen in vivo in organism such as yeast or in vitro using techniques such as ribosome display or mRNA display. The overall process can be efficiently performed in a high throughput and automated manner, thus mimicking the natural process of antibody affinity maturation.
-
Citations
8 Claims
-
1. A method for screening a library of humanized antibody in yeast, comprising:
-
expressing a library of humanized antibodies in yeast cells;
expressing a specific target protein in the yeast cells expressing the humanized antibodies; and
selecting those yeast cells in which a reporter gene is expressed, the expression of the reporter gene being activated by binding of the humanized antibody to the target protein. - View Dependent Claims (2, 3)
-
-
4. A method for screening a library of humanized antibody in vitro, comprising:
-
expressing a library of humanized antibodies in vitro;
displaying the library of humanized antibodies on ribosome;
contacting the ribosome displaying the library of humanized antibodies with a target antigen; and
selecting those ribosomes that bind to the target antigen, - View Dependent Claims (5, 6)
-
-
7. A method for screening a library of humanized antibody in vitro, comprising:
-
transcribing a library of cDNA encoding a library of humanized antibodies in vitro to produce a library of mRNA;
contacting the library of mRNA with a peptidyl acceptor linker to produce a library of modified mRNA;
translating the library of modified mRNA in vitro to produce a library of humanized antibody, wherein the translated humanized antibody is covalently linked to the modified mRNA by the peptidyl acceptor linker; and
contacting the library of humanized antibodies linked with the modified mRNA with a target antigen; and
selecting the humanized antibody that bind to the target antigen, - View Dependent Claims (8)
-
Specification