Methods for producing active scfv antibodies and libraries therefor
AssigneesInstitut National De La Sante Et De La Recherche Medical (Inserm), Centre National De La Recherche Scientifiq Ue (C.N.R.S), Universite De Strasbourg
InventorsPierre Emile Ulysse Martineau, Etienne Weiss
Life of this patent
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Abstract
The present disclosure describes scFv antibody libraries, antibodies isolated from the libraries, and methods of producing and using the same.
Claims18
An antibody library that comprises at least about 106 unique scFv clones, wherein at least about 20% of the scFv clones encode an antibody that can detectably specifically bind a target antigen within a cell when the antibody encoded by the scFv clone is expressed within the cell.
An antibody library, wherein the antibody library comprises at least about 106 unique scFv antibody clones, wherein at least about 20% of the scFv antibody clones can be expressed within an E. coli cell to produce soluble antibody at a level of at least about 5 mgs per liter of E. coli cells, wherein the E coli cells have been grown to an OD600nm of about 5.
An antibody library comprising at least about 106 unique scFv antibody clones, wherein each unique scFv antibody clone encodes a unique scFv antibody comprising at least one of a unique CDR3 VH sequence and a unique CDR3 VL sequence, and wherein the unique scFv antibody clones encode a framework sequence substantially identical to a framework sequence encoded by scFv 13R4.
A method for preparing an scFv antibody library enriched for scFv antibody clones that can be expressed within a cell, comprising:
- a) providing a first collection of scFv antibody clones, wherein the first collection comprises clones comprising a unique sequence within a CDR3 loop of VH, wherein the first collection has been enriched for scFv antibody clones that can be detectably expressed when introduced into a cell;
- b) providing a second collection of scFv antibody clones, wherein the second collection comprises clones comprising a unique sequence within a CDR3 loop of VL, wherein the second collection has been enriched for scFv antibody clones that can be detectably expressed when introduced into a cell;
- c) joining VH domains from scFv antibody clones of the first collection with VL domains from scFv antibody clones of the second collection to obtain a third collection of scFv antibody clones, wherein the third collection contains scFv antibody clones comprising a unique sequence within the CDR3 loop of VH and a unique sequence within the CDR3 loop of VL,
- thereby preparing the scFv antibody library enriched for scFv antibody clones that can be expressed within a cell.
A method for constructing an antibody library comprising:
- a) selecting an scFv antibody framework;
- b) introducing sequence diversity into a VH CDR3 region of the scFv antibody framework to generate a first library comprising scFv antibody clones comprising a unique VH CDR3 region;
- c) introducing sequence diversity into a VL CDR3 region of the scFv antibody framework to generate a second library comprising scFv antibody clones comprising a unique VL CDR3 region;
- d) removing, from the first library, clones that do not detectably express scFv antibody;
- e) removing, from the second library, clones that do not detectably express scFv antibody;
- and
- f) recombining the first and second libraries to generate a final library comprising scFv antibody clones comprising a unique VH CDR3 region and a unique VL CDR3 region, thereby constructing the antibody library.
Dependencies are inferred from the claim text.