A method for rapid, ligation-independent reformatting of recombinant monoclonal antibodies.
Recombinant monoclonal antibodies currently dominate the protein biologics marketplace. The path from target antigen discovery and screening, to a recombinant therapeutic antibody can be time-consuming and laborious. We describe a set of expression vectors, termed mAbXpress, that enable rapid and sequence-independent insertion of antibody variable regions into human constant region backbones. This method takes advantage of the In Fusion cloning system from Clontech, which allows ligation-free, high-efficiency insertion of the variable region cassette without the addition of extraneous amino acids. These modular vectors simplify the antibody reformatting process during the preliminary evaluation of therapeutic or diagnostic candidates. The resulting constructs can be used directly for transient or amplifiable, stable expression in mammalian cells. The effectiveness of this method was demonstrated by the creation of a functional, fully human anti-human CD83 monoclonal antibody.
Australian Institute for Bioengineering and Nanotechnology (AIBN), Bldg. 75 Cnr College and Cooper Rd., The University of Queensland, Brisbane 4072, Australia.
Martina L
ML
Jones
Therese
T
Seldon
Matthew
M
Smede
Ashleigh
A
Linville
David Y
DY
Chin
Ross
R
Barnard
Stephen M
SM
Mahler
David
D
Munster
Derek
D
Hart
Peter P
PP
Gray
Trent P
TP
Munro
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D000920:Antibody-Dependent Cell Cytotoxicity; D015703:Antigens, CD / Q000276:immunology*; D001666:Binding Sites, Antibody; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D003412:Cricetulus; D005434:Flow Cytometry; D005822:Genetic Vectors; D006801:Humans; D007127:Immunoglobulin Constant Regions / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D007136:Immunoglobulins / Q000276:immunology*; D015979:Killer Cells, Lymphokine-Activated / Q000276:immunology; D008562:Membrane Glycoproteins / Q000276:immunology*; D019151:Peptide Library; D011994:Recombinant Proteins / Q000276:immunology; D013997:Time Factors; D014162:Transfection; D000097802:CD83 Antigen
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D007127:Immunoglobulin Constant Regions; D007135:Immunoglobulin Variable Region; D007136:Immunoglobulins; D008562:Membrane Glycoproteins; D019151:Peptide Library; D011994:Recombinant Proteins; D000097802:CD83 Antigen