Abstract
Bispecific miniantibodies were constructed by genetically fusing the C(H)1 domain of an IgG1 to the C-terminus of a single-chain Fv fragment (scFv-425), specific for the EGF receptor, and fusing the C(L) domain of a kappa light chain to the C-terminus of a scFv specific for CD2 (scFv-M1). An efficient dicistronic gene arrangement for functional expression in Escherichia coli was constructed. Immunoblots demonstrated correct domain assembly and the formation of the natural C(H)1-C(L) disulfide bridge. Gel filtration confirmed the correct size, sandwich ELISAs demonstrated bispecific functionality, and SPR biosensor measurements determined binding to EGF-R in comparison to bivalent constructs. Bispecific anti-EGF-R/anti-CD2 miniantibodies are candidates for the immunotherapy of cancer.
Publication types
- Research Support, Non-U.S. Gov't
MeSH terms
- Amino Acid Sequence
- Antibodies, Bispecific / biosynthesis
- Antibodies, Bispecific / chemistry*
- Antibody Specificity
- Base Sequence
- Binding Sites, Antibody
- CD2 Antigens / immunology*
- Cloning, Molecular
- Dimerization
- ErbB Receptors / immunology*
- Escherichia coli
- Genes, Immunoglobulin
- Humans
- Immunoglobulin Constant Regions / biosynthesis
- Immunoglobulin Constant Regions / chemistry*
- Immunoglobulin G / biosynthesis
- Immunoglobulin G / chemistry*
- Immunoglobulin Heavy Chains / biosynthesis
- Immunoglobulin Heavy Chains / chemistry*
- Immunoglobulin kappa-Chains / biosynthesis
- Immunoglobulin kappa-Chains / chemistry*
- Models, Molecular
- Molecular Sequence Data
- Polymerase Chain Reaction
- Protein Conformation*
- Recombinant Proteins / biosynthesis
- Recombinant Proteins / chemistry
- Restriction Mapping
Substances
- Antibodies, Bispecific
- CD2 Antigens
- Immunoglobulin Constant Regions
- Immunoglobulin G
- Immunoglobulin Heavy Chains
- Immunoglobulin kappa-Chains
- Recombinant Proteins
- ErbB Receptors