title: Identification of human antibody fragment clones specific for tetanus toxoid in a bacteriophage lambda immunoexpression library.
abstract: We have applied a molecular biology approach to the identification of human monoclonal antibodies. Human peripheral blood lymphocyte mRNA was converted to cDNA and a select subset was amplified by the polymerase chain reaction. These products, containing coding sequences for numerous immunoglobulin heavy- and kappa light-chain variable and constant region domains, were inserted into modified bacteriophage lambda expression vectors and introduced into Escherichia coli by infection to yield a combinatorial immunoexpression library. Clones with binding activity to tetanus toxoid were identified by filter hybridization with radiolabeled antigen and appeared at a frequency of 0.2% in the library. These human antigen binding fragments, consisting of a heavy-chain fragment covalently linked to a light chain, displayed high affinity of binding to tetanus toxoid with equilibrium constants in the nanomolar range but did not cross-react with other proteins tested. We estimate that this human immunoexpression library contains 20,000 clones with high affinity and specificity to our chosen antigen.
authors: Stratagene, La Jolla, CA 92037.
authors: R L
authors: RL
authors: Mullinax
authors: E A
authors: EA
authors: Gross
authors: J R
authors: JR
authors: Amberg
authors: B N
authors: BN
authors: Hay
authors: H H
authors: HH
authors: Hogrefe
authors: M M
authors: MM
authors: Kubitz
authors: A
authors: A
authors: Greener
authors: M
authors: M
authors: Alting-Mees
authors: D
authors: D
authors: Ardourel
authors: J M
authors: JM
authors: Short
mesh_terms: D000595:Amino Acid Sequence; D000918:Antibody Specificity; D010582:Bacteriophage lambda / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay; D015723:Gene Library; D005821:Genetic Techniques; D006801:Humans; D007128:Immunoglobulin Fragments / Q000235:genetics* / Q000276:immunology; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D012333:RNA, Messenger / Q000235:genetics; D015183:Restriction Mapping; D013745:Tetanus Toxoid / Q000276:immunology*
chemical_list: D007128:Immunoglobulin Fragments; D012333:RNA, Messenger; D013745:Tetanus Toxoid