A high-affinity macaque antibody Fab with human-like framework regions obtained from a small phage display immune library.
Tetanus toxoid was used as a model antigen for the immunization of Macaca fascicularis. Using peripheral blood as a template, specific Fab-encoding genes were amplified by PCR on the fourth day after the final boost, and cloned in a phagemidic vector (pComb3X) as a small immune library (5 x 10(5) clones). A high-affinity Fab (Kd = 4 x 10(-10) M), 6-ATT, was isolated from this library by panning. The genes encoding 6-ATT were found to be similar to human immunoglobulin germline genes, and were assigned to subgroups of human V, (D) or J genes by IMGT/V-QUEST. Overall, the Fab variable domain framework regions were 93% identical to the representative genes and alleles of the human subgroups, this level of identity being similar to that between genes of the same human subgroup. This strategy could be used for the isolation of high-affinity, human-like Fab fragments directed against numerous antigens.
Chassagne
Stéphanie
S
Immunobiologie, Département de biologie des agents transmissibles (DBAT), Centre de recherche du Service de santé des armées, 24, av. des maquis du Gresivaudan, La Tronche, P.O. box 38702, France.
Laffly
Emmanuelle
E
Drouet
Emmanuel
E
Hérodin
Francis
F
Lefranc
Marie-Paule
MP
Thullier
Philippe
P
D000595:Amino Acid Sequence; D000818:Animals; D000915:Antibody Affinity; D003001:Cloning, Molecular; D006801:Humans; D007114:Immunization; D007140:Immunoglobulin Fab Fragments / Q000235:genetics / Q000276:immunology*; D008252:Macaca fascicularis / Q000276:immunology*; D019151:Peptide Library; D017434:Protein Structure, Tertiary; D017385:Sequence Homology; D013744:Tetanus Toxin / Q000008:administration & dosage / Q000276:immunology
D007140:Immunoglobulin Fab Fragments; D019151:Peptide Library; D013744:Tetanus Toxin