Immunological applications of single-domain llama recombinant antibodies isolated from a naïve library.
We describe the rapid isolation of single-domain recombinant antibodies in VHH format from a pre-immune llama library created in our laboratory. Such naïve library has demonstrated to be a versatile tool and enabled the isolation of several different antibodies for any of the six proteins panned in parallel. The binders specific for human fibroblast growth factor receptor 1 (FGFR1) were successively analyzed in more detail and resulted suitable for both western blot and immunofluorescence analyses. Several milligrams per liter of antibodies were purified by affinity chromatography and used for kinetic and thermodynamic characterization. Their K(D) was in the nanomolar range and they apparently bound a FGF receptor 1 domain not overlapping the region recognized by its physiological ligand FGF. Altogether, the collected data indicate that the new library can enable the recovery of binders of high affinity, specificity and functionality in the conventional immunological tests, avoiding the necessity of further maturation steps. Such results confirmed recent reports of high affinity pre-immune IgNARs and supported the choice of using large single-domain recombinant antibody naïve libraries as an alternative to the preparation of immune libraries for selecting monoclonal antibodies, at convenient cost and time conditions.
Monegal
Ana
A
Consortium for Genomic Technologies, Biochemistry Unit, IFOM-IEO Campus, Via Adamello 16, 20139 Milano, Italy.
Ami
Diletta
D
Martinelli
Chiara
C
Huang
He
H
Aliprandi
Marisa
M
Capasso
Paola
P
Francavilla
Chiara
C
Ossolengo
Giuseppe
G
de Marco
Ario
A
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000737:chemistry / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D001483:Base Sequence; D002161:Camelids, New World / Q000276:immunology*; D005346:Fibroblast Growth Factors / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007128:Immunoglobulin Fragments / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D008969:Molecular Sequence Data; D051496:Receptor, Fibroblast Growth Factor, Type 1 / Q000276:immunology / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D016415:Sequence Alignment; D020349:Surface Plasmon Resonance
D000906:Antibodies; D007128:Immunoglobulin Fragments; D007143:Immunoglobulin Heavy Chains; D011994:Recombinant Proteins; D005346:Fibroblast Growth Factors; C496348:FGFR1 protein, human; D051496:Receptor, Fibroblast Growth Factor, Type 1