Rational screening of antibodies and design of sandwich enzyme linked immunosorbant assay on the basis of a kinetic model.
A rational strategy for the rapid establishment of a sensitive sandwich enzyme linked immunosorbant assay was developed. The kinetic properties required for the solid-phase and enzyme-conjugated antibodies of sandwich ELISA were determined rationally on the basis of a kinetic model describing antibody-antigen interaction. Some antibodies possessing the required kinetic properties against a model antigen, C-reactive protein (CRP), were successfully isolated from a phage antibody library under the screening conditions that were designed on the basis of simulation results. The best combination of solid-phase and enzyme-conjugated antibodies that gives the most sensitive sandwich ELISA was determined by simulation on the basis of the apparent association and dissociation rate constants of the isolated antibodies. It was confirmed by experiment that the sandwich ELISA using the best combination of antibodies was actually the most sensitive one. Our strategy would be useful for the rapid establishment of sensitive sandwich ELISAs compared with the traditional hybridoma method in which the best condition is determined by trial and error.
Choi
Dong Hwan
DH
Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan.
Katakura
Yoshio
Y
Ninomiya
Kazuaki
K
Shioya
Suteaki
S
D000595:Amino Acid Sequence; D000906:Antibodies / Q000302:isolation & purification*; D000937:Antigen-Antibody Reactions; D002097:C-Reactive Protein / Q000276:immunology; D003198:Computer Simulation; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D007700:Kinetics; D008954:Models, Biological; D008969:Molecular Sequence Data; D019151:Peptide Library
D000906:Antibodies; D019151:Peptide Library; D002097:C-Reactive Protein