Antibody Fab display system that can perform open-sandwich ELISA.
Previously, immunological detection of small haptens or peptides was only possible in a competitive format, which needed competitor antigen either labeled by a reporter or attached to a carrier protein. Beside this, open-sandwich immunoassay (OS-IA) is a simple but powerful immunoassay that can noncompetitively determine monovalent antigen concentration by measuring the antigen-dependent increase in V(H)/V(L) interaction of an antibody. However, the procedure to obtain suitable assay reagents for OS-IA for a target antigen has not been straightforward because of the lack of easy-to-use antibody selection/manipulation methods. Here we devise a new Fab antibody phage display system that is useful for rapidly evaluating and selecting suitable antibody Fv fragments to OS-IA. The system is based on a phagemid vector in which two identical restriction sites were incorporated into both ends of a human constant region domain. After selection of the M13 phage displaying a Fab fragment, the vector can be easily converted to the vector that can simultaneously produce the V(H)-displaying phage and the light chain in the culture supernatant, which can be directly used for OS-ELISA. The successful results of model selection as well as conversion to OS format show the potential in developing various OS-IA for clinically and environmentally important targets.
Dong
Jinhua
J
Department of Bioengineering, School of Engineering, The University of Tokyo, Bunkyo-ku, Tokyo, Japan.
Ihara
Masaki
M
Ueda
Hiroshi
H
D000941:Antigens / Q000032:analysis*; D017104:Bacteriophage M13 / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay / Q000295:instrumentation / Q000379:methods*; D005822:Genetic Vectors; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000276:immunology*; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D008722:Methods; D012107:Research Design
D000941:Antigens; D007140:Immunoglobulin Fab Fragments; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains