Bionanocapsule-based enzyme-antibody conjugates for enzyme-linked immunosorbent assay.
Macromolecules that can assemble a large number of enzyme and antibody molecules have been used frequently for improvement of sensitivities in enzyme-linked immunosorbent assays (ELISAs). We generated bionanocapsules (BNCs) of approximately 30nm displaying immunoglobulin G (IgG) Fc-binding ZZ domains derived from Staphylococcus aureus protein A (designated as ZZ-BNC). In the conventional ELISA using primary antibody and horseradish peroxidase-labeled secondary antibody for detecting antigen on the solid phase, ZZ-BNCs in the aqueous phase gave an approximately 10-fold higher signal. In Western blot analysis, the mixture of ZZ-BNCs with secondary antibody gave an approximately 50-fold higher signal than that without ZZ-BNCs. These results suggest that a large number of secondary antibody molecules are immobilized on the surface of ZZ-BNCs and attached to antigen, leading to the significant enhancement of sensitivity. In combination with the avidin-biotin complex system, biotinylated ZZ-BNCs showed more significant signal enhancement in ELISA and Western blot analysis. Thus, ZZ-BNC is expected to increase the performance of various conventional immunoassays.
Iijima
Masumi
M
Institute of Scientific and Industrial Research, Osaka University, Ibaraki, Osaka 567-0047, Japan.
Matsuzaki
Takashi
T
Kadoya
Hiroyasu
H
Hatahira
Satoko
S
Hiramatsu
Shingo
S
Jung
Giman
G
Tanizawa
Katsuyuki
K
Kuroda
Shun'ichi
S
D000906:Antibodies / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D001360:Avidin / Q000378:metabolism; D001710:Biotin / Q000378:metabolism; D015153:Blotting, Western / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D055767:Immobilized Proteins / Q000737:chemistry / Q000378:metabolism; D007141:Immunoglobulin Fc Fragments / Q000378:metabolism; D053769:Nanocapsules / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D013205:Staphylococcal Protein A / Q000737:chemistry* / Q000378:metabolism
D000906:Antibodies; D055767:Immobilized Proteins; D007141:Immunoglobulin Fc Fragments; D053769:Nanocapsules; D013205:Staphylococcal Protein A; D001360:Avidin; D001710:Biotin