Utilization of the biotin/avidin system to amplify the sensitivity of the enzyme-linked immunosorbent assay (ELISA).
The biotin/avidin system was incorporated into the enzyme-linked immunosorbent assay (ELISA) technique to increase the sensitivity of the standard ELISA for the detection of mouse antibody to hepatitis B surface antigen ((anti-HBs and HBsAg, respectively). Two biotin/avidin ELISA designs were studied. In both assays, 96-well polystyrene plates were coated with HBsAg, post-coated with 0.5% gelatin and incubated with dilutions of mouse anti-HBs. In the biotin/avidin (BA) ELISA, reagents were added to antibody reacted wells in the following sequence: biotinylated goat anti-mouse IgG (b-GAMG), avidin-alkaline phosphatase (Av-AP) and substrate. The order of reactants after mouse antibody in the biotin/avidin/biotin (BAB) ELISA was b-GAMG, avidin, biotinylated alkaline phosphatase (b-AP) and substrate. The sensitivities of BA ELISA, BAB ELISA and a standard ELISA using a glutaraldehyde conjugated goat anti-mouse enzyme were compared to AUSAB (a commercial radioimmunoassay) using a panel of 23 mouse anti-HBs sera. All 3 ELISAs were more sensitive than AUSAB; the standard ELISA, BAB ELISA and BA ELISA were respectively 50, 1173 and 4134 times more sensitive than AUSAB for detection of mouse anti-HBs activity.
D000818:Animals; D000914:Antibodies, Viral / Q000032:analysis*; D001360:Avidin / Q000378:metabolism*; D001710:Biotin / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006041:Goats; D006510:Hepatitis B Antibodies / Q000032:analysis*; D006514:Hepatitis B Surface Antigens / Q000276:immunology; D006801:Humans; D051379:Mice; D010047:Ovalbumin / Q000031:analogs & derivatives*; D011863:Radioimmunoassay
D000914:Antibodies, Viral; D006510:Hepatitis B Antibodies; D006514:Hepatitis B Surface Antigens; D001360:Avidin; D001710:Biotin; D010047:Ovalbumin