New technique for solid-phase immunoassay: application to hepatitis B surface antigen.

Abstract

I describe a new technique for improving the sensitivity of the solid-phase "sandwich" assay, by using the through-passage receptacle in a novel flow-communication device. The technique allows a large volume of serum to flow through the antibody-coated receptacle repeatedly during the incubation and is thus termed the "flow-through large-volume incubation" method. Binding of 125I-labeled hepatitis B surface antigen to its corresponding antibody on a solid-phase by this method was more rapid and persistent than binding by the conventional method. When the method was applied to the first incubation of the sandwich assay, the test for the antigen was rendered four-, eight-, and 32-fold as sensitive as an accepted third-generation test for the antigen, by incubating 5-mL volumes of serum at (a) room temperature for 18 h, (b) 45 degrees C for 8 h, or (c) room temperature for seven days, respectively.