Immunoassay element

An immunoassay element for quantitatively analyzing an antigen by determining a change in enzymatic activity caused by any of followings:

• 1) a reaction between the antigen and an enzyme-labelled antibody, the enzyme-labelled antibody being a conjugate of an antibody and a labelling enzyme, the antibody being capable to react and bind specifically with the antigen, wherein the antigen sterically hinders enzymatic activity in a specific binding complex of the antigen and the enzyme-labelled antibody;
• 2) a reaction between the antigen, an antibody and an enzyme-labelled antigen, the enzyme-labelled antigen being a conjugate of an antigen and a labelling enzyme, wherein the antibody sterically hinders enzymatic activity in a specific binding complex of the antibody and the enzyme-labelled antigen;
• and
• 3) a reaction between the antigen, an enzyme-labelled antibody and a conjugate of the antigen with a high molecular weight compound, the enzyme-labelled antibody being a conjugate of an antibody and a labelling enzyme, wherein the conjugate of the antigen with a high molecular weight compound sterically hinders enzymatic activity in a specific binding complex of the conjugate of the antigen with a high molecular weight compound and the enzyme-labelled antibody;
• said element comprising:
• a substrate layer containing a non-diffusible substrate composed of polysaccharide which is capable of being fragmented by said labelling enzyme into a diffusible glucose oligomer which migrates from said substrate layer;
• and
• a reagent layer containing a fragmenting enzyme for further fragmenting said diffusible glucose oligomer, which has migrated from said substrate layer, into a detectable glucose monomer, said substrate layer being superposed on said reagent layer;
• wherein said labelling enzyme is α-amylase, and said fragmenting enzyme is glucoamylase or α-glucosidase;
• and
• wherein said non-diffusible substrate is carboxylmethylated starch restrictively decomposed in advance with an exo-active glucosidase from the non-reducing terminal glucose site through the carboxylmethyl-modified glucose unit site or the glucose chain branching site so that said non-diffusible substrate reacts solely with said labelling enzyme and avoids reacting with said fragmenting enzyme.