Amplified array analysis method and system

Abstract

The present invention concerns an array-based analytical system and method having an enhanced sensitivity which allows for simple and rapid analysis of relative unmodified samples which comprises an analytical system of the type having a plurality of different first members of a specific binding pair affixed in an array thereupon, a mixture including at least one second member of a specific binding pair capable of binding to one of the first members so as to form a specific binding pair which is affixed to the support member, and a reporter system that produces a detectable signal indicative of the presence of the specific binding pair on the support member and wherein the reporter system includes an amplified reporter system that is independent of layering.

1 . An analytical system comprising: 
 a support member having a plurality of chemically active, phenolic or proteinaceous receptor sites thereupon;    an array of different first members of a specific binding pair disposed on said support, said first members being selected from the group consisting of: protein, peptide, polysaccharide, cell fragments, cells, tissue, organometallics, metal ion chelating organic ligands, and combinations thereof, said first members each being capable of binding to a complementary second member of said specific binding pair, whereby when said array is contacted with a plurality of analyte complementary second members, a plurality of specific binding pairs will be formed on said support;    a peroxidase enzyme;    a coupling agent operative to couple said enzyme to said specific binding pairs; and    a conjugate of a labeling agent and a substituted phenol substrate for said enzyme, said substrate being activatable by said enzyme so as to cause said substrate to bind to one of said chemically active receptor sites whereby said substrate and said labeling agent are immobilized upon said support.    
     
     
         2 . An analytical system as in  claim 1 , wherein said coupling agent is operative to couple said enzyme to the second member of said specific binding pair.  
     
     
         3 . An analytical system as in  claim 1 , wherein said peroxidase enzyme comprises horseradish peroxidase.  
     
     
         4 . An analytical system as in  claim 1 , wherein said substituted phenol substrate comprises tyramine.  
     
     
         5 . An analytical system as in  claim 1 , wherein said labeling agent comprises a fluorescent cyanine dye.  
     
     
         6 . An analytical system as in  claim 1 , wherein said more than one of said first members of said specific binding pair comprise protein.  
     
     
         7 . An analytical system as in  claim 1 , wherein said more than one of said first members of said specific binding pair comprise peptides.  
     
     
         8 . An analytical system as in  claim 1 , wherein said more than one of said first members of said specific binding pair comprise cells.  
     
     
         9 . An analytical system as in  claim 1 , wherein said more than one of said first members of said specific binding pair comprise cell fragments.  
     
     
         10 . An analytical system as in  claim 1 , wherein said more than one of said first members of said specific binding pair comprise tissues.  
     
     
         11 . An analytical system as in  claim 1 , wherein said more than one of said first members of said specific binding pair comprise organometallic.  
     
     
         12 . An analytical system as in  claim 1 , wherein said more than one of said first members of said specific binding pair comprise metal ion chelating organic ligand.  
     
     
         13 . A commercial reagent package comprising a support member having a plurality of chemically active, phenolic or proteinaceous receptor sites thereon and an array of different first members of a specific binding pair disposed on said support, said first members being selected from the group consisting of: protein, peptide, polysaccharide, cell fragments, cells, tissue, organometallics, metal ion chelating organic ligands, DNA, cDNA, RNA, oligonucleotides and combinations thereof; a peroxidase enzyme; a coupling agent operative to couple said enzyme to said specific binding pairs; and a conjugate of a labeling agent and a substituted phenol substrate for said enzyme; and instructions for the use thereof as an analytical system for a second member capable of being bound to said first members to form said specific binding pairs.  
     
     
         14 . An analytical process for detection comprising the steps of: 
 immobilizing a first member of a specific binding pair on a substrate member having a plurality of chemically active, phenolic or proteinaceous receptor sites thereupon;    contacting said substrate with a mixture comprising a second member of said specific binding pair;    coupling an enzyme to said second member of said binding pair to form a binding pair-enzyme species;    contacting said binding pair-enzyme species with a labeled conjugate, said conjugate comprising a substrate for said enzyme, and a label;    allowing sufficient time for said labeling conjugate to react with said enzyme to form an activated conjugate that binds said plurality of receptor sites; and    detecting said label bound to one of said plurality of receptor sites.    
     
     
         15 . The process of  claim 14  wherein said first member is selected from a group consisting of: DNA, cDNA, RNA, oligonucleotides, protein, peptides, polysaccharides, cell fragments, cells, tissue, organometallics and metal ion chelating organic ligands.  
     
     
         16 . The process of  claim 14  wherein said enzyme is a peroxidase.  
     
     
         17 . The process of  claim 14  wherein said label is fluorescent, haptenic, radioactive, chemically reactive or color-forming.  
     
     
         18 . The process of  claim 14  wherein said enzyme is coupled to said second member prior to formation of said specific binding pair.