Membrane affinity concentration immunoassay

A method for determining the presence and/or concentration of an analyte in a biological fluid sample, comprising:

• conjugating a first reactant, which specifically binds with said analyte, to a monomer which comprises N-alkylacrylamides, N-arylacrylamides, alkyl acrylates or aryl acrylates, and combinations thereof to form a monomer/reactant conjugate;
• copolymerizing said monomer/reactant conjugate with additional monomers to yield a copolymer/reactant conjugate;
• conjugating a second reactant, which specifically binds with said analyte, to a reporter to form a reporter/reactant conjugate;
• admixing in solution in the substantial absence of unconjugated polymer said copolymer/reactant conjugate, reporter/reactant conjugate, and said biological fluid sample suspected of containing said analyte to form a reaction mixture, such that specific binding occurs between said first and second reactants and said analyte, thereby forming a ternary complex;
• contacting the reaction mixture with a solid phase comprising cellulose acetate or cellulose esters which selectively binds by noncovalent means said polymer/reactant conjugate and said ternary complex, thereby removing said complex from the reaction mixture;
• and
• measuring the amount of reporter activity in the bound complex or in solution and therefrom determining the presence and/or concentration of said analyte.

A method for determining the presence and/or concentration of an analyte in a biological fluid sample, comprising:

• in the substantial absence of unconjugated polymer, contacting a polymer/reactant conjugate, which specifically binds with said analyte and wherein said polymer is comprised of monomers selected from N-alkylacrylamides, N-arylacrylamides, alkyl acrylates and aryl acrylates, and combinations thereof, with a nonhydrophobic solid phase comprising cellulose acetate or a cellulose ester which binds by noncovalent means with said polymer/reactant;
• contacting the polymer/reactant-solid phase with a biological fluid sample suspected of containing said analyte such that specific binding occurs between said polymer/reactant and said analyte;
• contacting said solid phase having the polymer/reactant/analyte complex bound thereto with a reporter/reactant conjugate which specifically binds with said analyte, such that specific binding occurs between said reporter/reactant and said analyte;
• and
• measuring the amount of reporter activity in the bound complex, and therefrom determining the presence and/or concentration of said analyte.

A method for determining the presence and/or concentration of an analyte in a biological fluid sample, comprising:

• conjugating a first reactant to a polymer comprising monomers selected from N-alkylacrylamides, N-arylacrylamides, alkyl acrylates, aryl acrylates, and combinations thereof, to form a polymer/reactant conjugate;
• conjugating a second reactant to a reporter to form a reporter/reactant conjugate, said second reactant which specifically binds with said analyte and said first reactant;
• admixing in solution in the substantial absence of unconjugated polymer said polymer/reactant conjugate, reporter/reactant conjugate and said biological fluid sample suspected of containing said analyte to form a reaction mixture, such that competitive binding occurs between said polymer/reactant conjugate and said analyte for said second reactant;
• contacting said reaction mixture with a solid phase comprising cellulose acetate or an ester of cellulose, which selectively binds by noncovalent means the polymer/reactant conjugate and the polymer/reactant-reporter/reactant complex, thereby removing said complex and said polymer/reactant from the reaction mixture;
• and
• measuring the amount of reporter activity in the bound complex or in the solution and therefrom determining the presence and/or concentration of the analyte.

A method for conducting multiple analyses on a single biological fluid sample suspected of containing one or more analytes, comprising:

• conjugating a plurality of selected first reactants which specifically bind with a different analyte to a polymer comprising monomers selected from N-alkylacrylamides, N-arylacrylamides, alkyl acrylates, aryl acrylates or combinations thereof;
• conjugating a plurality of selected second reactants which specifically bind with a different analyte to one or more selected reporters to form multiple reporter/reactant conjugates;
• admixing in solution in the substantial absence of unconjugated polymer the multiple polymer/reactant, multiple reporter/reactant, and the biological fluid sample suspected of containing one or more analytes, such that specific binding occurs between the reactants and the analytes, thereby forming a plurality of ternary complexes;
• contacting the admixed polymer/reactant conjugate, reporter/reactant conjugate, and analyte containing solution with a solid phase comprising cellulose acetate or a cellulose ester which selectively binds by noncovalent means the multiple polymer/reactant conjugates and the ternary complexes, thereby removing the complexes from the reaction mixture;
• and
• measuring the activity of each selected reporter associated with the bound complexes or in the solution and therefrom determining the presence and/or concentration of each of the analytes.

A method for conducting multiple analyses on a biological fluid sample suspected of containing one or more analytes, comprising:

• conjugating a plurality of selected first reactants which bind to different analytes to a plurality of polymers comprising monomers selected N-alkylacrylamides, N-arylacrylamides, alkyl acrylates, aryl acrylates or combinations thereof, having varied specific affinities for a non-hydrophobic solid phase comprising cellulose acetate or esters of cellulose to form multiple polymer/reactant conjugates, each of said first reactants which specifically binds with one of the analytes;
• conjugating a plurality of selected second reactants which specifically bind with different analytes to one or more reporters to form multiple reporter/reactant conjugates;
• admixing in solution in the substantial absence of unconjugated polymer the multiple polymer/reactant conjugate, multiple reporter/reactant conjugates, and the biological fluid sample suspected of containing one or more analytes, such that specific binding occurs between the reactants and the analytes, thereby forming a plurality of ternary complexes;
• contacting the admixed polymer/reactant conjugate, reporter/reactant conjugates, and analyte sample-containing solution with the solid phase which selectively binds by noncovalent means the unbound multiple polymer/reactants and the ternary complexes, thereby removing the complexes from the reaction mixture;
• selectively eluting said ternary complexes from the solid phase;
• and
• measuring the reporter activity associated with each of the eluted complexes, and therefrom determining the presence and/or concentration of each of the analytes.

A method for determining the presence and/or concentration of an analyte in a biological fluid sample, comprising:

• conjugating a first reactant which specifically binds the analyte to a polymer comprising monomers selected from N-alkylacrylamides, N-arylacrylamides, alkyl acrylates, aryl acrylates or combinations thereof, to form a polymer/reactant conjugate;
• conjugating a second reactant to a reporter to form a reporter/reactant conjugate, said second reactant being a competitor for specific binding to the first reactant with the analyte;
• admixing in solution in the substantial absence of unconjugated polymer said polymer/reactant conjugate, reporter/reactant conjugate and said biological fluid sample suspected of containing said analyte to form a reaction mixture, such that competitive binding occurs between said reporter/reactant conjugate and said analyte for said first reactant;
• contacting said reaction mixture with a solid phase comprising cellulose acetate or an ester of cellulose which selectively binds by noncovalent means the polymer/reactant-reporter/reactant complexes and the polymer/reactant conjugate from the reaction mixture to form a resultant solution;
• immobilizing a third reactant to a second solid phase to form a reactant-activated solid phase, said third reactant which specifically binds the reporter in the reporter/reactant conjugate;
• contacting the resultant solution with the reactant-activated solid phase, thereby removing the reporter/reactant conjugate from the resultant solution;
• and
• measuring the reporter activity in the bound reporter/reactant conjugate and therefrom determining the presence and/or concentration of the analyte.