Fluoroimmunoassaying

Claim 1

A method of enhancing the fluorescent level of a fluoroimmunoassay, comprising the steps of: (a) tagging a long-chain carrier selected from proteins, cellulose, cellulose derivatives, polymers or polysaacharides, said carrier having fluorophore binding sites and capable of treatment to release fluorophores bound thereto, with a plurality of fluorophores, said fluorophores selected from the group consisting of fluorescein, dansyl, rhodamine, fluorescamine, pyrene, acradine, 2-methoxy-2,4-diphenyl-3(2H)-furanone, and fluorescein mercuric acetate until a critical spacing distance of less than 100 Å is reached between said fluorophores to induce self-quenching therebetween; (b) conjugating one reactant of said fluoro-immunoassay with said fluorescently-quenched carrier; (c) reacting said conjugated reactant in a known amount with a competing immunoreactant sample and a known amount of a complementary-reactant-containing solid phase binding agent to form a mixture of bound and unbound reaction products, said solid phase binding agent selected from magnetic particulate cellulose, glass, latexes, plastics, agaroses, sepharoses and sephadexes; (d) separating said bound and unbound reaction products; (e) liberating the fluorophores from said carrier following said separation in either one of the bound or unbound reaction product by breaking up the carrier molecule by a lysing or chemical treatment; and (f) measuring the enhanced fluorescent level of said liberated fluorophores.

Claim 2

A method of enhancing the fluorescent level of a fluoroimmunoassay, comprising the steps of: (a) tagging a long-chain carrier selected from proteins, cellulose, cellulose derivatives, polymers or polysaacharides, said carrier having fluorophore binding sites and capable of treatment to release fluorophores bound thereto, with a plurality of fluorophores, said fluorophores selected from the group consisting of fluorescein, dansyl, rhodamine, fluorescamine, pyrene, acradine, 2-methoxy-2,4-diphenyl-3(2H)-furanone, and fluorescein mercuric acetate until a critical spacing distance of less than 100 Å is reached between said fluorophores to induce self-quenching therebetween; (b) conjugating one reactant of said fluoro-immunoassay with said fluorescently-quenched carrier; (c) reacting said conjugated reactant in a known amount with a competing immunoreactant sample and a known amount of a complementary-reactant-containing solid phase binding agent to form a mixture of bound and unbound reaction products, said solid phase binding agent selected from magnetic particulate cellulose, glass, latexes, plastics, agaroses, sepharoses and sephadexes; (d) separating said bound and unbound reaction products; (e) liberating the fluorophores from said carrier following said separation in either one of the bound or unbound reaction product by breaking up the carrier molecule by a lysing or chemical treatment; and (f) measuring the enhanced fluorescent level of said liberated fluorophores.

Claim 6

A method of fluoroimmunoassaying a fluid sample for an immunological reactant, comprising the steps of: (a) forming a mixture of a sample containing an immunological reactant to be assayed with a known amount of a fluorescently self-quenched, fluorophore tagged competing immunological reactant wherein the spacing distance between fluorophores is less than 100 Å, and a known amount of a complementary-reactant-containing solid phase binding agent that equilibrates with both said sample reactant and said competing reactant; (b) equilibrating said mixture to form a first portion comprising reactants bound to said binding agent, and a second portion comprising reactants which are not bound to said binding agent; (c) separating the mixture into said first and second portions, respectively; (d) liberating fluorophores from said tagged immunological reactant in either one of said first or second portions by a lysing or chemical treatment; and (e) measuring the fluorescent level of said liberated fluorophores.

Claim 7

A method of fluoroimmunoassaying a fluid sample for an immunological reactant, comprising the steps of: (a) forming a mixture of a sample containing an immunological reactant to be assayed with a known amount of a fluorescently self-quenched, fluorophore tagged competing immunological reactant wherein the spacing distance between fluorophores is less than 100 Å, and a known amount of a complementary-reactant-containing solid phase binding agent that equilibrates with both said sample reactant and said competing reactant; (b) equilibrating said mixture to form a first portion comprising reactants bound to said binding agent, and a second portion comprising reactants which are not bound to said binding agent; (c) separating the mixture into said first and second portions, respectively; (d) liberating fluorophores from said tagged immunological reactant in either one of said first or second portions by a lysing or chemical treatment; and (e) measuring the fluorescent level of said liberated fluorophores.