Energy absorbing particle quenching in light emitting competitive protein binding assays

Claim 1

A method for determining the presence of an analyte in a sample suspected of containing said analyte, wherein said analyte is a member of a specific binding pair (hereafter "mip"). wherein employed as reagents are: (1) a water insoluble particle of at least 50nm capable of light emission upon activation having functionalities capable of light emission distributed within said particle and a plurality of mips bound to said particle to provide a mip-bound-light emitting particle; and (2) a quencher particle of at least 50nm to which a plurality of mips are bound to provide a mip-bound-quencher particle, wherein when said mip-bound-quencher particle is bound to said mip-bound-light emitting particle through a mip bridge, there is substantial reduction in light emission of said mip-bound-light emitting particle, and wherein said mips of said analyte, mip-bound-light emitting particle and mip-bound-quencher particle include at least one complementary pair; said method comprising: in an aqueous assay medium, combining said analyte, said mip-bound-light emitting particle and said mip-bound-quencher particle, wherein the amount of mip-bound-quencher particle which becomes bound to said mip-bound-light emitting particle is related to the amount of analyte in said medium; and determining the amount of light emission in said medium as compared to an assay medium having a known amount of analyte.

Claim 2

A method for determining the presence of an analyte in a sample suspected of containing said analyte, wherein said analyte is a member of a specific binding pair (hereafter "mip"). wherein employed as reagents are: (1) a water insoluble particle of at least 50nm capable of light emission upon activation having functionalities capable of light emission distributed within said particle and a plurality of mips bound to said particle to provide a mip-bound-light emitting particle; and (2) a quencher particle of at least 50nm to which a plurality of mips are bound to provide a mip-bound-quencher particle, wherein when said mip-bound-quencher particle is bound to said mip-bound-light emitting particle through a mip bridge, there is substantial reduction in light emission of said mip-bound-light emitting particle, and wherein said mips of said analyte, mip-bound-light emitting particle and mip-bound-quencher particle include at least one complementary pair; said method comprising: in an aqueous assay medium, combining said analyte, said mip-bound-light emitting particle and said mip-bound-quencher particle, wherein the amount of mip-bound-quencher particle which becomes bound to said mip-bound-light emitting particle is related to the amount of analyte in said medium; and determining the amount of light emission in said medium as compared to an assay medium having a known amount of analyte.

Claim 5

A method according to Claims 1, 2 or 3, wherein the light emitting particle is an addition polymer fluorescent particle.

Claim 6

A method according to any one of the preceding Claims, wherein said analyte is a protein.

Claim 7

A method according to any one of the preceding Claims, wherein the light emitting particle is a fluorescent particle which absorbs light of wavelength greater than about 350nm and emits light of wavelength greater than about 400nm and is different from the quencher particle.

Claim 8

A method for determining the presence of an analyte in a sample suspected of containing said analyte, wherein said analyte is a member of a specific binding pair-mip-, wherein employed as reagents are: (1) a fluorescent water insoluble particle of a size in the range of about 100nm to 20µ and having fluorescent functionalities substantially uniformly distributed within said particle and a plurality of mips bound to said particle to provide a mip-bound-fluorescent particle; and (2) a carbon quencher particle to which a plurality of mips are bound to provide a mip-bound-quencher particle, wherein when said mip-bound-quencher particle is bound to said mip-bound-fluorescent particle through a mip bridge, there is substantial reduction of fluorescence of said mip-bound-fluorescent particle, and wherein said mips of said analyte, mip-bound-fluorescent particle and mip-bound-quencher particle include at least one complementary pair: said method comprising: in an aqueous assay medium, combining said analyte, said mip-bound-fluorescent particle and said mip-bound-quencher particle, wherein the amount of mip-bound-quencher particle which becomes bound to said mip-bound-fluorescent particle is related to the amount of analyte in said medium; and determining the amount of fluorescence in said medium as compared to an assay medium having a known amount of analyte.

Claim 9

A method for determining the presence of an analyte in a sample suspected of containing said analyte, wherein said analyte is a member of a specific binding pair-mip-, wherein employed as reagents are: (1) a fluorescent water insoluble particle of a size in the range of about 100nm to 20µ and having fluorescent functionalities substantially uniformly distributed within said particle and a plurality of mips bound to said particle to provide a mip-bound-fluorescent particle; and (2) a carbon quencher particle to which a plurality of mips are bound to provide a mip-bound-quencher particle, wherein when said mip-bound-quencher particle is bound to said mip-bound-fluorescent particle through a mip bridge, there is substantial reduction of fluorescence of said mip-bound-fluorescent particle, and wherein said mips of said analyte, mip-bound-fluorescent particle and mip-bound-quencher particle include at least one complementary pair: said method comprising: in an aqueous assay medium, combining said analyte, said mip-bound-fluorescent particle and said mip-bound-quencher particle, wherein the amount of mip-bound-quencher particle which becomes bound to said mip-bound-fluorescent particle is related to the amount of analyte in said medium; and determining the amount of fluorescence in said medium as compared to an assay medium having a known amount of analyte.

Claim 12

A method for determining the presence of an analyte in a sample suspected of containing said analyte wherein said analyte is a member of a specific binding pair-mip-, wherein employed as a reagent is a fluorescent water insoluble particle of a size in the range of about 100nm to 20p and having fluorescent functionalities substantially uniformly distributed within said particle and a plurality of mips bound to said particle to provide a mip-bound-fluorescent particle, wherein when two or more of said mip-bound-fluorescent particles are brought together through mip bridges by binding to analyte or as a result of having complementary mips on mip-bound-fluorescent particles, there is a substantial diminution in fluorescence, and wherein said mips of said analyte and mip-bound-fluorescent particles include at least one complementary pair; said method comprising: in an aqueous assay medium, combining said analyte and said mip-bound-fluorescer particles, wherein the amount of mip-bound-fluorescer particles which are brought into close proximity by mip bridges is related to the amount of analyte in said medium; and determining the amount of fluorescence in said medium as compared to an assay medium having a known amount of analyte.

Claim 13

A method for determining the presence of an analyte in a sample suspected of containing said analyte wherein said analyte is a member of a specific binding pair-mip-, wherein employed as a reagent is a fluorescent water insoluble particle of a size in the range of about 100nm to 20p and having fluorescent functionalities substantially uniformly distributed within said particle and a plurality of mips bound to said particle to provide a mip-bound-fluorescent particle, wherein when two or more of said mip-bound-fluorescent particles are brought together through mip bridges by binding to analyte or as a result of having complementary mips on mip-bound-fluorescent particles, there is a substantial diminution in fluorescence, and wherein said mips of said analyte and mip-bound-fluorescent particles include at least one complementary pair; said method comprising: in an aqueous assay medium, combining said analyte and said mip-bound-fluorescer particles, wherein the amount of mip-bound-fluorescer particles which are brought into close proximity by mip bridges is related to the amount of analyte in said medium; and determining the amount of fluorescence in said medium as compared to an assay medium having a known amount of analyte.