Patent ID: 8318509

Claim:
A method of performing a two-phase optical assay of one or more than one analyte without intrinsic optical contrast in a sample, comprising: (a) providing functionalized microparticles, the functionalized microparticles each comprising immobilized two or more than two functional components, wherein the two or more than two functional components are selected from the group consisting of: (i) a first ligand; (ii) a color forming substrate; (iii) an enzyme for generating an intermediate reagent; and (iv) a fluorochrome, and wherein at least one of the two or more than two functional components is the first ligand; (b) providing a buffer solution and one or more than one functional component, the one or more than one functional component being the same or different from the two or more than two functional components immobilized to the microparticle in step (a) and being selected from the group consisting of: (i) an optical signal amplifier attached to a second ligand; (ii) a color forming substrate; (iii) a co-substrate; (iv) a blocked substrate or a blocked coupler; (v) a precursor substrate; (vi) a coupler; (vii) an enzyme for generating an intermediate reagent; and (viii) a fluorochrome; (c) admixing a sample comprising one or more than one analyte without intrinsic optical contrast with the buffer solution, the one or more than one functional component, and the functionalized microparticles in a container to form a two-phase suspension with the functionalized microparticles suspended in the buffer solution; (d) allowing the one or more than one analyte to bind the first ligand and/or the second ligand attached to the optical signal amplifier and to develop an optical signal in the microparticles; (e) measuring the optical signal of the two-phase suspension having the functionalized microparticles suspended in the buffer to obtain a first measurement; (f) allowing the functionalized microparticles to settle to the bottom of the container to obtain a microparticle-rich phase at the bottom of the container and a microparticle-free phase above the microparticle-rich phase; (g) measuring the optical signal of the microparticle-rich phase to obtain a second measurement; and (h) calculating an algorithmic relationship between the first and second measurements and comparing the calculated algorithmic relationship to a standard curve to determine the concentration of the one or more than one analyte.