METHODS FOR DETERMINING THE CONCENTRATION OF AN ANALYTE IN SOLUTION
First Claim
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1. A method of determining the concentration of an analyte in a fluid sample to be tested, the method comprising the steps of:
- (a) partitioning at least a portion of the analyte molecules in the fluid sample across a plurality of reaction vessels so that a statistically significant fraction of the reaction vessels contain analyte and a statistically significant fraction of the reaction vessels contain no analyte;
(b) determining the presence or absence of the analyte in each reaction vessel to identify the number of reaction vessels that contain analyte and/or to identify the number of reaction vessels that contain no analyte; and
(c) determining the concentration of the analyte in the fluid sample from the number of reaction vessels that contain the analyte.
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Abstract
Disclosed is a method for measuring the concentration of an analyte or analytes in a solution. Although the methods can be conducted using a number of different assay formats, in one embodiment, the assays are conducted in reaction vessels defined, at least in part, by the distal ends of fiber optic strands.
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Citations
51 Claims
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1. A method of determining the concentration of an analyte in a fluid sample to be tested, the method comprising the steps of:
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(a) partitioning at least a portion of the analyte molecules in the fluid sample across a plurality of reaction vessels so that a statistically significant fraction of the reaction vessels contain analyte and a statistically significant fraction of the reaction vessels contain no analyte; (b) determining the presence or absence of the analyte in each reaction vessel to identify the number of reaction vessels that contain analyte and/or to identify the number of reaction vessels that contain no analyte; and (c) determining the concentration of the analyte in the fluid sample from the number of reaction vessels that contain the analyte. - View Dependent Claims (39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51)
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2-11. -11. (canceled)
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12. A method of determining the concentration of an analyte in a fluid sample to be tested, the method comprising the steps of:
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(a) exposing the fluid sample to a plurality of reaction vessels under conditions so that analyte is captured in a statistically significant fraction of the reaction vessels, wherein each reaction vessel comprises a microwell and each reaction vessel defines a binding surface that has a capture component immobilized thereon; (b) determining the presence or absence of the analyte in each reaction vessel so as to identify the number of reaction vessels that contain captured analyte and/or the number of reaction vessels that do not contain captured analyte; and (c) determining the concentration of the analyte in the fluid sample to be tested from the number of reaction vessels that contain and/or do not contain the analyte.
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13-16. -16. (canceled)
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17. A method of determining the concentration of an analyte in a fluid sample to be tested, the method comprising the steps of:
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(a) partitioning at least a portion of the analyte molecules in the fluid sample into a plurality of reaction vessels, so that, for substantially all of the reaction vessels, each reaction vessel contains either no analyte or a single molecule of the analyte; (b) determining the presence or absence of the analyte in each reaction vessel to provide a number of the reaction vessels that contain the analyte; and (c) determining the concentration of the analyte in the fluid sample from the number of reaction vessels that contain the analyte.
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18-20. -20. (canceled)
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21. A method of determining the concentration of an analyte in a fluid sample to be tested, the method comprising the steps of:
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(a) partitioning the sample to be tested into a plurality of second, smaller fluid samples of equal volume so that a statistically significant fraction of the second, smaller fluid samples contain either a single molecule of the analyte or no analyte; (b) determining the presence or absence of the analyte in each of the second samples so as to identify the number of second samples that contain the analyte; and (c) determining the concentration of the analyte in the sample to be tested from the number of second samples that contain the analyte.
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22-35. -35. (canceled)
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36. A method of detecting analyte molecules or particles in a fluid sample, comprising:
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partitioning a fluid sample containing or suspected of containing analyte molecules or particles across an array comprising a plurality of reaction vessels, so that at least some of the reaction vessels contain no analyte molecules or particles and at least some of the reaction vessels contain at least one analyte molecule or particle; immobilizing within each reaction vessel containing at least one analyte molecule or particle, a first binding ligand comprising a label; applying an enzyme to the array and capturing the enzyme with the label; contacting the enzyme with an enzymatic substrate, wherein the enzymatic substrate produces a detectable product upon contact with the enzyme; detecting the detectable product; and determining from detecting the presence of the detectable product the number of reaction vessels which contain an analyte molecule or particle.
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37. A method of detecting analyte molecules or particles in a fluid, comprising:
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partitioning a fluid containing or suspected of containing analyte molecules or particles across an array comprising a plurality of reaction vessels, so that at least some of the reaction vessels contain no analyte molecules or particles and at least some of the reaction vessels contain at least one analyte molecule or particle; exposing the reaction vessels to an enzymatic substrate, wherein the enzymatic substrate produces a detectable product in reaction vessels containing at least one analyte molecule or particle; detecting the detectable product; and determining from detecting the presence of the detectable product the number of reaction vessels which contain at least one analyte molecule or particle.
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38. A method of detecting analyte molecules or particles in a fluid, comprising:
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immobilizing with respect to each analyte molecules or particles, an enzyme molecule; partitioning the enzyme molecules across an array comprising a plurality of reaction vessels, so that at least some of the reaction vessels contain no enzyme molecules and at least some of the reaction vessels contain at least one enzyme molecule; exposing the reaction vessels to an enzymatic substrate, wherein the enzymatic substrate produces a detectable product in reaction vessels containing at least one enzyme molecule; and determining from detecting the presence of the detectable product the number of reaction vessels which contain at least one enzyme molecule.
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Specification