Aptamer-Based Multiplexed Assays
AssigneeSomaLogic, Inc.
InventorsGlenn Sanders, Stephan Kraemer, Evaldas Katilius, EDGAR OLIVAS
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Abstract
The present disclosure describes methods, devices, reagents, and kits for the detection of one or more target molecules that may be present in a test sample. The described methods, devices, kits, and reagents facilitate the detection and quantification of a non-nucleic acid target (e.g., a protein target) in a test sample by detecting and quantifying a nucleic acid (i.e., an aptamer) where the aptamer-aptamer interactions are significantly reduced or eliminated while maintaining the aptamer-target interaction.
Claims32
A method for detecting a target molecule that may be present in a test sample comprising:
- exposing an aptamer having specific affinity for the target molecule to a first solid support, wherein the aptamer comprises a first tag and the first solid support comprises a first capture element, and wherein the first tag has affinity for the first capture element;
- allowing the first tag to interact with the first capture element to immobilize the aptamer on the first solid support;
- washing the first solid support with one or more solutions that dissociate aggregated aptamers;
- contacting the immobilized aptamer with a test sample, wherein an aptamer-target affinity complex is formed if the target molecule is present in the test sample;
- removing one or more components not immobilized on the first solid support;
- attaching a second tag to the target molecule in the aptamer-target affinity complex, wherein the second tag has an affinity to a second capture element;
- releasing the aptamer-target affinity complex from said first solid support;
- exposing the released aptamer-target affinity complex to a second solid support comprising a second capture element and allowing the second tag to interact with said second capture element to immobilize the aptamer-target affinity complex on the second solid support;
- removing one or more components not immobilized on the second solid support eluting the aptamer from the second solid support with one or more buffered solutions comprising a chaotropic salt;
- detecting the presence of said target molecule in the test sample by detecting the aptamer of said aptamer-target affinity complex;
- and
- wherein the target molecule is a protein.
The method of 1, wherein the aptamer comprises a detectable moiety is selected from the group consisting of a dye, a quantum dot, a radiolabel, an electrochemical functional group, and an enzyme plus a detectable enzyme substrate.
A kit comprising:
- a) one or more aptamers, wherein each of the one or more aptamers has specific binding affinity for one or more targets;
- b) one or more solid supports;
- c) one or more partitioning reagents;
- d) one or more reagents for the release of an aptamer from an aptamer-target affinity complex;
- e) one or more buffer solutions comprising an organic solvent;
- and
- f) one or more buffer solutions comprising a chaotropic salt.
A method comprising:
- contacting an aptamer having a specific affinity for a target molecule with a test sample, wherein an aptamer-target affinity complex is formed if the target molecule is present in the test sample, and wherein the aptamer is immobilized on a first solid support and washed with one or more solutions that dissociate aggregated aptamers;
- removing one or more components not immobilized on the first solid support;
- attaching a second tag to the target molecule in the aptamer-target affinity complex, wherein the second tag has an affinity to a second capture element;
- releasing the aptamer-target affinity complex from said first solid support;
- exposing the released aptamer-target affinity complex to a second solid support comprising a second capture element and allowing the second tag to interact with said second capture element to immobilize the aptamer-target affinity complex on the second solid support;
- removing one or more components not immobilized on the second solid support;
- and
- eluting the aptamer from the second solid support with one or more solutions comprising a chaotropic salt;
- wherein, the target molecule is a protein.
Dependencies are inferred from the claim text.