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US 11,761,955 B2Granted
Multiplexed catalyzed reporter deposition
AssigneeUltivue, Inc.
InventorsMark Bobrow, Stephanie Rae Hennek, Mael Manesse
Life of this patent
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Abstract
A method for testing a sample for the presence of one or more targets comprises multiplexed catalyzed reporter deposition (CARD) is provided.
Claims14
A method for testing a sample for the presence of one or more targets comprising:
- (1) contacting a sample being tested for the presence of one or more targets with one or more target-specific binding partners, wherein each target-specific binding partner is linked to a nucleic acid strand and wherein target-specific binding partners of different specificity are linked to different nucleic acid strands;
- (2) optionally removing unbound target-specific binding partners;
- (3) contacting the sample from step (1) or optionally step (2) with an enzyme linked to a nucleic acid strand complementary to the nucleic acid strand linked to the target-specific binding partner
- (4) optionally removing unbound enzyme linked to complementary nucleic acid strands;
- (5) contacting the sample from step (3) or optionally step (4) with a substrate conjugate comprised of a nucleic acid strand-bound substrate, where the substrate and nucleic acid strand are linked optionally with a first releasable linker;
- (6) optionally removing unbound substrate conjugates;
- (7) optionally deactivating the bound enzymes or releasing the bound enzymes;
- (8) contacting the sample from step (5) or optionally steps (6)-(7) with a nucleic acid strand bound detectable label, linked optionally with a second releasable linker, where the nucleic acid strand is a specific binding pair member to the nucleic acid strand in step (5), and wherein either or both the first and second releasable linkers are present;
- (9) imaging the sample to detect the bound labels;
- (10) releasing the bound label by releasing the first releasable linker or the second releasable linker or both;
- and
- (11) optionally repeating steps (1)-(10) or any subset thereof and optionally if more than one target-specific binding partner is used, repeating steps (1)-(8) or any subset thereof prior to step (9) of imaging.
9 more elements
A method for testing a sample for the presence of one or more targets comprising:
- (1) contacting a sample being tested for the presence of one or more targets with one or more target-specific binding partners, wherein each target-specific binding partner is linked to a nucleic acid strand and wherein target-specific binding partners of different specificity are linked to different nucleic acid strands;
- (2) optionally removing unbound target-specific binding partners;
- (3) contacting the sample from step (1) or optionally step (2) with an enzyme linked to a nucleic acid strand complementary to the nucleic acid strand linked to a first target-specific binding partner;
- (4) optionally removing unbound enzyme linked to complementary nucleic acid strands;
- (5) contacting the sample from step (3) or optionally step (4) with a substrate conjugate comprised of a nucleic acid strand-bound substrate, where the substrate and nucleic acid strand are linked with a first releasable linker;
- (6) optionally removing unbound substrate conjugates;
- (7) deactivating the bound enzymes or releasing the bound enzymes;
- (8) optionally repeating steps (3)-(7) for a next target-specific binding partner, resulting in two or more distinct substrate conjugates bound to the sample;
- (9) contacting the sample from step (8) with one or more nucleic acid strand bound detectable labels, wherein each nucleic acid strand and detectable label are linked with a second releasable linker, and each nucleic acid stand is a specific binding pair member to the nucleic acid strand of the substrate conjugate specific for the same target-specific binding partner,
- (10) imaging the sample to detect the bound labels;
- (11) optionally releasing the bound labels from step (8) by releasing the first releasable linker or the second releasable linker or both;
- (12) optionally repeating steps (1)-(11) or any subset thereof.
9 more elements
Dependencies are inferred from the claim text.