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US 2023/0407371 A1Published application
Multiplexed Imaging Using Strand Displacement
AssigneeUltivue, Inc.
InventorsXi Chen, Stephanie Hennek
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Abstract
The present disclosure describes various improved methods for imaging at least one target in a sample, including methods employing an adapter strand oligonucleotide and a bridge strand oligonucleotide. Some methods also employ bouncer oligonucleotides and/or blocker oligonucleotides. Some methods also use two partial docking strands to detect proximity of the partial docking strands to each other.
Claims21
A method for imaging of at least two targets in a sample comprising:
- a. providing at least two target-recognizing antibodies bound to a docking strand oligonucleotide;
- b. providing a first imager strand oligonucleotide capable of hybridizing to the docking strand on the first target-recognizing antibody and providing a first blocking strand capable of hybridizing to the docking strand on the second target-recognizing antibody, wherein the imager strand is labeled with a detectable label and the blocking strand is not labeled with a detectable label and wherein optionally additional blocking strands are used if more than two target-recognizing antibodies are present;
- c. imaging the first target and removing the imager strand and blocking strand with at least one bouncer strand;
- d. providing a second imager strand oligonucleotide capable of hybridizing to the docking strand on the second target-recognizing antibody and providing a second blocking strand capable of hybridizing to the docking strand on the first target-recognizing antibody, wherein the imager strand is labeled with a detectable label and the blocking strand is not labeled with a detectable label and wherein optionally additional blocking strands are used if more than two target-recognizing antibodies are present;
- e. imaging the second target;
- f. optionally removing the imager strand and blocking strand with at least one bouncer strand;
- and
- g. optionally repeating steps b-f for any additional targets.
5 more elements
A method for imaging of at least one target in a sample comprising:
- a. providing at least one target-recognizing antibody bound to an oligonucleotide complementary to a docking strand but not over the full length, wherein a portion of docking strand that is not complementary to the nucleic acid sequence comprises a constant domain and a target-specific domain;
- b. providing a blocker strand that hybridizes to the docking strand, wherein the blocker strand comprises a constant domain and a target-specific domain that are complementary to the constant domain and the target-specific domain of the docking strand respectively,
- c. providing an imager strand that hybridizes to the docking strand oligonucleotide at least in the region of the target-specific region, wherein the imager strand is labeled with a detectable label and allowing the imager strand to hybridize to the docking strand, displacing the blocker strand;
- d. optionally providing a bouncer strand that hybridizes to the constant domain of the blocker domain,
- e. optionally, if multiple targets are being imaged, providing blocker strands for all other targets, excluding the blocker strand containing the target-specific domain used in step b;
- f. imaging the at least one target;
- g. optionally removing the imager strand from the docking strand by providing the blocker strand used in step b;
- h. optionally repeating steps a-g or any subset thereof if multiple targets are being imaged.
5 more elements
Dependencies are inferred from the claim text.