Affinity reagents having enhanced binding and detection characteristics

Patent metadata

Field	Value
Publication number	US-11993807-B2
Application number	US-202318179924-A
Country	US
Kind code	B2
Filing date	Mar 7, 2023
Priority date	Nov 11, 2020
Publication date	May 28, 2024
Grant date	May 28, 2024

Abstract

An affinity reagent, having: (a) a retaining component such as a structured nucleic acid particle; and (b) one or both of (i) one or more label components attached to the retaining component, and (ii) one or more binding components attached to the retaining component.

First claim

What is claimed is:

1. A method of detecting an analyte, comprising:
(a) contacting an array of analytes with a detectable probe, wherein the array of analytes comprises individual analytes each attached to sites that are physically isolated from all other analytes in the array of analytes,
wherein the detectable probe comprises:
(i) a retaining component comprising nucleic acid origami,
(ii) two or more label components attached to the retaining component, and
(iii) two or more binding components attached to the retaining component, wherein the two or more binding components recognize the same epitope of the analyte,

wherein the detectable probe binds to the analyte via non-covalent binding of at least one of the two or more binding components to the epitope; and
(b) acquiring signals from the two or more label components of the detectable probe bound to the analyte, thereby detecting the analyte.

2. The method of claim 1 , wherein the nucleic acid origami comprises a scaffold nucleic acid and a staple nucleic acid hybridized to two regions of the scaffold nucleic acid, wherein the two regions of the scaffold nucleic acid are separated by an intervening region that does not hybridize to the staple nucleic acid.

3. The method of claim 2 , wherein the scaffold nucleic acid comprises a circular nucleic acid.

4. The method of claim 2 , wherein the nucleic acid origami comprises a plurality of staple nucleic acids hybridized to the scaffold nucleic acid.

5. The method of claim 1 , wherein the two or more label components comprise fluorophores that emit fluorescence at the same wavelength, and wherein the signals comprise the fluorescence.

6. The method of claim 5 , wherein the retaining component maintains the at least two label components at a spacing of at least 15 nm.

7. The method of claim 1 , wherein the analyte is attached to the site via a structured nucleic acid particle.

8. The method of claim 1 , wherein the analyte comprises a polypeptide.

9. The method of claim 8 , wherein the epitope is a trimer amino acid sequence.

10. The method of claim 8 , wherein the epitope is a tetramer amino acid sequence.

11. The method of claim 8 , wherein the polypeptide is denatured.

12. The method of claim 1 , wherein the two or more binding components comprise antibodies or functional fragments thereof.

13. The method of claim 1 , wherein the two or more binding components comprise nucleic acid aptamers.

14. The method of claim 1 , wherein the structures of the two or more binding components are the same.

15. The method of claim 1 , wherein the sites are separated by at least 100 nm.

16. The method of claim 1 , wherein the detectable probe is bound to the analyte via non-covalent binding of only one binding component of the two or more binding components to the epitope.

17. The method of claim 1 , wherein the retaining component comprises a first nucleic acid sequence that is complementary to a nucleic acid that is coupled to one of the binding components.

18. The method of claim 17 , wherein the retaining component further comprises a second nucleic acid sequence that is complementary to a nucleic acid that is coupled to one of the label components, wherein the first nucleic acid sequence differs from the second nucleic acid sequence.

19. The method of claim 1 , wherein the array of analytes comprises at least 1×10 4 different analytes each attached to sites that are physically isolated from all other analytes in the array of analytes.

20. The method of claim 1 , wherein the structures of the two or more label components are the same.

21. The method of claim 1 , further comprising:
(c) contacting the array of analytes with a second detectable probe,
wherein the second detectable probe comprises:
(i) a second retaining component comprising nucleic acid origami,
(ii) a second label component attached to the second retaining component, and
(iii) a second binding component attached to the second retaining component, wherein the second binding component of the second detectable probe is different from the two or more binding components of the detectable probe,

wherein the second detectable probe binds to the analyte via non-covalent binding of the second binding component to an epitope of the analyte; and
(d) acquiring signals from the second label component of the second detectable probe bound to the analyte, thereby detecting the analyte.

22. The method of claim 21 , further comprising, prior to step (c), removing the detectable probe of step (a) from the analyte.

23. The method of claim 22 , wherein the analyte comprises a polypeptide and wherein the method further comprises (e) identifying the polypeptide based on the detecting of steps (b) and (d).

Assignees

• Nautilus Subsidiary Inc

Inventors

• Aksel Tural
• Rinker Torri
• Burns Markus
• Dorwart Michael
• Galimidi Rachel
• Gremyachinskiy Dmitriy
• Hendricks Stephen
• Ikwa Elvis
• Kapp Gregory
• Klein Joshua Simon
• Robinson Julia
• Stawicki Cassandra
• Tonapi Sonal
• Mallick Parag

Classifications

• C12Q1/6832 Primary

  Enhancement of hybridisation reaction · CPC title

• C12Q1/6834

  Enzymatic or biochemical coupling of nucleic acids to a solid phase · CPC title

• C12Q1/6876

  Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes · CPC title

• C12Q1/6811 Primary

  Selection methods for production or design of target specific oligonucleotides or binding molecules · CPC title