Measurement of protein expression using reagents with barcoded oligonucleotide sequences

Patent metadata

Field	Value
Publication number	US-11460468-B2
Application number	US-201715715028-A
Country	US
Kind code	B2
Filing date	Sep 25, 2017
Priority date	Sep 26, 2016
Publication date	Oct 4, 2022
Grant date	Oct 4, 2022

Abstract

Some embodiments disclosed herein provide a plurality of compositions each comprising a protein binding reagent conjugated with an oligonucleotide. The oligonucleotide comprises a unique identifier for the protein binding reagent it is conjugated with, and the protein binding reagent is capable of specifically binding to a protein target. Further disclosed are methods and kits for quantitative analysis of a plurality of protein targets in a sample and for simultaneous quantitative analysis of protein and nucleic acid targets in a sample. Also disclosed herein are systems and methods for preparing a labeled biomolecule reagent, including a labeled biomolecule agent comprising a protein binding reagent conjugated with an oligonucleotide.

First claim

What is claimed is:

1. A plurality of compositions, comprising at least 100 protein-binding compositions each comprising a protein binding reagent associated with an oligonucleotide, wherein the oligonucleotide comprises a target region and a unique identifier sequence for the protein binding reagent that it is associated therewith, wherein the target region is at the 3′ region of the oligonucleotide and the protein binding reagent is capable of specifically binding to a protein target, wherein the unique identifier sequence comprises a nucleotide sequence of less than 30 nucleotides in length, wherein at least one of the 100 protein-binding compositions comprises a second protein binding reagent which is not associated with an oligonucleotide, and wherein the protein binding reagent and second protein binding reagent of the at least one of the 100 protein-binding compositions are the same.

2. The plurality of compositions of claim 1 , wherein the unique identifier sequence comprises a nucleotide sequence of 15 nucleotides in length.

3. The plurality of compositions of claim 1 , wherein at least 10 of the at least 100 protein-binding compositions comprise different unique identifier sequences.

4. The plurality of compositions of claim 1 , wherein at least 100 of the at least 100 protein-binding compositions comprise different unique identifier sequences.

5. The plurality of compositions of claim 1 , wherein the protein binding reagent comprises an antibody, an aptamer, or a combination thereof.

6. The plurality of compositions of claim 1 , wherein the oligonucleotide is associated with the protein binding reagent through a linker.

7. The plurality of compositions of claim 1 , wherein the oligonucleotide comprises a molecular label sequence, a poly(A) sequence, or a combination thereof.

8. The plurality of compositions of claim 1 , further comprising one or more additional protein-binding compositions that each comprises a second protein binding reagent which is not associated with an oligonucleotide.

9. The plurality of compositions of claim 8 , where at least one of the protein binding reagents of the at least 100 protein-binding compositions and at least one of the second protein binding reagents of the one or more additional protein-binding compositions are the same.

10. A kit for simultaneous quantitative analysis of a plurality of protein targets and a plurality of nucleic acid target molecules in a sample, comprising
a plurality of compositions, wherein the plurality of compositions comprises at least 100 protein-binding compositions each comprising a protein binding reagent associated with an oligonucleotide, wherein the oligonucleotide comprises a target region and a unique identifier sequence for the protein binding reagent that it is associated therewith, wherein the unique identifier sequence comprises a nucleotide sequence of less than 30 nucleotides in length, wherein the target region is at the 3′ region of the oligonucleotide and the protein binding reagent is capable of specifically binding to a protein target, wherein at least 10 of the at least 100 protein-binding compositions comprise different unique identifier sequences, wherein at least one of the 100 protein-binding compositions comprises a second protein binding reagent which is not associated with an oligonucleotide, and wherein the protein binding reagent and second protein binding reagent of the at least one of the 100 protein-binding compositions are the same; and
a plurality of oligonucleotide probes, wherein each of the plurality of oligonucleotide probes comprises a molecular label sequence and a target binding region capable of binding to a target region of the plurality of compositions, wherein the molecular label sequence is from a diverse set of unique molecular label sequences.

11. The plurality of compositions of claim 1 , wherein the target region comprises a poly(A) sequence.

12. The plurality of compositions of claim 1 , wherein the oligonucleotide associated with the protein binding reagent in each of the at least 100 protein-binding compositions comprises a molecular label sequence selected from a set of at least 100 different molecular label sequences.

13. The plurality of compositions of claim 1 , wherein the protein target is a cell-surface protein, a cell marker, a B-cell receptor, a T-cell receptor, an antibody, a major histocompatibility complex, a tumor antigen, a receptor, or any combination thereof.

14. The plurality of compositions of claim 1 , wherein at least 100 of the at least 100 protein-binding compositions comprise different protein binding reagents.

15. The plurality of compositions of claim 1 , wherein each of the at least 100 protein-binding compositions comprises a different protein binding reagent.

16. The plurality of compositions of claim 1 , wherein each of the at least 100 protein-binding compositions comprises a different unique identifier sequence.

17. The plurality of compositions of claim 1 , wherein at least 10 of the at least 100 protein-binding compositions comprise different protein binding reagents.

18. The plurality of compositions of claim 17 , wherein each—protein binding reagent is associated with one or more oligonucleotides comprising at least one molecular label sequence selected from a set of at least 10 different molecular label sequences.

Assignees

• Cellular Res Inc
• Becton Dickinson Co

Inventors

• Fan Christina
• Chang Christina
• Shum Eleen
• Zoellner Olaf
• Bansal Nidhanjali
• Ghadiali James
• Lam Gretchen Yinbon
• Jensen Eric
• Rohrer Jurg
• Gaylord Brent
• Martin Jody
• Vidal Jason G

Classifications

• G01N33/68 Primary

  involving proteins, peptides or amino acids {(involving lipoproteins G01N33/92)} · CPC title

• C12Q1/6851

  Quantitative amplification · CPC title

• G01N33/566 Primary

  using specific carrier or receptor proteins as ligand binding reagents {where possible specific carrier or receptor proteins are classified with their target compounds} · CPC title

• B01J2219/00725

  Peptides · CPC title

• G01N33/54313

  the carrier being characterised by its particulate form · CPC title