Assays and other reactions involving droplets

Patent metadata

Field	Value
Publication number	US-9850526-B2
Application number	US-201514721558-A
Country	US
Kind code	B2
Filing date	May 26, 2015
Priority date	Mar 7, 2007
Publication date	Dec 26, 2017
Grant date	Dec 26, 2017

Abstract

The present invention generally relates to droplets and/or emulsions, such as multiple emulsions. In some cases, the droplets and/or emulsions may be used in assays, and in certain embodiments, the droplet or emulsion may be hardened to form a gel. In some aspects, a heterogeneous assay can be performed using a gel. For example, a droplet may be hardened to form a gel, where the droplet contains a cell, DNA, or other suitable species. The gel may be exposed to a reactant, and the reactant may interact with the gel and/or with the cell, DNA, etc., in some fashion. For example, the reactant may diffuse through the gel, or the hardened particle may liquefy to form a liquid state, allowing the reactant to interact with the cell. As a specific example, DNA contained within a gel particle may be subjected to PCR (polymerase chain reaction) amplification, e.g., by using PCR primers able to bind to the gel as it forms. As the DNA is amplified using PCR, some of the DNA will be bound to the gel via the PCR primer. After the PCR reaction, unbound DNA may be removed from the gel, e.g., via diffusion or washing. Thus, a gel particle having bound DNA may be formed in one embodiment of the invention.

First claim

What is claimed is:

1. A composition comprising a plurality of aqueous droplets in a water-in-oil emulsion, wherein the plurality of aqueous droplets is monodisperse, wherein the plurality of aqueous droplets comprises a plurality of particles, wherein a given droplet of the plurality of aqueous droplets comprises a given particle from the plurality of particles, and wherein the given particle comprises one or more reagents for carrying out a nucleic acid amplification reaction on a nucleic acid molecule.

2. The composition of claim 1 , wherein the one or more reagents are coupled to the given particle.

3. The composition of claim 1 , wherein the one or more reagents are encapsulated within the given particle.

4. The composition of claim 1 , wherein the one or more reagents are selected from the group consisting of nucleic acids, proteins, and peptides.

5. The composition of claim 1 , wherein the one or more reagents comprise nucleic acids.

6. The composition of claim 5 , wherein the nucleic acids comprise nucleic acid primers.

7. The composition of claim 5 , wherein the nucleic acids are coupled to the given particle through an acrydite linkage.

8. The composition of claim 1 , wherein the given particle is dissolvable.

9. The composition of claim 1 , wherein at least 75% of the plurality of aqueous droplets each comprises at least one particle from the plurality of particles.

10. The composition of claim 9 , wherein at least 90% of the plurality of aqueous droplets each comprises at least one particle from the plurality of particles.

11. The composition of claim 1 , wherein some of the plurality of aqueous droplets are unoccupied by a particle from the plurality of particles.

12. The composition of claim 1 , wherein the plurality of aqueous droplets is monodisperse with standard deviations less than 2%.

13. The composition of claim 1 , wherein the given particle is a gel particle, and wherein the gel particle comprises the one or more reagents for carrying out the nucleic acid amplification reaction on the nucleic acid molecule.

14. The composition of claim 13 , wherein the gel particle comprises a hydrogel.

15. The composition of claim 13 , wherein the gel particle comprises a polyacrylamide polymer.

16. The composition of claim 15 , wherein the polyacrylamide polymer comprises a cross-linked polyacrylamide polymer.

17. The composition of claim 1 , wherein the nucleic acid amplification reaction is whole genome amplification.

18. The composition of claim 1 , wherein the nucleic acid amplification reaction is reverse transcription.

19. The composition of claim 1 , wherein the nucleic acid amplification reaction is polymerase chain reaction.

20. The composition of claim 1 , further comprising aqueous droplets that do not comprise a particle from the plurality of particles.

Assignees

• Harvard College

Inventors

• Agresti Jeremy
• Chu Liang-Yin
• Weitz David A
• Kim Jin-Woong
• Rowat Amy
• Sommer Morten
• Dantas Gautam
• Church George

Classifications

• B01J13/0065

  containing an organic phase · CPC title

• C12Q1/6834

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

• C12Q1/686

  Polymerase chain reaction [PCR] · CPC title

• G01N2015/1006

  for cytology · CPC title

• C12Q1/6848 Primary

  characterised by the means for preventing contamination or increasing the specificity or sensitivity of an amplification reaction · CPC title