Microfluidic devices, solid supports for reagents and related methods

Patent metadata

Field	Value
Publication number	US-12077808-B2
Application number	US-202217735509-A
Country	US
Kind code	B2
Filing date	May 3, 2022
Priority date	May 25, 2012
Publication date	Sep 3, 2024
Grant date	Sep 3, 2024

Abstract

A microfluidic device includes a plurality of reaction wells; and a plurality of solid supports, and each of the solid supports has a reagent attached thereto. The reagent is attached to the solid support via a labile reagent/support bond such that the reagent is configured to be cleaved from the support via a cleaving operation.

First claim

What is claimed is:

1. A method of conducting a reaction using solid supports in a microfluidic device, the microfluidic device comprising: a substrate having a plurality of reaction wells that are sized and configured to capture a plurality of solid supports, wherein each of the solid supports has a reagent attached via a labile reagent/support bond and the reagent is configured to be cleaved from the solid support via a cleaving operation, and further wherein the plurality of reaction wells and the plurality of solid supports are sized and configured such that only a single one of the plurality of solid supports is retained in a corresponding single one of the plurality of reaction wells, the method comprising:
loading the solid supports in the plurality of reaction wells;
loading a sample comprising DNA in the plurality of reaction wells, wherein some number of reaction wells may be double loaded with solid supports;
loading a solution comprising a PCR master mix and a detectable label in the plurality of reaction wells; and
after loading the solid supports, the sample, and the solution in the plurality of reaction wells, flowing a sealing fluid comprising at least one of oil, liquid polymer, or fluorocarbon liquid over the plurality of reaction wells to seal the plurality of reaction wells, wherein the sealing fluid fluidically isolates each of the plurality of reaction wells during a reaction.

2. The method of claim 1 , further comprising performing a cleaving operation to release the reagent into the reaction well.

3. The method of claim 2 , wherein the cleaving operation is an addition of a chemical, the addition of an enzyme, application of an electric potential, and/or an application of light, or ionizing radiation to the labile reagent/support bond.

4. The method of claim 2 , wherein the cleaving operation comprises a thermal operation.

5. The method of claim 4 , wherein the labile reagent/support bond comprises a streptavidin-biotin bond and/or avidin-biotin bond.

6. The method of claim 5 , wherein the bond is a streptavidin-biotin bond, and further wherein the streptavidin-biotin bond is configured to release the reagent from the solid support when the solid supports are incubated at about 50-99° C.

7. The method of claim 1 , wherein the reagent comprises a nucleic acid sequence for a nucleic acid transcription and/or amplification reaction.

8. The method of claim 7 , wherein the reagent comprises a DNA primer or probe for a PCR reaction.

9. The method of claim 1 , wherein the solid supports are microbeads.

10. The method of claim 9 , wherein the microbeads are a polymer, magnetic material, or a combination thereof.

11. The method of claim 9 , wherein the microbeads are about 3.3 um in diameter.

12. The method of claim 1 , wherein the solid support comprises a marker.

13. The method of claim 12 , wherein the marker comprises a predefined support shape, a predefined support size, a magnetic property, and/or optical property of the support.

14. The method of claim 12 , wherein the marker comprises a fluorescence marker.

15. The method of claim 12 , wherein the marker comprises a Qdot.

16. The method of claim 1 , wherein the plurality of solid supports comprises at least a first plurality of solid supports having a first reagent attached thereto and a second plurality of solid supports having a second reagent attached thereto, wherein the first and second plurality of solid supports further comprise respective first and second markers configured to identify a property of the solid support and/or reagent.

17. The method of claim 16 , further comprising decoding the device by identifying whether each of the plurality of solid support containment regions is occupied by the first plurality of solid supports or the second plurality of solid supports.

18. The method of claim 17 , wherein decoding the device comprises identifying the first and second markers of the plurality of solid supports.

19. The method of claim 1 , wherein the plurality of solid supports comprises at least:
a first plurality of solid supports having a first reagent attached thereto;
a second plurality of solid supports having a second reagent attached thereto; and
and a third plurality of solid supports having a third reagent attached thereto;

wherein the first, second, and third plurality of solid supports further comprise respective first, second, and third markers configured to identify a property of the solid support and/or reagent.

20. The method of claim 19 , wherein at least one of the markers may be an optical marker.

21. The method of claim 20 , wherein at least one of the markers may be the absence of an optical marker.

22. The method of claim 1 , wherein the plurality of solid supports comprises at least:
a first plurality of solid supports having a first reagent attached thereto;
a second plurality of solid supports having a second reagent attached thereto;
a third plurality of solid supports having a third reagent attached thereto; and
a fourth plurality of solid supports having a fourth reagent attached thereto;

wherein the first, second, third, and fourth plurality of solid supports further comprise respective first, second, third, and fourth markers configured to identify a property of the solid support and/or reagent.

23. The method of claim 1 , wherein the loading steps occur in the following order:
first loading the solid supports in the plurality of reaction wells; and

second loading a sample comprising DNA and a solution comprising a PCR master mix and a detectable label in the plurality of reaction wells at the same time.

Assignees

• Univ North Carolina Chapel Hill

Inventors

• Ramsey John Michael
• Henley William
• Oblath Emily

Classifications

• C12Q1/686

  Polymerase chain reaction [PCR] · CPC title

• B01L2300/0864

  comprising only one inlet and multiple receiving wells, e.g. for separation, splitting · CPC title

• B01L2300/0829

  Multi-well plates; Microtitration plates · CPC title

• B01L2300/0636

  Integrated biosensor, microarrays · CPC title

• B01L2200/16

  Reagents, handling or storing thereof · CPC title