Partition-based method of analysis

Patent metadata

Field	Value
Publication number	US-12090480-B2
Application number	US-202318362530-A
Country	US
Kind code	B2
Filing date	Jul 31, 2023
Priority date	Sep 23, 2008
Publication date	Sep 17, 2024
Grant date	Sep 17, 2024

Abstract

Methods of partition-based analysis. In an exemplary method, a device having a port fluidically connected to a chamber may be selected. A sample-containing fluid may be placed into the port. The sample-containing fluid may be moved from the port to the chamber. Partitions of the sample-containing fluid may be formed. A monolayer of the partitions in the chamber may be created. At least a portion of the monolayer may be imaged.

First claim

We claim:

1. A method for quality control in digital PCR, the method comprising the steps of:
preparing a reaction mixture comprising a target nucleic acid, a first dye, a second dye, and nucleic acid amplification reagents;
generating a plurality of partitions comprising the reaction mixture; and
assessing the first dye or and the second dye to indicate variation among the partitions, the first dye providing a test signal and the second dye providing a reference signal and indicating a partition volume of the partitions.

2. The method of claim 1 , wherein the assessing step comprises colorimetric measurement of the first dye or the second dye.

3. The method of claim 1 , wherein at least one of the partitions does not contain the reaction mixture.

4. The method of claim 1 , wherein the assessing step comprises assessing a partition volume for members of the plurality.

5. The method of claim 1 , further comprising the step of identifying a member of the plurality below a predefined partition volume.

6. The method of claim 1 , wherein reference signal indicates amplification of a control nucleic acid target.

7. A method for quality control in a digital PCR, the method comprising:
distributing a reaction mixture comprising a target nucleic acid, a first dye, a second dye, and nucleic acid amplification reagents into a plurality of partitions; and
assessing the first dye or and the second dye to indicate presences of the reaction mixture within members of the plurality, the first dye providing a test signal and the second dye providing a reference signal and indicating a partition volume of the partitions.

8. The method of claim 7 , wherein the assessing step comprises colorimetric measurement of the first dye or the second dye.

9. The method of claim 7 , wherein at least one of the partitions does not contain the reaction mixture.

10. The method of claim 7 , wherein the dye or the second dye is indicative of a volume of the reaction mixture in members of the plurality.

11. The method of claim 7 , wherein the assessing step comprises assessing a partition volume for members of the plurality.

12. The method of claim 7 , further comprising the step of identifying a member of the plurality of partitions below a predetermined partition volume.

13. The method of claim 7 , wherein the reference signal indicates amplification of a control nucleic acid target.

14. The method of claim 1 , wherein the dye or the second dye is indicative of a volume of the reaction mixture in members of the plurality.

15. The method of claim 1 , wherein the second dye is not coupled to an amplification reaction and provides a passive reference.

16. The method of claim 1 , further comprising transforming the test signal based on the reference signal to reduce variation.

17. The method of claim 7 , wherein the second dye is not coupled to an amplification reaction and provides a passive reference.

18. The method of claim 7 , further comprising transforming the test signal based on the reference signal to reduce variation.

Assignees

• Bio Rad Laboratories Inc

Inventors

• Masquelier Donald A
• Ness Kevin D
• Hindson Benjamin J
• Colston Jr Billy W

Classifications

• G01N2015/1481

  Optical analysis of particles within droplets (sorting particles within droplets G01N15/1492) · CPC title

• G01N35/08

  using a stream of discrete samples flowing along a tube system, e.g. flow injection analysis · CPC title

• C12Q1/686

  Polymerase chain reaction [PCR] · CPC title

• B01L2400/049

  vacuum · CPC title

• B01L2300/0861

  Configuration of multiple channels and/or chambers in a single devices · CPC title