System and method for temperature referencing for melt curve data collection

• US 9,393,566 B2
• Filed: 06/23/2008
• Issued: 07/19/2016
• Est. Priority Date: 06/23/2008
• Status: Active Grant

• Current Assignee: Canon USA, Inc. (Canon Inc.)
• Original Assignee: Canon USA, Inc. (Canon Inc.)
• Inventors: Hasson, Kenton C., Inoue, Hiroshi
• Primary Examiner(s): CROW, ROBERT THOMAS
• Application Number:
  US12/144,307
• Publication Number:
  US 20090318306A1
• Time in Patent Office: 2,948 Days
• Field of Search: None
• US Class Current: 1/1
• CPC Class Codes: B01L 2200/147 Employing temperature sensors
• CPC Class Codes: B01L 2300/0627 Sensor or part of a sensor is integrated
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2300/1827 using resistive heater
• CPC Class Codes: B01L 3/5025 for parallel transport of multiple samples
• CPC Class Codes: B01L 3/502784 specially adapted for droplet or plug flow, e.g. digital microfluidics automatic analysis using a stream of discrete samples in a tube system G01N35/08
• CPC Class Codes: B01L 7/52 with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples amplification or hybridisation processes per se C12Q1/68; controlling sequential reactions for synthesis B01J19/0046
• CPC Class Codes: C12Q 1/68 involving nucleic acids
• CPC Class Codes: C12Q 1/6825 Nucleic acid detection involving sensors
• CPC Class Codes: C12Q 2527/107 Temperature of melting, i.e. Tm
• CPC Class Codes: C12Q 2565/629 being a microfluidic device
• CPC Class Codes: G01N 2021/6439 with indicators, stains, dyes, tags, labels, marks
• CPC Class Codes: G01N 21/6428 Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" in vivo A61B5/00; immunoassay G01N33/53
• CPC Class Codes: G01N 21/645 Specially adapted constructive features of fluorimeters
• CPC Class Codes: G01N 2201/06113 Coherent sources; lasers
• CPC Class Codes: G01N 25/4866 by using a differential method
• CPC Class Codes: G01N 25/4893 by using a differential method

Abstract

The present invention relates to systems and methods of temperature referencing for melt curve data collection. More specifically, the present invention relates to systems and methods for collecting DNA melt curve data for a DNA sample and a temperature reference material.

Claims

1. A method of performing melt curve data collection comprising:

(a) providing a container which comprises at least two chambers that are in close thermal connection;

(b) introducing a DNA sample to be tested into at least one of the chambers and a temperature reference material exhibiting a known optical property change as a function of temperature into at least one of the other chambers, wherein the temperature reference materials bracket the DNA sample in space;

(c) heating the chambers from a first temperature to a second temperature;

(d) simultaneously measuring a detectable property emanating from the DNA sample and a detectable property emanating from the temperature reference material during step (c), wherein the detectable property of the DNA sample indicates an extent of denaturation of the DNA in the sample and the detectable property of the temperature reference material is correlated to the temperature of the chamber where the temperature reference material is located; and

,

(e) comparing the detectable property of the DNA sample with the detectable property of the temperature reference material to determine the actual melt properties of the DNA sample, wherein the detectable property emanating from the temperature reference materials is measured to determine a spatial temperature gradient and any temporal fluctuation and a temperature at the location of the DNA sample is estimated by interpolating the results from temperature reference materials that surround the location.

2. The method of claim 1, wherein the container contains at least two spatially separated chambers having temperature reference materials and the detectable property emanating from the temperature reference materials is measured to determine a spatial temperature gradient.

3. The method of claim 1, wherein the temperature reference material is mixed with the DNA sample.

4. The method of claim 1, wherein the detectable property is measured with an optical detection system.

5. The method of claim 1, wherein the detectable property from the temperature reference material provides feedback to a heating system.

6. The method of claim 1, wherein the container is a microfluidic chip and the chambers are microfluidic channels in the chip.

7. The method of claim 1, wherein the container comprises at least three chambers wherein at least two of the chambers are spatially separated from each other and contain temperature reference materials and the detectable property emanating from the temperature reference materials is measured to determine a spatial temperature gradient.

8. The method of claim 7, wherein at least two chambers containing DNA samples are located between two chambers containing the temperature reference material.

9. The method of claim 1, wherein the temperature reference material is a known DNA mixture.

10. The method of claim 1, wherein the temperature reference material is a thermochromatic material.

11. A method of performing melt curve data collection comprising:

(a) providing a microfluidic chip which comprises at least two channels, wherein the channels are in close thermal connection;

(b) introducing a DNA sample to be tested into at least one of the channels and a temperature reference material exhibiting a known optical property change as a function of temperature into at least one of the channels, wherein the temperature reference materials bracket the DNA sample in space;

(c) heating the channels from a first temperature to a second temperature; and

(d) simultaneously measuring a detectable property emanating from the DNA sample and a detectable property emanating from the temperature reference material during step (c), wherein the detectable property of the DNA sample indicates an extent of denaturation of the DNA and the detectable property of the temperature reference material is correlated to the temperature of the chamber where the temperature reference material is located; and

,

(e) comparing the detectable property of the DNA sample with the detectable property of the temperature reference material to determine the actual melt properties of the DNA sample, wherein the detectable property emanating from the temperature reference materials is measured to determine a spatial temperature gradient and any temporal fluctuation and a temperature at the location of the DNA sample is estimated by interpolating the results from temperature reference materials that surround the location.

12. The method of claim 11, wherein the detectable property is measured with an optical detection system.

13. The method of claim 11, wherein a detectable property from the temperature reference material provides feedback to a heating system.

14. The method of claim 11, wherein the DNA sample and the temperature reference material are alternated in the same channel.

15. The method of claim 11, wherein the DNA sample and the temperature reference material are mixed, wherein the temperature reference material has a detection signature that is discernible from that of the DNA sample and wherein data from both the sample and the temperature reference material are collected at the same place and the same time.

16. The method of claim 11, wherein the temperature reference material is also a flow marker.

17. The method of claim 11 wherein the microfluidic chip comprises at least three channels wherein at least two of the channels are spatially separated and contain temperature reference materials and the detectable property emanating from the temperature reference materials is measured to determine a spatial temperature gradient.

18. The method of claim 17, wherein at least two channels containing DNA samples are located between two channels containing the temperature reference material.

19. The method of claim 11, wherein the microfluidic chip comprises at least three channels wherein at least two of the channels are spatially separated and contain a temperature reference material to determine a spatial temperature gradient.

20. The method of claim 19, wherein at least two channels containing DNA samples are located between two channels containing the temperature reference material.

21. The method of claim 11, wherein the temperature reference material is a known DNA mixture.

22. The method of claim 11, wherein the temperature reference material is a thermochromatic material.

23. The method of claim 11, wherein the at least two channels are refillable without moving the microfluidic chip.