Method for simultaneously performing multiple amplification reactions

• US 8,569,020 B2
• Filed: 09/30/2008
• Issued: 10/29/2013
• Est. Priority Date: 05/01/1998
• Status: Expired due to Fees

• Current Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Original Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Inventors: Ammann, Kelly G., Burns, Ralph E., Hansberry, Ernest V., Horner, Glenn A., Jakub, Cheryl A., Kling, John E., Nieglos, Donald J., Schneider, Robert E., Smith, Robert J.
• Primary Examiner(s): Lu, Frank
• Application Number:
  US12/241,350
• Publication Number:
  US 20090029871A1
• Time in Patent Office: 1,855 Days
• Field of Search: 435/6.1, 435/6.11, 435/6.12, 435/91.1, 435/91.2, 435/183, 436/94, 436/501, 536/23.1, 536/24.3, 536/24.33, 536/25.3, 536/25.32
• US Class Current: 435/91.2
• CPC Class Codes: B01F 29/10 with receptacles rotated about two different axes, e.g. receptacles having planetary motion
• CPC Class Codes: B01F 29/30 Mixing the contents of individual packages or containers, e.g. by rotating tins or bottles
• CPC Class Codes: B01F 29/322 of two or more containers supported for simultaneous mixing, e.g. for bottles in crates
• CPC Class Codes: B01F 29/40354 arranged for planetary motion
• CPC Class Codes: B01L 2200/147 Employing temperature sensors
• CPC Class Codes: B01L 2300/1822 using Peltier elements
• CPC Class Codes: B01L 2300/1827 using resistive heater
• 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: B01L 7/5255 by moving sample containers
• CPC Class Codes: B01L 9/06 Test-tube stands; Test-tube holders
• CPC Class Codes: B03C 1/282 with associated accumulation indicator, e.g. Hall sensor
• CPC Class Codes: B03C 1/288 disposed at the outer circumference of a recipient
• CPC Class Codes: B03C 1/30 Combinations with other devices, not otherwise provided for
• CPC Class Codes: B03C 2201/26 for use in medical or biological applications
• CPC Class Codes: C12Q 1/6813 Hybridisation assays
• CPC Class Codes: C12Q 1/6832 Enhancement of hybridisation reaction
• CPC Class Codes: C12Q 1/6834 Enzymatic or biochemical coupling of nucleic acids to a solid phase
• CPC Class Codes: C12Q 2537/10 the purpose or use of
• CPC Class Codes: G01N 2035/00356 Holding samples at elevated temperature (incubation)
• CPC Class Codes: G01N 2035/00524 Mixing by agitating sample carrier
• CPC Class Codes: G01N 2035/0437 : Cleaning cuvettes or reaction vessels
• CPC Class Codes: G01N 2035/0455 : Coaxial carousels
• CPC Class Codes: G01N 2035/103 : using disposable tips
• CPC Class Codes: G01N 35/0098 : involving analyte bound to insoluble magnetic carrier, e.g. using magnetic separation magnetic particles used in immunoassays G01N33/54326; magnetic separation in general B03C
• CPC Class Codes: G01N 35/0099 : comprising robots or similar manipulators robots per se B25J
• CPC Class Codes: G01N 35/025 : having a carousel or turntable for reaction cells or cuvettes
• CPC Class Codes: G01N 35/1065 : Multiple transfer devices
• CPC Class Codes: Y10T 156/1105 : Delaminating process responsive to feed or shape at delamination
• CPC Class Codes: Y10T 156/1906 : Delaminating means responsive to feed or shape at delamination
• CPC Class Codes: Y10T 29/53657 : to apply or remove a resilient article [e.g., tube, sleeve, etc.]
• CPC Class Codes: Y10T 29/53683 : Spreading parts apart or separating them from face to face engagement
• CPC Class Codes: Y10T 29/53843 : Tube, sleeve, or ferrule inserting or removing
• CPC Class Codes: Y10T 436/11 : Automated chemical analysis
• CPC Class Codes: Y10T 436/111666 : Utilizing a centrifuge or compartmented rotor
• CPC Class Codes: Y10T 436/113332 : with conveyance of sample along a test line in a container or rack
• CPC Class Codes: Y10T 436/114998 : with treatment or replacement of aspirator element [e.g., cleaning, etc.]
• CPC Class Codes: Y10T 436/119163 : with aspirator of claimed structure
• CPC Class Codes: Y10T 436/2575 : Volumetric liquid transfer

Abstract

An automated method for performing multiple nucleic acid amplification procedures in multiple receptacles of a set of receptacles contained in an analyzer.

Claims

1. A method for detecting the presence of a nucleic acid in each of multiple samples, the method comprising performing within a housing of an analyzer the automated steps of:

(a) contacting each of multiple samples containing a nucleic acid with a solid support, such that a complex comprising the nucleic acid and the solid support is formed in each of the samples, wherein the solid support comprises a magnetically-responsive particle, and wherein the complex formed in each of the samples is suspended in a fluid component of the sample;

(b) after step (a), subjecting the complex in each of the samples to a magnetic field;

(c) during step (b), purifying the nucleic acid in each of the samples by accessing and removing at least a portion of the fluid component of each of the samples from the complex;

(d) forming multiple mixtures, each of the mixtures comprising the purified nucleic acid of one of the samples of step (c) and reagents for performing a nucleic acid amplification, wherein the mixtures are separately formed in multiple receptacles of a set of receptacles;

(e) moving the set of receptacles to a temperature-controlled environment;

(f) in each of the multiple receptacles, performing nucleic acid amplification, whereby amplification products are enzymatically synthesized, each of the amplification products comprising (i) a target sequence contained in the nucleic acid or (ii) the complement of the target sequence;

(g) in each of the multiple receptacles, selectively hybridizing a detectably labeled probe to an amplification product that is one of the amplification products synthesized in step (f), thereby forming a hybrid that comprises the probe and the amplification product, wherein the amplification product comprises the target sequence or its complement, and wherein the probe hybridizes to the target sequence or its complement in the amplification product; and

(h) in each of the multiple receptacles, detecting the presence of the hybrid, wherein the presence of the hybrid in each of the multiple receptacles is an indication of the presence of the nucleic acid in each of the samples,

wherein the method is performed without human intervention.

2. The method of claim 1, further comprising performing within the housing of the analyzer, prior to step (h) and after step (g), a step of providing a selection reagent to each of the multiple receptacles, whereby a label of an unhybridized, detectably labeled probe in each of the multiple receptacles is selectively hydrolyzed, wherein the label is an acridinium ester.

3. The method of claim 1, further comprising performing within the housing of the analyzer an automated step of washing the solid support of each of the samples one or more times after step (c) and prior to step (d).

4. The method of claim 1, wherein each receptacle of the set of receptacles comprises a cylindrical tube.

5. The method of claim 1, wherein the set of receptacles is an integrally formed set of receptacles.

6. The method of claim 1, wherein each of the amplification products consists of a nucleic acid sequence having the target sequence or its complement.

7. The method of claim 1, wherein the amplification products include nucleic acid sequences having a sequence that is the same as the target sequence and nucleic acid sequences having a sequence that is the complement of the target sequence.

8. The method of claim 1, wherein step (f) is carried out in a temperature- controlled incubator maintained at a substantially uniform temperature.

9. The method of claim 1, wherein step (f) is performed in a temperature-controlled chamber of a first incubator, and wherein step (g) is performed in a temperature-controlled chamber of a second incubator, the first and second incubators being in a spaced-apart relationship.

10. The method of claim 1, wherein the probe is detectably labeled with a fluorescent or chemiluminescent label.

11. The method of claim 1, wherein the hybrid formed in each of the multiple receptacles is in solution in steps (g) and (h).

12. The method of claim 1, wherein the method is performed at multiple locations of a processing deck contained within the housing.

13. The method of claim 1, wherein the housing remains closed during the method.

14. The method of claim 5, wherein the integrally formed receptacles are arranged in a linear array.

15. A method for detecting the presence of a nucleic acid in each of multiple samples, the method comprising performing within a housing of an analyzer the automated steps of:

(a) contacting each of multiple samples containing a nucleic acid with a solid support, such that a complex comprising the nucleic acid and the solid support is formed in each of the samples, wherein the solid support comprises a magnetically-responsive particle, and wherein the complex formed in each of the samples is suspended in a fluid component of the sample;

(b) after step (a), subjecting the complex in each of the samples to a magnetic field;

(c) during step (b), purifying the nucleic acid in each of the samples by contacting the fluid component of each of the samples with a fluid aspirator and aspirating at least a portion of the fluid component from the complex;

(d) forming multiple mixtures, each of the mixtures comprising the purified nucleic acid of one of the samples of step (c) and reagents for performing a nucleic acid amplification, wherein the mixtures are separately formed in multiple receptacles of a set of receptacles;

(e) moving the set of receptacles to a temperature-controlled environment;

(f) in each of the multiple receptacles, performing nucleic acid amplification, amplification products are enzymatically synthesized, each of the amplification products comprising (i) a target sequence contained in the nucleic acid or (ii) the complement of the target sequence;

(g) in each of the multiple receptacles, selectively hybridizing a detectably labeled probe to an amplification product that is one of the amplification products synthesized in step (f), thereby forming a hybrid that comprises the probe and the amplification product, wherein the amplification product comprises the target sequence or its complement, wherein the probe hybridizes to the target sequence or its complement in the amplification product, and wherein the hybrid is formed in solution; and

(h) in each of the multiple receptacles, detecting the presence of the hybrid while the hybrid is in solution, wherein the presence of the hybrid in each of the multiple receptacles is an indication of the presence of the nucleic acid in each of the samples,

wherein the method is performed without human intervention.

16. The method of claim 15, further comprising performing within the housing of the analyzer, prior to step (h) and after step (g), a step of providing a selection reagent to each of the multiple receptacles, whereby a label of an unhybridized, detectably labeled probe in each of the multiple receptacles is selectively hydrolyzed, wherein the label is an acridinium ester.

17. The method of claim 15, wherein step (f) is performed in a temperature-controlled chamber of a first incubator, and wherein step (g) is performed in a temperature-controlled chamber of a second incubator, the first and second incubators being in a spaced-apart relationship.

18. The method of claim 15 further comprising performing within the housing of the analyzer an automated step of washing the solid support of each of the samples one or more times after step (c) and prior to step (d).

19. The method of claim 15, wherein each receptacle of the set of receptacles comprises a cylindrical tube.

20. The method of claim 15, wherein the set of receptacles is an integrally formed set of receptacles.

21. The method of claim 20, wherein the integrally formed receptacles are arranged in a linear array.

22. The method of claim 15, wherein the fluid aspirator comprises an aspirator tube in frictional engagement with a contamination-limiting element configured to permit the passage of the fluid component of the specimen therethrough, and wherein the contamination- limiting element contacts the fluid component of the specimen in step (c).

23. The method of claim 22, wherein the contamination-limiting element is a tiplet.

24. The method of claim 15, wherein each of the amplification products consists of a nucleic acid sequence having the target sequence or its complement.

25. The method of claim 15, wherein the amplification products include nucleic acid sequences having a sequence that is the same as the target sequence and nucleic acid sequences having a sequence that is the complement of the target sequence.

26. The method of claim 15, wherein step (f) is carried out in a temperature-controlled incubator maintained at a substantially uniform temperature.

27. The method of claim 15, wherein the probe is detectably labeled with a fluorescent or chemiluminescent label.

28. The method of claim 15, wherein the method is performed at multiple locations of a processing deck contained within the housing.

29. The method of claim 15, wherein the housing remains closed during the method.

30. The method of claim 15, wherein the hybird formed in each of the multiple receptacles is in solution.