Method for performing an assay with a nucleic acid present in a specimen

• US 8,569,019 B2
• Filed: 10/31/2007
• 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 W
• Application Number:
  US11/931,722
• Publication Number:
  US 20080241837A1
• Time in Patent Office: 2,190 Days
• Field of Search: 435/6, 435/91.1, 435/91.2, 435/183, 435/6.1, 435/6.11, 435/6.12, 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 process for isolating and amplifying a target nucleic acid in a self-contained analyzer.

Claims

1. A method for detecting the presence of a nucleic acid in a specimen, the method comprising the steps of:

(A) providing a specimen tube containing specimen to a self-contained, standard-alone analyzer;

(B) performing an automated process within a housing of the analyzer that comprises;

(1) transferring at least a portion of the specimen from the specimen tube to a receptacle, wherein the transferred portion of the specimen is dispensed into the receptacle with a robotic pipettor, and wherein the transferred portion of the specimen contains a nucleic acid;

(2) immobilizing the nucleic acid on a solid support within the receptacle, the solid support comprising a magnetically-responsive particle;

(3) applying a magnetic field to the receptacle and its contents;

(4) during step (3), purifying the nucleic acid immobilized in step (2) by accessing and removing at least a portion of a fluid component of the specimen from the receptacle while the nucleic acid remains immobilized on the solid support within the receptacle;

(5) forming a reaction mixture comprising the purified nucleic acid of step (4) and all reagents required to perform a nucleic acid amplification;

(6) performing a nucleic acid amplification with the reaction mixture formed in step (5), the nucleic acid amplification resulting in the enzymatic synthesis of amplification products, each of the amplification products comprising (i) a target sequence contained in the nucleic acid or (ii) the complement of the target sequence;

(7) selectively hybridizing a probe having a label to the target sequence or its complement of an amplification product that is one of the amplification products synthesized in step (6), thereby forming a hybrid in a mixture that comprises the probe and the amplification product; and

(8) in the mixture, detecting [the label as an indication of] the formation of the hybrid, wherein the formation of the hybrid in the mixture is an indication of the presence of the nucleic acid in the specimen.

2. The method of claim 1 further comprising, prior to step (A), a step of loading the specimen tube onto a specimen tube rack.

3. The method of claim 1 further comprising, prior to step (A), a step of providing to the analyzer all reagents required to perform a nucleic acid amplification, wherein the specimen tube and the reagents required to perform a nucleic acid amplification are separately provided to the analyzer.

4. The method of claim 1, wherein the receptacle is a cylindrical tube.

5. The method of claim 1, wherein the receptacle is one of a plurality of integrally formed receptacles.

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

7. The method of claim 1, wherein the nucleic acid is immobilized on the solid support with a capture probe in step (2).

8. The method of claim 1, wherein the automated process further comprises performing within the housing of the analyzer a step of washing the solid support one or more times after step (4) and prior to step (5).

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

10. 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 and sequences having a sequence that is the complement of the target sequence.

11. The method of claim 1, wherein step (6) is carried out in a temperature-controlled incubator maintained at a substantially uniform temperature.

12. The method of claim 1, wherein step (6) is performed in a temperature-controlled chamber of a first incubator, and wherein step (7) is performed in a temperature-controlled chamber of a second incubator, the first and second incubators being in a spaced-apart relationship.

13. The method of claim 12, wherein the hybrid is in solution.

14. The method of claim 1, wherein the label is a fluorescent or chemiluminescent label.

15. The method of claim 1, wherein the housing remains closed during the automated process.

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

17. A method for detecting the presence of a nucleic acid in a specimen, the method comprising the steps of:

(A) providing a specimen tube containing a specimen to a self-contained, standard-alone unit analyzer;

(B) performing an automated process within a housing of the analyzer that comprises;

(1) transferring at least a portion of the specimen from the specimen tube to a receptacle, wherein the transferred portion of the specimen is dispensed into the receptacle with a robotic pipettor, and wherein the transferred portion of the specimen contains a nucleic acid;

(2) immobilizing the nucleic acid on a solid support within the receptacle, the solid support comprising a magnetically-responsive particle;

(3) applying a magnetic field to the receptacle and its contents;

(4) during step (3), purifying the nucleic acid immobilized in step (2) by contacting a fluid component of the specimen with a fluid aspirator and aspirating at least a portion of the fluid component from the receptacle while the nucleic acid remains immobilized on the solid support within the receptacle;

(5) forming a reaction mixture comprising the purified nucleic acid of step (4) and all reagents required to perform a nucleic acid amplification;

(6) performing a nucleic acid amplification procedure with the reaction mixture formed in step (5), the nucleic acid amplification resulting in the enzymatic synthesis of amplification products, each of the amplification products comprising (i) a target sequence contained in the nucleic acid or (ii) the complement of the target sequence;

(7) selectively hybridizing a probe having a label to the target sequence or it complement of an amplification product that is one of the amplification products synthesized in step (6), thereby forming a hybrid in a mixture that comprises the probe and the amplification product; and

(8) in the mixture, detecting [the label as an indication of] the formation of the hybrid in the mixture is an indication of the presence of the nucleic acid in the specimen.

18. The method of claim 17, wherein step (6) is performed in a temperature-controlled chamber of a first incubator, and wherein step (7) is performed in a temperature-controlled chamber of a second incubator, the first and second incubators being in a spaced-apart relationship.

19. The method of claim 17 further comprising, prior to step (A), a step of loading the specimen tube onto a specimen tube rack.

20. The method of claim 17 further comprising, prior to step (A), a step of providing to the analyzer all reagents required to perform a nucleic acid amplification, wherein the specimen tube and the reagents required to perform a nucleic acid amplification are separately provided to the analyzer.

21. The method of claim 17, wherein the receptacle is a cylindrical tube.

22. The method of claim 17, wherein the receptacle is one of a plurality of integrally formed receptacles.

23. The method of claim 22, wherein the integrally formed receptacles are arranged in a linear array.

24. The method of claim 17, wherein the nucleic acid is immobilized on the solid support with a capture probe in step (2).

25. The method of claim 17, wherein the automated process further comprises performing within the housing of the analyzer a step of washing the solid support one or more times after step (4) and prior to step (5).

26. The method of claim 17, 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 (4).

27. The method of claim 26, wherein the contamination-limiting element is a tiplet.

28. The method of claim 17, wherein each of the amplification products consists of a nucleic acid sequence having the sequence contained in the nucleic acid or its complement.

29. The method of claim 17, 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.

30. The method of claim 17, wherein step (6) is carried out in a temperature-controlled incubator maintained at a substantially uniform temperature.

31. The method of claim 17, wherein the label is a fluorescent or chemiluminescent label.

32. The method of claim 17, wherein the housing remains closed during the automated process.

33. The method of claim 17, wherein the automated process is performed at multiple locations of a processing deck contained within the housing.

34. The method of claim 17, wherein the hybrid is in solution.