Automated process for isolating and amplifying a target nucleic acid sequence

• US 6,890,742 B2
• Filed: 11/01/2001
• Issued: 05/10/2005
• Est. Priority Date: 05/01/1998
• Status: Expired due to Term

• Current Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Original Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Inventors: Horner, Glenn A., Smith, Robert J., Burns, Ralph E., Schneider, Robert E., Ammann, Kelly G., Hansberry, Ernest V., Nieglos, Donald J., Kling, John E., Jakub, Cheryl A.
• Primary Examiner(s): Horlick, Kenneth R.
• Assistant Examiner(s): LU, FRANK WEI MIN
• Application Number:
  US09/985,064
• Publication Number:
  US 20020028489A1
• Time in Patent Office: 1,286 Days
• Field of Search: 435/6, 435/91.1, 435/91.2, 435/183, 436/94, 536/23.1, 536/24.3, 536/24.33, 536/25.3
• 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 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 analyzer for performing multiple diagnostic assays simultaneously includes multiple stations, or modules, in which discrete aspects of the assay are performed on fluid samples contained in reaction receptacles. The analyzer includes stations for automatically preparing a specimen sample, incubating the sample at prescribed temperatures for prescribed periods, preforming an analyte isolation procedure, and ascertaining the presence of a target analyte. An automated receptacle transporting system moves the reaction receptacles from one station to the next. The analyzer further includes devices for carrying a plurality of specimen tubes and disposable pipette tips in a machine-accessible manner, a device for agitating containers of target capture reagents comprising suspensions of solid support material and for presenting the containers for machine access thereto, and a device for holding containers of reagents in a temperature controlled environment and presenting the containers for machine access thereto. A method for performing an automated diagnostic assay includes an automated process for isolating and amplifying a target analyte. The process is performed by automatically moving each of a plurality of reaction receptacles containing a solid support material and a fluid sample between stations for incubating the contents of the reaction receptacle and for separating the target analyte bound to the solid support from the fluid sample. An amplification reagent is added to the separated analyte after the analyte separation step and before a final incubation step.

Claims

1. A process for isolating and amplifying a target sequence contained in a target nucleic acid present in a fluid sample, the process comprising the steps of:

separating the target nucleic acid from other material present in the fluid sample in a separation station;

transporting a reaction receptacle containing the separated target nucleic acid from the separation station to an amplification incubation station comprising one or more temperature-controlled incubators; and

incubating the contents of the reaction receptacle, to which one or more amplification reagents have been provided, in the amplification incubation station for a period of time and under conditions sufficient to permit the target sequence to be amplified, wherein the separation and amplification incubation stations are contained within a housing, and wherein each of the separating, transporting and incubating steps is automated.

2. The process of claim 1 further comprising providing to the reaction receptacle a probe for a period of time and under conditions sufficient to permit the probe to hybridize to the target sequence or an amplicon thereof.

3. The process of claim 2, wherein the probe includes a detectable label.

4. The process of claim 3, wherein the label is a fluorescent dye or a chemiluminescent compound.

5. The process of claim 2 further comprising detecting the presence or absence of the probe hybridized to the target sequence, or an amplicon thereof, as an indication of the presence or absence of members of a target group of organisms or viruses in the fluid sample, wherein the target group consists of at least one organism or virus.

6. The process of claim 5, wherein the detecting step comprises determining the amount of light emitted from the reaction receptacle as an indication of the presence or absence of members of the target group of organisms or viruses in the fluid sample.

7. The process of claim 5 further comprising determining the amount of members of the target group of organisms or viruses in the fluid sample.

8. The process of claim 5 further comprising transporting the reaction receptacle from the amplification incubation station to a hybridization incubation station within the housing prior to providing the probe to the reaction receptacle, wherein the hybridization incubation station comprises one or more temperature-controlled incubators, and wherein the step of transporting the reaction receptacle from the amplification incubation station to the hybridization incubation station is automated.

9. The process of claim 8, wherein the probe is provided to the reaction receptacle after transporting reaction receptacle amplification incubation station to the hybridization incubation station.

10. The process of claim 8 further comprising transporting the reaction receptacle from the hybridization incubation station to a detection station within the housing prior to the detecting step, wherein the presence or amount of hybridized probe is determined in the detection station, and wherein the step of transporting the reaction receptacle from the hybridization incubation station to the detection station is automated.

11. The process of claim 10, wherein the detection station includes a luminometer for determining the amount of light emitted by the contents of the reaction receptacle.

12. The process of claim 1 further comprising raising or lowering the temperature of the contents of the reaction receptacle prior to transporting the reaction receptacle to the amplification incubation station.

13. The process of claim 8 further comprising raising or lowering the temperature of the contents of the reaction receptacle prior to transporting the reaction receptacle to at least one of the amplification and hybridization incubation station.

14. The process of claim 10 further comprising providing to the reaction receptacle a deactivating reagent for destroying nucleic acids present in the reaction receptacle.

15. The process of claim 14, wherein the deactivating reagent comprises a chemical agent selected from the group consisting of solutions of potassium permanganate, formic acid, solutions of sodium hypochiorite, hydrazine, and dimethyl sulfate.

16. The process of claim 14 further comprising transporting the reaction receptacle from the detection station to a deactivation station within the housing prior to providing the deactivating reagent to the reaction receptacle, wherein the step of transporting the reaction receptacle from the detection station to the deactivation station is automated.

17. The process of claim 1 further comprising providing to the fluid sample a solid support for a period of time and under conditions sufficient to permit the target nucleic acid to be immobilized, directly or indirectly, on the solid support prior to the separating step.

18. The process of claim 17, wherein the solid support comprises a magnetically responsive particle having a polynucleotide bound thereto.

19. The process of claim 18, wherein the target nucleic acid is hybridized to a capture probe prior to immobilizing the target nucleic acid on the solid support.

20. The process of claim 18, wherein the fluid sample is subjected to a magnetic field during the separating step.

21. The process of claim 20 further comprising providing to the reaction receptacle a probe for a period of time and under conditions sufficient to permit the probe to hybridize to the target sequence or an amplicon thereof.

22. The process of claim 21 further comprising detecting the presence or absence of the probe hybridized to the target sequence, or an amplicon thereof, as an indication of the presence or absence of members of a target group of organisms or viruses in the fluid sample, wherein the target group consists of at least one organism or virus.

23. The process of claim 22 further comprising transporting the reaction receptacle from the amplification incubation station to a hybridization incubation station within the housing prior to providing the probe to the reaction receptacle, wherein the hybridization incubation station comprises one or more temperature-controlled incubators, and wherein the step of transporting the reaction receptacle from the amplification incubation station to the hybridization incubation station is automated.

24. The process of claim 23, wherein the probe is provided to the reaction receptacle after transporting the reaction receptacle from the amplification incubation station to the hybridization incubation station.

25. The process of claim 23 further comprising transporting the reaction receptacle from the hybridization incubation station to a detection station within the housing prior to the detecting step, wherein the presence or amount of hybridized probe is determined in the detection station, and wherein the step of transporting the reaction receptacle from the hybridization incubation station to the detection station is automated.

26. The process of claim 17 further comprising providing to the reaction receptacle a deactivating reagent for destroying nucleic acids present in the reaction receptacle.

27. The process of claim 26 further comprising transporting the reaction receptacle from the detection station to a deactivation station within the housing prior to providing the deactivating reagent to the reaction receptacle, wherein the step of transporting the reaction receptacle from the detection station to the deactivation station is automated.

28. The process of claim 1, wherein the transporting step is performed with a receptacle transport mechanism.

29. The process of claim 1, wherein the reaction receptacle comprises a plurality of receptacle vessels formed as an integral array, each receptacle vessel containing the same or a different fluid sample for simultaneously separating and amplifying the same or different target sequences present in the fluid samples of the receptacle vessels.

30. The process of claim 1, wherein the housing defines a self-contained, stand alone analyzer unit.

31. The process of claim 30, wherein said analyzer unit is movable.

32. The process of claim 1, wherein the material includes non-target nucleic acid.