Continuous process for performing multiple nucleic acid amplification assays

• US 10,006,862 B2
• Filed: 05/19/2017
• Issued: 06/26/2018
• Est. Priority Date: 03/10/2005
• Status: Active Grant

• Current Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Original Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Inventors: Lair, Gary D., Nguyen, Thanh N., Li, Haitao, Li, Florence, Knight, Byron J., Heinz, Robert E., Macioszek, Jerzy A., Davis, Christopher B., Scalese, Robert F.
• Primary Examiner(s): BOWERS, NATHAN ANDREW
• Application Number:
  US15/600,245
• Publication Number:
  US 20170261431A1
• Time in Patent Office: 403 Days
• 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 1/6813 Hybridisation assays
• CPC Class Codes: C12Q 1/6851 Quantitative amplification
• CPC Class Codes: C12Q 2537/165 Mathematical modelling, e.g. logarithm, ratio
• CPC Class Codes: G01N 2021/6419 Excitation at two or more wavelengths
• CPC Class Codes: G01N 2021/6421 Measuring at two or more wavelengths
• CPC Class Codes: G01N 2021/6432 Quenching
• CPC Class Codes: G01N 2021/6439 with indicators, stains, dyes, tags, labels, marks
• CPC Class Codes: G01N 2021/6441 with two or more labels
• CPC Class Codes: G01N 2035/00346 Heating or cooling arrangements
• 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 2035/0097 monitoring reactions as a function of time
• CPC Class Codes: G01N 2035/0441 for samples
• CPC Class Codes: G01N 2035/0443 for reagents
• CPC Class Codes: G01N 2035/0444 for cuvettes or reaction vessels
• CPC Class Codes: G01N 2035/0455 Coaxial carousels
• CPC Class Codes: G01N 2035/0491 Position sensing, encoding; closed-loop control
• CPC Class Codes: G01N 2035/103 using disposable tips
• 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 21/6454 : using an integrated detector array
• CPC Class Codes: G01N 21/76 : Chemiluminescence; Bioluminescence
• CPC Class Codes: G01N 2201/0415 : Carrusel, sequential
• CPC Class Codes: G01N 35/00693 : Calibration
• CPC Class Codes: G01N 35/0092 : Scheduling
• 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 35/026 : having blocks or racks of reaction cells or cuvettes
• CPC Class Codes: G01N 35/028 : having reaction cells in the form of microtitration plates
• CPC Class Codes: G01N 35/04 : Details of the conveyor system G01N35/021 - G01N35/028 take precedence
• CPC Class Codes: G01N 35/1002 : Reagent dispensers
• CPC Class Codes: Y10S 435/808 : Optical sensing apparatus
• CPC Class Codes: Y10S 435/809 : Incubators or racks or holders for culture plates or containers
• CPC Class Codes: Y10T 436/113332 : with conveyance of sample along a test line in a container or rack
• CPC Class Codes: Y10T 436/25 : including sample preparation

Abstract

A continuous process for performing multiple nucleic acid amplification assays, where at least a portion of a second subset of reaction mixtures are transferred to a heater while a first subset of reaction mixtures are being subjected to conditions for performing a nucleic acid amplification assay. During the process, a plurality of reaction mixtures from the first and second subsets of reaction mixtures are simultaneously subjected to conditions sufficient to perform multiple nucleic acid amplification assays in the reaction mixtures. The presence or absence of a target nucleic acid in the first subset of reaction mixtures is determined while the reaction mixtures are in the heater.

Claims

1. A method for determining the presence or absence of target nucleic acid sequences in a plurality of samples, the method comprising the steps of:

a) loading a plurality of samples onto a self-contained, stand-alone analyzer having a view window situated above a generally horizontal work surface, each sample being contained within one of a plurality of receptacles, and each receptacle having an associated label containing machine-readable data;

b) reading each associated label with a label reading device, wherein the label reading device is in communication with a computer, and wherein the computer associates each sample with one of at least two assays, the at least two assays including a first nucleic acid amplification reaction for amplifying a first target nucleic acid sequence and a second nucleic acid amplification reaction for amplifying a second target nucleic acid sequence, and the first and second target nucleic acid sequences being different from each other;

c) exposing each sample to a solid support material, the solid support material being capable of immobilizing nucleic acids present in the samples;

d) isolating the solid support material within each sample and separating a fluid component of the samples from the solid support material;

e) after step d), washing the solid support material with a wash solution, thereby purifying the nucleic acids immobilized on the solid support material;

f) forming a plurality of reaction mixtures, each reaction mixture being associated with one of the samples and comprising the nucleic acids from the associated sample that are purified in step e), if any, a first subset of the reaction mixtures comprising a first amplification reagent for performing the first nucleic acid amplification reaction, and a second subset of the reaction mixtures comprising a second amplification reagent for performing the second nucleic acid amplification reaction, wherein the first subset of reaction mixtures does not contain an amplification reagent for amplifying the second target nucleic acid sequence, and wherein the second subset of reaction mixtures does not contain an amplification reagent for amplifying the first target nucleic acid sequence;

g) transferring the reaction mixtures to a heater disposed on the work surface;

h) in the heater, simultaneously subjecting a plurality of reaction mixtures from the first and second subsets of reaction mixtures to conditions sufficient to perform the first and second nucleic acid amplification reactions; and

i) determining the presence or absence of the first target nucleic acid sequence in each of the first subset of reaction mixtures and the presence or absence of the second target nucleic acid sequence in each of the second subset of reaction mixtures,

wherein a plurality of the reaction mixtures from the second subset of reaction mixtures are provided to the heater while a plurality of the reaction mixtures from the first subset of reaction mixtures are being subjected to conditions sufficient to perform the first nucleic acid amplification reaction,

wherein the determining step is performed during the subjecting step for the first subset of reaction mixtures, and

wherein steps b)-i) are automated steps performed on the analyzer.

2. The method of claim 1, wherein each associated label is a bar code.

3. The method of claim 1, wherein the solid support material comprises magnetically-responsive particles or beads.

4. The method of claim 3, wherein the isolating step comprises exposing the samples to a magnetic field.

5. The method of claim 4, wherein the magnetic particles or beads are present in the reaction mixtures during the subjecting step.

6. The method of claim 5, wherein each reaction mixture is exposed to a magnetic field during the subjecting step.

7. The method of claim 1, wherein a plurality of the reaction mixtures from the first subset of reaction mixtures are removed from the heater while a plurality of the reaction mixtures from the second subset of reaction mixtures are being subjected to conditions sufficient to perform the second nucleic acid amplification reaction.

8. The method of claim 7, wherein the plurality of reaction mixtures from the first subset of reaction mixtures are contained in a first set of integrally formed receptacles and the plurality of reaction mixtures from the second subset of reaction mixtures are contained in a second set of integrally formed receptacles.

9. The method of claim 1, wherein the conditions are the same for the plurality of reaction mixtures during the subjecting step.

10. The method of claim 9, wherein the reaction mixtures are maintained at a constant temperature during the subjecting step.

11. The method of claim 9, wherein the first nucleic acid amplification reaction is a real-time nucleic acid amplification reaction.

12. The method of claim 11, wherein the second nucleic acid amplification reaction is an end-point nucleic acid amplification reaction.

13. The method of claim 9, wherein the second nucleic acid amplification reaction is an end-point nucleic acid amplification reaction.

14. The method of claim 1, wherein the determining step is performed outside the heater for the second subset of reaction mixtures.

15. The method of claim 1, wherein the determining step is performed with a luminometer for the second subset of reaction mixtures.

16. The method of claim 1, wherein at least one reaction mixture from the first subset of reaction mixtures and at least one reaction mixture from the second subset of reaction mixtures are contained in an integrally formed set of receptacles.

17. The method of claim 1, wherein the heater comprises an enclosure having one or more closable openings for receiving the receptacles therethrough.

18. The method of claim 17, wherein the one or more closable openings are situated on a side wall of the heater.

19. The method of claim 18, wherein the reaction mixtures are carried on a rotatable carousel within the heater during the subjecting step.

20. The method of claim 1, wherein at least a portion of the second subset of reaction mixtures are transferred to the heater prior to transferring all of the first subset of reaction mixtures to the heater.

21. The method of claim 20, wherein the heater comprises an enclosure having one or more closable openings for receiving the receptacles therethrough.

22. The method of claim 21, wherein the one or more closable openings are situated on a side wall of the heater.

23. The method of claim 22, wherein the reaction mixtures are carried on a rotatable carousel within the heater during the subjecting step.

24. The method of claim 1, wherein the analyzer comprises at least two view windows situated above the work surface.

25. The method of claim 1, wherein the determining step is performed for the first subset of reaction mixtures with one or more fluorometers operatively associated with the heater.