Systems and methods for detecting multiple optical signals

• US 8,663,922 B2
• Filed: 06/01/2010
• Issued: 03/04/2014
• Est. Priority Date: 03/10/2005
• Status: Active Grant

• Current Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Original Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Inventors: Knight, Byron J., Li, Haitao, Scalese, Robert F., Hayes, Matthew J.
• Primary Examiner(s): BOWERS, NATHAN ANDREW
• Application Number:
  US12/791,390
• Publication Number:
  US 20100240063A1
• Time in Patent Office: 1,372 Days
• Field of Search: 435/288.7, 435/287.2, 435/287.1, 435/808, 435/4, 435/6.1, 435/6.11, 435/6.12
• US Class Current: 435/6.12
• 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

To minimize cross talk in systems and methods for detecting two or more different optical signals emitted from each of a plurality of reaction receptacles, an excitation signal associated with each of the optical signals has a known excitation frequency, and any detected signal having a frequency that is inconsistent with the excitation frequency is discarded. The receptacles are moved relative to optical sensors configured to detect each unique optical signal from an associated receptacle, and to further minimize cross talk, the optical sensors are arranged so that only one reaction receptacle at a time is in a signal detecting position with respect to one of its associated optical sensors, and the optical sensors are grouped by the optical signal they are configured to detect so that a first optical signal is detected from each of the reaction receptacles before a second optical signal is detected from the reaction receptacles.

Claims

1. A method for detecting two or more different optical emission signals emitted from each of a plurality of reaction receptacles, said method comprising, for each optical emission signal to be detected, the steps of:

(a) modulating an optical excitation signal at a known excitation signal frequency, the excitation signal having a known excitation wavelength that excites an associated optical emission signal having a known emission wavelength;

(b) directing the optical excitation signal at each receptacle;

(c) filtering an optical signal, including any optical emission signal emitted by the contents of each receptacle, to transmit a portion of the optical signal that is at the emission wavelength;

(d) detecting the portion of the optical signal transmitted in step (c);

(e) determining an emission signal frequency of the optical signal detected in step (d); and

(f) discarding any portion of the optical signal detected in step (d) having an emission signal frequency determined in step (e) that is inconsistent with the known excitation signal frequency to determine the portion of the optical signal detected in step (d) that corresponds to the associated optical emission signal.

2. The method of claim 1, wherein the excitation frequency for one optical emission signal to be detected is different from the excitation frequency for at least one other optical emission signal to be detected.

3. The method of claim 2, wherein three or more different optical emission signals are detected from each of the reaction receptacles.

4. The method of claim 3, wherein the excitation frequencies are the same for two or more, but less than all, of the optical emission signals to be detected.

5. The method of claim 1, wherein the optical emission signal is a fluorescent emission.

6. The method of claim 1, wherein each different optical emission signal is generated by a different fluorescent dye contained in the reaction receptacle.

7. The method of claim 6, wherein the fluorescent dyes comprise rhodamine dyes in combination with a DABCYL quencher.

8. The method of claim 7, wherein the fluorescent dyes further comprise fluorescein dyes in combination with a DABCYL quencher.

9. The method of claim 8, wherein the dyes comprise two or more dyes selected from the group consisting of tetramethyl-6-rhodamine (“

TAMRA”

), tetrapropano-6-carboxyrhodamine (“

ROX”

), 6-carboxyfluorescein (“

FAM”

), and 2′

,7′

-dimethoxy-4′

,5′

-dichloro-6-carboxyrhodamine (“

JOE”

).

10. The method of claim 1,

wherein the two or more different optical emission signals comprise first, second, and third optical emission signals,

wherein the first optical emission signal comprises a first range of optical wavelengths, the second optical emission signal comprises a second range of optical wavelengths, and the third optical emission signal comprises a third range of optical wavelengths,

wherein the first and second range of optical wavelengths at least partially overlap each other, the second and third range of optical wavelengths at least partially overlap each other, but the first and third range of optical wavelengths do not overlap each other, and

wherein the excitation frequencies of the excitation signals associated with the first and third optical emission signals are the same and the excitation frequency of the excitation signal associated with the second optical emission signal is different from the excitation frequencies of the excitation signals associated with the first and third optical emission signals.

11. The method of claim 1, wherein the reaction receptacles are disposed in a fixed arrangement with respect to each other, wherein said method is performed using an instrument comprising optical sensors configured to detect one of two or more optical emission signals from each of a plurality of reaction receptacles, wherein each optical sensor is associated with one of the reaction receptacles and each reaction receptacle has associated therewith one optical sensor for each optical emission signal to be detected, and wherein the optical sensors are arranged so that only one of the plurality of reaction receptacles at a time can be in a signal-detecting position with respect to one of its associated optical sensors, and the optical sensors are grouped so that each of the reaction receptacles is placed in a signal-detecting position with respect to an associated optical sensor configured to detect one optical emission signal before any of the reaction receptacles is placed in signal-detecting position with respect to an associated optical sensor configured to detect a different optical emission signal, and wherein said method further comprises the steps of:

effecting relative movement between the plurality of reaction receptacles and the optical sensors to sequentially place each reaction receptacle of the fixed arrangement of reaction receptacles into a signal-detecting position with respect to one of its associated optical sensors configured to detect one of the two or more optical emission signals; and

sequentially measuring an amount of the one optical emission signal emitted by the contents of each reaction receptacle as each reaction receptacle is placed in the signal detecting position with respect to its associated optical sensor configured to detect the one optical signal.

12. The method of claim 11, wherein the plurality of optical sensors are disposed in fixed positions, and effecting relative movement comprises moving the reaction receptacles into signal-detecting positions with respect to the fixed optical sensors.

13. The method of claim 12, wherein moving the reaction receptacles comprises moving each of the reaction receptacles in a circular path.

14. The method of claim 13, wherein the optical sensors are arranged at different angular positions with respect to the circular path of the associated reaction receptacle.

15. The method of claim 13, wherein all of the reaction receptacles are moved in a circular path having a common axis of rotation and each of the reaction receptacles is positioned at a different radial position with respect to the axis of rotation, and wherein the optical sensors associated with each reaction receptacle are positioned at different angular locations and at the radial position corresponding to the associated reaction receptacle.

16. The method of claim 15, wherein each of the reaction receptacles comprises a test tube, and wherein the test tubes are connected to each other in an aligned arrangement and an upright orientation and further wherein moving the reaction receptacles in a circular path comprises carrying the connected arrangement of test tubes in a generally radial orientation with respect to the axis of rotation.

17. The method of claim 11, wherein the instrument comprises three optical sensors associated with each reaction receptacle, each of the three optical sensors being configured to detect a different one of three different optical emission signals.

18. The method of claim 11, wherein each of the reaction receptacles comprises a test tube, and wherein the test tubes are connected to each other in an aligned arrangement and an upright orientation.

19. The method of claim 11, wherein each of the optical sensors is optically isolated from the other optical sensors.

20. The method of claim 11, wherein each of the optical sensors comprises a fluorometer constructed and arranged to detect fluorescent emissions from the reaction receptacles.

21. The method of claim 11, wherein the two or more different optical emission signals are optical signals having different wavelengths.

22. The method of claim 1, wherein an amount of each different optical emission signal emitted by the contents of each reaction receptacle is indicative of the presence or amount of a different target nucleic acid within the contents of the reaction receptacle, and wherein the method further comprises the step of determining the presence or amount, within the contents of each reaction receptacle, of the target nucleic acid associated with each optical emission signal based upon the optical emission signal determined in step c).

23. The method of claim 22, further comprising the step of periodically performing steps a) through c) to generate periodic optical emission data for each optical emission signal and wherein an amount of the target nucleic acid associated with each optical emission signal is determined based upon the periodic optical emission data.