SYSTEMS AND METHODS FOR DISTINGUISHING OPTICAL SIGNALS OF DIFFERENT MODULATION FREQUENCIES IN AN OPTICAL SIGNAL DETECTOR

• US 20120221252A1
• Filed: 02/24/2012
• Published: 08/30/2012
• Est. Priority Date: 02/24/2011
• Status: Active Grant

• Current Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Original Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Inventors: HEINZ, Robert E., OPALSKY, David, NEWELL, Dennis, RHUBOTTOM, Jason
• Granted Patent: US 8,718,948 B2
• Time in Patent Office: Days
• US Class Current: 702/19
• 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 21/6486 Measuring fluorescence of biological material, e.g. DNA, RNA, cells G01N21/6428 takes precedence

Abstract

Systems and method for detecting optical signals, and for discriminating optical signals emitted by an emission moiety that is excited by an associated excitation signal from background signals and other optical noise, employing digital techniques for determining the portion of a detected optical signal having a modulation frequency corresponding to a modulation of the associated excitation signal.

Claims

1. A method for detecting multiple different optical signals emitted from the contents of each of two or more receptacles, each different optical signal indicating the presence of a different analyte of interest or amplification products thereof, said method comprising:

(a) forming amplification products of an analyte of interest within each receptacle;

(b) while forming the amplification products, directing an excitation signal at each receptacle, the excitation signal having a predetermined excitation wavelength that excites an emission moiety, which emits an emission signal that is associated with the excitation wavelength and has a predetermined emission wavelength, wherein each excitation signal has a predetermined excitation frequency;

(c) with a signal detecting device, detecting an optical signal including an emission signal emitted from each receptacle and generating detection data from the detected signal;

(d) executing a computer-implemented algorithm for digitizing the detection data;

(e) executing a computer-implemented algorithm for determining from the digitized detection data an amplitude of the detection data at the predetermined excitation frequency to ascertain the portion of the detected signal that corresponds to the associated emission signal; and

(f) repeating steps (b)-(e) for each of the multiple different optical signals to be detected,

wherein the predetermined excitation frequency for an excitation signal associated with the emission signal of one emission moiety is different from the predetermined excitation frequency for at least one other emission moiety in the receptacle.

2. The method of claim 1, wherein detecting an optical signal comprises measuring an intensity of an optical signal at each of a plurality of discrete times, and wherein the measured intensity of the optical signal at each discrete time comprises an average intensity measured over a range of time including the discrete time.

3. The method of claim 1, wherein steps (b)-(c) are repeated two or more times for each receptacle prior to step (d).

4. The method of claim 1, wherein three or more different optical signals are detected from each of the receptacles.

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

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

7. The method of claim 1, wherein the emission moiety comprises a fluorescent dye, and each different optical signal is generated by a different fluorescent dye contained in the receptacle.

8. The method of claim 1,

wherein the at least one optical signal comprises first, second, and third optical signals,

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

wherein the spectra of the first and second range of optical wavelengths at least partially overlap each other, the spectra of the second and third range of optical wavelengths at least partially overlap each other, but the spectra of 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 signals are the same and the excitation frequency of the excitation signal associated with the second optical signal is different from the excitation frequencies of the excitation signals associated with the first and third optical signals.

9. The method of claim 1, wherein the two or more receptacles are arranged in a linear, side-by-side arrangement and wherein the predetermined excitation frequencies for adjacent receptacles are different.

10. The method of claim 9, wherein each receptacle is releasably fixed within the side-by-side arrangement.

11. The method of claim 9, wherein the predetermined excitation frequencies for alternate receptacles are the same.

12. The method of claim 1, wherein step (e) comprises executing a Goertzel signal processing technique.

13. The method of claim 1, further comprising executing a computer-implemented algorithm for determining the amount of an analyte present within each receptacle from data relating to the portion of the detected signal that corresponds to the associated emission signal determined in step (e).

14. The method of claim 1, wherein the detection data is digitized at a frequency that is at least twice the excitation frequency of an associated excitation signal.

15. The method of claim 14, wherein the excitation frequency is within the range of 200-350 Hz, and the digitization frequency is about 4 kHz.

16. The method of claim 1, further comprising the formation of amplification products of a control analyte and performing steps (b)-(f) for the control analyte.

17. The method of claim 16, wherein the control analyte comprises a known analyte unrelated to the analyte of interest.

18. The method of claim 16, further comprising adding a control probe having specificity for the control analyte to each receptacle, wherein the control probe comprises an emission moiety that emits an emission signal having an emission wavelength that is different from the emission wavelength of the emission moiety for the analyte of interest.

19. The method of claim 16, further comprising executing a computer-implemented algorithm for determining the amount of the control analyte from data relating to the portion of the detected signal that corresponds to the associated emission signal determined in step (e).

20. The method of claim 18, further comprising alternately exposing the contents of the receptacle to light energy at different excitation wavelengths and detecting the presence or absence of a signal having the emission wavelength of the emission moiety of the control probe as in indication of the successful formation of amplification products.

21. A system for detecting an optical signal emitted from a receptacle, said system comprising:

an excitation device constructed and arranged to (a) generate an excitation signal having a predetermined excitation wavelength and a predetermined excitation frequency that excites an emission moiety which emits an emission signal that is associated with the excitation wavelength and having a predetermined emission wavelength, and (b) direct the excitation signal at the receptacle;

a signal detecting device constructed and arranged to detect an optical signal including an emission signal emitted from the receptacle and to store detection data relating to the detected signal on a computer-readable medium;

a processor for executing an algorithm to digitize the stored detection data; and

a processor for executing an algorithm for determining from the digitized detection data an amplitude of the emission signal at the predetermined excitation frequency to ascertain the portion of the detected signal corresponding to the associated emission signal.

22. The system of claim 21, further comprising a processor for executing an algorithm for determining the amount of an analyte present within the receptacle based on data relating to the portion of the detected signal corresponding to the associated emission signal.

23. The system of claim 21, further comprising:

a plurality of excitation devices in fixed positions with respect to each other, each excitation device being constructed and arranged to (a) generate an excitation signal having a predetermined excitation wavelength and a predetermined excitation frequency that excites an emission moiety that emits an associated emission signal having a predetermined emission wavelength, and (b) direct the excitation signal at a different receptacle; and

a plurality of signal detecting devices in fixed positions with respect to each other, each signal detecting device being constructed and arranged to detect an emission signal emitted from a receptacle and to store detection data relating to the detected signal on a computer-readable medium,

wherein each excitation device corresponds with an associated detecting device such that the generated excitation signal and detected emission signal of each associated excitation and detecting device are directed at, and emitted from, an associated receptacle,

wherein the processor is configured to digitize the stored detection data associated with each signal detecting device; and

wherein the processor is configured to determine from the digitized detection data associated with each signal detecting device an amplitude of the emission signal at the associated predetermined excitation frequency and to ascertain the portion of the associated detected signal that corresponds to the associated emission signal.

24. The system of claim 23, wherein the different associated receptacles comprise a plurality of receptacles arranged in a linear, side-by-side arrangement and wherein the associated predetermined excitation frequencies for adjacent receptacles are different.

25. The system of claim 24, wherein each receptacle is releasably fixed within the side-by-side arrangement.

26. The system of claim 24, wherein the associated predetermined excitation frequencies for alternate receptacles are the same.

27. The system of claim 23, further comprising a receptacle moving apparatus constructed and arranged to move the different associated receptacles relative to the plurality of excitation and detecting devices.

28. The system of claim 23, comprising two or more sets of the plurality of excitation and associated detecting devices, wherein each set is spatially distinct from each other set within the system.

29. The system of claim 28, comprising three to six sets of the plurality of excitation and associated detecting devices.

30. The system of claim 28, wherein each set of the plurality of excitation and associated detecting devices is arranged radially around a central axis.

31. A method for detecting an optical signal emitted from the contents of a receptacle in the presence of multiple, different optical signals originated from the same receptacle or a different receptacle, said method comprising:

(a) forming amplification products of an analyte of interest within the receptacle;

(b) while forming the amplification products, directing an excitation signal at the receptacle, the excitation signal having a predetermined excitation wavelength that excites an emission moiety, which emits an emission signal that is associated with the excitation wavelength and has a predetermined emission wavelength, wherein the excitation signal has a predetermined excitation frequency;

(c) with a signal detecting device, detecting an optical signal including an emission signal emitted from the receptacle and generating detection data from the detected signal;

(d) executing a computer-implemented algorithm for digitizing the detection data; and

(e) executing a computer-implemented algorithm for determining from the digitized detection data an amplitude of the detection data at the predetermined excitation frequency to ascertain the portion of the detected signal that corresponds to the associated emission signal.

32. The method of claim 31, further comprising detecting a second optical signal emitted from the contents of the receptacle by:

while forming the amplification products, directing a second excitation signal at the receptacle, the second excitation signal having a second predetermined excitation wavelength that excites an emission moiety, which emits a second emission signal that is associated with the second excitation wavelength and has a predetermined second emission wavelength, wherein the second excitation signal has a predetermined second excitation frequency;

with the same or a different signal detecting device, detecting an optical signal including an emission signal emitted from the receptacle and generating second detection data from the detected signal;

executing a computer-implemented algorithm for digitizing the second detection data; and

executing a computer-implemented algorithm for determining, from the second digitized detection data, an amplitude of the second detection data at the predetermined second excitation frequency to ascertain the portion of the detected signal that corresponds to the associated second emission signal.

33. The method of claim 32, wherein the excitation frequencies are different for at least two optical signals to be detected.

34. The method of claim 31, wherein the emission signal is a fluorescent emission.

35. The method of claim 31, wherein the emission moiety comprises a fluorescent dye.

36. The method of claim 31, wherein first, second, and third optical signals are detected from the receptacle;

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

wherein the spectra of the first and second range of optical wavelengths at least partially overlap each other, the spectra of the second and third range of optical wavelengths at least partially overlap each other, but the spectra of 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 signals are the same and the excitation frequency of the excitation signal associated with the second optical signal is different from the excitation frequencies of the excitation signals associated with the first and third optical signals.

37. The method of claim 31, wherein step (e) comprises executing a Goertzel signal processing technique.

38. The method of claim 31, further comprising executing a computer-implemented algorithm for determining the amount of an analyte present within the receptacle from data relating to the portion of the detected signal that corresponds to the associated emission signal determined in step (e).

39. The method of claim 31, wherein the detection data is digitized at a frequency that is at least twice the excitation frequency of the associated excitation signal.

40. The method of claim 39, wherein the excitation frequency is within the range of 200-350 Hz, and the digitization frequency is about 4 kHz.