sweep fluorometer

Abstract

The swept-frequency fluorometer has a signal processor or processing module configured to: receive a signal containing information relating to light reflected from one or more fluorescent substances of interest in a liquid sample, the liquid sample being scanned by light having a variable frequency range, the information including a characteristic optical frequency corresponding to the fluorescent substance of interest in the liquid, and a characteristic / lifetime optical frequency corresponding to the unique fluorescence lifetime of the fluorescent substance of interest in an excited state; and, based on the received signal, provide a corresponding signal containing information relating to the identity of the fluorescent substance of interest, using the characteristic / lifetime optical frequency, the fluorescent substance of interest is detected and distinguished from overlapping fluorescent substances in the liquid.

Description

[0001] Cross-reference to related applications

[0002] This application claims the benefit of provisional patent application serial number 63 / 027,587 (911-023.9-1-1 / N-YSI-0045US01) filed on May 20, 2020; 63 / 028,013 (911-023.010-1-1 / N-YSI-0046US02) filed on May 21, 2020; and 63 / 028,723 (911-023.011-1-1 / N-YSI-0047US02) filed on May 22, 2020, the entire contents of which are incorporated herein by reference. Technical Field

[0003] The present invention relates to a fluorometer for identifying substances in liquids, including cases where the liquid contains overlapping substances. Background Technology

[0004] Traditional fluorescence sensing techniques are often hampered by broad spectral characteristics, especially when the fluorescence characteristics of multiple substances overlap, which often results in ambiguous identification of water substances.

[0005] Specifically, a traditional (single-channel) fluorometer typically includes a single excitation source, which is used to optically excite the water parameter of interest (at a specific optical wavelength) and to re-emit the optical light specific to the water parameter of interest (at a longer optical wavelength). Such sensors typically utilize lock-in detection, which selectively measures sinusoidal signals at a fixed (in-band) frequency and rejects signals with other frequencies (out-of-band frequencies).

[0006] There is a need in the field for better fluorometers, for example, for identifying substances in liquids containing overlapping substances. Summary of the Invention

[0007] In summary, the sensor according to the present invention differs from a conventional fluorometer in that the locking frequency for this sensor is "scanned," that is, from a certain initial modulation frequency f. i Gradually increase to a certain final modulation frequency f fEach fluorescent substance possesses its own distinct fluorescence "lifetime" (FLT), within which the excited fluorophore (excited by the excitation optical wavelength) remains in the excited state for a finite time (e.g., lifetime [seconds]) before returning to the ground state and undergoing subsequent emission. When the modulation frequency of the signal sweeps across the substance's characteristic frequency f = (1 / lifetime) [Hz], in addition to a decrease in signal amplitude, the signal's phase undergoes a total relative phase shift of 90 degrees. The midpoint of the relative phase transition (45 degrees) precisely corresponds to the fluorescent substance's characteristic / lifetime frequency. In short, if there is uncertainty regarding the identity of overlapping fluorescent substances, the knowledge of lifetimes can be further described between the two substances.

[0008] In fact, the fluorescence lifetime (excited by frequency sweeping) ensures another dimension of information, increasing the depth of the water's "fingerprint".

[0009] The fluorometer according to the present invention measures: Ex / Em+ to provide fluorescence lifetime for enhanced detection techniques.

[0010] Furthermore, a key aspect of this invention lies in the optomechanical configuration of a wide linear array along the length of the pseudo-collimated light.

[0011] The design presented in this paper is compatible with turbidity-based, non-intensity-based determination.

[0012] For example, this invention can be applied to environmental water quality monitoring.

[0013] Example Implementation

[0014] As an example and according to some embodiments, the present invention may include or employ means characterized by a signal processor or processing module, the signal processor or processing module being configured to:

[0015] The signaling receives information relating to light reflected from one or more fluorescent substances of interest in a liquid sample, the liquid sample being scanned by light with a variable frequency range. The information includes characteristic optical frequencies corresponding to the fluorescent substances of interest in the liquid, and characteristic / lifetime optical frequencies corresponding to unique fluorescence lifetimes in which the fluorescent substances of interest remain in an excited state.

[0016] Based on the received signaling, a corresponding signaling is provided, which contains information related to the identity of the fluorescent substance of interest. Using characteristic / lifetime optical frequencies, the fluorescent substance of interest is detected and distinguished from overlapping fluorescent substances in the liquid.

[0017] The device may include one or more of the following additional features:

[0018] The device may include a light sensor configured to sense light across a variable frequency range and to provide signaling received by a signal processor or processing module.

[0019] A light sensor can be a linear sensor array configured to sense light across a variable frequency range along the length of a light source.

[0020] The device may include a light source configured to scan excitation light having an excitation optical wavelength across a variable frequency range to excite one or more fluorescent substances of interest.

[0021] The light source can be a frequency-sweeping light source, which includes a pseudo-collimated light source.

[0022] The variable frequency range may include a locked frequency, which is achieved by gradually increasing the excitation light from an initial modulation frequency f1 to a final modulation frequency f. f And then it was scanned.

[0023] The variable frequency range can include the midpoint frequency corresponding to the characteristic / lifetime optical frequency.

[0024] The device may include a swept-frequency fluorometer or take the form of a swept-frequency fluorometer.

[0025] Sweep fluorometer

[0026] As a further example, and according to some embodiments, the present invention may take the form of a swept-frequency fluorometer characterized by a light source, a light sensor, and a signal processor or processing module.

[0027] The light source can be configured to provide excitation light on a liquid sample having one or more fluorescent species of interest, the excitation light having a variable frequency range having an initial frequency f. i and final frequency f f .

[0028] The optical sensor can be configured to sense light reflected from one or more fluorescent substances of interest in a liquid sample and to provide signaling containing information relating to the light reflected from the one or more fluorescent substances of interest in the liquid sample, which is scanned with excitation light having a variable frequency range. This information includes characteristic optical frequencies corresponding to the fluorescent substances of interest in the liquid and characteristic / lifetime optical frequencies corresponding to the unique fluorescence lifetime of the fluorescent substances of interest in the excited state.

[0029] The signal processor or processing module can be configured as follows:

[0030] Receive signaling, and

[0031] Based on the received signaling, a corresponding signaling is provided, which contains information related to the identity of the fluorescent substance of interest. Using characteristic / lifetime optical frequencies, the fluorescent substance of interest is detected and distinguished from overlapping fluorescent substances in the liquid.

[0032] method

[0033] As yet another example, and according to some embodiments, the present invention may include a method characterized in that:

[0034] A signal processor or processing module receives signaling containing information related to light reflected from one or more fluorescent substances of interest in a liquid sample, the liquid sample being scanned by light with a variable frequency range, the information including characteristic optical frequencies corresponding to the fluorescent substances of interest in the liquid, and characteristic / lifetime optical frequencies corresponding to the unique fluorescence lifetime of the fluorescent substances of interest in the excited state; and

[0035] Using a signal processor or processing module, a corresponding signaling is provided based on the received signaling. This corresponding signaling contains information related to the identity of the fluorescent substance of interest. Using characteristic / lifetime optical frequencies, the fluorescent substance of interest is detected and distinguished from overlapping fluorescent substances in the liquid.

[0036] The method may also include one or more of the features mentioned above.

[0037] Computer-readable storage media

[0038] As yet another example, and according to some embodiments of the invention, the invention can also take the form of a computer-readable storage medium having computer-executable components for performing the steps of the foregoing methods. The computer-readable storage medium may also include one or more of the features described above. Attached Figure Description

[0039] Attached figures that are not necessarily drawn to scale include Figures 1-5 ,as follows:

[0040] Figure 1 This includes graphs of excitation, emission, and fluorescence lifetimes, and provides a conceptual diagram of a fluorometer according to the invention, in which three different but overlapping substances can be measured within the same Ex / Em bandpass region. Conventional fluorometers do not have the ability to distinguish between the three substances, but according to the invention, this can be achieved by utilizing the rendering of fluorescence lifetimes.

[0041] Figure 2 includes Figure 2A and Figure 2BThe images are excitation emission matrix (EEM) plots for two overlapping materials (TrisBipy Ru(II) and Pt Porphyrin). For clarity, these plots are stacked (not overlapping). The axes and scales are identical in both frames.

[0042] Figure 3 It is a graph or line plot of the relative phase angle (degrees) of the corresponding substances (TrisBipy Ru(II) and Pt Porphyrin) against the sweep frequency modulation excitation frequency (Hz), for example, showing how the phase response is completely distinguishable, but their corresponding EEM markers occupy the same area of ​​the EEM plot, resulting in blurring.

[0043] Figure 4 This is a block diagram of a swept-frequency fluorometer according to some embodiments of the present invention. The swept-frequency fluorometer has, for example, a light source, a light sensor, and a signal processor or processing module for performing signal processing functions.

[0044] Figure 5 This is a diagram of an optical sensor in the form of a linear sensor array (e.g., having multiple rows and columns of optical elements) according to some embodiments of the present invention.

[0045] To reduce clutter in the accompanying figures, each figure need not include every figure label for every element shown therein. Detailed Implementation

[0046] Figure 4 An apparatus 10 including a swept-frequency fluorometer according to the invention is shown, the apparatus 10 having, for example, a light source 20 such as a quasi-collimated light source, a light sensor 30 such as a linear sensor array, and a signal processor or processing module 40.

[0047] Light source 20 can be configured to provide light onto a liquid sample having one or more fluorescent species of interest, the light having a variable frequency range having an initial frequency f. i and final frequency f f According to some embodiments, the light source 20 can be configured to respond to a light source control signal, for example, provided by a signal processor or processing module 40, and to provide light with a variable frequency range.

[0048] The light sensor 30 can be configured to: sense light reflected from a fluorescent substance of interest in a liquid sample; and provide signaling containing information relating to light reflected from one or more fluorescent substances of interest in the liquid sample, the liquid sample being scanned by light having a variable frequency range, the information including: a characteristic optical frequency corresponding to the fluorescent substance of interest in the liquid; and a characteristic / lifetime optical frequency corresponding to the unique fluorescence lifetime of the fluorescent substance of interest while it remains in an excited state. According to some embodiments, the light sensor 30 can be configured to: respond to a light sensor control signal, for example, provided by a signal processor or processing module 40, and sense the reflected light.

[0049] The signal processor or processing module 40 can be configured as follows:

[0050] Receive signaling; and

[0051] Based on the received signaling, a corresponding signaling is provided, which contains information related to the identity of the fluorescent substance of interest, which is detected and distinguished from overlapping fluorescent substances in the liquid using characteristic / lifetime optical frequencies.

[0052] According to some embodiments, the signal processor or processing module 40 may be configured to, for example, provide light source control signals and / or light sensor control signals to the light source 20 and / or light sensor 30.

[0053] Light source 20

[0054] As an example, the light source 20 can be configured to: extend along the corresponding length of the linear sensor array 30 (e.g., as shown in the image). Figure 5 (As shown) Light is provided, including pseudo-collimated light, which passes through a liquid sample arranged relative to the light source 20 and the linear sensor array 30, such that light from one or more fluorescent species of interest in the liquid sample being monitored or tested is reflected onto the linear sensor array 30.

[0055] As those skilled in the art will understand, pseudo-collimated light sources are known in the art, and the scope of the invention is not intended to be limited to any particular type or kind now known or to be developed in the future.

[0056] Linear sensor array 30

[0057] As an example, the linear sensor array 30 may include, for example, a linear photodiode array, a linear charge-coupled device (CCD) array, or a linear CMOS array. Specifically, the linear sensor array 30 may include arrays similar to... Figure 5The example shown is, for instance, a two-dimensional array of rows and columns of individually addressable optical elements (r1, c1; r1, c2; r1, c3; r1, c4; r1, c5; r1, c6; r1, c7; r1, c8; ...; r1, cn; r2, c1; r2, c2; r2, c3; r2, c4; r2, c5; r2, c6; r2, c7; r2, c8; ...; r2, cn; r3, c1; r3, c2; r3, c3; r3, c4; r3, c5; r3, c6; r3, c7; r3, c8; ...; r3, cn; ...; rn, c1; rn, c2; rn, c3; rn, c4; rn, c5; rn, c6; rn, c7; rn, c8; ...; rn, cn). Linear sensor arrays are known in the art, and the scope of the present invention is not limited to any particular type or kind now known or to be developed in the future.

[0058] For example, linear sensor arrays are disclosed in the following U.S. Patent Nos. 9,020,202; 8,022,349; 7,956,341; 7,040,538; 5,252,818 and 4,193,057, all of which are incorporated herein by reference.

[0059] Signal processor or processing module 40

[0060] For example, the signal processor or processing module 40 may be configured to: determine one or more fluorescent substances of interest based on such a determined frequency, and then determine the turbidity concentration of the liquid based on the such determined frequency. Techniques for processing signaling containing information related to the sensed optical frequency (e.g., regarding the turbidity concentration in the liquid) are known in the art, and the scope of the invention is not intended to be limited to any particular type or kind now known or to be developed in the future.

[0061] How to implement signal processing functions

[0062] For example, the functionality of the signal processor or processing module 40 can be implemented using hardware, software, firmware, or a combination thereof. In a typical software implementation, the signal processor 40 will include an architecture based on one or more microprocessors, having, for example, at least one signal processor or microprocessor. Those skilled in the art will be able to program implementations such as microcontroller-based or microprocessor-based implementations with appropriate program code to perform the signal processing functions disclosed herein without much experimentation.

[0063] The scope of this invention is not intended to be limited to any particular implementation using techniques now known or developed in the future. The scope of this invention is intended to include implementing the functionality of (multiple) signal processors as independent processors, signal processors or signal processor modules, as well as individual processors or processor modules and some combination thereof.

[0064] As an example, as those skilled in the art will understand, the device 10 may also include other signal processor circuitry or components, such as those generally indicated by 50, including random access memory or memory modules (RAM) and / or read-only memory (ROM), input / output devices and controls, and data and address buses and / or at least one input processor and at least one output processor connected thereto.

[0065] As another example, the signal processor may include or take the form of a combination of a signal processor and at least one memory, the at least one memory including computer program code, wherein the signal processor and at least one memory are configured to cause the system to perform the functions of the present invention, such as responding to received signaling and determining corresponding signaling based on the received signaling.

[0066] Scope of the Invention

[0067] While the invention has been described with reference to exemplary embodiments, those skilled in the art will understand that various changes can be made without departing from the scope of the invention, and equivalents can be used to replace its elements. Additionally, modifications can be made to adapt particular situations or materials to the teachings of the invention without departing from the basic scope of the invention. Therefore, the invention is not limited to the specific embodiments disclosed herein as the best mode for carrying out the invention.

Claims

1. An apparatus for detecting and distinguishing fluorescent substances of interest in a liquid sample containing overlapping fluorescent substances, comprising: The signal processor or processing module is configured as follows: receiving signaling, the signaling including information related to excitation light reflected from one or more fluorescent substances of interest in a liquid sample scanned with light having a variable frequency range, the information including a characteristic optical frequency corresponding to a fluorescent substance of interest in the liquid sample, and a characteristic / lifetime optical frequency corresponding to a unique fluorescent lifetime in which the fluorescent substance of interest remains in an excited state, wherein the variable frequency range includes a lock-in frequency, the lock-in frequency being swept from an initial modulation frequency f i up to a final modulation frequency f f , the variable frequency range including a mid-point frequency corresponding to a mid-point of the characteristic / lifetime optical frequency and a relative phase change at 45 degrees; and Based on the received signaling, corresponding signaling is provided, the corresponding signaling containing information related to the identity of the fluorescent substance of interest, and using the characteristic / lifetime optical frequency, the fluorescent substance of interest is detected and distinguished from the overlapping fluorescent substances in the liquid sample.

2. The apparatus of claim 1, wherein the apparatus includes a light sensor configured to sense the excitation light across the variable frequency range and provide the signaling.

3. The apparatus of claim 2, wherein the optical sensor is a linear sensor array configured to sense the excitation light across the variable frequency range along the length of the light source providing the excitation light.

4. The apparatus of claim 1, wherein the apparatus includes a light source configured to: scan the excitation light having an excitation optical wavelength across the variable frequency range to excite the one or more fluorescent substances of interest.

5. The apparatus according to claim 4, wherein the light source is a swept frequency light source, and the swept frequency light source includes a pseudo-collimated light source.

6. The apparatus according to claim 1, wherein the apparatus is a swept-frequency fluorometer.

7. A method for detecting and distinguishing fluorescent substances of interest in a liquid sample containing overlapping fluorescent substances, comprising: A signal processor or processing module receives signaling containing information relating to excitation light reflected from one or more fluorescent substances of interest in a liquid sample, the liquid sample being scanned by the excitation light having a variable frequency range. The information includes characteristic optical frequencies corresponding to the fluorescent substances of interest in the liquid sample, and characteristic / lifetime optical frequencies corresponding to unique fluorescence lifetimes during which the fluorescent substances of interest are maintained in an excited state. The variable frequency range includes a locking frequency, which is achieved by modulating the excitation light from an initial modulation frequency f. i Gradually increase to the final modulation frequency f f The variable frequency range being scanned includes a midpoint frequency corresponding to the characteristic / lifetime optical frequency and the midpoint of the relative phase transition at 45 degrees; and Using the signal processor or processing module, based on the received signaling, a corresponding signaling is provided, the corresponding signaling containing information related to the identity of the fluorescent substance of interest, and using the characteristic / lifetime optical frequency, the fluorescent substance of interest is detected and distinguished from the overlapping fluorescent substances in the liquid sample.

8. The method of claim 7, wherein the method comprises: The optical sensor is configured to sense the excitation light across the variable frequency range and to provide the signaling.

9. The method of claim 8, wherein the method comprises: The optical sensor is configured as a linear sensor array to sense the excitation light across the variable frequency range along the length of the light source providing the excitation light.

10. The method of claim 7, wherein the method comprises: The light source is configured to scan excitation light with an excitation optical wavelength across the variable frequency range to excite the one or more fluorescent substances of interest.

11. The method of claim 10, wherein the method comprises: The light source is configured as a frequency-sweeping light source, which includes a pseudo-collimated light source.

12. The method of claim 7, wherein the method includes configuring the signal processor or signal processing module as part of a swept-frequency fluorometer.

13. A swept-frequency fluorometer, comprising: A light source is configured to provide excitation light onto a liquid sample containing one or more fluorescent substances of interest, the excitation light having a variable frequency range, wherein the variable frequency range includes a locked frequency, the locked frequency being adjusted by modulating the excitation light from an initial modulation frequency f. i Gradually increase to the final modulation frequency f f And being scanned, and the variable frequency range includes a midpoint frequency corresponding to the characteristic / lifetime optical frequency and the midpoint of the relative phase transition at 45 degrees; An optical sensor is configured to sense the excitation light reflected from the fluorescent substance of interest in the liquid sample and to provide signaling containing information relating to the excitation light reflected from the one or more fluorescent substances of interest in the liquid sample, the liquid sample being scanned by the excitation light having the variable frequency range, the information including a characteristic optical frequency corresponding to the fluorescent substance of interest in the liquid sample and a characteristic / lifetime optical frequency corresponding to a unique fluorescence lifetime in which the fluorescent substance of interest remains in an excited state; as well as The signal processor or processing module is configured as follows: Receive the signaling, and Based on the received signaling, corresponding signaling is provided, the corresponding signaling containing information related to the identity of the fluorescent substance of interest, using characteristic / lifetime optical frequencies, the fluorescent substance of interest is detected and distinguished from overlapping fluorescent substances in the liquid sample.

14. The swept-frequency fluorometer according to claim 13, wherein... The light source is a frequency-sweeping light source, which includes a quasi-collimated light source; and The optical sensor is a linear sensor array configured to sense the excitation light across the variable frequency range along the length of the light source providing the excitation light.

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