A micro-area sample multi-spectral measurement system and measurement method thereof

Abstract

The present invention relates to the field of optical instrument technology, and provides a micro-area sample multi-spectral measurement system and measurement method thereof, comprising: a first light source, a second light source, an optical path switching device, a microscope, a light selection device, a spectrometer, a CCD camera, a single-photon detector, a counter, and a computer; the first light source or the second light source emits excitation light, the optical path switching device selects the first light source or the second light source to pass through, and allows the excitation light to enter the microscope; the excitation light enters the microscope to excite the sample to generate sample fluorescence, which is emitted through the microscope and enters the light selection device; after the light selection device performs light selection, the selected sample fluorescence enters the spectrometer; the spectrometer has a first optical outlet and a second optical outlet, the first optical outlet corresponding to the CCD camera, and the second optical outlet corresponding to the single-photon detector. The present invention can realize multiple spectral measurements of fluorescence excitation spectrum, fluorescence emission spectrum, and fluorescence lifetime spectrum.

Description

Technical Field

[0001] The present invention relates to the technical field of optical instruments, and in particular to a micro-area sample multi-spectral measurement system and a measurement method thereof. Background Art

[0002] A typical micro-area sample spectrum testing system includes a microscope, a xenon lamp spectroscopic light source, a laser light source, optical components, and a detection device.

[0003] Existing technical solutions require separate measurement equipment for PLE, PL, and TRPL spectroscopy, resulting in inconvenient testing, low test efficiency, and high equipment costs. Furthermore, there are currently no products for measuring PLE in micro-areas under high pressure, only solutions for PLE testing of conventional samples.

[0004] Furthermore, the microscopes currently used in spectral measurement systems are standard commercial microscopes, which have limited sample space and cannot accommodate presses or other heavy cryogenic equipment such as liquid helium and liquid nitrogen. Furthermore, the microscopes are integrated, making them difficult to modify. Summary of the Invention

[0005] The present invention mainly solves the technical problems that the existing technology has no high-voltage PLE testing solution, and that PLE, PL and TRPL spectral measurements require different instruments and equipment, resulting in inconvenient testing, low test efficiency and high equipment costs. A micro-area sample multi-spectral measurement system and measurement method are proposed to meet the needs of PLE testing under high pressure, while realizing multi-spectral measurement requirements at one time, improving measurement convenience and improving test work efficiency.

[0006] The present invention provides a micro-area sample multi-spectral measurement system, comprising: a first light source, a second light source, an optical path switching device, a microscope, a light selection device, a spectrometer, a CCD camera, a single photon detector, a counter and a computer;

[0007] The first light source or the second light source emits excitation light, and the light path switching device selects to allow the first light source or the second light source to pass through and allows the excitation light to enter the microscope;

[0008] The microscope has a sample placement position, and the sample is placed on the sample placement position by a high-pressure press;

[0009] The excitation light enters the microscope to excite the sample to generate sample fluorescence, and the sample fluorescence is emitted through the microscope and enters the light selection device;

[0010] After the light selection device performs light selection, the sample fluorescence after light selection enters the spectrometer;

[0011] The spectrometer has a first optical outlet and a second optical outlet, the first optical outlet corresponds to a CCD camera, and the second optical outlet corresponds to a single photon detector;

[0012] The single-photon detector collects the number of fluorescence photons of the sample after light selection and the corresponding fluorescence photon arrival time information, and the CCD camera collects the fluorescence intensity and wavelength information of the sample after light selection;

[0013] The single photon detector and the pulse laser are respectively connected to the counter signal; the monochromator, the spectrometer, the CCD camera and the counter are respectively connected to the computer signal.

[0014] Preferably, the first light source uses a xenon lamp and a monochromator;

[0015] The monochromator has an optical inlet and an optical outlet. The light of the xenon lamp enters the monochromator perpendicularly to the optical inlet and exits the monochromator from the optical outlet as a monochromatic light beam.

[0016] Preferably, the optical inlet and the optical outlet of the monochromator are respectively in the form of slits.

[0017] Preferably, the second light source is a pulsed laser;

[0018] A first optical adjustment device is provided between the optical path switching device and the pulse laser.

[0019] Preferably, the optical path switching device includes an adjustment seat and a reflector;

[0020] The reflector is mounted on an adjustment seat at an angle of 45 degrees, and the adjustment seat can cause the reflector to fall down or rise up.

[0021] Preferably, a second optical adjustment device is provided between the optical path switching device and the microscope.

[0022] Preferably, the microscope comprises: a support base, a light source support frame, an illumination light source, an excitation module, a focusing frame, an objective lens, a three-dimensional moving stage and a sample support frame;

[0023] A light source support frame is provided on the support seat; an illumination light source and a plurality of excitation modules are provided on the light source support frame;

[0024] A focusing frame is provided on the upper part of the light source support frame, and an objective lens is provided on the focusing frame;

[0025] A three-dimensional moving platform is provided below the light source support frame, and a sample support frame is provided on the three-dimensional moving platform; the sample support frame has a sample placement position;

[0026] Through holes are provided at positions corresponding to the excitation module on the light source support frame and at positions corresponding to the objective lens on the focusing frame;

[0027] After the microscope is adjusted, the excitation module, the objective lens and the sample are placed in the same straight line;

[0028] A trinocular tube and a camera are sequentially arranged above the multiple excitation modules;

[0029] An eyepiece is arranged on the trinocular tube.

[0030] Correspondingly, the present invention further provides a measurement method of a micro-area sample multispectral measurement system according to any embodiment of the present invention, comprising the following process:

[0031] Step 1, confirming the test light path: the computer receives the test instruction input by the operator, and according to the test type of the test instruction, the computer controls the first light source or the second light source to emit excitation light, and controls the light path switching device to switch the light path, so that the first light source or the second light source enters the test light path, and the CCD camera or the single photon detector enters the test light path;

[0032] Step 2, performing a test: excitation light enters the microscope to excite the sample to generate sample fluorescence, and the sample fluorescence is emitted through the microscope and enters the light selection device; after the light selection device performs light selection, the sample fluorescence after light selection enters the spectrometer; the CCD camera collects the fluorescence intensity and wavelength information of the sample after light selection, or the single photon detector collects the number of fluorescence photons and corresponding time information of the sample after light selection;

[0033] Step 3: When the excitation light wavelength a signal reaches the wavelength threshold, the computer controls the light selection device to switch to another filter and repeats step 2 for measurement;

[0034] Step 4: The computer calculates and processes the collected data.

[0035] Preferably, in step 1, if a fluorescence excitation spectrum test or a fluorescence emission spectrum test is performed, the first light source is selected to emit excitation light, the light path switching device selects to allow the first light source to pass, and the test light path is composed of a xenon lamp, a monochromator, a light path switching device, a microscope, a light selection device, a spectrometer, and a CCD camera in sequence;

[0036] If a fluorescence lifetime spectrum test is performed, a second light source is selected to emit excitation light, and the optical path switching device selects to allow the second light source to pass. The test optical path is composed of a pulsed laser, an optical path switching device, a microscope, a light selection device, a spectrometer, and a single-photon detector in sequence.

[0037] Preferably, in step 4, if a fluorescence excitation spectrum test is performed, the excitation light wavelength information a is selected as the x-coordinate and the sample fluorescence intensity information c is selected as the y-coordinate to obtain fluorescence excitation spectrum data;

[0038] If a fluorescence emission spectrum test is performed, the sample fluorescence wavelength information b is selected as the x-coordinate and the sample fluorescence intensity information c is selected as the y-coordinate to obtain the fluorescence emission spectrum data;

[0039] If a fluorescence lifetime spectrum test is performed, the relative time difference Δt is selected as the x-coordinate, and the number of fluorescence photons A0 corresponding to the relative time difference Δt is selected as the y-coordinate to obtain matrix data, which is the fluorescence lifetime spectrum data.

[0040] The present invention provides a micro-area sample multispectral measurement system and a measurement method thereof, which have the following advantages over the prior art:

[0041] 1. Regarding measurement methods, a high-pressure PLE testing scheme for samples is proposed, combining a high-pressure press with a micro-area PLE microscope. This simulates the high-pressure environment of the Earth's crust and allows for the study of the photoinduced properties of materials. The microscope provided by this invention meets the requirements for focusing an external laser spot onto a sample within the high-pressure press, achieving a micron-scale spot size, while also collecting the fluorescence signal generated by this spot, enabling PLE testing of micro-areas under high pressure.

[0042] 2. The present invention adds PLE, and corresponding detection devices for PL and TRPL spectral measurements, and simultaneously switches the optical path, combining PLE, PL and TRPL spectral measurements into one instrument, thereby realizing a multi-spectral measurement system for micro-area samples that is compatible with PLE, PL and TRPL spectral measurements.

[0043] 3. In terms of system composition, the optics are designed, the software control and calling logic are determined, and the obtained data are processed. This enables various spectral measurements such as fluorescence excitation spectrum (PLE), fluorescence emission spectrum (PL), and fluorescence lifetime spectrum (TRPL).

[0044] 4. The microscope provided by the present invention can achieve fixed optical path above and introduce spatial light source; the space below is large, which can realize the placement and measurement of high-pressure press, is easy to modify, and has strong regional modification of sample position, such as adding a low-temperature device; the position of the objective lens can be coarsely and finely adjusted up and down, retaining the focusing function of the microscope; the long working distance objective lens, the high-pressure press is placed below, and the sample is placed in the high-pressure press, which can meet the needs of high-pressure sample testing. BRIEF DESCRIPTION OF THE DRAWINGS

[0045] Figure 1 It is a structural schematic diagram of the micro-area sample multi-spectral measurement system provided by the present invention;

[0046] Figure 2 It is a structural schematic diagram of the microscope provided by the present invention;

[0047] Figure 3 This is a schematic diagram of the test results of fluorescence emission spectrum measurement performed by the micro-area sample multi-spectral measurement system provided by the present invention;

[0048] Figure 4It is a schematic diagram of the test results of fluorescence lifetime spectrum measurement performed by the micro-area sample multi-spectral measurement system provided by the present invention.

[0049] Figure numerals: 1. Xenon lamp; 2. Monochromator; 3. Optical path switching device; 4. Pulsed laser; 5. Microscope; 6. Light selection device; 7. Spectrometer; 8. CCD camera; 9. Single-photon detector; 10. Counter; 11. Computer; 501. Support base; 502. Light source support frame; 503. Sample; 504. Illumination light source; 505. Excitation module; 506. Eyepiece; 507. Camera; 508. Focusing frame; 509. Objective lens; 510. Three-dimensional moving stage; 511. Sample support frame. DETAILED DESCRIPTION

[0050] To make the technical problems solved, the technical solutions adopted, and the technical effects achieved by the present invention more clearly apparent, the present invention is further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the present invention and are not intended to limit the present invention. It should also be noted that, for ease of description, the accompanying drawings only illustrate portions relevant to the present invention, rather than all of the contents.

[0051] like Figure 1 As shown, the micro-area sample multi-spectral measurement system provided by the embodiment of the present invention includes: a first light source, a second light source, an optical path switching device 3, a microscope 5, a light selection device 6, a spectrometer 7, a CCD camera 8, a single photon detector 9, a counter 10 and a computer 11.

[0052] The first light source comprises a xenon lamp 1 and a monochromator 2. The monochromator 2 has an optical inlet and an optical outlet. Light from the xenon lamp 1 enters the monochromator 2 perpendicularly to the optical inlet and exits the monochromator 2 as a monochromatic beam. The optical inlet and optical outlet of the monochromator 2 are each in the form of a slit. The xenon lamp 1 and monochromator 2 are fixed in position.

[0053] The second light source adopts a pulse laser 4; a first optical adjustment device is set between the optical path switching device 3 and the pulse laser 4. The first optical adjustment device can adopt a reflector to adjust the light beam and transmit the excitation light emitted by the pulse laser 4 to the optical path switching device 3 through the reflector.

[0054] The first light source or the second light source emits excitation light, and the optical path switching device 3 selects the first light source or the second light source to pass through, allowing the excitation light to enter the microscope 5. A second optical adjustment device is provided between the optical path switching device 3 and the microscope 5. The second optical adjustment device can be a reflector to adjust the light beam and transmit the excitation light to the microscope 5 through the reflector.

[0055] Specifically, the optical path switching device 3 includes an adjustment seat and a reflector; the reflector is mounted on the adjustment seat at a 45-degree angle, and the adjustment seat can tilt or lift the reflector. The tilting or lifting of the reflector of the optical path switching device 3 can be controlled manually or automatically by the computer 11. If the optical path switching device 3 allows the first light source to pass through, its reflector is tilted down without obstruction, and the excitation light of the first light source can be directly projected into the microscope 5. If the optical path switching device 3 allows the second light source to pass through, the reflector reflects the excitation light of the second light source into the microscope 5 through its reflector.

[0056] The microscope 5 has a sample placement position, and the sample 503 is placed on the sample placement position via a high-pressure press. Excitation light enters the microscope 5, exciting the sample 503 to generate sample fluorescence, which is emitted through the microscope 5 and enters the light selection device 6. The high-pressure press is a common device in the high-pressure field. Its principle is a pair of diamonds facing each other. During use, the sample is placed on the sharp surface of the diamond, and the mechanical thread squeezes the diamond. Because the sharp surface of the diamond is very small (usually on the order of hundreds of microns), according to the principle of pressure P = pressure F / force-bearing area S, a sample is obtained under high pressure.

[0057] like Figure 2 As shown, the microscope 5 includes: a support base 501, a light source support frame 502, an illumination light source 504, an excitation module 505, a focusing frame 508, an objective lens 509, a three-dimensional moving stage 510 and a sample support frame 511.

[0058] A light source support frame 502 is provided on the support seat 501; an illumination light source 504 and a plurality of excitation modules 505 are provided on the light source support frame 502; a focusing frame 508 is provided on the upper part of the light source support frame 502, and an objective lens 509 is provided on the focusing frame 508; the focusing frame 508 can be raised and lowered and adjusted, and there are various adjustment methods, which are not limited in this application. One feasible method is given, for example, by installing a gear on the rotating shaft, engaging the gear with the rack on the focusing frame 508, and providing a knob on the rotating shaft, and rotating the knob can realize the raising and lowering of the focusing frame 508.

[0059] A trinocular tube and a camera 507 are sequentially arranged above the multiple excitation modules 505; an eyepiece 506 is arranged on the trinocular tube.

[0060] A three-dimensional moving platform 510 is provided below the light source support frame 502 , and a sample support frame 511 is provided on the three-dimensional moving platform 510 ; the sample support frame 511 has a sample placement position; the three-dimensional moving platform 510 is a mature product on the market and can realize movement in three directions of XYZ.

[0061] Through holes are provided on the light source support frame 502 at positions corresponding to the excitation module 505 and the objective lens 509 on the focusing frame 508. After the microscope 5 is adjusted, the excitation module 505, the objective lens 509, and the sample are placed in a straight line. The operator can observe the sample 503 through the camera 507, the excitation module 505, and the objective lens 509, facilitating focusing and observation.

[0062] The excitation light can enter from the bottom excitation module 505 , pass through the objective lens 509 , and irradiate the sample 503 , causing the sample 503 to generate sample fluorescence; the sample fluorescence passes through the objective lens 509 and multiple excitation modules 505 , and is emitted from the top excitation module 505 .

[0063] After the light selection device 6 performs light selection, the sample fluorescence enters the spectrometer 7. The light selection device 6 has multiple filters and can switch between filters to filter different wavelengths. The light selection device 6 can be implemented in a variety of ways, which are not limited by the present invention. One possible implementation method is described. For example, different filters can be installed along the circumference of a rotating disk. By rotating the rotating disk, one filter is placed in the test light path, allowing the filter to be switched.

[0064] The spectrometer 7 has a first optical outlet and a second optical outlet, the first optical outlet corresponds to a CCD camera 8, and the second optical outlet corresponds to a single-photon detector 9; the single-photon detector 9 collects the number of fluorescence photons of the sample after light selection and the corresponding time information, and the CCD camera 8 collects the fluorescence intensity and wavelength information of the sample after light selection;

[0065] The single photon detector 9 and the pulse laser 4 are respectively connected to the counter 10 for signal transmission; the monochromator 2 , the spectrometer 7 , the CCD camera 8 and the counter 10 are respectively connected to the computer 11 for signal transmission and are controlled by the computer 11 .

[0066] Correspondingly, the present invention also provides a measurement method of a micro-area sample multispectral measurement system, comprising the following steps:

[0067] Step 1, confirm the test light path: the computer 11 receives the test instruction input by the operator, and the computer 11 controls the first light source or the second light source to emit excitation light according to the test type of the test instruction, and controls the light path switching device 3 to switch the light path, so that the first light source or the second light source enters the test light path, and the CCD camera 8 or the single-photon detector 9 enters the test light path.

[0068] After the optical path switching device 3 switches the optical path, each instrument in the test optical path feeds back an electrical signal to the computer 11 to complete the corresponding optical path switching and test the optical path conduction.

[0069] If a fluorescence excitation spectrum (PLE) test or a fluorescence emission spectrum (PL) test is performed, the first light source is selected to emit excitation light, and the optical path switching device 3 selects to let the first light source pass. The test light path is composed of a xenon lamp 1, a monochromator 2, an optical path switching device 3, a microscope 5, a light selection device 6, a spectrometer 7, and a CCD camera 8 in sequence;

[0070] If a fluorescence lifetime spectroscopy (TRPL) test is performed, a second light source is selected to emit excitation light, and the optical path switching device 3 selects to allow the second light source to pass. The test light path is composed of a pulse laser 4, an optical path switching device 3, a microscope 5, a light selection device 6, a spectrometer 7, and a single-photon detector 9 in sequence.

[0071] Step 2, test: excitation light enters the microscope 5 to excite the sample 503 to generate sample fluorescence, and the sample fluorescence is emitted through the microscope 5 and enters the light selection device 6; after the light selection device 6 performs light selection, the sample fluorescence after light selection enters the spectrometer 7; the CCD camera 8 collects the fluorescence intensity and wavelength information of the sample after light selection, or the single photon detector 9 collects the number of fluorescence photons of the sample after light selection and the corresponding fluorescence photon arrival time information;

[0072] During the fluorescence excitation spectrum (PLE) test and fluorescence emission spectrum (PL) test, the monochromator 2 feeds back the excitation light wavelength information a to the computer 11; the spectrometer 7 feeds back the sample fluorescence wavelength information b after light selection to the computer 11, and the 8ccd camera feeds back the sample fluorescence light intensity information c after light selection to the computer 11;

[0073] During the fluorescence lifetime spectrum (TRPL) test process, the single-photon detector 9 collects the arrival time d of the sample fluorescence photons after light selection, and the corresponding number of fluorescence photons e. The single-photon detector 9 transmits the data d and e to the counter 10. At the same time, the pulse laser 4 also transmits the time information f of each pulse light to the counter 10. The counter 10 performs data calculation to obtain the relative time difference Δt between the arrival time d of the sample fluorescence photons after light selection and the time information f of each pulse light, and the number of fluorescence photons A0 within the corresponding relative time difference Δt.

[0074] Step 3: When the excitation light wavelength a signal reaches a wavelength threshold, the computer 11 controls the light selection device 6 to switch to another filter and repeats step 2 for measurement. The wavelength threshold is, for example, 500 nm.

[0075] Step 4: The computer 11 calculates and processes the collected data, and the software displays the corresponding test data.

[0076] If performing a fluorescence excitation spectrum (PLE) test, select the excitation light wavelength information a as the x-coordinate and the sample fluorescence light intensity information c as the y-coordinate to obtain the fluorescence excitation spectrum data. This data is the fluorescence excitation spectrum (PLE). During the above process, the sample fluorescence wavelength information b is also recorded. At this time, this data is a fixed value and serves as the test condition for this PLE test.

[0077] If performing a fluorescence emission spectrum (PL) test, the sample's fluorescence wavelength information (b) is used as the x-coordinate, and the sample's fluorescence intensity information (c) is used as the y-coordinate to obtain fluorescence emission spectrum data. This data is known as the fluorescence emission spectrum (PL). The above process also records the excitation light wavelength information (a). At this time, this data is a fixed value, which serves as the test condition for the PL test.

[0078] If performing a fluorescence lifetime spectroscopy (TRPL) test, the relative time difference Δt is used as the x-coordinate, and the number of fluorescence photons A0 within the corresponding relative time difference Δt is used as the y-coordinate to generate matrix data. This data is the fluorescence lifetime spectroscopy (TRPL) data. During this process, the excitation light wavelength information a and the sample fluorescence wavelength information b are simultaneously recorded. At this time, these data are fixed values, which constitute the TRPL test conditions.

[0079] Under the condition that the excitation light wavelength information a is 400nm, the fluorescence emission spectrum (PL) of a sample is measured and the following is obtained: Figure 3 The test results shown in the figure reflect that the sample can emit red fluorescence and that energy transfer occurs during the luminescence process.

[0080] Fluorescence lifetime spectroscopy (TRPL) measurements were performed to obtain Figure 4 The test result diagram shown here includes the sample fluorescence time and intensity information, indicating the duration of the sample's fluorescence. The longer the time scale of the test result, the longer the sample's fluorescence duration.

[0081] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the above embodiments, those skilled in the art should understand that modifications to the technical solutions described in the above embodiments, or equivalent replacement of some or all of the technical features therein, do not deviate the essence of the corresponding technical solutions from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A micro-area sample multispectral measurement system, characterized in that: include: A first light source, a second light source, an optical path switching device (3), a microscope (5), a light selection device (6), a spectrometer (7), a CCD camera (8), a single photon detector (9), a counter (10), and a computer (11); The first light source adopts a xenon lamp (1) and a monochromator (2); the monochromator (2) has an optical inlet and an optical outlet, and the light of the xenon lamp (1) enters the monochromator (2) perpendicularly to the optical inlet and exits the monochromator (2) as a monochromatic light beam from the optical outlet; The second light source adopts a pulse laser (4); a first optical adjustment device is provided between the optical path switching device (3) and the pulse laser (4); The optical path switching device (3) comprises an adjustment seat and a reflector; the reflector is mounted on the adjustment seat at a 45° angle, and the adjustment seat can cause the reflector to fall down or rise up; The microscope (5) comprises: a support base (501), a light source support frame (502), an illumination light source (504), an excitation module (505), a focusing frame (508), an objective lens (509), a three-dimensional moving stage (510) and a sample support frame (511); A light source support frame (502) is provided on the support base (501); an illumination light source (504) and a plurality of excitation modules (505) are provided on the light source support frame (502); A focusing frame (508) is provided on the upper portion of the light source support frame (502), and an objective lens (509) is provided on the focusing frame (508); A three-dimensional moving platform (510) is provided below the light source support frame (502), and a sample support frame (511) is provided on the three-dimensional moving platform (510); a sample placement position is provided on the sample support frame (511); Through holes are provided at positions corresponding to the excitation module (505) on the light source support frame (502) and at positions corresponding to the objective lens (509) on the focusing frame (508); After the microscope (5) is adjusted, the excitation module (505), the objective lens (509) and the sample placement position are on the same straight line; A trinocular tube and a camera (507) are sequentially arranged above the multiple excitation modules (505); An eyepiece (506) is provided on the trinocular tube; The first light source or the second light source emits excitation light, and the light path switching device (3) selects to allow the first light source or the second light source to pass through and allows the excitation light to enter the microscope (5); The microscope (5) has a sample placement position, and the sample (503) is placed on the sample placement position by a high-pressure press; The excitation light enters the microscope (5) to excite the sample (503) to generate sample fluorescence, and the sample fluorescence is emitted through the microscope (5) and enters the light selection device (6); After the light selection device (6) performs light selection, the sample fluorescence after light selection enters the spectrometer (7); The spectrometer (7) has a first optical outlet and a second optical outlet, the first optical outlet corresponds to a CCD camera (8), and the second optical outlet corresponds to a single-photon detector (9); The single-photon detector (9) collects the number of fluorescence photons of the sample after light selection and the corresponding fluorescence photon arrival time information, and the CCD camera (8) collects the fluorescence intensity and wavelength information of the sample after light selection; The single-photon detector (9) and the pulse laser (4) are respectively connected to the counter (10) for signal connection; the monochromator (2), the spectrometer (7), the CCD camera (8), and the counter (10) are respectively connected to the computer (11) for signal connection.

2. The micro-area sample multispectral measurement system according to claim 1, characterized in that: The optical inlet and the optical outlet of the monochromator (2) are respectively in the form of slits.

3. The micro-area sample multispectral measurement system according to claim 1, characterized in that: A second optical adjustment device is provided between the light path switching device (3) and the microscope (5).

4. A measurement method of a micro-area sample multi-spectral measurement system according to any one of claims 1 to 3, characterized in that: The following processes are included: Step 1, confirming the test light path: the computer (11) receives the test instruction input by the operator, and the computer (11) controls the first light source or the second light source to emit excitation light according to the test type of the test instruction, and controls the light path switching device (3) to switch the light path, so that the first light source or the second light source enters the test light path, and the CCD camera (8) or the single photon detector (9) enters the test light path; In step 1, if a fluorescence excitation spectrum test or a fluorescence emission spectrum test is performed, a first light source is selected to emit excitation light, and the light path switching device (3) selects to allow the first light source to pass through. The test light path is composed of a xenon lamp (1), a monochromator (2), a light path switching device (3), a microscope (5), a light selection device (6), a spectrometer (7), and a CCD camera (8) in sequence; If a fluorescence lifetime spectrum test is performed, a second light source is selected to emit excitation light, and the light path switching device (3) selects to allow the second light source to pass through. The test light path is composed of a pulse laser (4), a light path switching device (3), a microscope (5), a light selection device (6), a spectrometer (7), and a single photon detector (9). Step 2, performing a test: the excitation light enters the microscope (5) to excite the sample (503) to generate sample fluorescence, and the sample fluorescence is emitted through the microscope (5) and enters the light selection device (6); after the light selection device (6) performs light selection, the sample fluorescence after light selection enters the spectrometer (7); the CCD camera (8) collects the fluorescence intensity and wavelength information of the sample after light selection, or the single photon detector (9) collects the number of fluorescence photons of the sample after light selection and the corresponding time information; Step 3, when the excitation light wavelength a signal reaches the wavelength threshold, the computer (11) controls the light selection device (6) to switch to another filter, and repeats step 2 to perform measurement; Step 4: The computer (11) calculates and processes the collected data.

5. The measuring method according to claim 4, characterized in that In step 4, if a fluorescence excitation spectrum test is performed, the excitation light wavelength information a is selected as the x-coordinate and the sample fluorescence light intensity information c is selected as the y-coordinate to obtain the fluorescence excitation spectrum data; If a fluorescence emission spectrum test is performed, the sample fluorescence wavelength information b is selected as the x-coordinate and the sample fluorescence intensity information c is selected as the y-coordinate to obtain the fluorescence emission spectrum data; If a fluorescence lifetime spectrum test is performed, the relative time difference Δt is selected as the x-coordinate, and the number of fluorescence photons A0 corresponding to the relative time difference Δt is selected as the y-coordinate to obtain matrix data, which is the fluorescence lifetime spectrum data.

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