Calibration Method of Fluorescence Detector and Fluorescence Detector

Abstract

The present invention relates to the calibration of a fluorescence detector, and particularly to a calibration method and a fluorescence detector for a fluorescence detector. The calibration method of the fluorescence detector comprises the following steps: Step 1, using the fluorescence detector to be calibrated to detect a calibration sample to obtain a detected value a; Step 2, if the detected value a > a set value b, according to the detected value, reducing the light intensity of the excitation light source in a closed-loop manner through the control device of the fluorescence detector to be calibrated, so that the detected value obtained by detection = the set value b; if the detected value a < the set value b, according to the detected value, increasing the light intensity of the excitation light source in a closed-loop manner through the control device of the fluorescence detector to be calibrated, so that the detected value obtained by detection = the set value b. The fluorescence detector can adjust the excitation light source of the fluorescence detector according to the detected value, so that the light intensity of the excitation light source is reduced or increased; the above solution can solve the problems of difficult assembly and debugging during the calibration of the existing fluorescence detector and difficult control of the CV value.

Description

Technical Field

[0001] The present invention relates to the calibration of fluorescence detectors, and particularly to a calibration method for fluorescence detectors and fluorescence detectors. Background Art

[0002] Fluorescence immunoassay technology is a commonly used rapid detection technology in current biomedical tests. Its main advantages include strong specificity, high sensitivity, and fast speed, etc. Its basic principle is to combine the sensitive measurability of fluorescence with the highly specific reaction of antigen and antibody. Fluorescein is used to label Ab (antibody) or Ag (antigen), which binds to the corresponding Ag or Ab in the test specimen. By detecting fluorescence, it is determined whether there is the corresponding Ab or Ag in the specimen. Under the action of excitation light with a specific wavelength, the fluorescent substance can absorb light energy and enter the excited state, and release the previously absorbed light energy in the form of electromagnetic radiation to generate fluorescence. Specific fluorescence can be used directly, or can be received by a photoelectric converter and converted into an electrical signal for further processing, which can accurately, sensitively, and quickly locate and detect some trace or ultra-trace substances, and is widely used in many fields such as medicine, biology, and environmental protection.

[0003] An existing fluorescence detector, such as a fluorescence detector disclosed in a Chinese patent document with the publication number CN207850924U, includes a chassis unit and an optical path system. The optical path system is arranged in the chassis unit and includes an excitation light source and a CCD data acquisition module, and also includes a dichroic mirror. One side of the dichroic mirror is provided with an excitation optical path, and the other side is provided with a receiving optical path and a sampling convex lens. An aperture and an excitation filter are provided on the excitation optical path. The aperture is used to control the amount of light beam passing through to obtain a thin excitation beam, and the excitation filter is used to filter out stray light in the excitation light. An emission filter and a receiving-side convex lens are provided on the receiving optical path.

[0004] However, in actual production, the characteristics of optical detectors are that they generally have high requirements for the processing accuracy and assembly accuracy of optical components. Affected by factors such as the processing accuracy and assembly accuracy of components, even when detecting the same reagent strip, the excitation light intensity irradiated on the reagent strip by different fluorescence detectors varies greatly (a small excitation light intensity will cause the fluorescence not to be fully excited). Even if the fluorescence intensity excited is the same, the intensity of the same-intensity fluorescence collected by the CCD data acquisition module will also vary greatly. This will affect the CV value (coefficient of variation) of batch production. For a standard reagent strip, the smaller the CV value of the fluorescence detector, the better. The smaller the CV value, the higher the accuracy of the instrument calibration. Currently, during mass production, usually, the processing accuracy of components is appropriately improved, and manual grinding of components and debugging means are supplemented to ensure the consistency of detection results among multiple detectors. However, this method is difficult to assemble and debug and has a high cost.

[0005] A Chinese patent document with the publication number CN108535471A discloses an immunofluorescence detection optical path mechanism, an immunofluorescence detector and its calibration method. The focusing lens assembly facing the sample to be measured during use is set to be axially adjustable in position to reduce the requirements for the manufacturing precision of components, and at the same time reduce the difficulty of assembly and debugging, so as to achieve the purpose of reducing the assembly cost and debugging cost. However, the solution in the above patent document requires a precise drive mechanism to control the position of the focusing lens assembly, resulting in a relatively high cost of components, and the drive mechanism needs to occupy a certain space, resulting in a relatively large size of the fluorescence detector. Summary of the Invention

[0006] The object of the present invention is to provide a fluorescence detector to solve the problems of difficult assembly and debugging and difficult control of the CV value during calibration of the existing fluorescence detector. The object of the present invention is to provide a calibration method for a fluorescence detector, which can conveniently realize the calibration of the fluorescence detector and the control of the CV value.

[0007] The calibration method of the fluorescence detector in the present invention adopts the following technical solution:

[0008] The calibration method of the fluorescence detector includes the following steps:

[0009] Step 1: Use the fluorescence detector to be calibrated to detect the calibration sample, and obtain the detection value a according to the fluorescence signal collected by the CCD data acquisition module;

[0010] Step 2: If the detection value a > the set value b, according to the detection value, reduce the light intensity of the excitation light source in a closed-loop manner through the control device of the fluorescence detector to be calibrated, so that the detected detection value = the set value b;

[0011] If the detection value a < the set value b, according to the detection value, increase the light intensity of the excitation light source in a closed-loop manner through the control device of the fluorescence detector to be calibrated, so that the detected detection value = the set value b;

[0012] The set value b is a value less than the standard detection value c, and the standard detection value c is the detection value that should be detected when the calibration sample is fully excited.

[0013] Beneficial effects: By adopting the above technical solution, the detected value a is obtained based on the fluorescence signal collected by the CCD data acquisition module, and the light intensity of the excitation light source is reduced in a closed-loop manner through the control device of the fluorescence detector to be calibrated, so that the detected value obtained by detection is equal to the set value, thereby enabling the fluorescence detectors to have consistent detection results, ensuring the CV value of mass production. Compared with the prior art methods of improving the processing accuracy of components and supplementing with manual grinding of components for debugging, and adjusting the position of the focusing lens assembly, calibration can be achieved only through software control, which is easier to calibrate, beneficial to cost reduction, and can avoid the problems of high component cost and large space occupation caused by setting a driving mechanism to control the position of the focusing lens assembly.

[0014] As a preferred technical solution: Before adjusting the light intensity of the excitation light source, ensure that the light intensity of the excitation light source at the calibration sample < the light intensity required for the calibration sample to be fully excited.

[0015] Beneficial effects: Adopting the above technical solution is beneficial to flexibly adopt the adjustment method of increasing or decreasing the light intensity when adjusting the light intensity of the excitation light source, making the operation more convenient and easier to meet the CV value control requirements of different fluorescence detectors.

[0016] As a preferred technical solution: 0.85 × standard detected value c < set value b < 0.95 × standard detected value c.

[0017] Beneficial effects: Adopting the above technical solution can better ensure the excitation of counterfeits, and at the same time facilitate the smooth adjustment of increasing or decreasing the brightness of the excitation light source.

[0018] As a preferred technical solution: The set value b = 0.9 × standard detected value c.

[0019] As a preferred technical solution:

[0020] The standard detected value c is obtained by the following method:

[0021] Set a standard fluorescence detector, the structure of the standard fluorescence detector is the same as that of the fluorescence detector to be calibrated, and the processing accuracy and assembly accuracy of the components all meet the design accuracy requirements;

[0022] Use this standard fluorescence detector to detect the calibration sample, fully excite the calibration sample, and the detected value obtained is the standard detected value c.

[0023] Beneficial effects: Adopting the above technical solution can make the standard detected value c more matched with the fluorescence detector to be calibrated, which is beneficial to enabling the fluorescence detector to be calibrated to better achieve the design performance.

[0024] The fluorescence detector in the present invention adopts the following technical solution:

[0025] Fluorescence detector, comprising:

[0026] A dichroic mirror, with an excitation light source and a CCD data acquisition module respectively arranged on both sides of the dichroic mirror;

[0027] A control device, connected to the excitation light source and the CCD data acquisition module;

[0028] When using the fluorescence detector to be calibrated to detect a calibration sample:

[0029] If the detected value a obtained from the fluorescence signal collected by the CCD data acquisition module is greater than the set value b, the control device can adjust the excitation light source of the fluorescence detector to reduce the light intensity of the excitation light source;

[0030] If the detected value a is less than the set value b, the control device can adjust the excitation light source of the fluorescence detector to increase the light intensity of the excitation light source.

[0031] Advantageous effects: By adopting the above technical solution, the detected value a is obtained according to the fluorescence signal collected by the CCD data acquisition module, and the light intensity of the excitation light source is reduced in a closed-loop manner through the control device of the fluorescence detector to be calibrated, so that the detected value obtained by detection is equal to the set value, thereby enabling each fluorescence detector to have consistent detection results, ensuring the CV value of batch production. Compared with the prior art methods of improving the processing accuracy of components and supplementing with manual grinding of components for debugging, and the method of adjusting the position of the focusing lens assembly, calibration can be achieved only through software control, which is easier to calibrate, conducive to reducing costs, and can avoid the problems of high component costs and large space occupation caused by setting a driving mechanism to control the position of the focusing lens assembly.

[0032] As a preferred technical solution: The light intensity of the excitation light source at the calibration sample can be adjusted to be less than the light intensity required for the calibration sample to be fully excited.

[0033] Advantageous effects: By adopting the above technical solution, it is beneficial to flexibly adopt the adjustment methods of increasing or decreasing the light intensity when adjusting the light intensity of the excitation light source, which is more convenient to operate and easier to meet the CV value control requirements of different fluorescence detectors. Description of the Drawings

[0034] Figure 1 is a schematic structural diagram of Embodiment 1 of a fluorescence detector in the present invention;

[0035] The names of the components corresponding to the corresponding reference numerals in the figure are: 11, dichroic mirror; 21, excitation light source; 22, excitation-side convex lens; 31, receiving-side convex lens; 32, CCD data acquisition module; 41, sample-side convex lens; 42, slit element; 51, calibration sample; 61, control device. Detailed implementation manners

[0036] In order to make the objectives, technical solutions and advantages of the present invention more clear and understandable, the present invention will be further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention, rather than to limit the present invention, that is, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. Generally, the components of the embodiments of the present invention described and illustrated in the drawings here can be arranged and designed in various different configurations.

[0037] Therefore, the detailed description of the embodiments of the present invention provided in the drawings below is not intended to limit the scope of the claimed present invention, but only represents the selected embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative efforts belong to the scope of protection of the present invention.

[0038] It should be noted that the relative terms such as "first" and "second" that may appear in the specific implementation manners of the present invention are only used to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Moreover, the terms "including", "comprising" or any other variants thereof that may appear are intended to cover non-exclusive inclusion, so that a process, method, article or device including a series of elements not only includes those elements, but also includes other elements not explicitly listed, or also includes elements inherent to such process, method, article or device. Without further limitation, the element defined by the statement "including one..." does not exclude the existence of another identical element in the process, method, article or device including the said element.

[0039] In the description of the present invention, unless otherwise clearly defined and limited, the terms "installed", "connected" and "coupled" that may appear should be understood in a broad sense. For example, it may be a fixed connection, a detachable connection, or an integral connection; it may be a mechanical connection or an electrical connection; it may be a direct connection or an indirect connection through an intermediate medium, and it may be the communication inside two elements. For those skilled in the art, the specific meanings of the above terms in the present invention can be understood through specific situations.

[0040] In the description of the present invention, unless otherwise clearly defined and limited, the term "provided with" that may appear should be understood in a broad sense. For example, the object of "provided with" may be a part of the body, or may be arranged separately from the body and connected to the body, and this connection may be a detachable connection or a non-detachable connection. For those skilled in the art, the specific meanings of the above terms in the present invention can be understood through specific situations.

[0041] The present invention will be further described in detail below in conjunction with embodiments.

[0042] Embodiment 1 of the fluorescence detector in the present invention:

[0043] As Figure 1 shown, the fluorescence detector includes a housing (not shown in the figure), and an optical path system and a control device 61 are provided inside the housing. The optical path system includes a dichroic mirror 11. On one side of the dichroic mirror 11, an excitation light source 21 and an excitation-side convex lens 22 are provided. The excitation-side convex lens 22 is located on the excitation optical path and constitutes a part of the excitation optical path for the excitation light to pass through. On the other side of the dichroic mirror 11, a receiving-side convex lens 31 and a CCD data acquisition module 32 are provided. The receiving-side convex lens 31 is located on the receiving optical path and constitutes a part of the receiving optical path.

[0044] On the side of the dichroic mirror 11 where the CCD data acquisition module 32 is provided, a sample-side convex lens 41 and a slit element 42 are also provided. The sample-side convex lens 41 faces the calibration sample 51 during use. The slit element 42 is provided on the side of the sample-side convex lens 41 facing away from the dichroic mirror 11 and is used to form a slit. The slit element 42 plays a role in beam shaping, preventing external stray light from entering the detection optical path, matching the test line and the quality control line on the reagent strip, and is beneficial to improving the detection accuracy. The slit element 42 is provided near the calibration sample 51 of the fluorescence detector, which is convenient for assembly and adjustment.

[0045] The optical path from the dichroic mirror 11 to the calibration sample 51 is a common optical path for the excitation light and the fluorescence.

[0046] After the fluorescence detector is assembled, it needs to be calibrated and the CV value control in mass production is realized. First, during mass production, strictly control the machining accuracy and assembly accuracy of the parts of the first set of fluorescence detector products, so that both the machining accuracy and assembly accuracy of the parts meet the design accuracy requirements. For example, during assembly, the assembly accuracy is ensured by grinding. Then, use this set of fluorescence detectors as the standard fluorescence detectors. The standard fluorescence detectors have the same structure as the fluorescence detectors to be calibrated. Use the standard fluorescence detectors to detect the standard reagent strip used as the calibration sample, increase the brightness of the excitation light source, and completely excite the calibration sample. The obtained detection value is the standard detection value c. Among them, the standard reagent strip is a reagent strip added with a sample of a certain concentration. For samples with different concentrations, the standard reagent strip has specific fluorescence detection values. For example, a sample of 2 g / ml corresponds to one fluorescence detection value, and a sample of 3 g / ml corresponds to another fluorescence detection value. However, the calibration sample is only used to calibrate the remaining mass-produced products to control the CV value, so its concentration will not affect the detection results of the calibrated products.

[0047] When calibrating the remaining fluorescence detectors, place the standard reagent strip, which serves as a calibration sample, on the sample-bearing part of the fluorescence detector to be calibrated. The ultraviolet excitation beam emitted by the excitation light source 21 converges at the calibration sample 51 after passing through the dichroic mirror 11, the sample-side convex lens 41, and the slit element 42. The fluorescent agent attached to the calibration sample 51 is excited by the monochromatic ultraviolet light and emits fluorescence. After the fluorescence passes through the slit element 42, the sample-side convex lens 41, and the dichroic mirror 11, it is converged by the receiving-side convex lens 31 and the excitation light other than the fluorescence is filtered out, and then irradiates the CCD data acquisition module 32. The CCD data acquisition module 32 converts the received optical signal into an electrical signal and transmits it to the control device 61 to obtain the detection value a. The control device 61 outputs a power adjustment signal to the excitation light source 21 according to the detection value a based on the embedded algorithm to adjust the light intensity of the light source, forming a closed-loop control. Specifically, if the detection value a > the set value b, the light intensity of the excitation light source is reduced until the detection value of the CCD data acquisition module = the set value b; if the detection value a < the set value b, the light intensity of the excitation light source is increased until the detection value of the CCD data acquisition module = the set value b. In this embodiment, the set value b = 0.9 × the standard detection value c, and the control device 61 uses a microprocessor. In other embodiments, the detection value obtained when other fluorescence detectors detect the standard reagent strip can also be determined as the set value b.

[0048] To facilitate the consistency of the detection results of different fluorescence detectors for the same calibration sample, the light intensity of the excitation light source at the calibration sample can be adjusted to be less than the light intensity required for the calibration sample to be fully excited. For example, a light intensity detection device can be used to detect the light intensity at the calibration sample. Before adjusting the light intensity of the excitation light source, first make the light intensity of the excitation light source at the calibration sample < the light intensity required for the calibration sample to be fully excited. In this way, when detecting the calibration sample, according to the detection result, the detection value can be made equal to the set value b either by reducing the light intensity of the excitation light source or by increasing the light intensity of the excitation light source. In addition, taking the standard detection value that should be detected when the calibration sample is fully excited as c, the set value b < the standard detection value c.

[0049] Embodiment 2 of the fluorescence detector in the present invention:

[0050] The difference between this embodiment and Embodiment 1 is that in Embodiment 1, the light intensity of the excitation light source at the calibration sample can be adjusted to be less than the light intensity required for the calibration sample to be fully excited; while in this embodiment, before detecting the calibration sample, the light intensity of the excitation light source at the calibration sample can be adjusted to be greater than the light intensity required for the calibration sample to be fully excited. At this time, the detection value a can be adjusted by reducing the light intensity of the excitation light source.

[0051] Embodiment 1 of the method for calibrating the fluorescence detector of the present invention:

[0052] The calibration method comprises the following steps:

[0053] Step 1: Use a light intensity detection device to detect the light intensity at the calibration sample to ensure that the light intensity of the excitation light source at the calibration sample before calibration is less than the light intensity required when the calibration sample is fully excited;

[0054] Then, a standard reagent strip is selected as a calibration sample, and the calibration sample is detected using a fluorescence detector to be calibrated, and a detection value a is obtained according to the fluorescence signal collected by a CCD data acquisition module; the CCD data acquisition module is connected to a control device of the fluorescence detector;

[0055] Step 2: During calibration, the standard detection value c that should be detected when the calibration sample is fully excited is selected as the target value that is smaller than the quasi-detection value c as the set value b; in this embodiment, the set value b=0.9×standard detection value c.

[0056] If the detection value a> the set value b, according to the detection value, the light intensity of the excitation light source is reduced in a closed-loop manner through the control device of the fluorescence detector to be calibrated, so that the detection value obtained by the detection is equal to the set value b;

[0057] If the detection value a is less than the set value b, the light intensity of the excitation light source is increased in a closed-loop manner through the control device of the fluorescence detector to be calibrated according to the detection value, so that the detection value obtained by the detection is equal to the set value b.

[0058] The set value b is a value smaller than the standard detection value c, and the standard detection value c is the detection value that should be detected when the calibration sample is fully excited.

[0059] The above standard test value c is obtained by the following method:

[0060] First, during batch production, strictly control the processing accuracy and assembly accuracy of the components of the first set of fluorescence detector products, so that the processing accuracy and assembly accuracy of the components meet the design accuracy requirements. For example, the assembly accuracy is ensured by grinding during assembly. Then, use this set of fluorescence detectors as standard fluorescence detectors. The standard fluorescence detector has the same structure as the fluorescence detector to be calibrated. Use this standard fluorescence detector to detect the standard reagent strip used as the calibration sample, increase the brightness of the excitation light source, so that the calibration sample is fully excited, and the obtained detection value is the standard detection value c. Among them, the standard reagent strip is a reagent strip with a certain concentration of sample added. For samples of different concentrations to be tested, the standard reagent strip has a specific fluorescence detection value.

[0061] According to the above calibration method, the CV value of the batch can be guaranteed by using the same calibration sample or calibration samples with the same characteristics to detect each fluorescence detector to be calibrated in the batch.

[0062] Example 2 of the calibration method for the fluorescence detector in the present invention:

[0063] The difference between this example and Example 1 is that in Example 1, the light intensity of the excitation light source at the calibration sample was adjusted to be less than the light intensity required for the calibration sample to be fully excited; while in this example, before detecting the calibration sample, the light intensity of the excitation light source at the calibration sample was adjusted to be greater than the light intensity required for the calibration sample to be fully excited. At this time, the detected value a can be adjusted by reducing the light intensity of the excitation light source.

[0064] As mentioned above, the above are only the preferred embodiments of the present application and are not intended to limit the present application. The patent protection scope of the present application is subject to the claims. Any equivalent structural changes made by using the description and drawings of the present application should similarly be included in the protection scope of the present application.

Claims

1. A calibration method for a fluorescence detector, characterized in that, Including the following steps: Step 1: Use the fluorescence detector to be calibrated to detect the calibration sample, and obtain the detection value a according to the fluorescence signal collected by the CCD data acquisition module; Step 2: If the detection value a > the set value b, according to the detection value, reduce the light intensity of the excitation light source in a closed-loop manner through the control device of the fluorescence detector to be calibrated, so that the detected detection value = the set value b; If the detection value a < the set value b, according to the detection value, increase the light intensity of the excitation light source in a closed-loop manner through the control device of the fluorescence detector to be calibrated, so that the detected detection value = the set value b; The range of the set value b is 0.85 × standard detection value c < set value b < 0.95 × standard detection value c, and the standard detection value c is the detection value that should be detected when the calibration sample is fully excited.

2. The calibration method of the fluorescence detector according to claim 1, characterized in that Before adjusting the light intensity of the excitation light source, ensure that the light intensity of the excitation light source at the calibration sample < the light intensity required for the calibration sample to be fully excited.

3. The calibration method of the fluorescence detector according to claim 1, characterized in that The set value b = 0.9 × standard detection value c.

4. The calibration method of the fluorescence detector according to claim 1 or 2, characterized in that, The standard detection value c is obtained by the following method: Set a standard fluorescence detector, which has the same structure as the fluorescence detector to be calibrated, and the processing accuracy and assembly accuracy of its components both meet the design accuracy requirements; Use this standard fluorescence detector to detect the calibration sample to fully excite the calibration sample, and the obtained detection value is the standard detection value c.

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