Inspection device and inspection method

Abstract

Provided is an inspection device and an inspection method that can rapidly measure microorganisms. [Solution] The testing device 1 for testing a target microorganism X in a specified specimen 100 has an antibody Y that undergoes an antigen-antibody reaction with the target microorganism X, a fluorescent unit 11 that causes the specified specimen 100 to fluoresce after the target microorganism X reacts with the antibody Y that has been pre-bound to a specified fluorescent reagent Z, and a fluorescence amount measuring unit 12 that measures the amount of fluorescence of a specific wavelength emitted from the specimen 100 that has fluoresced due to the fluorescent unit 11.

Description

[Technical field]

[0001] The present invention relates to a testing device and a testing method, and more particularly to a testing device and a testing method for testing a target microorganism in a given sample. [Background technology]

[0002] Conventionally, for example, a culture method has been used for qualitative and quantitative measurement of microorganisms such as viruses, bacteria, and fungi. For such a culture method, the technology disclosed in Patent Document 1 can be referred to. [Prior art documents] [Patent documents]

[0003] [Patent Document 1] JP 2015-198598 A Summary of the Invention [Problem to be solved by the invention]

[0004] Incidentally, quantitative analysis of bacteria, for example, among microorganisms, requires a culture time of 24 to 48 hours. Quantitative analysis of fungi (mold, yeast) requires a culture time of about 2 to 7 days. Furthermore, qualitative analysis by species requires an additional 7 days of identification culture or identification under a microscope. In other words, it takes about 2 weeks to determine the results of qualitative and quantitative analysis of microorganisms.

[0005] The present invention has been made in consideration of the above circumstances, and has an object to provide an inspection device and an inspection method that can rapidly measure microorganisms. [Means for solving the problem]

[0006] In order to achieve the above-mentioned object, the testing device of the present invention is a testing device for testing a target microorganism in a specified specimen, and has a fluorescent unit that makes the specified specimen fluoresce after reacting the target microorganism with an antibody that undergoes an antigen-antibody reaction with the target microorganism and has been pre-bound to a specified fluorescent reagent, and a fluorescence amount measuring unit that measures the amount of fluorescence of a specific wavelength emitted from the specimen that has become fluorescent due to the fluorescent unit.

[0007] According to the present invention, the apparatus has a fluorescent unit that causes the specified specimen to fluoresce after the target microorganism is reacted with the antibody that undergoes an antigen-antibody reaction with the target microorganism and that has been bound to a specified fluorescent reagent in advance, and a fluorescence amount measuring unit that measures the amount of fluorescence of a specific wavelength emitted from the specimen that has been fluorescentized by the fluorescent unit, thereby making it possible to measure the amount of fluorescence of a specific wavelength emitted from the specimen through the antigen-antibody reaction between the target microorganism and the antibody. As a result, the microorganisms can be measured quickly without the need for culturing the microorganisms.

[0008] The fluorescent part is a fluorescent part that causes a target microorganism that has undergone an antigen-antibody reaction with an antibody that has been bound in advance to the specified fluorescent reagent in the specified specimen to fluoresce, and does not cause microorganisms other than the target microorganism to fluoresce.

[0009] The antibody is an antibody that undergoes the antigen-antibody reaction with the target microorganism, but does not undergo the antigen-antibody reaction with microorganisms other than the target microorganism.

[0010] The device has a correlation memory unit that stores data indicating the correlation between the amount of fluorescence of the specific wavelength emitted from the target microorganism and the number of the target microorganisms, and a target microorganism number calculation unit that calculates the number of the target microorganisms based on the amount of fluorescence of the specific wavelength emitted from the specified specimen measured by the fluorescence amount measuring unit, which is the amount of fluorescence of the specific wavelength emitted from the target microorganisms, and the data indicating the correlation stored in the correlation memory unit, thereby making it possible to calculate the number of target microorganisms from the amount of fluorescence of the specific wavelength emitted from the target microorganisms.

[0011] The predetermined fluorescent reagent is pre-bound to the antibody while being incorporated into the antibody.

[0012] The fluorescence amount measuring unit has a light amount sensor using a photomultiplier tube and / or a semiconductor, and can measure the amount of fluorescence by the photomultiplier tube and / or the light amount sensor using the semiconductor.

[0013] The antibody is preferably an antibody extracted by immunizing a mammal with the target microorganism or a protein antigen having the same structure as the protein structure of the target microorganism.

[0014] The antibody is preferably a monoclonal or polyclonal antibody.

[0015] The predetermined fluorescent reagent is preferably at least one of merocyanine, perylene, acridine, luciferin, pyranine, stilbene, and rhodamine.

[0016] In order to achieve the above-mentioned object, the testing method of the present invention is a testing method for testing a target microorganism in a specified specimen, which includes a fluorescence step of making the specified specimen fluoresce after reacting the target microorganism with an antibody that undergoes an antigen-antibody reaction with the target microorganism and which has been pre-bound to a specified fluorescent reagent, and a fluorescence amount measurement step of measuring the amount of fluorescence of a specific wavelength emitted from the specimen that has become fluorescent in the fluorescence step.

[0017] According to the present invention, the method includes a fluorescence step of making the specified specimen fluoresce after reacting the target microorganism with an antibody that undergoes an antigen-antibody reaction with the target microorganism and that has been bound to a specified fluorescent reagent in advance, and a fluorescence amount measurement step of measuring the amount of fluorescence of a specific wavelength emitted from the specimen that has become fluorescent in the fluorescence step, thereby making it possible to measure the amount of fluorescence of a specific wavelength emitted from the specimen through the antigen-antibody reaction between the target microorganism and the antibody. As a result, microorganisms can be measured quickly without culturing the microorganisms.

[0018] The fluorescence step is a step of causing a target microorganism in the specified specimen that has undergone an antigen-antibody reaction with an antibody that has been bound in advance to the specified fluorescent reagent to fluoresce, and causing microorganisms other than the target microorganism not to fluoresce.

[0019] The antibody is an antibody that undergoes the antigen-antibody reaction with the target microorganism, but does not undergo the antigen-antibody reaction with microorganisms other than the target microorganism.

[0020] The method has a correlation storage step for storing data showing the correlation between the amount of fluorescence of the specific wavelength emitted from the target microorganism and the number of the target microorganisms, and a target microorganism number calculation step for calculating the number of the target microorganisms based on the amount of fluorescence of the specific wavelength emitted from the specified specimen measured in the fluorescence amount measurement step, which is the amount of fluorescence of the specific wavelength emitted from the target microorganisms, and the data showing the correlation stored in the correlation storage step, thereby making it possible to calculate the number of target microorganisms from the amount of fluorescence of the specific wavelength emitted from the target microorganisms.

[0021] The predetermined fluorescent reagent is pre-bound to the antibody while being incorporated into the antibody.

[0022] The fluorescence amount measuring step may include a step of having a light amount sensor using a photomultiplier tube and / or a semiconductor, and measuring the amount of fluorescence with the light amount sensor using the photomultiplier tube and / or the semiconductor.

[0023] The antibody is preferably an antibody extracted by immunizing a mammal with the target microorganism or a protein antigen having the same structure as the protein structure of the target microorganism.

[0024] The antibody is preferably a monoclonal or polyclonal antibody.

[0025] The predetermined fluorescent reagent is preferably at least one of merocyanine, perylene, acridine, luciferin, pyranine, stilbene, and rhodamine. Effect of the Invention

[0026] According to the present invention, microorganisms can be measured quickly. [Brief description of the drawings]

[0027] [Figure 1] 1 is a diagram showing a configuration of an inspection device according to an embodiment of the present invention. [Diagram 2] FIG. 2 is a diagram showing a configuration of an inspection unit of the inspection device. [Diagram 3] FIG. 2 is a diagram showing the configuration of a calculation processing unit of the inspection device. [Figure 4] FIG. 2 is a diagram illustrating a sample that is an object to be inspected by the inspection device. [Diagram 5] FIG. 1 is a schematic diagram of an antibody. [Figure 6] FIG. 2 is a diagram illustrating a fluorescent reagent. [Figure 7] FIG. 1 is a diagram showing the state in which an antibody and a fluorescent reagent are bound to each other. [Figure 8] FIG. 2 is a diagram showing the state in which an antigen-antibody reaction occurs between a target microorganism and an antibody. [Figure 9] FIG. 2 is a diagram showing a configuration of a computer of the inspection device. [Figure 10] 10 is a flowchart for explaining an inspection method in the inspection device. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0028] Hereinafter, an embodiment of the present invention will be described in detail with reference to the drawings. Fig. 1 is a diagram showing the configuration of a testing device according to an embodiment of the present invention, Fig. 2 is a diagram showing the configuration of a testing unit of the testing device, Fig. 3 is a diagram showing the configuration of a calculation processing unit of the testing device, Fig. 4 is a diagram showing a sample to be tested by the testing device, Fig. 5 is a diagram showing an antibody, Fig. 6 is a diagram showing a fluorescent reagent, Fig. 7 is a diagram showing a state in which an antibody and a fluorescent reagent are bound, Fig. 8 is a diagram showing a state in which an antigen-antibody reaction has occurred between a target microorganism and an antibody, Fig. 9 is a diagram showing the configuration of a computer of the testing device, and Fig. 10 is a flow chart for explaining a testing method in the testing device.

[0029] The inspection device 1 according to the embodiment of the present invention can inspect target microorganisms X in a predetermined specimen 100. As shown in FIG. 1, the inspection device 1 has an inspection section 10 and a calculation processing section 20. As shown in FIG. 2, the inspection section 10 further has a fluorescent section 11 and a fluorescence amount measuring section 12. As shown in FIG. 3, the calculation processing section 20 further has a correlation storage section 21 and a target microorganism number calculation section 22. The inspection device 1 can measure the amount of fluorescence of a predetermined specimen 100 shown in FIG. 4 and calculate the number of target microorganisms X in the predetermined specimen 100. The target microorganisms X are target microorganisms, and are microorganisms to be inspected by the inspection device 1.

[0030] In this embodiment, antibody Y shown in Fig. 5 and fluorescent reagent Z shown in Fig. 6 are used to measure the amount of fluorescence from a predetermined sample 100. As shown in Fig. 7, the predetermined fluorescent reagent Z is bound to antibody Y in advance. That is, the predetermined fluorescent reagent Z is incorporated into antibody Y and bound to antibody Y in advance. Antibody Y is an antibody Y that uses target microorganism X as an antigen and undergoes an antigen-antibody reaction with target microorganism X, but does not undergo an antigen-antibody reaction with microorganisms other than target microorganism X. Antibody Y can be selected appropriately for each target microorganism X.

[0031] Here, the antibody Y is preferably an antibody Y extracted by immunizing a mammal with the target microorganism X or a protein antigen having the same structure as the protein structure of the target microorganism X. More specifically, the antibody Y is preferably an antibody Y extracted from the serum or chicken egg of a mammal immunized with the target microorganism X or a protein antigen having the same structure as the protein structure of the target microorganism X (the antibody Y is an antibody Y extracted from the serum or chicken egg of a mammal immunized with the target microorganism X by immunizing a mammal such as a mouse or chicken, or the antibody Y is an antibody Y extracted from the serum or chicken egg of a mammal immunized with the protein antigen having the same structure as the protein structure of the target microorganism X by immunizing a mammal such as a mouse or chicken), and the antibody Y is preferably a monoclonal antibody or a polyclonal antibody. In addition, the predetermined fluorescent reagent Z is preferably at least one of merocyanine, perylene, acridine, luciferin, pyranine, stilbene, and rhodamine.

[0032] That is, the fluorescent unit 11 is an antibody Y that undergoes an antigen-antibody reaction with the target microorganism X, and can cause a predetermined specimen 100 to fluoresce after the antibody Y, which has been bound in advance to a predetermined fluorescent reagent Z, reacts with the target microorganism X. The fluorescent unit 11 can cause the predetermined specimen 100 to fluoresce by irradiating the predetermined specimen 100 with excitation light (e.g., 488 nm±10 nm).

[0033] In addition, the fluorescent unit 11 is a fluorescent unit 11 that causes a target microorganism X that has undergone an antigen-antibody reaction with an antibody Y that has been pre-bound to a specified fluorescent reagent Z in a specified specimen 100 to fluoresce, and does not cause a microorganism X' other than the target microorganism X to fluoresce.

[0034] 8, in a given specimen 100, a target microorganism X that has undergone an antigen-antibody reaction with an antibody Y that has been bound in advance to a given fluorescent reagent Z can be made to fluoresce by applying the given fluorescent reagent Z. The fluorescent unit 11 is configured to have a cell holder filled with the specimen 100 inside a casing that blocks 100% of external light, and excitation light can be irradiated from the sides (first to fourth sides) of the cell holder.

[0035] The fluorescence amount measuring unit 12 can measure the amount of fluorescence of a specific wavelength (for example, 512 nm±5 nm) emitted from the specimen 100 that has been fluorescently emitted by the fluorescent unit 11. The fluorescence amount measuring unit 12 has a light amount sensor using a photomultiplier tube and / or a semiconductor, and can measure the amount of fluorescence using the light amount sensor using a photomultiplier tube and / or a semiconductor. The amount of fluorescence measured by the fluorescence amount measuring unit 12 can be displayed digitally.

[0036] The correlation storage unit 21 can store data showing the correlation between the amount of fluorescence of a specific wavelength emitted from the target microorganism X and the number of the target microorganisms X. The data showing the correlation includes data correlating the amount of fluorescence of a specific wavelength emitted from the target microorganism X and the number of the target microorganisms X, mathematical formula data of the amount of fluorescence of a specific wavelength emitted from the target microorganism X and the number of the fluorescent target microorganisms X, and the like.

[0037] The target microorganism count calculation unit 22 can calculate the number of target microorganisms X based on the amount of fluorescence of a specific wavelength emitted from a specified sample 100 measured by the fluorescence amount measurement unit 12, which is the amount of fluorescence of a specific wavelength emitted from the target microorganism X, and data indicating the correlation stored in the correlation memory unit 21.

[0038] The amount of fluorescence measured by the fluorescence amount measuring unit 12 and the number of target microorganisms X calculated by the target microorganism number calculating unit 22 can be displayed digitally.

[0039] The inspection device 1 has a general configuration as a computer. That is, as shown in Fig. 9, the inspection device 1 has a central processing unit (CPU, GPU, DSP) 1B, a storage device (ROM, RAM, hard disk, cache memory) 1C, an input device (keyboard, touch panel, mouse) 1D, a display device (liquid crystal display) 1E, etc., which are mutually connected via a bus 1A, and these can function as an arithmetic processing unit 20 by interacting with each other. The storage device 1C functions as a computer-readable storage medium.

[0040] The inspection method using the inspection device 1 configured as above will be described with reference to the flowchart in Fig. 10. In the following, the correlation storage unit 21 will be described as having performed the correlation storage step and pre-stored data showing the correlation between the amount of fluorescence of a specific wavelength emitted from the target microorganisms X and the number of the target microorganisms X.

[0041] First, in step S10, a predetermined specimen 100 is prepared. That is, an antibody Y shown in Fig. 5 and a fluorescent reagent Z shown in Fig. 6 are prepared, and the predetermined fluorescent reagent Z and the antibody Y are preliminarily bound to each other as shown in Fig. 7. The predetermined fluorescent reagent Z is incorporated into the antibody Y and preliminarily bound to the antibody Y.

[0042] Next, in step S20, the fluorescent unit 11 irradiates excitation light onto a specified specimen 100 after the reaction between the target microorganism X and antibody Y, which undergoes an antigen-antibody reaction with the target microorganism X and has been previously bound to a specified fluorescent reagent Z, causing the specified specimen 100 to fluoresce (fluorescence step).

[0043] Next, in step S30, the fluorescence amount measuring section 12 measures the amount of fluorescence of a specific wavelength emitted from the specimen 100 that has been fluoresced by the fluorescent section 11 (fluorescence amount measuring step).

[0044] Next, in step S40, the target microorganism count calculation unit 22 calculates the number of target microorganisms X based on the amount of fluorescence of a specific wavelength emitted from a specific specimen 100 measured by the fluorescence amount measurement unit 12, which is the amount of fluorescence of a specific wavelength emitted from the target microorganism X, and the data indicating the correlation stored in the correlation memory unit 21 (target microorganism count calculation step).

[0045] As described above, the testing device 1 of the present invention has the fluorescent unit 11 that fluoresces the specified specimen 100 after the antibody Y that causes an antigen-antibody reaction with the target microorganism X and that is bound in advance to a specified fluorescent reagent Z is reacted with the target microorganism X, and the fluorescent amount measuring unit 12 that measures the amount of fluorescence of a specific wavelength emitted from the specimen 100 that has been fluorescentized by the fluorescent unit 11, thereby making it possible to measure the amount of fluorescence of a specific wavelength emitted from the specimen 100 through the antigen-antibody reaction between the target microorganism X and the antibody Y. As a result, it is possible to rapidly measure microorganisms without culturing the microorganisms. (Note that, as a means for performing qualitative and quantitative analysis using an antigen-antibody reaction, a dye determination using two antibodies by the ELISA method has generally been used in the past, but it takes about 5 hours, is very difficult to operate, requires a large-scale measuring device, and has high running and initial costs. It is possible to operate it in the medical field, but in the food industry, where the need for rapid measurement is greatest, operation is difficult, and it is considered that the present invention is in wide use.)

[0046] In addition, the device has a correlation memory unit 21 that stores data showing the correlation between the amount of fluorescence of a specific wavelength emitted from the target microorganism X and the number of the target microorganisms X, and a target microorganism number calculation unit 22 that calculates the number of the target microorganisms X based on the amount of fluorescence of a specific wavelength emitted from a specific specimen 100 measured by a fluorescence amount measuring unit 12, which is the amount of fluorescence of a specific wavelength emitted from the target microorganism X, and the data showing the correlation stored in the correlation memory unit 21, thereby making it possible to calculate the number of the target microorganisms X from the amount of fluorescence of a specific wavelength emitted from the target microorganisms X.

[0047] Furthermore, the testing method of the present invention includes a fluorescence step of making a predetermined specimen 100 fluoresce after the target microorganism X reacts with antibody Y that undergoes an antigen-antibody reaction with the target microorganism X and is previously bound to a predetermined fluorescent reagent Z, and a fluorescence amount measurement step of measuring the amount of fluorescence of a specific wavelength emitted from the specimen 100 that has become fluorescent in the fluorescence step, thereby making it possible to measure the amount of fluorescence of a specific wavelength emitted from the specimen 100 through the antigen-antibody reaction between the target microorganism X and antibody Y. As a result, microorganisms can be measured quickly without culturing the microorganisms.

[0048] In addition, the method has a correlation storage step for storing data showing the correlation between the amount of fluorescence of a specific wavelength emitted from the target microorganism X and the number of the target microorganisms X, and a target microorganism number calculation step for calculating the number of the target microorganisms X based on the amount of fluorescence of a specific wavelength emitted from a specified specimen 100 measured in a fluorescence measurement step, which is the amount of fluorescence of a specific wavelength emitted from the target microorganism X, and the data showing the correlation stored in the correlation storage step, thereby making it possible to calculate the number of the target microorganisms X from the amount of fluorescence of a specific wavelength emitted from the target microorganisms X. [Explanation of symbols]

[0049] X: Target microorganism X´: A microorganism other than the target microorganism X Y: Antibody 1: Inspection equipment 1A: Bus 1B: Central processing unit 1C: Storage device 1D: Input Device 1E:Display device 10: Inspection Department 11: Fluorescent part 12: Fluorescence measurement unit 20: Processing unit 21: Correlation memory section 22: Target microorganism count calculation section 100: Sample

Claims

1. 1. A testing device for testing a target microorganism in a given sample, comprising: a fluorescent unit that makes the predetermined specimen fluorescent after reacting with the target microorganism and an antibody that undergoes an antigen-antibody reaction with the target microorganism and that has been bound to a predetermined fluorescent reagent in advance, and a fluorescence amount measuring unit that measures the amount of fluorescence of a specific wavelength emitted from the specimen that has been fluorescentized by the fluorescent unit, the fluorescent unit is a fluorescent unit that causes a target microorganism that has undergone an antigen-antibody reaction with an antibody that has been bound to the predetermined fluorescent reagent in advance in the predetermined specimen to fluoresce, and does not cause microorganisms other than the target microorganism to fluoresce, and the fluorescent unit causes the predetermined specimen to fluoresce by irradiating the predetermined specimen with excitation light of 488 nm±10 nm. Furthermore, the fluorescent unit has a cell holder filled with the predetermined specimen in a casing that blocks 100% of external light, and irradiates the excitation light, the antibody undergoes the antigen-antibody reaction with the target microorganism, but does not undergo the antigen-antibody reaction with microorganisms other than the target microorganism; a correlation storage unit that stores data showing a correlation between the amount of fluorescence of the specific wavelength emitted from the target microorganism and the number of the target microorganisms; a target microorganism number calculation unit that calculates the number of the target microorganisms based on the amount of fluorescence at a specific wavelength emitted from the specified specimen measured by the fluorescence amount measurement unit, which is the amount of fluorescence at the specific wavelength emitted from the target microorganisms, and data indicating the correlation stored in the correlation storage unit; The predetermined fluorescent reagent is incorporated into the antibody and is pre-bound to the antibody; Furthermore, the fluorescence amount measuring unit measures the amount of fluorescence having a wavelength of 512 nm±5 nm emitted from the predetermined specimen by the fluorescent unit, the antibody is an antibody extracted from serum of a mammal or from a chicken egg by immunizing a mammal with the target microorganism or a protein antigen having the same structure as the protein structure of the target microorganism, The inspection device according to the present invention is characterized in that the predetermined fluorescent reagent is at least one of perylene, luciferin, pyranine, and stilbene.

2. 1. A method for testing a target microorganism in a given sample, comprising: a fluorescence step of making the predetermined specimen fluorescent after reacting the target microorganism with an antibody that undergoes an antigen-antibody reaction with the target microorganism and that has been bound to a predetermined fluorescent reagent in advance, and a fluorescence amount measuring step of measuring the amount of fluorescence of a specific wavelength emitted from the specimen that has become fluorescent in the fluorescence step, the fluorescent step is a step of making a target microorganism that has undergone an antigen-antibody reaction with an antibody that has been bound to the predetermined fluorescent reagent in advance in the predetermined specimen fluoresce, and not making microorganisms other than the target microorganism fluoresce; and the fluorescence step is a step of irradiating the predetermined specimen with excitation light of 488 nm±10 nm to cause the predetermined specimen to fluoresce, Furthermore, the fluorescence step is a step of irradiating the excitation light with a cell holder filled with the predetermined specimen in a casing that blocks 100% of external light, the antibody undergoes the antigen-antibody reaction with the target microorganism, but does not undergo the antigen-antibody reaction with microorganisms other than the target microorganism; a correlation storage step of storing data showing a correlation between the amount of fluorescence of the specific wavelength emitted from the target microorganism and the number of the target microorganisms; a target microorganism number calculation step for calculating the number of the target microorganisms based on the amount of fluorescence at the specific wavelength emitted from the specified specimen measured in the fluorescence amount measurement step, which is the amount of fluorescence at the specific wavelength emitted from the target microorganisms, and data indicating the correlation stored in the correlation storage step; The predetermined fluorescent reagent is incorporated into the antibody and is pre-bound to the antibody; Furthermore, the fluorescence amount measuring step is a step of measuring the amount of fluorescence having a wavelength of 512 nm±5 nm emitted from the predetermined specimen that has been fluorescently emitted in the fluorescence step, the antibody is an antibody extracted from serum of a mammal or from a chicken egg by immunizing a mammal with the target microorganism or a protein antigen having the same structure as the protein structure of the target microorganism, The inspection method, wherein the predetermined fluorescent reagent is at least one of perylene, luciferin, pyranine, and stilbene.

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