JP2010279335A - Microorganism detection method and apparatus - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method and an apparatus enabling an efficient detection of microorganisms with a simple means in a microorganism-detection technique to bond a magnetic fine particle to the microorganism and detect light-emission of the microorganism-magnetic fine particle complex. <P>SOLUTION: The apparatus is provided with means 5, 7 for generating a magnetic force on the front end face 31 of the optical fiber to make the front end face 31 of an optical fiber 3 magnetically adsorb the microorganism-magnetic fine particle complex obtained by bonding the microorganism with the magnetic fine particle and labeled with a luminescent enzyme or a fluorescent dye, a means for inducing light emission of the luminescent enzyme or the fluorescent dye, and a means 8 for collecting the light emitting from the microorganism-magnetic fine particle complex through the same optical fiber 3 and detecting the light. <P>COPYRIGHT: (C)2011,JPO&INPIT

Description

  The present invention relates to a method for detecting microorganisms and an apparatus therefor.

The technology to detect microorganisms has increased in importance, and in particular, it is indispensable to detect various bacteria such as Escherichia coli in food and beverages from the aspect of ensuring food safety and hygiene management. It is. Conventionally, detection of these microorganisms, particularly bacteria, has been performed by detection of colonies grown on a plate agar medium and observation using various microscopes.
In recent years, proteins such as antibodies and lectins are immobilized in advance on magnetic particles with a diameter of about 1 mm or less, and the microorganisms to be detected are bound using their specific binding to the microorganism surface antigen. Thus, collecting target microorganisms by magnetic force has been used in some bioanalytical chemistry.
In addition, a technique for detecting the presence or absence of microorganisms and the amount of microorganisms from the light by causing the microorganisms to emit light has been developed.
For example, in Patent Document 1, after binding magnetic fine particles to specific Escherichia coli in a sample such as food, these complexes are collected by magnetic force, and a fluorescent molecule is bound to the specific Escherichia coli among the collected ones. The presence or absence of specific Escherichia coli is detected by detecting the specific degree of fluorescence deflection in the emitted fluorescence.

Japanese Patent Laid-Open No. 10-211000

However, in the technique of Patent Document 1, the collection by the magnetic force of the microorganism and the detection of the microorganism by collecting the light emitted from the microorganism are performed by different means. Therefore, various devices are required. At the same time, the operation of each process takes time and cannot be said to be efficient.
The present invention has been made in order to solve the above-described problems. For the detection of microorganisms, the present invention provides a simple means for detecting a microorganism / magnetic particle complex by binding magnetic particles to a microorganism. It is an object of the present invention to provide a method and an apparatus capable of performing efficient detection.

  The microorganism detection method according to the present invention includes a step of binding a microorganism and magnetic fine particles, a step of labeling the microorganism with a luminescent enzyme or a fluorescent dye, a magnetic force generated on the front end surface of the optical fiber, and the microorganism / magnetic fine particles in which the microorganism and the magnetic fine particles are combined. A step of magnetically adsorbing the complex to the front end surface of the optical fiber, a step of emitting the luminescent enzyme or fluorescent dye of the microorganism / magnetic fine particle complex, collecting the light through the same optical fiber, and detecting the light. It is characterized by including.

According to the present invention, magnetic microparticles are bound to a predetermined microorganism to form a microbe / magnetic microparticle complex, and the microbe is labeled with a luminescent enzyme or a fluorescent dye.
Then, by generating a magnetic force on the front end face of the optical fiber, the complex of the microorganism to be detected and the magnetic fine particles can be magnetically adsorbed on the front end face of the optical fiber, and the target microorganism can be selectively collected. it can.
Furthermore, the luminescent enzyme or fluorescent dye of the microorganism / magnetic fine particle complex adsorbed on the front end face of the optical fiber is caused to emit light, the light is collected through the same optical fiber, and the light is detected, thereby detecting the microorganisms to be detected. Presence / absence and its amount can be detected.
At that time, the microorganism / magnetic fine particle complex is adsorbed and collected by magnetic force, and the light from the microorganism / magnetic fine particle complex is collected by the same optical fiber, and the function of the magnetic fine particle as a magnetic substance is also achieved. In the light detection step, it is used as it is for retaining microorganisms, so that it can be an efficient microorganism detection step.

In addition, the present invention is characterized in that the binding between the microorganism and the magnetic fine particles, and the labeling of the luminescent enzyme or fluorescent dye to the microorganism is performed via an antibody, a lectin, or polymyxin B.
According to the present invention, the binding between the microorganism and the magnetic fine particle, and the binding between the microorganism and the luminescent enzyme or fluorescent dye utilize specific binding based on binding by an antibody antigen reaction or sugar chain binding of a lectin. Therefore, a complex with magnetic fine particles, luminescent enzyme or fluorescent dye can be easily formed with the target microorganism.
The complex can be used for collecting, detecting and quantifying microorganisms using the same optical fiber.

The luminescent enzyme emits light by immersing the microbial / magnetic fine particle complex adsorbed on the front end face of the optical fiber in a luminescent reaction solution.
According to the present invention, the microorganism / magnetic fine particle complex adsorbed on the front end face of the optical fiber is made to emit light only by being immersed in the luminescence reaction solution, and the light emitted from the microorganism / magnetic fine particle complex as it is through the same optical fiber. The presence or amount of microorganisms can be detected by collecting the light and detecting the light.

Further, the present invention is characterized in that the fluorescent dye emits light by irradiating the microorganism / magnetic fine particle complex adsorbed on the front end face of the optical fiber with excitation light.
According to the present invention, the light emitted from the microorganism / magnetic fine particle complex through the same optical fiber is emitted as it is emitted by irradiating the microorganism / magnetic fine particle complex adsorbed on the front end surface of the optical fiber with excitation light. By collecting the light and detecting the light, it is possible to detect the presence or amount of the microorganism to be detected.

The microorganism detecting apparatus according to the present invention also includes an optical fiber for magnetically adsorbing a microorganism / magnetic fine particle complex in which microorganisms and magnetic fine particles are bonded to the front end surface of the optical fiber and labeled with a luminescent enzyme or a fluorescent dye. A means for generating a magnetic force on the front end surface; a means for emitting a luminescent enzyme or a fluorescent dye; and a means for collecting the light emitted from the microorganism / magnetic fine particle complex through the same optical fiber and detecting the light. To do.
According to the present invention, a magnetic force is generated on the front end face of the optical fiber, and the microorganism / magnetic fine particle complex in which the magnetic fine particles are bound to the target microorganism is adsorbed there, and the microorganism / magnetic fine particle composite is passed through the same optical fiber as it is. By collecting the light emitted from the body and detecting the light, it is possible to easily detect the presence and amount of the microorganism to be detected.
Thus, both the same optical fiber and the magnetic fine particles can function in the collection of the microorganism / magnetic fine particle complex and the subsequent light detection.

  If the means for generating a magnetic force on the front end face of the optical fiber comprises a conductive coil wound around the outer peripheral surface of the optical fiber and a power supply means for the conductive coil, the conductive coil wound around the outer peripheral face of the optical fiber is energized. Thus, a magnetic force can be generated on the front end face of the optical fiber. With such a simple and compact configuration, the same optical fiber can be provided with functions of selective collection and collection of the microorganism / magnetic fine particle complex.

  In addition, when a magnetic shielding material is disposed on the outer peripheral surface near the front end of the optical fiber, the microorganism / magnetic fine particle complex can be prevented from being adsorbed by the magnetic force on the outer peripheral surface near the front end of the optical fiber. Can prevent the detection of microorganisms.

  In addition, when the optical fiber is provided with means for condensing the light emitted from the microorganism / magnetic fine particle composite and irradiating the microorganism / magnetic fine particle composite with excitation light, the microorganism / magnetic fine particle composite is selected on the same optical fiber. In addition to the functions of collecting and condensing, it is possible to provide an excitation light irradiation function.

  According to the present invention, in order to detect a target microorganism, in a technique for binding magnetic microparticles to a microbe and collecting the microbe / magnetic microparticle complex, efficient microbe detection is performed by simple means. It is possible to provide a method and apparatus that can be used.

1 is a conceptual diagram of a microorganism / magnetic fine particle complex 1 in the present invention. It is a conceptual diagram of the microorganisms detection apparatus in the 1st Embodiment of this invention. It is explanatory drawing of the microorganisms detection method in the 1st Embodiment of this invention. It is a conceptual diagram of the microorganisms detection apparatus in the 2nd Embodiment of this invention. It is a perspective view of the optical fiber in this invention. It is a figure which shows the result of the confirmation test with the optical fiber in this invention. It is explanatory drawing of the confirmation test in the optical fiber in this invention. It is a conceptual diagram of the Example of the 1st Embodiment of this invention.

Hereinafter, a method and apparatus for detecting a microorganism according to an embodiment of the present invention will be described with reference to the accompanying drawings. In the description, the same reference numerals are used for the same elements, and redundant descriptions are omitted.
The microorganisms to be detected in the present embodiment are bacteria, viruses, etc., as long as they can bind to magnetic particles, but typically, Escherichia coli, staphylococci, Salmonella, Vibrio parahaemolyticus in foods and beverages are used. Bacteria such as bacteria. On the other hand, the magnetic fine particles are particles mainly composed of a magnetic substance and have a diameter of about 1 mm or less, preferably 100 μm or less, more preferably 10 μm or less, and may be commercially available. For example, as a component of the magnetic material, there are _{Fe 3 O 4, γ-Fe} 2 O 3, Co-γ-Fe 2 O 3 or the like.

FIG. 1 shows a microorganism / magnetic fine particle complex 1 in which such a microorganism, in particular, bacteria A and magnetic fine particles B1 are bound, and a luminescent enzyme C1 is bound to bacteria A.
First, for example, when detecting specific bacteria A in foods and beverages, if the food is solid, it is liquefied by adding an appropriate liquid, and even if it is liquid, there are many impurities If it is, remove them and make an aqueous solution to make a sample.
In such a sample, if bacteria A to be detected are present by adding magnetic particles B1, luminescent enzyme C1, and antibodies for binding them to bacteria A, the microorganism / magnetic particle complex 1 is constructed.
A first antibody B2 that specifically binds to and has a complementarity with the surface antigen a of the bacterium A to be detected is immobilized on the magnetic microparticle B1, thereby constituting the first antibody-binding magnetic microparticle B. .
Therefore, the first antibody-binding magnetic fine particles B are bound to the detection target bacteria A by an antigen-antibody reaction.
Next, the second antibody / luminescent enzyme complex C in which the luminescent enzyme C1 such as peroxidase and the second antibody C2 are bound is similarly bound to the bacterium A by the antigen-antibody reaction, and the luminescent enzyme C1 is bound to the bacterium A. Label.

In addition, the first antibody and the second antibody may be a protein such as a lectin having a specific binding activity by a sugar chain with the surface antigen a of bacteria A, instead of the antibody.
Further, the surface antigen a to which the first antibody binds and the surface antibody a to which the second antibody binds may be different types of surface antigens in the same bacterium. In that case, the surface antigen a of the bacterium A to be detected is saturated only by the binding with either the magnetic fine particle B1 or the luminescent enzyme C1, and the binding with the other becomes impossible, and the detection of the bacterium A is inhibited. Can be prevented.
In addition, the binding with the first antibody B2, the second antibody C2, and the lectin instead thereof is not limited to binding at one step, but may be binding at multiple steps as in the examples described later. .
Alternatively, the second antibody C2 labeled with a fluorescent dye may be bound to the bacterium A using a fluorescent dye such as commercially available fluorescent nanoparticles instead of the luminescent enzyme C1.

Next, a first embodiment of the microorganism detection apparatus 2 for detecting bacteria A in aqueous solution samples of foods and beverages will be described with reference to FIGS.
This embodiment is an apparatus configuration in which a luminescent enzyme C1 is used as a light emission source of light to be detected.
An iron pipe 4 is attached to the tip of the optical fiber 3. Although there is no restriction | limiting in particular in the optical fiber 3, Since a detection target object is made to adsorb | suck and collect on the front end surface 31, a thing with a certain large diameter is desirable, and diameters 2-20 mm are preferable. For this purpose, a bundle fiber in which fiber strands are bundled is desirable.
The tip of the iron tube 4 reaches the front end surface 31 of the optical fiber 3 in the drawing, but it may be shorter than that, and the tip of the iron tube 4 may be limited to a length that is not immersed in the aqueous solution of the sample.
A coil 5 is formed by winding a conductive wire from above the iron pipe 4. The coil 5 may be wound in a single layer or multiple layers, and the length can be appropriately designed so that a necessary magnetic flux density can be obtained at the front end face 31 of the optical fiber 3.
The coil 5 is connected to the power supply means 7, and when energized, a magnetic field is formed by the coil 5 and the iron pipe 4, and a magnetic force is generated on the front end face 31 of the optical fiber 3.

A silicon steel cylinder is attached to the outer periphery of the tip of the optical fiber 3 as the magnetic shielding material 6. Thereby, for example, even when a magnetic force is generated on the outer periphery of the distal end of the optical fiber 3 due to application of large electric power to generate a large magnetic force, the microorganism / magnetic fine particle complex 1 is Adsorption to the outer peripheral surface other than the surface 31 can be prevented. However, when there is no fear as described above, the magnetic shielding material 6 is not always necessary.
The rear end of the optical fiber 3 is optically connected to the light detection means 8. The light detection means 8 is a light detection device such as a photomultiplier tube (PMT), a photodiode, or photon counting, and receives and detects light L from the optical fiber 3.
The signal detected by the light detection means 8 is transmitted to the recording / judgment means 9, and the detected intensity and the like are recorded. In addition, a correspondence relationship between the detected light intensity and the number of bacteria A under a certain test condition is obtained in advance, and the number of bacteria A can be determined from the correspondence relationship.

Next, the operation of detecting bacteria A using the apparatus of FIG. 2 will be described with reference to FIG.
As shown in FIG. 3A, this is limited to the case where bacteria A to be detected are present in the suspension D in which the magnetic fine particles B1, the luminescent enzyme C1-labeled antibody C2, etc. are suspended in the aqueous sample solution. Thus, the surface antigen a of the bacterium A forms a microbial / magnetic fine particle complex 1 containing three bacteria A, magnetic fine particles B1, and luminescent enzyme C1. In addition, it is microorganisms etc. which are not object to have shown with the white circle.
Therefore, the tip of the optical fiber 3 is immersed in the suspension D in the container as shown in FIG. Then, a magnetic force is generated on the front end surface 31 of the optical fiber 3, the magnetic fine particles B 1 are adsorbed on the front end surface 31 by the magnetic force, and the entire microbial / magnetic fine particle complex 1 including the magnetic fine particles B 1 is adsorbed on the front end surface 31. Will be.

Next, as shown in FIG. 6C, the front end of the optical fiber 3 in which a current is passed through the coil 5 and a magnetic force is generated on the front end face 31 is caused to emit light in response to the luminescent enzyme C1, such as a luminol reaction solution. Immerse in the luminescent reaction solution E.
When the microorganism / magnetic fine particle complex 1 is adsorbed on the front end face 31 of the optical fiber 3, light L is generated by the reaction between the luminescent enzyme C1 and the luminescent reaction solution E, and thus the light detection shown in FIG. Detection by means 8 is performed. The front end face 31 of the optical fiber 3 may have adsorbed the magnetic fine particles B1 that are not bound to the bacteria A. In this case, the light is emitted because it is not bound to the luminescent enzyme C1. Is not mistaken for bacteria A.
The container containing the luminescent reaction liquid E and the tip of the optical fiber 3 in FIG. 3C are housed in a dark box for making a dark environment, and the luminescent reaction liquid E at the tip of the optical fiber 3 is externally operated. It is desirable for photodetection to be immersed in

Next, as a second embodiment, a microorganism detection apparatus 2 in the case of using a fluorescent dye instead of the luminescent enzyme C1 will be described with reference to FIG.
In the case of a fluorescent dye, when the fluorescent dye to be used is irradiated with excitation light having a predetermined wavelength, the fluorescent dye emits fluorescence having a specific wavelength.
Therefore, a part of the optical fiber 3 allows the excitation light LA from the excitation light source 11 to pass therethrough and irradiates the excitation light LA to the fluorescent dye in the microorganism / magnetic fine particle complex 1 adsorbed on the front end surface 31 of the optical fiber 3. Has been. Then, the fluorescent LB emitted from the fluorescent dye passes through other portions of the optical fiber 3 and is emitted and received by the light detection means 8. In order to separate the excitation light LA and the fluorescence LB and guide them to a predetermined route, a beam splitter 10 is disposed in the optical path.
In the configuration of the apparatus, points other than these are the same as those described in the first embodiment.
In the second embodiment, the wavelength of the fluorescence generated varies greatly depending on the type of fluorescent dye used, and if it is out of the visible light range, light may be detected even in a dark environment.
Further, in the second embodiment, the detection operation can be performed by irradiating the excitation light LA at the stage where the microorganism / magnetic fine particle complex 1 is adsorbed without moving the optical fiber 3 to the luminescence reaction liquid tank.
However, in both the first embodiment and the second embodiment, after the microorganism / magnetic fine particle complex 1 is once adsorbed and collected by a magnetic force, the microorganism / magnetic fine particle complex 1 is washed, and the front end of the optical fiber 3 is again formed. The microbe / magnetic fine particle complex 1 may be adsorbed by the surface 31 by a magnetic force, and light detection may be performed by causing a luminescent enzyme or a fluorescent dye to emit light in this state.

"Confirmation test"
A confirmation test for confirming that magnetic particles can be adsorbed by generating a magnetic force on the front end surface 31 of the optical fiber device as shown in FIG. 5 will be described.
An iron pipe 4 having an inner diameter of 7 mm and an outer diameter of 12 mm was prepared, and the bundle optical fiber 3 was inserted into the iron pipe 4 in a state where the tip portion protruded. Then, they were inserted into a PVC tube 12 (inner diameter: 12 mm, outer diameter: 14 mm) with the tip of the optical fiber 3 protruding.
A nichrome wire (diameter 0.7 mm) was wound about 50 times around the outer periphery of the polyvinyl chloride tube 12 to double it.
Each end of the nichrome wire was connected to the positive and negative ends of a constant current power source, and a constant direct current was applied. And the magnetic flux density in the front-end surface 31 of the optical fiber 3 was measured with the gauss meter (GM-301, the electronic magnetic industry make).
The measurement result is as shown in FIG. 6, and a predetermined magnetic flux density was obtained at the front end face 31 of the optical fiber 3. Further, as shown in the figure, a proportional relationship between the applied current and the generated magnetic field was recognized in the experimental range in both the single-layer winding and the two-layer winding of the coil.

Next, commercially available magnetic fine particles (Dynabeads M-280) often used for collecting microorganisms are suspended in a 0.1 M phosphate buffer (pH 7.0), and the magnetic fine particles are directed to the front end face 31 of the optical fiber 3. It was experimentally confirmed that the adsorption and dissociation of can be performed.
FIG. 7 shows a conceptual diagram at that time.
The concentration of the suspension D used is 2 × 10 ⁸ beads / ml (the number of magnetic fine particles per ml), and 2 ml of this suspension is put into a beaker having a diameter of about 20 mm to make the magnetic fine particles B1 uniform. After stirring and allowing to stand, immediately after immersion in the suspension D so that the front end face 31 of the optical fiber 3 was about 1 mm below the liquid level, an electric current was applied to the coil 5.
As a result, it was found that if the magnetic field of 50 mT (500 G) or more is present on the optical fiber front end face 31, the magnetic fine particles B1 in the suspension accumulate almost uniformly on the optical fiber front end face 31. Moreover, within this measurement range, the magnetic fine particles B1 have not accumulated in any part of the fiber part, the iron pipe 4 and the PVC pipe 12 other than in the liquid.
When the supply of current was stopped, the magnetic fine particles B1 were dissociated smoothly. In addition, although residual magnetization remains in the iron pipe 4, it did not become a problem in dissociation.

"Example"
A specific example of the first embodiment will be described below with reference to the conceptual diagram of FIG.
Add 50 ml of 50 mM phosphate buffer (pH 7.0) containing 0.1 mM MnSO ₄ and 0.1 mM MgCl ₂ to 50 g of commercially available ground beef, crush it with a blender, and centrifuge (1,000 × g E. coli (Escherichia coli ATCC 12740) is added (104 cells / ml) to the resulting extract. This is used as a model sample as an aqueous solution extracted from food.

Next, Dynabeads R-280 (Dynabeads R-280 Tosyl-activated) and the antibiotic polymyxin B (polymyxinB) conjugate (polymyxin B-conjugated Dynabeads R-280, Pol-R beads) Add 1.0 ml of the same phosphate buffer (2 × 10 ⁸ beads / ml) suspended in 1.0 ml of the above model sample and stir at room temperature for 2 hours. In addition, the detailed preparation method of ConA-Rbeads followed the manual attached to the product.
As a result, in FIG. 8, a complex is formed in which polymyxin B as B2 bound to Dynabeads R-280 as magnetic microparticles B1 is bound to the outer membrane part P1 of specific Escherichia coli A.

Next, the complex formed in this manner (hereinafter also referred to as “bead”) is obtained by a magnet, and the residue is discarded. The obtained beads are washed with a phosphate buffer similar to the above.
Next, in FIG. 8, a 50 mM phosphate buffer (as a blocking agent) containing anti-O127aIgG (derived from rabbit) as an antibody C21 that binds to the O127a antigen, which is the surface antigen a2 of the above-mentioned Escherichia coli A, at a concentration of 4 μg / ml. The obtained beads are suspended in 1.0 ml of 3% bovine serum albumin (containing 0.2% Tween 20) and stirred at room temperature for 1 hour. For the preparation of the anti-O127aIgG (rabbit derived) solution, the literature by the inventors of the present invention `` Masuko, M., Kataoka, T., Sugiyama, N., Uchiyama, S., Sugiyama, H., Tarui, K., Kamiya, K. & Kawai, T. (1992) Photochem Photobiol, 56, 107-111 ”.
As a result, the complex up to magnetic fine particle B1-polymyxin B B2-E. Coli A-antibody C21 in FIG. 8 is formed.

Next, the beads thus formed are obtained by a magnet, and the solution is discarded. The beads are then washed with 50 mM phosphate buffer (pH 7.0).
Next, a commercially available horseradish as luminescent enzyme C1, anti-rabbit IgG antibody solution as antibody C22 to which peroxidase is bound (diluted 200 times, buffer is 50 mM phosphate buffer (3 as blocking agent) %) Bovine serum albumin and 0.2% Tween20) are added, and the obtained beads are stirred at room temperature for 1 hour. For the preparation of such a solution, the above-mentioned document “Masuko, M., Kataoka, T., Sugiyama, N., Uchiyama, S., Sugiyama, H., Tarui, K., Kamiya, K. & Kawai, T. (1992) Photochem Photobiol, 56, 107-111 ".

Thereby, the microbe / magnetic microparticle complex 1 including magnetic microparticle B1-polymyxin B B2-E. Coli A-antibody C21-antibody C22-luminescent enzyme C1 in FIG. 8 is completed.
Next, the beads thus formed are obtained with a magnet and washed with 50 mM phosphate buffer (pH 7.0).
Then, this is suspended in 2.0 ml of a commercially available luminol reaction solution (RP2106, GE Healthcare) as the luminescence reaction solution E, and the peroxidase C1 in the beads adsorbed on the front end face 31 of the optical fiber 3 is caused to emit light. Below, the measurement of the light L as FIG.3 (C) is performed for a fixed time. The conceptual diagram in the luminescent reaction liquid E in that case is as FIG.
In this manner, the microorganism / magnetic fine particle complex 1 is adsorbed and collected on the front end face 31 of the optical fiber 3 and is used as a magnetic substance for retaining the microorganisms during light detection without removing the magnetic fine particles. The optical measurement for detecting bacteria can be performed with the same optical fiber while maintaining the above functions.

DESCRIPTION OF SYMBOLS 1 ... Microorganism / magnetic fine particle complex, A ... Bacteria, a ... Surface antigen, B ... First antibody binding magnetic fine particle, B1 ... Magnetic fine particle, B2 ... Polymyxin B, C ... Second antibody / luminescent enzyme complex, C1 ... Luminescent enzyme, C2 second antibody, D suspension, E luminescent reaction solution, L light, LA excitation light, LB fluorescence, microbial detection device, 3 optical fiber, 31 front end surface of optical fiber, 4. Iron pipe, 5. Coil, 6. Magnetic shielding material, 7. Power supply means, 8. Light detection means, 9. Recording / judgment means, 10. Beam splitter, 11. Excitation light source, 12. PVC pipe

Claims (8)

A step of binding microorganisms and magnetic fine particles,
Labeling the microorganism with a luminescent enzyme or a fluorescent dye,
A step of generating a magnetic force on the front end face of the optical fiber, and magnetically adsorbing the microorganism / magnetic fine particle complex in which the microorganism and the magnetic fine particles are bonded to the front end face of the optical fiber;
Emitting the luminescent enzyme or the fluorescent dye of the microorganism / magnetic fine particle complex, collecting the light through the same optical fiber, and detecting the light;
A method for detecting microorganisms, comprising:   The method for detecting a microorganism according to claim 1, wherein the binding between the microorganism and the magnetic fine particle, and the labeling of the luminescent enzyme or fluorescent dye to the microorganism is performed through an antibody, a lectin, or polymyxin B.   The microorganism detection method according to claim 1 or 2, wherein the luminescent enzyme emits light by immersing the microorganism / magnetic fine particle complex adsorbed on the front end face of the optical fiber in a luminescence reaction solution. .   3. The microorganism detection method according to claim 1, wherein the fluorescent dye emits light by irradiating the microorganism / magnetic fine particle complex adsorbed on the front end face of the optical fiber with excitation light. 4. Means for generating a magnetic force on the front end face of the optical fiber in order to magnetically adsorb the microorganism / magnetic fine particle complex in which microorganisms and magnetic fine particles are bonded to the front end face of the optical fiber and labeled with a luminescent enzyme or a fluorescent dye;
Means for emitting the luminescent enzyme or the fluorescent dye;
Means for collecting the light emitted from the microorganism / magnetic fine particle composite through the same optical fiber and detecting the light;
A microorganism detection apparatus comprising:   6. The microorganism detecting apparatus according to claim 5, wherein the means for generating a magnetic force on the front end surface of the optical fiber includes a conductive coil wound around an outer peripheral surface of the optical fiber, and a power supply means for the conductive coil. .   The microorganism detecting apparatus according to claim 5 or 6, wherein a magnetic shielding material is disposed on an outer peripheral surface in the vicinity of the front end of the optical fiber.   8. The optical fiber includes means for condensing light emitted from the microorganism / magnetic fine particle complex and irradiating the microorganism / magnetic fine particle complex with excitation light. 8. 2. The microorganism detection apparatus according to item 1.

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