TWI694249B - Light-trapping microbial identifying method - Google Patents

Abstract

A light-trapping microbial identifying method includes: trapping a known-type microorganism with an optical tweezer to contain a trapped microorganism therein; moving the optical tweezer and the trapped microorganism with at least one predetermined speed to measure an escape velocity (velocity range or average velocity) of the trapped microorganism; utilizing a specific sample of the escape velocity (velocity range or average velocity) of the trapped microorganism to compare with that of an unknown-type microorganism; and determining a type of the unknown-type microorganism according to a result comparing the escape velocity of the unknown-type microorganism with the specific sample.

Description

Light-trapping microorganism identification method

The invention relates to a light-trapping microbial identification method; in particular to a laser beam [laser beam] capturing microbial identification method; more particularly to an optical tweezer to capture microbes Category identification method; more particularly about a fiber optic optical tweezers to capture microorganism identification method; more particularly to a laser fiber optical tweezers to capture microorganism identification method.

For example, regarding the manufacturing method of conventional fiber optic lens [fiber microlens], for example: the patent for invention of "Method of Manufacturing Hyperbolic Fiber Optic Lens" of the Republic of China Patent Announcement No. TW-I241421, which discloses a method of manufacturing a hyperbolic fiber optic lens Method. The manufacturing method of the optical fiber lens includes the following steps: [a] stripping a coating layer of a predetermined length of an optical fiber to form an exposed part; [b] cleaning the exposed part; [c] fixing the optical fiber to be processed on an optical fiber In the fixed seat; [d] Provide a container, which includes a hydrofluoric acid layer, an oil layer and an intermediate mixed layer; [e] extend the exposed portion of the optical fiber into the hydrofluoric acid layer for etching to form A cone angle; [f] melting the cone angle using two arcs; and [g] adjusting the relative positions of the two arcs and the cone angle, using the uneven temperature field formed by the two arc discharges to obtain a hyperbolic fiber The lens and the set radius of curvature improve the coupling efficiency of the optical fiber.

However, the aforementioned No. TW-I241421 only discloses the manufacturing method of the fiber lens and the technology suitable for mass production. In addition, the fiber The lens manufacturing method also only discloses how to perfectly match the fiber lens and the laser light field, and adjust the relative positions of the cone angles of the two arcs and the fiber lens to obtain the optimal radius of curvature of the fiber lens and couple it The technology of light efficiency has a decisive influence.

Another conventional manufacturing method of fiber optic tweezers [fiber optic tweezers], for example: the invention patent of "Process of Fiber Optic Tweezers" No. TW-I474061 of the Republic of China Patent, which discloses a method of manufacturing optical tweezers. The manufacturing method of the optical tweezers includes the following steps: [a] stripping: cutting an optical fiber of an appropriate length, and stripping a coating layer of the optical fiber (coating) to leak a bare optical fiber, which includes a fiber coat layer [cladding ] And a core layer [core]; [b] cleaning: clean the optical fiber; [c] cutting: cutting and flattening the end surface of the bare fiber; [d] etching: fixing the optical fiber in a container Add an oxide buffer etching solution (buffered oxide etch, BOE), and then set the fixed optical fiber corresponding to the container, immerse the end of the optical fiber to be processed into the oxide buffer etching solution for etching, and the end of the optical fiber can be formed A tapered shape to make fiber microlenses or lensed fibers [fiber microlens]; the fiber ends of the tapered shape can also be further sintered with arc devices at both ends to form hemispherical fiber microlenses.

In addition, the aforementioned No. TW-I474061 uses the hemispherical lensed optical fiber of the fiber optic tweezers to achieve a simpler overall manufacturing and the effect of reducing the manufacturing cost. If its operation in the field of biotechnology or biomedical applications uses light or light energy to capture small objects or microorganisms in a non-contact manner, and it has high capture efficiency and low capture power.

However, the aforementioned No. TW-I474061 only discloses that the fiber optic tweezers are suitable for non-contact capture of tiny objects or microorganisms. In addition, the fiber optic tweezers are suitable for non-contact capture of small objects or microorganisms. There is also a potential need to provide future applications for other biotechnologies (for example, applications for microbial identification).

Another conventional manufacturing method of fiber optic tweezers [fiber optic tweezers], for example: the invention patent of "light-trapping cancer cell stage testing method" of US Patent No. US-9958663, which discloses a method for identifying stage of light-trapping cancer cells . The light-trapping cancer cell stage identification method includes: measuring the average escape speed or range of a first cancer cell and the average escape speed or range of a second cancer cell, and the cancer cells of the first cancer cell and the second cancer cell The stages are known to be different, and the first cancer cell and the second cancer cell are known types; then the average escape velocity of the first cancer cell and the average escape velocity of the second cancer cell are used to calculate an escape velocity ratio or range; Select a cancer cell to be identified, and measure the escape speed of a cancer cell to be identified, and the cancer cell to be identified is a known type but an unknown stage; use the ratio or range of the escape speed to compare with the escape speed of the cancer cell to be identified ; And determine a cancer cell stage based on the comparison result of the escape speed of the cancer cells to be identified.

However, the aforementioned US-9958663 only discloses a light-trapping cancer cell stage identification method, and the cancer cell stage identification method is not suitable for identifying other objects. In addition, the light-trapping cancer cell stage identification method is suitable for non-contact capture of small objects or microorganisms. There is also a potential need to provide future applications for other biotechnologies (for example, the identification of various types of microorganisms).

The aforementioned invention patents No. TW-I241421, No. TW-I474061 and No. US-9958663 are the invention patents only for reference to the technical background of the present invention and to illustrate the current state of technological development, which is not intended to limit the scope of the invention .

In view of this, in order to meet the above requirements, the present invention provides a light-trapping microorganism identification method, which uses a fiber optic tweezers to capture a microbial organism [a known type of microorganism], so as to form a captured on the fiber optic tweezers Microorganisms, and control the optical fiber optical tweezers and the captured microorganisms to move at a predetermined speed to measure the escape speed of a microorganism [or speed range or average escape speed], and optionally establish a microorganism Identify the standard database, and then use the escape speed of the microorganism [or speed range or average escape speed] to compare with the escape speed of a microorganism to be identified [microorganism of unknown type]. Microbial species to expand the scope of further application of conventional fiber optic tweezers.

The main objective of the preferred embodiment of the present invention is to provide a light-trapping microorganism identification method that uses a fiber optic tweezers to capture a microbial organism [a known type of microorganism] so as to form a captured microbe on the fiber optic tweezers , And control the optical fiber optical tweezers and the captured microorganisms to move at a predetermined speed to measure the escape speed of a microorganism [or speed range or average escape speed], and can choose to establish a database of microorganism identification standards, and then use the The microorganism escape speed [or speed range or average escape speed] is compared with the escape speed of a microorganism to be identified [microorganism of unknown type], so a microorganism type is determined according to the comparison result of the escape speed of the microorganism to be identified, so as to achieve accurate identification of microorganisms Kind of purpose.

In order to achieve the above objective, the light-trapping microorganism identification method according to the preferred embodiment of the present invention includes: capturing a microbial organism [a known type of microorganism] with a fiber optic tweezers, so as to form a captured microbe on the fiber optic tweezers ; Control the optical fiber optical tweezers and the captured microorganisms to move at a predetermined speed to measure a microorganism's escape speed [or speed range or average escape speed]; use the microorganism's escape speed [or speed range or average escape speed] and A comparison of the escape speed of the microorganisms to be identified [microorganisms of unknown type]; and the determination of a microorganism type according to the comparison result of the escape speeds of the microorganisms to be identified.

The preferred embodiment of the present invention selects various microorganisms as a reference microorganism category.

The preferred embodiment of the present invention establishes a database of microbe identification standards required by the light capture inspection method.

The optical fiber optical tweezers of the preferred embodiment of the present invention are used to measure a capturing force, a capturing efficiency, or a combination thereof.

The microorganism of the preferred embodiment of the present invention is placed in a buffer.

In the preferred embodiment of the present invention, the predetermined speed is a fixed speed.

In the preferred embodiment of the present invention, the microorganism escape speed is a microorganism escape speed range or an average microorganism escape speed.

In the preferred embodiment of the present invention, the microorganism escape speed range is between a maximum microorganism escape speed and a minimum microorganism escape speed.

In the preferred embodiment of the present invention, the escape speed of the microorganism to be identified is selected from measuring the escape speed five times and calculating the average result.

In the preferred embodiment of the present invention, the microorganism is selected from a bacterium, a fungus, a virus, an algae or a pathogenic bacterium.

In the preferred embodiment of the present invention, the microorganism is selected from a yeast (eg, natural yeast, bun yeast, beer yeast or wine yeast), a probiotic or a pathogenic bacteria.

The optical fiber optical tweezers of the preferred embodiment of the present invention is a laser optical fiber tweezers.

In the preferred embodiment of the present invention, the optical fiber optical tweezers fixture has a catching tilt angle or a working tilt angle, and the catching tilt angle or working tilt angle is 40° or 50°.

The optical fiber optical tweezers fixture of the preferred embodiment of the present invention has a single-mode/multimode fiber or a single-mode/multimode fiber microlens.

1‧‧‧ Computer system

2‧‧‧Image capture system

21‧‧‧Microscope device

22‧‧‧filter

3‧‧‧Laser light source device

31‧‧‧Laser light source

32‧‧‧Optical fiber coupler

33‧‧‧ Fiber

34‧‧‧ fiber optic microlens

5‧‧‧Working platform

50‧‧‧Microbial culture container to be identified

51‧‧‧Pan control device

52‧‧‧First electronic control platform

53‧‧‧stage

54‧‧‧ stage light source

55‧‧‧Second electronic control platform

9‧‧‧ microorganism

Figure 1: Schematic diagram of an optical tweezers operating system for the light-trapping microorganism identification method according to the preferred embodiment of the present invention.

Figure 2: Schematic diagram of the optical tweezers operating system according to the preferred embodiment of the present invention performing light capture microbial operations.

Figure 3: Schematic flow chart of the light-trapping microorganism identification method according to the preferred embodiment of the present invention.

Figure 3A: Schematic diagram of the construction process of the light-trapping microorganism identification standard database according to the preferred embodiment of the present invention.

FIG. 3B: A schematic flow chart of the method for identifying a light-trapping microorganism according to a preferred embodiment of the present invention.

Figure 4: A schematic diagram of a light refraction mechanism when the light beam is subjected to the force of the microorganisms when the microorganism capture method of the preferred embodiment of the present invention starts to capture the microorganisms.

Figure 5: A schematic diagram of a light-capturing mechanism generated by the light-trapping microorganism identification method of the preferred embodiment of the present invention when the microorganisms are captured by the reaction force of the light beam.

Figure 6: A schematic diagram of the light trapping microorganism identification method of the preferred embodiment of the present invention when the microorganisms have been captured and the microorganisms have been stably bound by the light beam.

Fig. 7A: A schematic diagram of a microscopic image of the light-trapping microorganism identification method used for measuring the first yeast according to the preferred embodiment of the present invention.

Fig. 7B: A schematic diagram of a microscopic image of a light-trapping microorganism identification method for measuring a second yeast according to a preferred embodiment of the present invention.

Figure 7C: A schematic diagram of a microscopic image of the third embodiment of the light-trapping microorganism identification method of the present invention for measuring the third yeast.

Figure 8: The light-trapping microorganism identification method according to the preferred embodiment of the present invention is used to measure the first, second and third yeasts to obtain an indication of their escape speed intention.

Figure 9: A schematic diagram of the light-trapping microorganism identification method according to the preferred embodiment of the present invention is used to measure the first, second, and third yeasts to obtain their capturing efficiency.

In order to fully understand the present invention, preferred embodiments will be exemplified below and described in detail in conjunction with the accompanying drawings, and they are not intended to limit the present invention.

The light-trapping microorganism identification method according to the preferred embodiment of the present invention can be applied to identify various microorganisms [bacteria are prokaryotes, fungi, algae, and protozoans are eukaryotes, and viruses have no cell structure], for example, including bacteria and pathogens. Bacteria [e.g. Salmonella, Vibrio enteritidis, Vibrio, Escherichia coli, Botox or Staphylococcus], probiotic cells [e.g. natural yeast, bun yeast, beer yeast or wine yeast] or other microorganisms Cells, but it is not used to limit the scope of application of the present invention; furthermore, the light-trapping microorganism identification method of the preferred embodiment of the present invention can use light of various suitable wavelength ranges, for example: laser light of various wavelengths [infrared laser light , Red laser, green laser, blue laser or purple laser, etc.], but it is not intended to limit the scope of the present invention.

The optical capturing microorganism identification method of the preferred embodiment of the present invention selects preset parameters of the optical fiber optical tweezers system. It uses an optical fiber optical tweezers to measure the escape speed and diameter of several known types of microorganisms. It uses statistical mathematical methods to calculate the average escape speed and escape speed range of each known type of microorganism, and selects various reference types of microorganisms to establish a standard database of microorganism identification required by the light capture inspection method. Then use the same optical fiber optical tweezers system parameters and experimental methods to measure the escape speed and diameter of a microorganism to be identified, and calculate the average value of the escape speed ratio and capture efficiency. Compare the ratio of the escape speed of the microorganism to be identified and the capture efficiency with the value of the same type of microorganism in the standard database, so as to determine a microorganism according to the comparison result of the ratio of the escape speed of the microorganism to be identified and the capture efficiency species.

FIG. 1 shows a schematic diagram of the structure of a light-trapping microorganism identification method using an optical tweezers operating system according to a preferred embodiment of the present invention. Please refer to FIG. 1, for example, in construction, the preferred embodiment of the invention uses an optical tweezers operating system including a computer system 1, an image capture system 2, a microscope device 21, and a laser light source device 3. A working platform 5 and a translation stage control device 51, which are selected to be properly connected to each other for assembly or corresponding configuration to form the optical tweezers operating system.

Please refer to FIG. 1 again. For example, the computer system 1 includes a computer host [computer device] and a display device [display device] or other peripheral devices [peripheral], and the computer system 1 is appropriately configured. At a predetermined position, the computer host of the computer system 1 is connected to the image acquisition system 2 or the microscope device 21. In addition, the computer system 1 also chooses to connect to the laser light source device 3, the working platform 5 or the translation stage control device 51.

Please refer to FIG. 1 again. For example, the image capturing system 2 has an image capturing processing unit, and the image capturing system 2 is connected to and combined with the microscope device 21, or the image capturing system 2 may Choose to connect any electron microscope device to capture at least one microscopic image (for example: microbial cell image) or a series of electron microscopic images from the microscope device 21. In addition, the image capture system 2 includes a filter 22 or other equipment.

Please refer to FIG. 1 again. For example, the microscope device 21 includes a CCD element [Charge Coupled Device] and a microscope or other functional devices [for example: a fixing device], and the CCD device is combined with the microscope In order to use the CCD device to take at least one microscopic image or a series of microscopic images through the microscope. During the assembly operation, the microscope device 21 corresponds to the working platform 5 so as to capture working images in the area above the working platform 5.

Please refer to FIG. 1 again. For example, the laser light source device 3 is selected from a predetermined wavelength laser light source device (for example: 650 nm or other wavelengths), and the laser light source device 3 optionally includes a laser light source [Laser source, for example: semiconductor laser or other lasers] 31, a fiber coupling device 32, a fiber 33 and a fiber microlens 34, and the laser light source 31 passes through the fiber coupler 32 is connected to the optical fiber 33, and a laser beam (laser beam) is appropriately emitted by the optical fiber 33 and its optical fiber microlens 34, so as to form an optical fiber optical tweezers. In the identification of microorganism types, the optical fiber optical tweezers are suitable for performing light capture microorganism operations. In the preferred embodiment of the present invention, the optical fiber 33 of the optical fiber optical tweezers has a single-mode/multimode optical fiber, and the optical fiber microlens 34 of the optical fiber optical tweezers has a single-mode/multimode optical fiber microlens.

Please refer again to FIG. 1, for example, the working platform [operational platform] 5 includes a translation stage control device 51, a first electric control platform [XY axis electric control platform] 52, a stage 53, A stage light source 54 and a second electric control platform [XYZ axis electric control platform] 55 or other automatic electronically controlled desktops, so as to choose to operate the working platform 5 in an automatic or electronically controlled manner. Place a microbial culture container or a device with a similar function on the stage 53 of the working platform 5, and use the first electronic control platform 52 of the working platform 5 to properly adjust the operation of the stage 53 and the stage The horizontal position of the light source 54, and the second electric control platform 55 of the working platform 5 is used to appropriately adjust the [capture] working tilt, working distance, horizontal position and vertical position of the optical fiber 33 and the optical fiber microlens 34. In addition, there is a predetermined irradiation angle between the stage 53 and the stage light source 54.

FIG. 2 shows a schematic diagram of the optical tweezers operating system according to the preferred embodiment of the present invention performing light capture microbial operations. Please refer to FIG. 2, for example, the optical fiber 33 of the optical tweezers operating system according to the preferred embodiment of the present invention selects an optical fiber diameter of 125 μm and a core diameter of 8 μm in the unetched section, which has been etched The fiber diameter of the section is about 14 μm, and the light The fiber microlens 34 is located at the tail end of the etched section, and the fiber microlens 34 is a cone, and it has a tapered angle α, which is between 90° and 115° or other angles Cone angle range.

Please refer to Figs. 1 and 2, for example, the laser light source device 3 chooses to adopt the capture power adjusted to 7mW or other appropriate capture power [for example: 5mW, 20mW or its interval], and the optical fiber etching cone 34 Corresponding to a microorganism cultivation container 50 to be identified, a microbial organism [microorganism to be identified] 9 is captured with light [laser]. At this time, the operator may choose to use the first electronic control platform 52 of the working platform 5 to properly adjust the angle and horizontal position of the stage 53 and the microorganism cultivation container 50 to be identified. The microorganism 9 of the preferred embodiment of the present invention is placed in a buffer solution.

Referring again to FIG. 2, for example, when the microorganism-cultivating container 50 to be identified performs light-trapping microorganism identification, it can be operated up to the tip of the end face of the fiber microlens 34 of the laser light source device 3 and microorganisms There is a predetermined working distance D or a predetermined working distance range between the bodies 9, and there is a predetermined working inclination angle between the longitudinal axis of the fiber microlens 34 of the laser light source device 3 and the microorganism 9 angle] θ or a predetermined range of working tilt angle, that is, the angle between the traveling direction of the light beam and the horizontal direction.

FIG. 3 shows a schematic flow chart of the light-trapping microorganism identification method according to the preferred embodiment of the present invention. Please refer to FIGS. 1, 2 and 3, for example, the light capturing microorganism identification method according to the preferred embodiment of the present invention includes a first step S1: First, the optical tweezers operating system or optical fiber tweezers are used to properly capture Measure a microorganism [a known type of microorganism] to form a captured microorganism on the optical fiber optical tweezers. The microorganism in the preferred embodiment of the present invention may also be selected from a bacterium, a fungus, a virus, an algae, a protozoan [protozoa] or a pathogenic bacterium.

Please refer again to Figures 1, 2 and 3, for example, this The method for identifying light-trapping microorganisms according to a preferred embodiment of the invention includes a second step S2: Next, the optical fiber tweezers and the captured microorganisms are controlled at least a predetermined speed and appropriate technical means [for example: dynamic measurement method [dynamic measurement method] 〔Or static measurement method〕to move to measure the escape speed of a microorganism [or speed range or average escape speed]. Use the optical tweezers operating system or optical fiber optical tweezers to select and measure the average escape speed or range of a first microorganism [e.g. Saccharomyces cerevisiae] and a second microorganism [e.g. Saccharomyces cerevisiae] One of the average escape speed of the second microorganism or its range.

Another preferred embodiment of the present invention establishes a database of microorganism identification standards required by the light capture inspection method to store the known average escape speed or range of the first microorganism and the average escape speed or range of the second microorganism, which can also be used Store known microorganism diameters. The average escape speed of the first microorganism [or the average escape speed of the second microorganism] is between a maximum average escape speed of the microorganism and a minimum average escape speed of the microorganism.

Please refer to FIGS. 1 and 2 again. For example, the preferred embodiment of the present invention selects various microorganisms as a reference microorganism category, and the optical fiber optical tweezers are used to measure a capture force [or lateral capture force, transverse trapping force], a trapping efficiency, or a combination thereof, and the predetermined speed may be selected as a fixed speed (ie, equal speed), as shown in the y-axis direction in FIG. 2.

Please refer to FIGS. 1 and 2 again. For example, the light-trapping microorganism identification method according to the preferred embodiment of the present invention includes a third step S3: Then, using the microorganism escape speed [or speed range or average escape speed] and A microorganism to be identified [a microorganism of unknown type] is compared for the escape speed, and a microorganism to be identified [or a microorganism to be tested] is selected, and the optical fiber optical tweezers can be used to choose to automatically or otherwise measure the escape speed of a microorganism to be identified. In the preferred embodiment of the present invention, the escape speed of the microorganism to be identified is selected from measuring the escape speed five times and calculating the average result.

Please refer to FIG. 1 and FIG. 2 again, for example, the light-trapping microorganism identification method according to the preferred embodiment of the present invention includes a fourth step S4: Then, a microorganism is identified and determined according to the comparison result of the escape speed of the microorganism to be identified species. Using the microorganism escape speed [or speed range or average escape speed] and the escape speed of the microorganism to be identified can be selected to be compared in an automatic or other manner. In the comparison of microbial identification, the light-trapping microbial identification method of the preferred embodiment of the present invention and the built standard database of microbial identification can choose to use various mathematical expressions or formulas for microbial identification.

FIG. 3A shows a schematic diagram of the construction process of the light-trapping microorganism identification standard database according to the preferred embodiment of the present invention. Please refer to FIG. 3A. For example, the construction of the standard database of light-trapping microorganism identification according to the preferred embodiment of the present invention includes: setting the parameters of the optical tweezers system; selecting several known types of yeast [or other microorganisms] ; Measure the diameter of various types of yeast cells [or other microorganisms] and their escape speed several times; calculate the average value of various types of yeast cells [or other microorganisms] escape speed; establish various types of yeast cells [or other microorganisms], average Database of escape speed and escape speed range; standard database for identification of light-capturing yeasts (or other microorganisms) required to complete the classification of live bacteria.

FIG. 3B shows a schematic flowchart of a method for identifying a light-trapping microorganism according to a preferred embodiment of the present invention. Please refer to FIG. 3B. For example, the light-trapping microorganism identification method according to the preferred embodiment of the present invention includes: setting optical tweezers system parameters according to a standard database, and setting the same experimental parameters according to the standard database; selecting unknown species Several yeasts (or other microorganisms); measure the diameters of various yeasts (or other microorganisms) and their escape speed several times; measure the diameters, average escape speeds and capture efficiency of each yeast (or other microorganisms) to be tested; Compare with the established standard database to complete the identification of each yeast (or other microorganism) to be tested.

Please refer again to Figures 1, 2 and 3A. For example, the preferred embodiment of the present invention is built on a standard database for identification of light-trapping microorganisms. In the above, the mathematical formulas or formulas for the identification of various types of microorganisms are used, and the selection of a mathematical formula is illustrated below.

For example, the light-trapping microorganism identification method of the preferred embodiment of the present invention uses the trapping force F as: F = 6 πηrv

Where η is the viscosity coefficient of the solution, r is the radius of the sample [similar to a sphere], and v is the escape velocity of the sample.

For example, the light-trapping microorganism identification method of the preferred embodiment of the present invention uses the capture efficiency Q as:

Where c is the speed of light in vacuum, n is the refractive index of the solution, and P is the capture power of the optical tweezers.

Please refer to Figures 1 and 2 again. For example, if the output optical power of the same fiber increases, the escape speed required by the captured object will also increase. Therefore, when the output optical power is fixed, each type of microorganism has its corresponding range of escape speed, so that the identification test of the type of microorganism can be achieved. Generally speaking, under the condition of the same light capture power, the escape speed of microorganisms of the same type but different diameters is relatively insignificant; and among different types of microorganisms, the difference in escape speed is relatively obvious. As for the escape speed of microorganisms, their various optical characteristics [refraction, reflection, transmission, absorption] are different according to the different biochemical composition of their organisms, which further results in their different escape speeds.

In general, the capturing principle of single-beam optical tweezers can be explained with geometric optics (ray optics). When light propagates through media with different refractive indexes, the direction of light travel will be bent due to the difference in light speed, and the phenomenon of light refraction [refraction] and the change of light momentum [momentum] will occur. The principle of optical tweezers is to use the principle or mechanism of light refraction and the change of light momentum Microorganisms [transparent and tiny objects] are optically captured. That is, when the laser light [light field] approaches or enters the microorganism to be captured, the light produces a refraction phenomenon within the microorganism and causes a change in the light momentum to produce a phenomenon of capturing the microorganism. If you encounter opaque microorganisms, you need to use a double-beam optical tweezers to capture, the principle of capture using radiation pressure or radiation pressure [radiation pressure] theory to explain.

FIG. 4 shows a schematic diagram of the light refraction mechanism when the light beam is subjected to the force of the microorganisms when the microorganism capturing method of the preferred embodiment of the present invention starts to capture the microorganisms. Please refer to FIGS. 1, 2 and 4, for example, the light capturing microorganism identification method according to the preferred embodiment of the present invention uses the light beam output from the fiber microlens 34 to start capturing a microorganism 9. The optical fiber optical tweezers is a laser optical fiber tweezers (for example: 650nm laser optical fiber tweezers). The optical fiber optical tweezers fixture has a catch tilt angle or a working tilt angle, and the catch tilt angle or working tilt angle is 40°, 50° or other suitable angles. The optical fiber optical tweezers fixture has a single-mode optical fiber or other suitable optical fiber, and the optical fiber is processed to form an optical fiber microlens.

Please refer to Fig. 4 again. When the light beam [laser beam] passing through the fiber microlens is incident on the surface of the microbial body 9, if it is not deflected, the penetrating light should travel a straight path [two symmetrical broken lines [Shown], but because the refractive index of the microorganism [refractive index] is different from the surrounding environment, the incident light enters the microorganism and is deflected [shown by two symmetric solid lines, that is, refracted light]. Both sides of the center of symmetry are deflected inwardly [toward the longitudinal axis of the fiber microlens 34] as if the internal material of the microorganism exerts a force F1 (shown by two symmetrical inward arrows) on the penetrating light, causing it to deviate The path shown by the original dotted line, that is, the change of the light momentum generated by the refracted light compared with the original incident light.

FIG. 5 shows a schematic diagram of the light capture mechanism when the microorganism capture method of the preferred embodiment of the present invention starts capturing microorganisms, and the microorganisms are subjected to the reaction force of the light beam. Please refer to figures 4 and 5 As shown, the internal substance of the microbial body 9 exerts a force F1 (shown by the two symmetrical inward arrows in Figure 4) on the incident light to deflect the incident light. According to Newton's third law of motion, the refracted light also Generates a force F2 of the same size but in the opposite direction (shown by two symmetric outward arrows in Figure 5) on the internal substance of the microbial body 9, that is, the photon [refracted light] also exerts a reaction force F2 on the microbial body 9. The microbial body 9 can be appropriately moved along the longitudinal axis of the optical fiber microlens 34 by the sum of the vertical vectors F3 of the reaction force F2 (shown by the vertical upward arrows in FIG. 5).

FIG. 6 shows a schematic diagram of the light trapping microorganism identification method of the preferred embodiment of the present invention when the microorganisms have been captured, and the microorganisms have been stably bound by the light beam. Please refer to Figures 5 and 6, when the photon [refracted light] applies a reaction force F2 to the microbial body 9 at the same time, the microbial body 9 is subjected to the sum of the vertical vectors of the reaction force F2 [vertical upward arrows in Figure 5 It is possible to produce proper motion along the longitudinal axis of the fiber microlens 34. When the microorganism 9 moves to the optical focus of the fiber microlens 34, a stable optical potential well is generated near the focus by the [laser] beam incident on the fiber microlens 34, and at A force balance point [a position where the total force is zero] is formed at the potential energy well, and the microbial body 9 is formed to trap and restrict the microorganism. At this time, once the optical fiber microlens 34 of the optical tweezers is moved, the microbial body 9 also moves, so that the light beam of the optical fiber microlens 34 can be used to control the movement of the microbial body 9.

FIG. 7A shows a schematic diagram of a microscopic image of the light capturing microorganism identification method used for measuring the first yeast according to the preferred embodiment of the present invention. Referring to FIG. 7A, the light-trapping microorganism identification method according to the preferred embodiment of the present invention is used to measure the first yeast selected from the saf-instant brand. The measurement results are shown in Tables 1 and 4.

FIG. 7B shows a schematic diagram of a microscopic image of the light-trapping microorganism identification method used for measuring the second yeast according to the preferred embodiment of the present invention. Referring to FIG. 7B, the light-trapping microorganism identification method according to the preferred embodiment of the present invention is used to measure the second yeast selected from the brown brand. The measurement results are shown in Tables 2 and 4.

FIG. 7C shows a schematic diagram of a microscopic image of the light-trapping microorganism identification method used for measuring the third yeast according to the preferred embodiment of the present invention. Referring to FIG. 7C, the light-trapping microorganism identification method according to the preferred embodiment of the present invention is used to measure the third yeast selected from the fermipan brand. The measurement results are shown in Tables 3 and 4.

As shown in Table 4, different types of yeast have different characteristics of escape speed and capture efficiency. Therefore, in the mixed state of the first, second and third yeasts, the mixed yeast can be captured by the optical tweezers system, and after obtaining its escape speed and capturing efficiency, the yeasts caught can be known [brand] species.

FIG. 8 shows a schematic diagram of the light-trapping microorganism identification method of the preferred embodiment of the present invention used in measuring the first, second, and third yeasts to obtain their escape speed. Please refer to Figure 8 and Table 4, the escape speed of the first yeast is 25.58 to 29.95 μm/s, the escape speed of the second yeast is 12.22 to 14.40 μm/s and the escape speed of the third yeast is 8.49 To 10.4μm/s.

FIG. 9 shows a schematic diagram of the light-trapping microorganism identification method according to the preferred embodiment of the present invention for measuring the first, second, and third yeasts to obtain their capturing efficiency. Please refer to Figure 9 and Table 4. The first yeast capture efficiency is 3.17 to 3.46%, the second yeast capture efficiency is 2.26 to 2.80% and the third yeast capture efficiency is 1.43 to 1.82%.

The above experimental data are preliminary experimental results obtained under specific conditions. They are only for easy understanding or reference to the technical content of the present invention, and other relevant experiments are still required. The experimental data and results are not intended to limit the scope of the present invention.

The foregoing preferred embodiment only exemplifies the present invention and its technical features, and the technology of this embodiment can still be appropriately implemented with various substantial equivalent modifications and/or replacements; therefore, the scope of the present invention is subject to the appended patent application The scope defined by the scope shall prevail. The copyright in this case is restricted to the use of patent applications in the Republic of China.

S1‧‧‧ First step

S2‧‧‧The second step

S3‧‧‧The third step

S4‧‧‧The fourth step

Claims (10)

A light-trapping microorganism identification method, comprising: using a fiber optic tweezers to capture a microorganism, so as to form a captured microbe on the fiber optic tweezers; controlling the fiber optic tweezers and the captured microbes with at least one Move at a predetermined speed to measure the escape speed of a microorganism; use the escape speed of the microorganism to compare with the escape speed of a microorganism to be identified; and determine a type of microorganism according to the comparison result of the escape speed of the microorganism to be identified. According to the light-trapping microorganism identification method described in item 1 of the patent scope, various microorganisms are selected as a reference microorganism category. According to the light-trapping microorganism identification method described in item 1 of the scope of patent application, a standard database of microorganism identification required by the light-trapping inspection method is established. According to the light-trapping microorganism identification method described in item 1 of the patent scope, wherein the optical fiber optical tweezers are used to measure a catching force, a catching efficiency, or a combination thereof. According to the light-trapping microorganism identification method described in item 1 of the patent application scope, wherein the predetermined speed is a fixed speed. According to the light-trapping microorganism identification method described in item 1 of the patent application scope, wherein the microorganism escape speed is a microorganism escape speed range or a microorganism average escape speed. According to the light-trapping microorganism identification method described in item 6 of the patent application scope, wherein the microorganism escape speed range is between a maximum microorganism escape speed and a minimum microorganism escape speed. According to the light-trapping microorganism identification method described in item 1 of the patent application scope, wherein the microorganism is selected from a bacterium, a fungus, a virus, an algae or a protozoan [protozoa]. The method for identifying light-trapping microorganisms according to item 1 of the patent application scope, wherein the microorganism is selected from a yeast, a probiotic or a pathogen bacteria. According to the light-trapping microorganism identification method described in item 1 of the patent application scope, wherein the optical fiber optical tweezers is a laser optical fiber optical tweezers.

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