JP4881779B2 - Fine particle arrangement method, device in which fine particles are arranged, and device for creating a device in which fine particles are arranged - Google Patents

Description

  The present invention relates to a fine particle arrangement method for arranging fine particles at predetermined positions on a substrate. For example, the present invention relates to a method for arranging a single microparticle holding a single biomolecule on a substrate, a device manufactured using the same, and a manufacturing apparatus thereof. More specifically, for example, the present invention relates to a device manufacturing method for decoding the base sequence of a nucleic acid such as DNA or RNA.

  The operation of decoding the DNA base sequence is called a DNA sequence. Currently, for DNA sequencing, a method combining a Sanger method, which is a clever DNA fragment preparation method, and capillary electrophoresis is widely used. However, from the viewpoint of tailor-made medical care and the like, development of a new DNA sequencing technique that is inexpensive and simple is underway worldwide. Typical methods for next-generation sequencing include the Parallel bead assay method and the Sequence by ligation method. Among these technologies, as seen in Non-Patent Documents 1 and 2, the single molecule sequencing method can count up to the number of DNA molecules, and at the same time is inexpensive because it does not use PCR. Is expected.

  In the single molecule sequencing method, as described in Non-Patent Document 3, a method is proposed in which DNA or the like is randomly fixed on a substrate and its base sequence is determined. A sample DNA fragment to be analyzed is randomly fixed to the surface of the substrate one molecule at a time, extended by about one base at a time, and the result is detected by fluorescence measurement to determine the base sequence.

  On the other hand, nanoparticles, especially metal nanoparticles, or metal particles are expected as nanomaterials. As reported in Non-Patent Document 4, a material having a crystal generally becomes lower in energy and stable when surrounded by the same kind of atoms. On the other hand, on the surface where there is no atom next to it, the hands of the atoms extended for bonding cannot find the bonding partner and become unstable. That is, since atoms present on the surface are in a state of being higher in energy than in the bulk, the metal powder becomes unstable when the ratio of the surface area to the internal energy increases. By gradually reducing the size and reaching the nano level, various physical properties that cannot be considered in the bulk have been predicted.

  In addition, although single-molecule fluorescence is weak, in recent years, it is known that surface plasmon waves are generated when an evanescent wave is coupled to a gold surface as a fluorescence enhancement method. Non-Patent Document 5 and the like report enhancement of single molecule fluorescence using this surface plasmon wave. Furthermore, when fluorescent molecules are immobilized on gold fine particles, which are isolated nanoparticles, a strong electromagnetic field is generated in the vicinity of the gold fine particles, and the fluorescence emission is enhanced 10 times or more.

  Patent Document 6 reports a method of immobilizing single-stranded oligo DNA molecules on one gold fine particle. The oligo DNA and gold fine particles are mixed, and the functional group at the oligo DNA end is covalently bonded to the gold fine particle surface. Thereafter, gel electrophoresis is performed to selectively collect gold fine particles on which one single-stranded oligo DNA is immobilized. By this method, it is possible to prepare gold fine particles on which one single-stranded oligo DNA is immobilized.

  In addition, oligo DNA microarrays manufactured using inkjet technology are on the market. The phosphoramidite reaction is performed on the slide by sequentially spotting the nucleotide synthesis reagent at the correct position on the slide glass. By repeating this 60 times, an oligo of 60 base length is synthesized in situ on a slide glass. It is possible to form 40,000 or more spots having different base sequences on the slide glass. The spot diameter is 20 μm. This technique is described in Non-Patent Document 7.

  On the other hand, in the ink jet method (piezo method or thermal method) currently in practical use, it is difficult to discharge a minute amount of liquid that is less than 4 picoliters (corresponding to a sphere volume with a diameter of about 20 μm). However, as disclosed in Patent Document 1, in recent years, it has become possible to discharge ultrafine fluid droplets by using ultrafine nozzles. These technical innovations enable ejection of ultra-fine droplets of femtoliters (corresponding to a sphere volume with a diameter of about 1 μm) that is 1/1000 or less compared to the prior art. With this technique, it has been reported that a metal nanoparticle paste stably dispersed is used as an ink, and direct drawing of a metal fine wiring having a width of several μm on a glass substrate or a polyimide substrate is reported.

  Moreover, dark field microscopy can be mentioned as a means for easily observing metal fine particles of about nm. The dark field microscope is described in detail in Non-Patent Document 8. Dark field microscopy is a method of observing scattered light and reflected light generated by irradiating a sample with light obliquely. In this method, contrary to the bright field microscope, the background of the visual field is black and the sample appears to shine. In addition, this method can be realized simply by inserting a dark field condenser into an ordinary optical microscope. Although it is not suitable for observing the fine structure of the object surface or inside, it is possible to observe the presence of an object smaller than the wavelength of visible light with high contrast. According to dark field microscopy, flagellar fibers with a diameter of 20 nm can be clearly observed. It is also possible to detect metal fine particles with a high refractive index of 4 nm in diameter using high-intensity illumination. However, when observing biological supramolecules composed of proteins in solution, the limit is about 15 nm in diameter.

JP2004165559 Nature, vol437, pp376,2005 Science, vol309, pp1728, 2005 Proc.Natl.Acad.Sci.USA, vol.100 (7), pp3960, 2003 “Inkjet fine wiring of metal nanoparticle paste”, pp.3, CMC Publishing, 2006 Physical Review Letters, vol97, pp017402, 2006 Nano Lett, vol1, pp32, 2001 “Lab Manual: DNA Chips and Real-Time PCR”, pp46, Kodansha Scientific, 2006 "Biological microscope approaching the limit", pp17, Society Press, 1991

  As a result of intensive studies by the present inventor, the following has been found.

  As described above, in the conventional single molecule sequencing method, sample DNA fragment molecules to be analyzed are randomly fixed on the substrate surface. Therefore, for example, when the distance between DNA molecules is adjusted to be 1 μm on average, the distance between the molecules may be larger than 1 μm or may be closer than 1 μm. Therefore, when DNA molecules are randomly fixed on the substrate surface, it is necessary to detect fluorescent images at an interval finer than the average interval between DNA molecules. In order to separate and detect DNA molecules fixed at random with an average interval of 1 μm from each other, it is necessary to measure at intervals of several tenths of 1 μm in terms of the substrate surface. That is, comparing the case where the DNA molecules are randomly fixed and the case where the DNA molecules are regularly and ideally fixed in a lattice shape, the former requires several hundred times the number of pixels for measurement. Since a two-dimensional sensor with a large number of pixels is more expensive than that with a small number of pixels, in order to reduce the cost of the two-dimensional sensor in the detection system, it is desirable to regularly fix DNA molecules to the substrate at intervals of about 1 μm. It is.

  Further, an EB method is conceivable as a method for regularly arranging the gold fine particles on the substrate. This is a method in which a resist having a size of about 10 to 100 nm is removed with an electron beam at intervals of 1 μm, the exposed surface is modified with a functional group, and covalently bonded to gold fine particles. However, since the EB method is one-stroke writing with an electron beam, it is difficult to improve the throughput and a problem that costs increase is also expected. It is considered difficult to use the EB method for mass production of a substrate in which gold fine particles are regularly fixed. Therefore, a method for arranging gold fine particles regularly and efficiently on a substrate at low cost has been demanded.

  There has been no report on a method for regularly, rapidly and inexpensively arranging metal fine particles on which a single biomolecule to be measured is fixed on a substrate at intervals of μm order one by one.

  An object of the present invention relates to the regular, high-speed and low-cost arrangement of fine particles or the like that can immobilize a single biomolecule or the like on a substrate or the like at intervals of μm order.

  The present invention relates to placing droplets containing fine particles or the like on a substrate or the like with a predetermined probability, determining the presence of the fine particles or the like at a predetermined position, and arranging the fine particles or the like at a predetermined position on the substrate or the like.

  More specifically, for example, there is a method in which a plurality of single fine particles holding a single molecule to be measured are regularly arranged on a light-transmitting substrate. A droplet containing metal fine particles to which a single biomolecule is fixed is discharged onto the substrate by an ink jet method and fixed. Further, the concentration of the solution containing fine particles is adjusted so that the number of fine particles contained in the discharged droplets is 0.1 to 1 or less. In addition, the number of fine particles in the droplets discharged on the substrate is measured in real time using dark field microscopy. Further, the substrate can be driven in the XY directions, and droplets can be ejected to any region of the substrate.

  In the device creation apparatus, for example, fine particles are discharged onto the substrate from above the substrate, and the number and presence of fine particles are measured from below the substrate with a dark field microscope. The substrate can be arbitrarily driven in the XY directions. From the obtained information on the number of fine particles, droplets are ejected again to the region where the fine particles are not fixed by the ink jet method. By repeating this, it is possible to regularly arrange a plurality of single fine particles holding a single molecule to be measured on the substrate.

  According to the present invention, fine particles and the like can be arranged on a substrate or the like at intervals of the order of μm at high speed and at low cost.

  The above and other novel features and effects of the present invention will be described below with reference to the drawings. The drawings are exclusively used for understanding the invention and do not limit the scope of rights.

In this embodiment, a method for regularly arranging a plurality of gold fine particles fixed with a single biomolecule on a substrate, a device manufactured using the gold fine particle, and a device manufacturing apparatus will be described with reference to FIG.

  The fine particle arrangement method of this embodiment prepares a solution containing fine particles, discharges droplets of the solution to a predetermined position of the substrate, determines the presence of the fine particles at the predetermined position of the substrate, and places the fine particles at the predetermined position of the substrate. To do.

  In addition, the device manufacturing apparatus of this embodiment includes a means for moving a substrate capable of holding fine particles holding a single molecule to be measured to an arbitrary distance in a two-dimensional direction, and one fine particle on the substrate. Means for arranging, means for measuring the presence / absence or number of fine particles on the substrate, and means for integrating and controlling these means to place a plurality of fine particles regularly at different positions on the substrate one by one And have.

Further, the method of arranging the fine particles on the substrate is based on an ink jet method, and a fluid droplet containing the fine particles is discharged onto the substrate by applying a voltage to the tip of an ultrafine nozzle.

  The substrate is light transmissive.

  In addition, it has means for measuring the presence / absence or number of fine particles contained in the fluid droplets discharged on the substrate by dark field microscopy.

  Also, each time a fluid droplet is placed on one droplet substrate, the substrate is moved a certain distance, and the operation of ejecting the fluid droplet again to a new position on the substrate is repeated, so that a plurality of microparticles are collected one by one. A means for regularly arranging a plurality of different positions on the substrate is provided.

  Further, the substrate is moved so that the nozzle is located at a position where the presence of the fine particles is not detected by dark field microscopy, and the fluid droplet is discharged again.

  Also, by repeatedly discharging fluid droplets until the presence of fine particles on the substrate is detected by dark field microscopy, and when the presence of fine particles is detected, the operation of moving the substrate by a certain distance is repeated. A plurality of fine particles are regularly arranged one by one at different positions on the substrate.

  The means for moving the substrate is a means constituted by a stepping motor or a piezo.

  In addition, the volume of fluid droplets ejected from an inkjet type ultrafine nozzle is about femtoliter, and the nozzle diameter is about 0.5 to 1.5 μm.

  Also, a plurality of ultrafine nozzles of ink jet type are arranged, and the ejection of fluid droplets by the nozzles is independently controlled.

  In addition, the fine particles are arranged on the substrate by a bond between biotin and streptavidin or a chemical covalent bond.

In this apparatus, an inkjet nozzle 101, a light-transmitting substrate 102, a dark field microscope objective lens 104, a dark field observation light source 108, a dichroic mirror 109,
It comprises a two-dimensional sensor camera 105, a control PC 106, a stage drive motor 110, an arbitrary waveform generator 111, a high voltage amplifier 107, and a pressure regulator 112. In this embodiment, the nozzle 101, the dark field microscope objective lens 104, and the two-dimensional sensor camera 105 are arranged on the same optical axis. The dark field observation light source 108 is disposed perpendicular to the optical axis.

  A discharge signal from the control PC 106 is sent to the arbitrary waveform generator 111. The voltage generated by the arbitrary waveform generator 111 is transmitted to the electrode in the nozzle 101 through the high voltage amplifier 107. The solution in the nozzle 101 is charged and droplets 103 are ejected onto the substrate.

The diameter of the ink jet type nozzle 101 is 0.01 μm to 25 μm. The nozzle 101 can discharge femtoliter, that is, a droplet 103 which is a fluid having a diameter of 0.1 μm to 1.5 μm, onto the substrate 102. The dark-field microscope objective lens 104 does not use the illumination light from the dark-field observation light source 108 for the image formation of the gold fine particles, and forms only the light scattered by the gold fine particles. For this purpose, the dark field microscope objective lens 104 has an aperture stop mechanism. The aperture stop is appropriately adjusted so that the illumination light does not directly enter the dark field microscope objective lens 104. In the present embodiment, the dark field observation light source 108 is a halogen lamp. However, a mercury lamp, a xenon lamp, or a laser beam can be used as the observation light source.

  A discharge signal from the control PC 106 is sent to the arbitrary waveform generator 111. The voltage generated by the arbitrary waveform generator 111 is transmitted to the electrode in the nozzle 101 through the high voltage amplifier 107. The solution in the nozzle 101 is charged and droplets 103 are ejected onto the substrate. In this embodiment, the average number of gold fine particles contained in the ejected droplet 103 is 0.1. This is achieved by adjusting the concentration of the gold fine particle solution filled in the nozzle 101. Further, the gold fine particles are uniformly dispersed in the aqueous solution in the nozzle 101.

In this embodiment, every time one droplet 103 is ejected onto the substrate 102, the substrate 102 is moved at regular intervals. This is achieved by driving the stage drive motor 110 by the control PC 106. After driving the substrate 102, the control PC 106 sends an ejection signal to the arbitrary waveform generator 111. The voltage generated by the arbitrary waveform generator 111 is transmitted to the electrode in the nozzle 101 through the high voltage amplifier 107. The solution in the nozzle 101 is charged and droplets 103 are ejected onto the substrate. In order to explain more specifically, it will be described below with reference to FIG. Droplets 202, 203, and 204 are continuously discharged from the nozzle 201. The substrate 205 is driven by a fixed interval every time the droplets 202, 203, 204 are discharged. The droplets 202, 203, 204 contain gold fine particles 206. A single single-stranded oligo DNA is fixed to the gold fine particle 206. As a result, a single gold fine particle 206 on which a single single-stranded oligo DNA is immobilized can be disposed on the substrate 205. In this embodiment, the base 206 is driven, but the present invention is not necessarily limited to this configuration, and the inkjet nozzle 201 may move.

  FIG. 3 illustrates a drying process of droplets containing gold fine particles on which a single single-stranded oligo DNA is immobilized. Gold droplets 303 are contained in the droplets 301 ejected on the substrate 304. A single single-stranded oligo DNA 302 is fixed to the gold fine particle 303. The discharged droplet 301 has a femtoliter volume and a diameter of about 1 μm. The diameter of the gold fine particles 303 is in the range of 1 nm to 100 nm. The droplets 301 land on the substrate 304, and the drying of the droplets 301 proceeds quickly on the substrate 304. The gold fine particles 303 are attracted to the contact point between the droplet 301 and the substrate 304 by the capillary force of the droplet 301 during drying. A plurality of gold fine particles 303 can be regularly arranged on the substrate 301 by the effect of this capillary force.

The substrate 304 is modified on the substrate surface with a thiol group, and the gold fine particles 303 are covalently bonded to the substrate 304 surface. In this example, a reaction between gold and a thiol group was used, but the reaction is not limited to this, and various known binding methods such as gold and amino group, maleimide group and amino group, biotin and streptavidin are used. May be.

  The volume ratio of the gold fine particles 303 to the droplets 301 is 1: 1000 or less, and the concentration of the gold fine particle solution filled in the inkjet nozzle is extremely thin. Specifically, it is 3 centipoise (the viscosity of water is 1 centipoise) or less. Accordingly, problems such as nozzle clogging, which are problems in the fine particle paste method by the ink jet method, do not occur in this embodiment.

  By repeating the operations for discharging the droplet 103 and moving the substrate 102 in this way, a plurality of droplets 103 can be discharged and arranged on the substrate 102. Further, the movement interval of the substrate 102 can be controlled in the range of 0.1 to 100 μm. In this example, the interval was set to 1 μm.

By the above method, 4 × 10 ⁷ droplets 103 are arranged on the substrate 102. Since the frequency of the electrical signal applied to the nozzle 103 is on the order of 10 kHz, the time required to arrange 4 × 10 ⁷ droplets 103 on the substrate 102 is estimated to be about 1 hour. In addition, by further increasing the frequency, it is possible to reduce the time required for substrate manufacture.

When the concentration of gold fine particles in the solution filled in the nozzle 101 is determined (when the number of gold fine particles contained per droplet 103 is determined), the number of gold fine particles contained in the droplet 103 is Poisson distribution. Follow. Therefore, by adjusting the concentration of the fine particle solution filled in the nozzle 101, the number of fine particles contained in the discharged droplet 103 can be controlled stochastically. The Poisson distribution formula is p (x) = λ / x!・ Exp (−λ)
(Λ: average number of fine particles contained in one droplet, x: number of fine particles contained in one droplet)
Given in. When a large number of droplets 103 containing an average of 0.1 gold fine particles per liquid are discharged onto the substrate, the probability that one gold fine particle is placed on the substrate 102 is 9.0%. Similarly, the probability of 0, 2, and 3 is estimated to be 90.5%, 0.5%, and 0.0%. This probability distribution is shown in Table 1. According to this probability distribution, 99.5% of the ejected droplet 103 is in a state where one gold fine particle is held therein or not.

  As a method of measuring the number of gold fine particles in the discharged droplet 103, dark field microscopy is used. The presence / absence of gold fine particles in the droplet 103 can be accurately determined using dark field microscopy. It is also possible to distinguish one gold fine particle from two or more gold fine particles that have aggregated. The intensity of scattered light from nanometer-order gold fine particles smaller than the wavelength of light is explained by Rayleigh scattering. The Rayleigh scattered light intensity is proportional to the square of the diameter of the gold fine particle aggregate. Accordingly, four times as many scattered lights as single gold fine particles are emitted from the two gold fine particle aggregates. Similarly, 9 times as many scattered lights as single gold fine particles are emitted from three gold fine particle aggregates. Therefore, by measuring the scattered light intensity of the bright spot, the number of gold fine particles can be accurately determined using dark field microscopy.

Further, the specific concentration of 0.1 to 1 gold fine particles in the droplet 103 is 10 ⁻⁶ to 10 ⁻¹⁵ %, although the concentration of the gold fine particles depends on the size of the gold fine particles. It is included in the range of w / v.

When performing dark field microscopy, specifically, the illumination light from the dark field observation light source 108 is reflected by the dichroic mirror 109 in the optical axis direction, and the substrate 102 is illuminated through the dark field microscope objective lens 104. When fine particles are present in the droplet 103, scattered light is generated and collected by the dark field microscope objective lens 104, and an image of the gold fine particles is determined on the two-dimensional sensor camera 105. When there is no fine particle in the droplet 103, no scattered light is generated and no light is detected on the two-dimensional sensor camera 105. By using this method, the number of gold fine particles in the droplet 103 on the substrate 102 can be detected in real time immediately after the droplet 103 is ejected from the nozzle 101. On the substrate 102, 4 × 10 ⁷ droplets 103 are arranged. This is the first arrangement of gold fine particles on the substrate 102. Information on the number of gold fine particles in each droplet 103 is recorded by the control PC 106.

  Further, the control PC 106 moves a region on the substrate 102 where gold fine particles are not present directly below the nozzle 101 and applies a voltage to the nozzle 101 to rearrange the droplet 103 on the substrate 102. Here, since the previously disposed droplet 103 has already evaporated, the state of the substrate 102 with respect to the droplet 103 is the same as the state in which the droplet 103 is discharged for the first time. After the droplet 103 is discharged, the number of fine particles on the substrate 102 is detected by the dark field microscope objective lens 104. Thereafter, the substrate 102 is moved by 1 μm, and the discharge of the droplet 103 is repeated. Thereby, the second gold fine particles are arranged on the substrate 102.

By repeating the arrangement of the droplet 103 up to the tenth time, one gold fine particle can be arranged for 60.1% of the 4 × 10 ⁷ positions where the gold fine particles are to be arranged on the substrate 102. Similarly, the probability of 0, 2, and 3 is 36.8%, 3.0%, and 0.1%. This probability distribution is shown in Table 2. The number of arrangements is not limited to 10 and can be arbitrarily set.

  In this embodiment, a single nozzle 101 is used. However, by arranging a plurality of nozzles in parallel, a plurality of droplets can be arranged in parallel on a plurality of locations or a plurality of substrates.

  Further, in this embodiment, after one droplet 103 is arranged on the substrate 102, the substrate 102 is moved, and the droplet 103 is ejected and arranged at a different position on the substrate 102. good.

  The droplet 103 is continuously discharged from the nozzle 101 at a target position on the substrate 102. At the same time that one gold fine particle is detected and confirmed by the objective lens 104 for the dark field microscope, the control PC 106 stops the output to the arbitrary waveform generator 111. Next, the control PC 106 drives the substrate 102 by 1 μm and starts discharging the droplet 103 again. By repeating these operations, a plurality of gold fine particles having a single biomolecule fixed thereon can be regularly arranged on the substrate.

  According to the present embodiment, the metal fine particles on which a single biomolecule is immobilized can be regularly arranged at a high speed and at a low cost on the substrate at intervals of μm. As a result, the number of pixels of the two-dimensional sensor required for fluorescence measurement can be reduced from 10 times to 30 times or less, and even several hundred times or less, compared to the conventional case. Accordingly, a significant improvement in measurement throughput can be achieved. Moreover, the cost required for a two-dimensional sensor can be reduced. Further, the S / N of fluorescence measurement can be increased by the single molecule fluorescence enhancement effect of the gold fine particles, and the reliability of data can be improved. Therefore, the measurement throughput can be greatly improved, the cost can be reduced, and the data reliability can be improved.

The schematic diagram which shows the basic composition of an Example. Explanatory drawing about the discharge method of the droplet on a board | substrate in an Example. Explanatory drawing about the fixing method of one gold microparticle which fixed the single single strand oligo DNA on the board | substrate in an Example.

Explanation of symbols

101, 201 Inkjet nozzles 102, 205, 304 Substrate 103, 202, 203, 204, 301 Droplet 104 Objective lens 105 for dark field microscope 105 Two-dimensional sensor camera 106 Control PC
107 High-voltage amplifier 108 Dark-field observation light source 109 Dichroic mirror 110 Stage drive motor 111 Arbitrary waveform generator 112 Pressure regulators 207 and 302 Single single-stranded single-stranded oligo DNA

Claims (21)

Prepare a solution containing fine particles,
Discharging droplets of the solution to a predetermined position on the substrate,
Determine the presence of particulates at a given location on the substrate,
A fine particle arrangement method for arranging fine particles at a predetermined position on a substrate,
A method of adjusting the concentration of a solution containing fine particles so that the number of fine particles contained in a discharged droplet is 0.1 to 1. The fine particle arrangement method according to claim 1,
A method of discharging droplets of a solution to a predetermined position of a substrate by an ink jet method. The fine particle arrangement method according to claim 2,
The ink-jet method is based on voltage application to the tip of a nozzle. The fine particle arrangement method according to claim 1,
The method is characterized in that the volume of the droplet is about femtoliter. The fine particle arrangement method according to claim 1,
A method of measuring the number of fine particles at a predetermined position of a substrate. The fine particle arrangement method according to claim 1,
A method of determining the presence of fine particles at a predetermined position on a substrate by a measurement method using light. The fine particle arrangement method according to claim 1,
A method for determining the presence of fine particles at a predetermined position of a substrate by dark field microscopy. The fine particle arrangement method according to claim 1,
A method wherein the substrate is light transmissive. The fine particle arrangement method according to claim 1,
The method wherein the substrate is glass or quartz glass. The fine particle arrangement method according to claim 1,
A method of discharging a droplet to a predetermined position of a substrate, moving the substrate by a predetermined distance, and discharging the droplet to a new position of the substrate. The fine particle arrangement method according to claim 1,
A method of determining the presence of fine particles at a predetermined position on a substrate, and discharging a droplet to the predetermined position again when no fine particles are present at the predetermined position. The fine particle arrangement method according to claim 1,
A method wherein the fine particles hold single-stranded oligo DNA. The fine particle arrangement method according to claim 1,
The method, wherein the microparticles hold nucleic acids, proteins, sugar chains, glycolipids, peptides, or oligos. The fine particle arrangement method according to claim 1,
The method, wherein the fine particles are metal fine particles having a diameter of 5 to 100 nm. The fine particle arrangement method according to claim 1,
A method, wherein the fine particles are arranged in a lattice pattern on a substrate. The fine particle arrangement method according to claim 1,
A method characterized in that an interval between fine particles arranged on a substrate is 0.1 to 10 μm. The fine particle arrangement method according to claim 1,
10. The method according to claim 1, wherein the fine particles are arranged on the substrate in the number of 5 to the 10th power. The fine particle arrangement method according to claim 1,
A method of moving the substrate by a stepping motor, a stepping motor with an encoder, a linear motor, a linear motor with an encoder, or a piezo. The fine particle arrangement method according to claim 1,
The method, wherein the fine particles are held on the substrate by binding of biotin and streptavidin. The fine particle arrangement method according to claim 1,
The method, wherein the fine particles are held on the substrate by chemical covalent bonding.   A device prepared by the method according to any one of claims 1 to 20, wherein fine particles are arranged at predetermined positions on a substrate.

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