JP3333244B2 - How to arrange fine objects - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for three-dimensionally arranging fine objects on a substrate.

[0002]

2. Description of the Related Art In various fields, for a variety of purposes, fine objects are fixed on a base in a desired arrangement.

For example, Japanese Patent Application Laid-Open No.
No. 11428 discloses a method of two-dimensionally arranging liposomes having a certain function as a fine substance on a substrate in order to obtain a biodevice imitating the function of a living body.
Specifically, the antibody is fixed to the base by the LB (Langmuir-Blodgett) method, and the fixed antibody is selectively irradiated with an electron beam or an ion beam to selectively remove unnecessary portions of the antibody from the base. Then, an arbitrary pattern of an antibody is formed on a substrate, and the liposome is fixed to the antibody pattern by using an antigen-antibody reaction.

For example, Japanese Patent Application Laid-Open No. 3-79774 discloses ultrafine particles of metal, ultrafine particles of a metal compound, and ultrafine particles of ceramics on a substrate as a base for obtaining various electronic devices. A method of regularly arranging ultrafine particles of an oxide superconductor or ultrafine particles of an organic substance two-dimensionally is disclosed. Specifically, antigens are regularly arranged two-dimensionally on a substrate, while a monoclonal antibody reacting with the antigen is bound to ultrafine particles, and the antibody-bound ultrafine particles are combined with the antigen sequence. And binding to the substrate by an antigen-antibody reaction.

[0005]

However, in any of the above-mentioned methods, only a two-dimensional arrangement of fine objects on a base was impossible, and a three-dimensional arrangement was not possible. In addition, since the sequence fixation of the fine substance was performed using the antigen-antibody reaction,
The bonding strength between the fine substance and the base was not always satisfactory.

In order to improve the bonding strength between a fine substance and a substrate, the applicant of the present application has filed Japanese Patent Application No.
In No. 62130, antigen - a specific binding reaction and streptavidin and biotin binding strength compared to the antibody response capable of specifically binding also strong and binding objects together 10 ⁶ times, avidin and biotin And proposed a method of two-dimensionally arranging a fine substance (specifically, fine gold particles, liposomes, and proteins) on the base using both or one of the specific binding reactions.

However, this Japanese Patent Application No. 4-62130 discloses
Even with the method proposed in Japanese Patent Application Laid-Open Publication No. H11-27139, it was possible to arrange only two-dimensional fine objects on the base, but not three-dimensionally.

If a desired fine object can be three-dimensionally arranged in a desired pattern, the realization of a more sophisticated device can be expected. Therefore, the establishment of such a method is desired.

SUMMARY OF THE INVENTION The present invention has been made in view of the above circumstances, and an object of the present invention is to provide a method for three-dimensionally arranging desired fine objects on a base.

[0010]

In order to achieve this object, the inventors of the present application have made various studies. As a result, Document I (Journal of Physics D:
Applied Physics (J. Phys. D: App
l. Phys. 24 (1991) p. 1443-145
Attention has been paid to combining the bivalent binding property of bisbiotin disclosed in (0) with the above-mentioned technology proposed in Japanese Patent Application No. 4-62130 of the present application. In reference I,
It discloses that the properties of a monolayer prepared by chemisorption are examined by surface potential measurement, and that the use of a specific binding property of bisbiotin and avidin in preparation of a sample for that purpose is disclosed.

Therefore, according to the method for arranging fine objects according to the present invention, when three-dimensionally arranging fine objects on a base,
Using a specific binding reaction between avidin and bisbiotin and / or a specific binding reaction between streptavidin and bisbiotin between the fine substances, and comprising a plurality of steps of binding, between these steps, Patterning is performed by selectively irradiating avidin or streptavidin bound to the fine substance on the underside with energy rays to make the irradiated area an inactive area and leave the unirradiated area as an active area. And binding bisbiotin only to the active region.

[0012] In the present invention, the underlayer can be any of various objects on which fine objects are to be arranged. For example, a semiconductor base such as a silicon base or a compound semiconductor base used in the manufacture of semiconductor devices, a glass substrate or a ceramic substrate frequently used in various fields, and a semi-finished product in which other components are incorporated in these substrates. And so on.

That is, the method for arranging fine substances according to the present invention comprises: (a) a step of binding biotin to the above-mentioned base via an amino group; Binding to the bound biotin using both or one of a specific binding reaction between avidin and biotin and / or a specific binding reaction between streptavidin and biotin, and two-dimensionally arranging fine substances on a substrate; ( c) a patterning step for three-dimensionally arranging the fine particles, wherein avidin or streptavidin that binds to the fine particles and can bind to bisbiotin in the step (d) performed following the step (c); On the other hand, it selectively irradiates energy beam, inactivates avidin or streptavidin in the region where the energy beam was irradiated, and removes it. A patterning step of leaving avidin or streptavidin active in a region where irradiation with rugi rays has not been performed; and (d) converting bisbiotin to active avidin or streptavidin by a specific binding reaction between avidin and bisbiotin; A step of binding using both or one of the specific binding reactions of streptavidin and bisbiotin, and (e) specific binding of avidin or streptavidin-bound fine substance to bisbiotin and avidin and bisbiotin A step of binding using both or one of a binding reaction and a specific binding reaction between streptavidin and bisbiotin,
Further, by repeating the series of steps (c) to (e) as many times as necessary, fine objects are three-dimensionally arranged on the base.

[0014]

[0015]

[0016]

Further, in the practice of the present invention, when biotin is bonded to the substrate, an amino group is bonded to the entire surface of the substrate, the bonded amino groups are patterned, and the amino group pattern obtained by the patterning is formed. It is preferable to bond biotin, or to bond an amino group to the entire surface of the underlayer, bind biotin to the amino group, and pattern the biotin group.

[0018] The term "fine material" as used herein may be any of various materials to be arranged on a base. For example, A
ultrafine particles of metals such as u, Ag, Ni, Co, Fe, Cu, Pd, WCl ₆ , W ₂ C, Fe ₃ O ₄ , γ-Fe ₂
Ultrafine particles of metal compounds such as O ₃ and CrO ₂ , BaTi
Ultra fine particles of ceramics such as O ₃ and MnFe ₂ O ₄ ,
Ultra-fine particles of oxide high-temperature superconductors such as YBa ₂ Ca ₃ O ₇ , ultra-fine particles of organic substances such as carbon, proteins having a certain function, or liposomes composed of both or one of a biological substance and / or a biologically similar substance Can be mentioned.

[0019]

According to the method for arranging fine substances according to the present invention, since bisbiotin has a bivalent binding property, the fine substances can be bound to both ends thereof via avidin or streptavidin. Therefore, after arranging the microscopic objects on the substrate two-dimensionally, the required number of new microscopic objects are sequentially bonded to these microscopic objects by the method of the present invention using bisbiotin, whereby the microscopic objects can be arbitrarily three-dimensionally arranged. Can be arranged.

[0020]

Embodiments of the method for arranging fine objects according to the present invention will be described below with reference to the drawings. It should be noted that the drawings used in the description merely schematically show the dimensions, shapes, and arrangements of the components so that the invention can be understood. In the following drawings, the same components are denoted by the same reference numerals, and description thereof is omitted. In addition, numerical conditions such as chemicals used, chemical concentration, chemical use amount, processing time, processing temperature, etc., and processing methods used in the following examples are only preferred examples within the scope of the invention. Therefore, it is to be understood that these inventions are not limited only to these conditions.

1. First Example A first example will be described with an example in which a fine substance is made of two kinds of gold fine particles having different particle diameters. The size of the base is 1c.
An mx 1 cm silicon substrate is used. It is to be understood that the use of two types of gold fine particles is for facilitating the observation of the quality of the three-dimensional array pattern and is not the essence of the invention (the same applies to the number of types of fine objects in the following examples). .).

1-1. Substrate-side treatment A silicon substrate with a size of 1 cm x 1 cm is 31% by weight.
Of hydrogen peroxide solution and 97% by weight of sulfuric acid at a volume ratio of 1: 4 for 1 hour. Next, the silicon substrate is sufficiently washed with pure water. Next, this silicon substrate is immersed in an aqueous solution of aminopropyltriethoxysilane containing aminopropyltriethoxysilane (manufactured by Shin-Etsu Silicone Co., Ltd., model number LS-3150) at a ratio of 1% by volume for 2 minutes. After the silicon substrate taken out of the aqueous solution is washed with pure water, the silicon substrate is heated at a temperature of 110 ° C. in a constant temperature bath.
Heat for a minute. By this series of processes, as schematically shown in FIG. 1, an amino group capable of binding to a biotin derivative to be used later is fixed on the surface of the silicon substrate 11.

Next, a solution prepared by dissolving 10 mg of biotin long arm-NHS (a biotin derivative manufactured by Funakoshi Pharmaceutical Co., Ltd.) in 100 ml of dimethylformamide and 10 m
The above-mentioned amino group-immobilized silicon substrate is placed in a sample bottle containing a mixture with 1 l of bicarbonate buffer (pH 8.5). Thereafter, the sample bottle is placed in an ice bath for 2 hours to allow the substrate and the biotin derivative to react. Through this series of processes, the biotin derivative 13 is bonded to the silicon substrate 11 via the amino group, as schematically shown in FIG.

Next, an ultraviolet ray having a main wavelength of 290 nm is selectively irradiated through a photomask as an energy ray to the silicon substrate bound with the biotin derivative. In this case, the ultraviolet light is supplied by an aligner (PLA-52 manufactured by Canon Inc.).
Irradiate at an intensity of 1.3 mW for 30 minutes using 1). A photomask having a pattern corresponding to 1.2 μm line and space is used. In this ultraviolet irradiation, biotin in the ultraviolet irradiation region is inactivated, while biotin activity in the non-irradiation region is maintained. Therefore, as shown schematically in FIG. 3, a large number of linear biotin active regions having a width of 1.2 μm are formed on the silicon substrate 11 at 1.2 μm intervals by a selective irradiation treatment with ultraviolet light. An array pattern 13a is formed.

1-2. Bonding of Gold Fine Particles to Streptavidin On the other hand, streptavidin is bonded to fine gold particles as follows. Gold fine particles are described in Academic paper by Demey et al. (Proc. Natl. Acad. Sci. USA. 79, p. 1898 (19
1982)), by reducing an aqueous solution of chloroauric acid with a yellow phosphorus saturated solution of diethyl ether and ascorbic acid or citric acid. In this case, first, gold fine particles having a diameter of 5 nm are obtained. Streptavidin is extracted from bacteria. In this case, Funakoshi Pharmaceutical Co., Ltd. is used. Then, the ultrafine gold particles and streptavidin were mixed with 50 mM Tris-HCl.
(PH 7.5) and a 50 mM NaCl solution, and allowed to react for 24 hours to bind the ultrafine gold particles with streptavidin. What bound ultrafine gold particles and streptavidin in this manner is hereinafter referred to as "5 nm gold colloid streptavidin" for convenience of explanation.

Separately, the diameter of the fine gold particles is 25 nm.
A gold nanoparticle having a diameter of 25 nm is bound to streptavidin in the same procedure as that for obtaining 5 nm gold colloid streptavidin, except that streptavidin was obtained. This conjugate is hereinafter referred to as 25 nm gold colloid streptavidin for convenience of explanation.

1-3. Next, 2 μl of the above-mentioned solution containing 5 nm gold colloid streptavidin was taken out, and this was taken as a phosphate buffer (pH
Dilute to 2 ml according to 7.4). The biotin-aligned substrate prepared in section 1-1 is immersed in the 5 nm gold colloid streptavidin diluted solution thus prepared for 1 hour at room temperature. As a result of this process, as schematically shown in FIG. 4, gold particles 15 having a diameter of 5 nm are bonded to the substrate 11 via streptavidin 17 and biotin 13, so that gold colloids having a diameter of 5 nm are two-dimensionally arranged on the substrate. You. To confirm such a two-dimensional array, 1-1
5 nm in diameter on the substrate by the same procedure as that of the section 1-3.
A sample in which two gold colloids are arranged two-dimensionally is prepared separately,
This sample is sampled for observation by a scanning electron microscope (SEM) and then observed by SEM. In this observation, it was confirmed that a 1.2 μm line-and-space pattern formed of gold fine particles was formed on the substrate.

1-4. Three-dimensional array of gold fine particles Next, for the sample prepared in section 1-3, which was not used as the SEM sample, the line direction of the line pattern was changed to the previous one using the photomask used in section 1-1. Ultraviolet rays are exposed through this photomask under the same conditions as those described in section 1-1, while being set so as to be in a direction orthogonal to the time of exposure. Due to this selective irradiation treatment with ultraviolet light, as shown schematically in FIG. 5, a 1.2 × 1.2 μm square streptavidin active region 19 is formed in the non-irradiated region on the silicon substrate 11, and Then, a 1.2 × 1.2 μm square streptavidin-inactive region 21 is formed in each of the irradiation regions.

Next, the substrate on which the active region 19 of streptavidin is formed is immersed in a phosphate buffer of pH 7.2 containing 1 mg of 1,12-bisbiotinamide dodecane as bisbiotin at room temperature for 20 minutes. After taking out the substrate from this solution, it is washed three times using 100 ml each of a phosphate buffer of pH 7.2. By this treatment, as shown schematically in FIG. 6, only the active region 19 of streptavidin on the substrate 11 has bisbiotin 23
Selectively bind. In addition, the bisbiotin used here is the same as that disclosed in the above-mentioned document I. Here, the one synthesized by the inventor of the present application is used.

Next, 2 μl of a solution of 25 nm gold colloid streptavidin prepared in the above section 1-2 is taken and diluted to 2 ml with a phosphate buffer of pH 7.4. The bisbiotin-bound substrate described above is immersed in the 25 nm gold colloid streptavidin diluted solution thus prepared for 1 hour at room temperature. By this treatment, gold fine particles having a diameter of 25 nm are bound to the region of the substrate where bisbiotin is bound by a specific binding reaction between bisbiotin and streptavidin. When a sample for SEM was prepared from this sample and observed by SEM, the substrate had a diameter of 5 nm as schematically shown in FIG.
It was found that the region 25a where the gold fine particles were bonded and the region 25b where the gold fine particles having a diameter of 25 nm were bonded were regularly formed. FIG. 8 schematically shows a portion obtained by cutting the P portion of FIG. 7 along the II line. 8, reference numeral 27 denotes gold fine particles having a diameter of 25 nm.

As is clear from the description of the first embodiment, according to the method for arranging fine objects of the present invention, it is understood that gold fine particles can be three-dimensionally arranged on a base.

2. Second Example Next, a second example will be described using an example in which a fine substance is made of two types of proteins. In this second embodiment, a silicon substrate having a size of 2 × 2 cm is used as a base as in the first embodiment.

First, the processing on the substrate side is described as 1-1 in the first embodiment.
The silicon substrate 1 by performing the same procedure as
A 1.2 μm line-and-space pattern of a biotin derivative is formed on 1 (see FIG. 3). One of the two proteins (first protein)
First, albumin was described in Document II (J. Cell Biol.) Vol2
3. p. 981-986) and conjugated with streptavidin. The thus bound product is hereinafter referred to as an albumin-streptavidin complex for convenience of explanation.

Next, 10 ml of a phosphate buffer containing 1 mg of this albumin-streptavidin complex (pH
In 7.2), the silicon substrate on which the array pattern of biotin has been formed is immersed at room temperature for 20 minutes. By this treatment, albumin 31 is arranged in the biotin-binding region of the substrate 11 by a specific binding reaction between biotin 13 and streptavidin 17, as schematically shown in FIG.

Next, in the same manner as in the first embodiment, 1,12-bisbiotinamidododecane as bisbiotin was added for 1 m.
The albumin-bound substrate is immersed in a phosphate buffer of pH 7.2 containing g at room temperature for 20 minutes. After taking out the substrate from this solution, it is washed three times using 100 ml each of a phosphate buffer of pH 7.2. By this process, as shown schematically in FIG. 10, bisbiotin 23 binds to only the region of the substrate 11 to which the albumin-streptavidin complex is bound via the streptavidin 17.

Next, β-phycoerytherin (beta-phycoerytherin), which is a fluorescent protein, was used as the second protein in the same manner as in the case of albumin described above.
(J. Cell Biol. Vol 23. p. 981-
986) to bind to streptavidin. This conjugate is hereinafter referred to as a beta-phycoerythrin-streptavidin complex for convenience of explanation.

Next, the above-mentioned silicon substrate (FIG. 10) on which the sequence pattern of bisbiotin was formed was placed in 10 ml of a phosphate buffer (pH 7.2) containing 1 mg of the beta-phycoerythrin-streptavidin complex. Soak at room temperature for 20 minutes. As a result of this treatment, beta-phycoerythrin 33 is added to the bisbiotin 23 binding region of the substrate 11 as shown schematically in FIG.
And streptavidin 17 by specific binding reaction. It should be noted that the formation of such an array pattern indicates that the above sample was obtained by using a fluorescence microscope (Optiphoto 2: manufactured by Nikon Corporation) and a high-sensitivity camera (DAS-512-G1:
This was confirmed by the fact that a fluorescence image derived from beta-phycoerythrin was observed in a 1.2 μm-wide line shape in the fluorescence image obtained using Imagista.

3. Third Embodiment Next, a third embodiment will be described using an example in which a fine substance is made into two types of liposomes. In the third embodiment, a silicon substrate having a size of 2 × 2 cm is used as a base as in the first embodiment.

3-1. Processing on Substrate Side First, processing on the substrate side is performed in the same procedure as described in section 1-1 of the first embodiment to form a 1.2 μm line and space pattern of a biotin derivative on the silicon substrate 11. (See FIG. 3).

3-2. Preparation of Avidinated Proteoliposome Suspension First, 5 ml of a DPPC chloroform solution containing 20 mg of dipalmitoyl phosphatidylcholine (hereinafter, “DPPC”: manufactured by NOF Corporation) in 1 ml of chloroform.
l and alamesitin (Sigma) in 1 ml of ethanol
Solution of alamecithin containing 1 mg of
is mixed in a 50 ml eggplant-shaped flask. Thereafter, the solvent is removed from this mixed solution under reduced pressure by a rotary evaporator.

Next, 10 ml of a phosphate buffer (pH 7.2) containing 5 mg of Texas Red (trade name, manufactured by Funakoshi Pharmaceutical Co., Ltd.) as a fluorescent dye was added to the flask.
The suspension in the flask is stirred at a temperature of 37 ° C. using a vortex mixer to obtain a proteoliposome suspension.
The presence of proteoliposomes in this suspension was confirmed using a fluorescence microscope.

Next, 10 ml of a phosphate buffer (pH 7.2) containing 2% of glutaraldehyde is added to the proteoliposome suspension and reacted at a temperature of 4 ° C. for 2 hours. Thereafter, the solution is dialyzed to remove unreacted glutaraldehyde.

Next, this proteoliposome suspension was
It is added to 10 ml of phosphate buffer (pH 7.2) containing 1 mg of streptavidin and reacted at a temperature of 4 ° C. for 20 minutes.

As a result of this series of treatments, as schematically shown in FIG. 12, a liposome in which a fluorescent dye 43 is encapsulated in a lipid bilayer 41 and alamethicin 45 is incorporated in the lipid bilayer (hereinafter referred to as “ A streptavidinated liposome in which streptavidin 17 is bound to the first liposome via alamesitin is obtained.

Separately, a liposome containing carboxyfluorescein (manufactured by Funakoshi Pharmaceutical Co., Ltd.) instead of Texas Red as a fluorescent dye (this is referred to as a “second liposome”) is prepared by the above-mentioned procedure, and further streptococcal. Avidin is allowed to bind.

3-3. Two-dimensional arraying After washing the substrate having the biotin sequence pattern prepared in section 3-1 using a phosphate buffer (pH 7.3), in this case, the substrate is placed in a suspension of the first liposome streptavidinated. Soak for 1 hour at room temperature. By this treatment, as schematically shown in FIG. 13, the first liposome 47 binds to the substrate 11 by a specific binding reaction between biotin and streptavidin. Dimensionally arranged.

3-4. Next, for the sample in which the first liposome was two-dimensionally arranged, the photomask used in the section 1-1 of the first embodiment was applied so that the line direction of the line pattern was orthogonal to that of the previous exposure. Then, ultraviolet rays are exposed through this photomask under the same conditions as those described in section 1-1. Since the activity of streptavidin in the ultraviolet irradiation region disappears in the selective irradiation treatment of ultraviolet light, as shown schematically in FIG.
Only the streptavidin 17 which binds to the first liposome 47 in the 1.2 μm square region at every 1.2 μm region maintains the activity.

Next, the substrate on which the active region of streptavidin has been formed is immersed in a phosphate buffer having a pH of 7.2 containing 1 mg of 1,12-bisbiotinamidododecane as bisbiotin at room temperature for 20 minutes. After taking out the substrate from this solution, it is washed three times using 100 ml each of a phosphate buffer of pH 7.2. By this process, as shown schematically in FIG. 15, the bisbiotin 23 is selectively bound only to the streptavidin active region of the substrate 11.

Next, the substrate to which the above-mentioned bisbiotin is bound is immersed in a suspension of the second liposome streptavidinated at room temperature for 1 hour. By this processing, FIG.
As schematically shown in FIG. 2, the second liposome 49 binds to the region of the substrate 11 to which bisbiotin is bound by a specific binding reaction between bisbiotin 23 and streptavidin 17.

3-5. Confirmation of pattern formation Confirmation of the formation of the arrangement as shown in FIG. 16 is performed by the following method. A sample was placed on a fluorescence microscope (Optiphoto 2:
Nikon Corporation) and high-sensitivity camera (DAS-512-G)
1: Imagista Co., Ltd.) and observation conditions corresponding to the above two types of fluorescent dyes (Texas Red, etc.): observation at an excitation wavelength of 490 nm at an observation wavelength of 520 nm, and observation at an excitation wavelength of 600 nm at an observation wavelength of 620 nm. Observation is performed under the two conditions described above and fluorescence images are obtained. From these fluorescent images, it was confirmed that a three-dimensional array structure of liposomes was obtained as schematically shown in FIG.

Although the embodiments of the method for arranging fine objects according to the present invention have been described above, the present invention is not limited to the above embodiments.

For example, in each of the above embodiments, two layers of fine objects are formed on a substrate. However, the number of layers of fine objects may be three or more. Even when three or more layers are stacked, the arrangement of the fine substances can be performed by repeating the procedure of the example, that is, by bonding the fine substances of the adjacent layers by a specific binding reaction between bisbiotin and streptavidin. .

In each of the above embodiments, two types of fine objects are used. However, as described above, the number of types may be one or three or more depending on the design.

In the third embodiment, alamecithin was used as a membrane protein for binding streptavidin to liposomes. However, any other protein that penetrates the lipid bilayer and exposes amino groups on the membrane surface can be used. Things are fine. For example, glycophorin and the like can be mentioned.

In each of the above embodiments, biotin and streptavidin were inactivated by irradiation with ultraviolet rays.
The same effect as that of the embodiment can be obtained by performing the irradiation with another energy ray such as a beam or laser beam.

In the above description, the binding between the substrate and the fine substance is carried out using a specific binding reaction between biotin and streptavidin, and the binding of the new fine substance to the fine substance fixed on the substrate is performed with bisbiotin. Although a specific binding reaction with streptavidin has been performed, avidin extracted from egg white may be used instead of streptavidin. If necessary, both the specific binding reaction between avidin and bisbiotin and the specific binding reaction between streptavidin and bisbiotin may be used.

The biotin and bisbiotin used are not limited to those of the embodiment, but may be other suitable ones.

[0058]

As is clear from the above description, according to the method for arranging fine objects of the present invention, fine objects can be three-dimensionally arranged in a desired arrangement. Therefore, when a fine object is made of, for example, metal fine particles, for example, the construction of a three-dimensional fine wiring structure can be expected. When a fine object is made of a liposome or a protein having a certain function, an information processing multifunctional bio imitating an organism An element can be expected to be constructed, and further, it can be expected to contribute to the construction of a biocomputer, its input device or its output device.

[Brief description of the drawings]

FIG. 1 is an explanatory diagram of substrate-side processing.

FIG. 2 is an explanatory view following FIG. 1 of substrate-side processing;

FIG. 3 is an explanatory view subsequent to FIG. 2 of substrate-side processing;

FIG. 4 is a diagram showing a state where gold fine particles are two-dimensionally arranged on a substrate.

FIG. 5 is a process chart of a three-dimensional array of gold fine particles.

FIG. 6 is a process drawing following FIG. 5 of a three-dimensional array of gold fine particles.

FIG. 7 is a process drawing following FIG. 6 of a three-dimensional array of gold fine particles.

FIG. 8 is an enlarged schematic view of a portion P in FIG. 7;

FIG. 9 is a process chart for explaining a second embodiment.

FIG. 10 is a process drawing following FIG. 9 of the second embodiment.

FIG. 11 is a process drawing following FIG. 10 of the second embodiment.

FIG. 12 is a process chart for explaining a third embodiment;

FIG. 13 is a process drawing following FIG. 12 for explaining the third embodiment;

FIG. 14 is a process drawing following FIG. 13 for describing a third embodiment;

FIG. 15 is a process drawing following FIG. 14 for describing a third embodiment;

FIG. 16 is a process drawing following FIG. 15 for describing a third embodiment;

[Explanation of symbols]

11: base (for example, a silicon substrate) 13: biotin 13a: sequence pattern of biotin 15: fine gold particles (5 nm in diameter) 17: streptavidin 19: active region of streptavidin 21: inactive region of streptavidin 23: bisbiotin 25a: Gold fine particle binding region with a diameter of 5 nm 25b: Gold fine particle binding region with a diameter of 25 nm 25: Gold fine particles with a diameter of 25 nm 31: First protein (albumin) 33: Second protein (β-phycoerythrin) 41: Lipid bilayer 43: Fluorescent dye 45: Membrane protein 47: First liposome (containing Texas Red) 49: Second liposome (containing carboxyfluorescein)

Continuation of the front page (72) Inventor Masakazu Kato 1-7-12 Toranomon, Minato-ku, Tokyo Examiner, Oki Electric Industry Co., Ltd. Kyoichi Saito (56) References JP-A-5-226637 (JP, A) JP-A-1-248570 (JP, A) JP-A-63-111428 (JP, A) D.M. Phys. D: Appl. Phys. 24, UK, 1443-1450 (58) Fields investigated (Int. Cl. ⁷ , DB name) H01L 49/00 H01L 51/00

Claims (4)

(57) [Claims] 1. A method for arranging microscopic objects on a lower ground using a specific binding reaction between avidin and bisbiotin and / or a specific binding reaction between streptavidin and bisbiotin. And a step of binding a plurality of times, between each of the steps, selectively irradiating an energy beam to avidin or streptavidin bound to the fine substance on the underlying side, the irradiation portion inactive region A patterning method for performing patterning for leaving unirradiated portions as active regions, and bonding bisbiotin only to the active regions. 2. A step of: (a) binding biotin to a substrate via an amino group; and (b) combining avidin or streptavidin-bound fine substance with avidin and biotin on the biotin bound to the substrate. A step of binding by using both or one of a specific binding reaction and a specific binding reaction between streptavidin and biotin to two-dimensionally arrange the fine substances on the substrate; and (c) aligning the fine substances three-dimensionally. Patterning step for selectively irradiating energy rays to avidin or streptavidin which binds to the fine substance and can bind to bisbiotin in the step (d) following the step (c). Deactivating the avidin or the streptavidin in the area where the energy beam irradiation has been performed, and performing the energy beam irradiation. A patterning step of leaving the avidin or the streptavidin in the active region in an active state; and (d) converting the bisbiotin to the active avidin or streptavidin by a specific binding reaction between avidin and bisbiotin and the streptavidin. A step of binding using both or one of the specific binding reactions with bisbiotin; and (e) binding the avidin or streptavidin-bound fine substance to the bisbiotin, specifically binding reaction between avidin and bisbiotin. And binding using streptavidin and / or bisbiotin using a specific binding reaction, and repeating the series of steps (c) to (e) as many times as necessary to form A method for arranging fine objects, wherein the fine objects are three-dimensionally arranged. 3. The method for arranging fine objects according to claim 2, wherein: (f) a patterning step for two-dimensionally arranging the fine objects, which is performed subsequent to the step (a); The bound biotin is selectively irradiated with an energy beam to inactivate biotin in a region where the energy beam has been irradiated and to activate biotin in a region where the energy beam has not been irradiated. The method further includes a patterning step of leaving the pattern as it is, and in the step (b) subsequent to the step (f),
Binding the active biotin to the fine substance to which avidin or streptavidin has been bound, using a specific binding reaction between avidin and biotin and / or a specific binding reaction between streptavidin and biotin. A method of arranging fine objects. 4. The method for arranging fine objects according to claim 1, wherein an ultraviolet ray is used as the energy ray.