JP5718673B2 - Element for molecular manipulation - Google Patents

Description

  The present invention relates to a molecular manipulation element for performing a separation operation on a molecular film disposed on a substrate.

  A technique for detecting molecules contained in a very small amount of sample is a very important technique for industrial application. According to this technique, for example, a molecule such as an environmental hormone that affects a living body can be detected even in a very small amount. In addition, the smaller the number of samples, such as biomolecules, can reduce the invasion of intake sources such as blood and tissues. Thus, the above-described molecular detection technique has practical advantages.

  Also, in basic research, a technique that can manipulate, fractionate, and detect molecules without impairing their functions is desired. Ultimately, it is desired to apply these techniques to a single molecule. Such technology is not only useful for basic molecular science research such as intermolecular interaction, intermolecular self-organization, and intermolecular reaction, but the knowledge obtained by applying these researches is, for example, disease. It will lead to drug discovery through elucidation of the mechanism of this and is expected to be applied in the future life science field.

  As a general trace detection technique, a chromatography technique represented by gas chromatography and liquid chromatography is most commonly used in the field of chemistry and biology. However, in these chromatographic techniques, the time required for one analysis is a few tens of units, and it is necessary to prepare a suitable time or a plurality of chromatographic instruments for the analysis of a large number of samples. The amount of processing per process cannot be increased (low throughput), and rapid analysis is not easy.

  On the other hand, with the recent development of MEMS (micro electromechanical system) technology, microfluidic devices and micro-TAS (micro total analysis system) called tube-shaped reactors that supply samples to microreactors and microreactors Devices with flow paths have been developed. In this element, medical applications such as detection of biomolecules and blood tests are among the major goals. Development of this type of device has led to a reduction in the amount of specimens required to reduce the amount of specimens and thus to less invasiveness to the human body, and to achieve high throughput by incorporating multiple microsensors. It has been.

  In the microfluidic device and the μTAS element, the required amount of the sample is significantly reduced as compared with the analysis technique that is generally used conventionally. However, in these elements, since a tube is used for transporting the solution, an increase in viscous resistance at the time of liquid feeding due to a decrease in the tube diameter is a limiting factor for further miniaturization.

  Under such circumstances, a novel microchannel device that does not use a three-dimensional reaction tank or a liquid feeding system has been reported (Non-Patent Document 1). These devices utilize the spontaneous development of lipid bilayers, where lipid molecules spontaneously form lipid bilayers at the solid-liquid interface, and use lipid bilayers as sample transport media instead of aqueous solutions. It is characterized by that. Although the sample needs to be bound to or embedded in the lipid bilayer membrane, the sample can be transported and reacted while confined in a two-dimensional region of about 5 nm thick membrane.

  Further, in a conventional microfluidic device or μTAS, it is necessary to dissolve or disperse a sample in an aqueous solution. For this reason, for example, it is impossible to inspect a sample that does not exhibit its function unless placed in a specific environment. There are many types of such samples, including a series of membrane proteins that are responsible for in-vivo information transfer. This is because these membrane proteins express their functions by changing the molecular shape, but this molecular shape cannot be maintained unless they are embedded in the cell membrane. A microchannel device using a lipid bilayer membrane as a sample transport medium is also a useful means for the purpose of detecting the function of a membrane protein.

  On the other hand, the lipid bilayer membrane supported on the solid surface itself has a fluid property. This is known as lateral diffusion, and has been proved by fluorescence recovery (fluorescence recovery after photobleaching; FRAP) experiments after fluorescence quenching (Non-patent Document 2).

  Regarding the fluidity of the film supported on the solid (substrate) surface, the situation is the same in the case of the monomolecular film. For example, the same lipid molecules as described above have been reported to form monolayers on hydrophobic surfaces. This monomolecular film exhibits the same spontaneous expansion phenomenon as described above, and the fluidity of molecules in the film has been proven (Non-patent Document 3).

  As a method of moving molecules bound or encapsulated in two-dimensional regions of molecules such as fluid bilayers or monolayers by external stimuli, the lipid bilayer supported on the substrate surface is externally attached. There is a method of applying an electric field. It has been shown that by applying an electric field in a supported lipid bilayer membrane confined in a specific region, dye molecules existing in the membrane can be unevenly distributed at specific sites in the membrane ( Non-patent document 4).

K. Furukawa et al., "Microchannel device using self-spreading lipid bilayer as molecule carrier", Lab Chip, vol.6, pp.1001-1006, 2006. J. Salafsky et al., "Architecture and Function of Membrane Proteins in Planar Supported Bilayers: A Study with Photosynthetic Reaction Centers", Biochemistry, vol.35, pp.14773-14781, 1996. I. Czolkos et al., "Controlled Formation and Mixing of Two-Dimensional Fluids", Vol.7, No.7, pp.1980-1984, 2007. JAY T. GROVES AND STEVEN G. BOXER, "Micropattern Formation in Supported Lipid Membranes", Acc. Chem. Res., Vol.35, pp.149-157, 2002.

  However, the above-described molecular manipulation technique by applying an external electric field involves applying an electric field by placing an electrode in an electrolyte solution for maintaining a lipid bilayer membrane supported on a substrate. The electric field can only be applied. Therefore, only one type of electric field is applied to the lipid bilayer membrane supported on the substrate surface at the same time. This makes it possible to manipulate the molecules bound or embedded in the support membrane in one direction, but in principle, no more complicated operations can be performed.

  Here, if a molecule bound or embedded in the membrane can be manipulated in the membrane by applying a plurality of electric fields locally and independently within one supporting membrane, a new molecular fractionation and molecular It may be applicable to detection technology. For example, by manipulating specific molecules in the support membrane, this may be separated and detected. Further, by limiting the movement of molecules to a two-dimensional region instead of a three-dimensional region, there is a possibility that operations such as separation and detection can be performed efficiently using a smaller amount of sample through manipulation of the position of the molecule. However, no method has been developed so far for a method of applying an electric field as described above and a molecular operation using the same in a film supported on a substrate.

  The present invention has been made to solve the above-described problems, and an object of the present invention is to enable more efficient manipulation of molecular positions in a molecular film.

The element for molecular manipulation according to the present invention includes an electrode formed on a substrate, an insulating layer formed on the substrate so as to cover the electrode, and formed on the insulating layer, to which molecules to be manipulated are bonded. Alternatively, it includes at least an embedded molecular film.

  The molecular manipulation element may be provided with a partition structure that is formed on the insulating layer and partitions the presence or absence of the molecular film on the insulating layer. The partition structure may be composed of, for example, a hydrophilic region and a hydrophobic region. In addition, the partition structure may be composed of unevenness and areas having different roughness.

  In the molecular manipulation element, the molecular film may be composed of a lipid bilayer. In addition, a plurality of electrodes each insulated and separated may be provided on the substrate, and different voltages may be applied to the plurality of electrodes.

  As described above, according to the present invention, at least an electrode formed on a substrate, an insulating layer formed on the substrate so as to cover the electrode, and a molecular film formed on the insulating layer are provided. As a result, it is possible to obtain an excellent effect that the manipulation of the position of the molecule in the molecular film can be performed more locally and efficiently.

FIG. 1A is a cross-sectional view showing the configuration of the molecular manipulation element according to Embodiment 1 of the present invention. FIG. 1B is a cross-sectional view showing the configuration of the molecular manipulation element according to Embodiment 1 of the present invention. FIG. 1C is a plan view showing the configuration of the molecular manipulation element according to Embodiment 1 of the present invention. FIG. 2 is a photograph showing the result of observing the molecular layer on the surface of the molecular manipulation element with a confocal laser microscope. FIG. 3A is a perspective view showing a partial configuration of the molecular manipulation element according to Embodiment 2 of the present invention. FIG. 3B is a perspective view showing a partial configuration of the molecular manipulation element according to Embodiment 2 of the present invention. FIG. 4A is a plan view showing a partial configuration of the molecular manipulation element according to Embodiment 2 of the present invention. FIG. 4B is a cross-sectional view showing a partial configuration of the molecular manipulation element according to Embodiment 2 of the present invention. FIG. 4C is a cross-sectional view showing a partial configuration of the molecular manipulation element according to Embodiment 2 of the present invention. FIG. 4D is a cross-sectional view showing a partial configuration of the molecular manipulation element according to Embodiment 2 of the present invention. FIG. 5 is a photograph showing a microscopic observation image of the pattern layer produced on the insulating layer formed so as to cover the electrodes. FIG. 6 is a photograph showing the result of observation of the state of molecular manipulation using the molecular manipulation element in Example 2 of Embodiment 2 of the present invention with a confocal laser microscope.

  Hereinafter, embodiments of the present invention will be described with reference to the drawings.

[Embodiment 1]
First, Embodiment 1 of the present invention will be described with reference to FIGS. 1A, 1B, and 1C. 1A and 1B are cross-sectional views showing the configuration of the molecular manipulation element according to Embodiment 1 of the present invention. FIG. 1C is a plan view showing the configuration of the element for molecular manipulation according to Embodiment 1. 1A shows a cross section taken along line aa ′ in FIG. 1C, and FIG. 1B shows a cross section taken along line bb ′ in FIG. 1C.

  This molecular operation element includes electrodes 131, 132, 133, and 134 formed on the substrate 101 so as to be isolated from each other, and insulation formed on the substrate 101 so as to cover the electrodes 131, 132, 133, and 134. At least a layer 104 and a molecular film 110 formed on the insulating layer 104 are provided. The molecular film 110 is, for example, a bimolecular film or a monomolecular film. In this embodiment mode, a plurality of electrodes 131, 132, 133, and 134 are provided, and each is insulated and separated by an insulating layer 102 formed on the substrate 101.

The substrate 101 only needs to be made of a solid material such as silicon. When a silicon substrate is used, the insulating layer 102 can be formed by thermally oxidizing the surface. Each electrode can be formed by patterning a metal such as Au by a so-called lift-off method. The insulating layer 104 may be formed, for example, by depositing SiO ₂ and SiN by sputtering or CVD.

  According to the present embodiment, since the molecular film is disposed on the electrode via the insulating layer, the region where the molecular film spreads by the electrode is not limited. Moreover, since an electric field can be applied to the molecule | numerator immediately above an electrode, an electric field can be locally applied with the large area matched with the electrode. Since an electric field can be applied in a larger area in this way, operations such as collecting specific molecules on the insulating layer can be performed efficiently. In addition, since an electric field can be applied locally, a complex molecular operation can be performed by arranging a plurality of electrodes and independently applying an electric field.

[Example 1]
Next, the first embodiment will be described in more detail using examples. First, preparation of a molecular manipulation element will be described. First, a circular silicon wafer having a diameter of 3 inches on which a thermal oxide layer having a thickness of 300 nm was formed was prepared, and a 200 nm SiN film was deposited on the thermal oxide layer of the silicon wafer by a plasma CVD apparatus.

  Next, the surface of the deposited SiN film was washed with a 0.5% hydrofluoric acid solution, and a photoresist was applied to the washed surface by a spin coating method. Next, the applied photoresist layer was exposed to light through a reticle on which an electrode pattern was drawn, and further developed to transfer the electrode pattern to the photoresist layer.

Next, the photoresist layer to which the electrode pattern was transferred was used as a mask pattern, and then the SiN film was selectively etched with sulfur hexafluoride gas. At this time, a condition in which the SiN film is etched at a high selection ratio with respect to the thermal oxide layer made of SiO ₂ is adopted, and the SiN film in the portion where the electrode is formed is completely removed, and the thermal oxide layer in this region Was not etched.

  Next, with the photoresist layer left, a film of titanium 50 nm, gold 100 nm, and platinum 20 nm was deposited using an electron beam evaporation apparatus. Thereafter, the photoresist layer was removed by a lift-off method, and an electrode made of the above-mentioned metal laminated film was formed in the region where the SiN film was removed. The surface was cleaned by oxygen plasma treatment (400 W, 10 minutes) and semi-coclean (organic alkaline cleaning solution manufactured by Furuuchi Chemical Co., Ltd.).

Next, a photoresist layer was formed on the SiN film on which the electrode was formed by spin coating to form a photoresist layer, and a pattern for protecting the electrode pad portion for external input / output was drawn on the photoresist layer. Next, 200 nm of SiO ₂ was deposited by a sputtering apparatus, and the photoresist portion was removed by a lift-off method. Through the above-described process, an electrode substrate is obtained which is composed of a plurality of electrodes formed by insulating and separating on the substrate and an insulating layer formed on the substrate so as to cover the electrodes. Further, the insulating layer covering the electrode is not formed on the electrode pad portion, and the electrode pad portion is exposed. This electrode substrate was used for the experiment.

  Next, production of a molecular film formed on the electrode substrate (insulating layer covering the electrode) will be described. First, a vesicle solution is prepared. Lipid molecule mixtures containing 1 mol% of Texas Red-dihexanoylphosphatidylcholine in egg yolk-derived phosphatidylcholine were prepared using each chloroform solution. The chloroform solution is prepared in a glass bottle. Next, nitrogen gas was blown onto the prepared chloroform solution to evaporate the chloroform, and a film composed of mixed lipid molecules was formed on the inner wall of the glass bottle. Further, it was dried overnight under vacuum to completely remove chloroform.

  Next, add 1.0 mL of phosphate buffer (PBS buffer, 0.1 M sodium phosphate, 0.15 M sodium chloride, pH 7.2) into the glass bottle, and sonicate for 10 minutes. Lipid vesicles were formed within. A solution obtained by diluting this solution 10 times with a PBS buffer (lipid concentration: 0.2 mg / mL) was prepared as a vesicle solution.

  Next, the surface of the electrode substrate described above was washed with pure water, then immersed in a 40% ammonium fluoride aqueous solution for 5 minutes, and washed again with pure water. Subsequently, this electrode substrate was dipped in the vesicle solution described above for 10 minutes, and further washed repeatedly with the phosphate buffer used for the preparation of the vesicle solution. By this process, a lipid bimolecular film is formed on the surface of the electrode substrate (insulating layer covering the electrode), and the molecular manipulation element is completed. The electrode substrate (element for molecular manipulation) after being immersed in the vesicle solution must always be kept covered with a buffer solution or water. This is because the lipid bilayer film formed on the surface of the electrode substrate collapses upon exposure to the atmosphere.

  Next, the molecular manipulation in the molecular manipulation device produced as described above will be described. Here, the state of molecular manipulation is observed with a confocal laser microscope. In this observation, the space between the lens of the confocal laser microscope and the sample substrate produced as a molecular manipulation element was filled with a phosphate buffer. The electrode pads used for external input / output were arranged so as not to come into contact with the phosphate buffer. As the observation condition of the confocal laser microscope, excitation light of 543 nm was used and fluorescence having a wavelength of 610 nm or more was recorded.

  2A and 2B are confocal laser microscope images before and after voltage application, in which FIG. 2A shows a state immediately before voltage application, and FIG. 2B shows a state 30 minutes after voltage application. In the microscopic image, four straight lines extending in the vertical direction of the drawing are electrodes. Each electrode can be connected to an external circuit. In FIG. 2, a voltage of 1 V was applied between the leftmost electrode and the rightmost electrode. The polarity is + on the left and-on the right.

  As shown in FIG. 2 (b), after 30 minutes of voltage application, a state in which fluorescence is emitted more strongly in the vicinity of the electrode to which the positive voltage is applied is observed. This is because the Texas Red-dihexanoyl phosphatidylcholine in the yolk-derived phosphatidylcholine support membrane moved in the support membrane (lipid bilayer) by the electric field in the direction of the electrode to which a positive voltage was applied. Conceivable. Thus, efficient molecular operation became possible by using the electrode arranged under the insulating layer.

[Embodiment 2]
Next, a second embodiment of the present invention will be described. 3A, FIG. 3B, FIG. 4A, FIG. 4B, FIG. 4C, and FIG. 4D show a partial configuration of the molecular manipulation element in the second embodiment. 3A and 3B are perspective views. In FIG. 3B, each layer is shown separately. 4A is a plan view, and FIGS. 4B, 4C, and 4D are cross-sectional views. 4B shows a cross section taken along line bb ′ of FIG. 4A, FIG. 4C shows a cross section taken along line cc ′ of FIG. 4A, and FIG. 4D shows a cross section taken along line dc ′ of FIG. In FIG. 3A, FIG. 3B, FIG. 4A, FIG. 4B, FIG. 4C, and FIG. 4D, the molecular film is omitted and only the electrode substrate is shown.

This molecular operation element includes electrodes 331, 332, 333, and 334 that are formed on the substrate 301 so as to be isolated from each other, and insulation that is formed on the substrate 301 so as to cover the electrodes 331, 332, 333, and 334 And at least a layer 304. In this embodiment mode, four electrodes 331, 332, 333, and 334 are provided, and each of them is insulated and separated by an insulating layer 302 and an insulating layer 302 a formed over the substrate 301. The insulating layer 302 is made of, for example, SiO ₂ , and the insulating layer 302a is made of, for example, SiN. Each electrode is formed in the recess of the electrode pattern formed in the insulating layer 302a. In addition, electrode pad portions 331a, 332a, 333a, and 334a connected to the electrodes 331, 332, 333, and 334 are formed on the insulating layer 302 where the insulating layer 304 is not formed. On the electrode pad portions 331a, 332a, 333a, 334a, an insulating layer or the like is not formed, and these are exposed.

In this embodiment mode, a pattern layer 305 is provided on the insulating layer 304 as a partition structure that partitions the presence or absence of a molecular film (not shown) on the insulating layer 304. The pattern layer 305 includes two opening regions 351 and 353, and a plurality of communication paths 352 that allow the opening region 351 and the opening region 353 to communicate with each other. In the present embodiment, four communication paths 352 are provided. For example, if the insulating layer 304 is made of SiO _2, the surface of the insulating layer 304 can be made hydrophilic, and if the pattern layer 305 is made of a photoresist material, the surface can be made hydrophobic. Thus, a partition structure including a hydrophilic region and a hydrophobic region can be formed on the insulating layer 304.

  Hereinafter, the fabrication of the molecular manipulation element in the present embodiment will be described.

For example, the substrate 301 may be a silicon substrate. If a silicon substrate on which a SiO ₂ film is formed is used, an insulating layer 302 is obtained on the substrate 301. Note that the substrate 301 may be formed of a solid material such as a semiconductor, glass, metal, plastic, or ceramics as long as it has an insulating surface such as an insulating layer 302 and is not particularly limited. For example, in the case where the substrate 301 is formed using a conductive material, for example, silicon oxide may be deposited to form the insulating layer 302 as described above.

  Next, electrodes 331, 332, 333, and 334 are formed over the insulating layer 302. First, SiN is deposited to form the insulating layer 302a. Next, this insulating layer 302a is patterned by a known lithography technique and etching technique. In this patterning, an opening is formed in a region where an electrode is to be formed. For this patterning, a pattern forming technique such as nanoimprinting can be applied. By depositing a metal material for forming an electrode in the opening region of the insulating layer 302a formed in this manner, each electrode and electrode pad portion can be formed. Note that in the insulating layer 302, a recess corresponding to a region where an electrode is to be formed may be formed, and each electrode and electrode pad portion may be formed in the recess.

  The insulating layer is not limited to SiN, and may be made of other insulating materials including organic polymers. An insulating layer made of these insulating materials may be patterned and etched to produce an electrode. Each electrode and electrode pad portion may be formed by patterning an electrode material layer formed by depositing an electrode material. As the electrode material, various materials having conductivity can be used.

After each electrode is formed as described above, an insulating layer 304 is formed on these electrodes. The insulating layer 304 may be made of, for example, SiO ₂ . The insulating layer 304 may be composed of other insulating materials such as organic polymers.

  Next, the pattern layer 305 is formed over the insulating layer 304. The pattern layer 305 can be formed by patterning it with a lithography technique using a known photosensitive resist. The pattern layer 305 may be formed by a pattern formation technique such as nanoimprint. In this manner, after the pattern layer 305 is formed, a molecular film such as a bimolecular film or a monomolecular film may be disposed in a portion where the insulating layer 304 in the opening region of the pattern layer 305 is exposed.

  According to the second embodiment, in a molecular film disposed on the insulating layer 304, molecules that are bound to or embedded in these films can be manipulated with high sensitivity and a very small sample. In addition, as shown to FIG. 4A, it is also possible to apply a different voltage between several electrodes by arrange | positioning several electrodes. Alternatively, when the molecular manipulation element is submerged in water / aqueous solution, it is possible to apply different voltages between the water / aqueous solution and the electrode.

  As described above, in this embodiment, an electrode is covered with an insulating layer, and a pattern is formed thereon to provide a bimolecular film or a monomolecular film. Since the electrode is buried under the insulating layer in this way, the electrode has an advantage in structure that the region where the bimolecular film or the monomolecular film spreads is not limited. In addition, since an electric field can be applied to molecules directly above the electrode, it is possible to apply an electric field locally in a large area that matches the electrode, and position operations such as collecting specific molecules on this large solid surface. It becomes possible.

[Example 2]
Next, the second embodiment will be described in more detail using examples. First, preparation of a molecular manipulation element will be described. First, a circular silicon wafer having a diameter of 3 inches on which a thermal oxide layer having a thickness of 300 nm was formed was prepared, and a 200 nm SiN film was deposited on the thermal oxide layer of the silicon wafer by a plasma CVD apparatus.

  Next, the surface of the deposited SiN film was washed with a 0.5% hydrofluoric acid solution, and a photoresist was applied to the washed surface by a spin coating method. Next, the applied photoresist layer was exposed to light through a reticle on which an electrode pattern was drawn, and further developed to transfer the electrode pattern to the photoresist layer.

Next, the photoresist layer to which the electrode pattern was transferred was used as a mask pattern, and then the SiN film was selectively etched with sulfur hexafluoride gas. At this time, a condition in which the SiN film is etched at a high selection ratio with respect to the thermal oxide layer made of SiO ₂ is adopted, and the SiN film in the portion where the electrode is formed is completely removed, and the thermal oxide layer in this region Was not etched.

  Next, with the photoresist layer left, a film of titanium 50 nm, gold 100 nm, and platinum 20 nm was deposited using an electron beam evaporation apparatus. Thereafter, the photoresist layer was removed by a lift-off method, and an electrode made of the above-mentioned metal laminated film was formed in the region where the SiN film was removed. Oxygen plasma treatment (400 W, 10 minutes) and cleaning with an organic alkaline cleaning solution were performed to clean the surface.

Next, a photoresist was applied on the SiN film on which the electrode was formed by spin coating to form a photoresist layer, and a pattern for protecting the electrode pad portion for external input / output was drawn on the photoresist layer. Next, 200 nm of SiO ₂ was deposited by a sputtering apparatus, the photoresist portion was removed by a lift-off method, and an insulating layer covering the electrode was formed. The insulating layer covering the electrode is not formed on the electrode pad portion, and the electrode pad portion is exposed.

  Next, a photoresist layer was formed by applying a photoresist on the insulating layer covering the electrodes by a spin coating method. Next, the formed photoresist layer was exposed to a pattern through a photomask, developed, and drawn (transferred) on the photoresist layer to form a pattern layer (partition region). FIG. 5 is a microscopic observation image of the pattern layer produced on the insulating layer formed so as to cover the electrodes. It can be seen that there are two upper and lower patterns that are connected between two 500 μm square opening areas by communication paths (16 lines) having a width of 10 μm and a length of 400 μm. In the photograph of FIG. 5, the communication path extends in the left and right direction on the paper surface. The opening region and the communication path are regions where there is no photoresist, and the surface of the hydrophilic insulating layer (silicon oxide) is exposed. Note that the four visible lines extending in the vertical direction in FIG. 5 are electrodes formed below the insulating layer, and are arranged with a line width of 10 μm and a line interval of 500 μm.

  Next, the production of the molecular film formed on the electrode substrate having the pattern layer as the partition region described above will be described. First, a lipid mixture is prepared. A lipid molecule mixture containing 1 mol% of fluorescein-dihexanoylphosphatidylcholine in egg yolk-derived phosphatidylcholine was prepared using each chloroform solution. The chloroform solution is prepared in a glass bottle. Next, nitrogen gas was blown onto the prepared chloroform solution to evaporate the chloroform, and a film composed of mixed lipid molecules was formed on the inner wall of the glass bottle. Further, it was dried overnight under vacuum to completely remove chloroform. A viscous solid was obtained as the final product.

  Next, formation of a molecular film using the prepared lipid mixture will be described. First, a glass capillary is prepared, and the viscous solid is scraped from the glass bottle at the tip of the capillary. Next, the scraped viscous solid was applied to the square open areas in the pattern layer. The applied viscous solid was a spot-like lump having a diameter of about 100 μm.

  The sample prepared as described above was made observable with a confocal laser microscope, and tris (hydroxymethyl) aminomethane (Tris) 10 mM adjusted to pH 7.6 between the objective lens for water immersion and the sample. And an electrolyte solution consisting of 100 mM NaCl was gently poured. Thereby, the applied lipid mass started to expand spontaneously. When about 30 minutes have elapsed from the start of the spontaneous deployment, the membranes that have started the spontaneous deployment from the two open regions come into contact with each other in the communication path having a width of 10 μm. The spontaneously developing lipid bilayer membrane was formed only on the hydrophilic surface where the silicon oxide surface was exposed, and when it spread over the entire hydrophilic surface, further development (development on the pattern layer) was stopped.

  Next, the molecular manipulation in the molecular manipulation device produced as described above will be described. Here, the state of molecular manipulation is observed with a confocal laser microscope in which a sample is arranged. In this observation, excitation light of 488 nm was used, and fluorescence having a wavelength of 505 to 525 nm and fluorescence having a wavelength of 610 nm or more were separated and recorded (observed) simultaneously. The state of observation is shown in FIG.

  6A shows an initial state, and FIG. 6B shows a voltage application state described below. In FIGS. 6A and 6B, lines passing through the portions of the characters “Black1,” “Black2,” “Red1,” and “Red2” in the vertical direction each indicate a part of the electrode. ing. In FIG. 6, there are four electrodes. Further, in FIG. 6, the stripe pattern extending in the horizontal direction of the paper, which is shown brighter in the drawing, indicates the communication port portion of the pattern layer, in other words, the region where the spontaneously developed lipid bilayer exists. . In this part, fluorescence of fluorescein is observed. On the other hand, in FIG. 6, the stripe pattern that exists in the lateral direction of the paper, which is shown darker in the drawing, is the portion of the pattern layer that exists so as to sandwich the communication port portion, in other words, the lipid bilayer membrane is It is an area that does not exist

  Under observation as described above, a voltage is applied to the electrodes. In this voltage application, a voltage of 1 V is applied between the “Black2” electrode and the “Red2” electrode. The state after this voltage application (after 420 seconds) is a photograph shown in FIG. When 1 V is applied and 430 seconds elapse, as shown in FIG. 6B, it can be seen that the lipid bilayer membrane in the “Black2” electrode portion is darker than the other regions. This is because the fluorescence intensity at the electrode portion of “Black2” decreased. This change in fluorescence intensity occurs because it is proportional to the amount of fluorescein-bound lipid molecules in the lipid bilayer. From this result, the concentration of the fluorescein-bound lipid molecule in the lipid bilayer membrane arranged in the insulating layer above the electrode is determined by applying a voltage to the electrode arranged via the insulating layer under the lipid bilayer membrane. It was shown that it can be controlled.

  As described above, according to the present invention, the position of molecules existing in a molecular film such as a monomolecular film or a bimolecular film can be controlled in the molecular film by applying an external voltage. Yes. As described above, according to the present invention, a specific molecule in the molecular film can be manipulated to a specific position, so that the specific molecule can be classified and detected based on this. The present invention is particularly applicable to molecular manipulation in a lipid bilayer membrane, which is the basic structure of a cell membrane, and is expected to be applied to concentration and detection of biomolecules such as membrane proteins and functional analysis. . Thereby, according to the element for molecular manipulation of this invention, biochip applications, such as a disease diagnostic device and a protein chip, are possible, for example.

The present invention is not limited to the embodiment described above, and many modifications and combinations can be implemented by those having ordinary knowledge in the art within the technical idea of the present invention. It is obvious. For example, in the above description, a hydrophobic region and a hydrophilic region are formed using a resist pattern to form a partitioned region, but the present invention is not limited to this. For example, a partition pattern region may be formed by forming a metal pattern such as gold on a layer made of a hydrophilic material such as SiO ₂ . Since the metal becomes hydrophobic, a partition structure composed of a hydrophobic region and a hydrophilic region can be formed. Alternatively, a partition structure may be obtained by simply forming irregularities, disposing a molecular film in the concave portion, and not having a molecular film in the convex portion. Moreover, you may form a division area structure from the area | region of each different roughness.

  DESCRIPTION OF SYMBOLS 101 ... Substrate, 102 ... Insulating layer, 104 ... Insulating layer, 110 ... Molecular film, 131, 132, 133, 134 ... Electrode.

Claims (6)

An electrode formed on a substrate;
An insulating layer formed on the substrate to cover the electrode;
A molecular manipulation element comprising at least a molecular film formed on the insulating layer and bonded or embedded with a molecule to be manipulated. The molecular manipulation device according to claim 1,
A molecular manipulation element comprising a partition structure formed on the insulating layer and configured to partition the presence or absence of the molecular film on the insulating layer. The element for molecular manipulation according to claim 2,
The element for molecular manipulation, wherein the partition structure is composed of a hydrophilic region and a hydrophobic region. The element for molecular manipulation according to claim 2,
The element for molecular manipulation characterized in that the partition structure is composed of irregularities and regions of different roughness. In the element for molecular manipulation of any one of Claims 1-4,
The molecular manipulation element is characterized in that the molecular membrane is composed of a lipid bilayer membrane. In the molecular manipulation element according to any one of claims 1 to 5,
A molecular manipulation element comprising a plurality of the electrodes each insulated and separated on the substrate, wherein different voltages are applied to the plurality of electrodes.