JP2008051766A - Method and device for manufacturing droplet forming substrate - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a manufacturing technology of a droplet forming substrate capable of reducing dispersion in spot size. <P>SOLUTION: The position of a dispensing vessel is set so that the perpendicular distance between the tip of a delivery port disposed in the bottom of the dispensing vessel storing liquid and a spot forming part of an object to be delivered becomes a predetermined gap not longer than the length in the dropping direction of the mass of liquid capable of being held in the delivery port. Liquid is pressurized to form the mass of liquid in the delivery port, the mass of liquid grown until it comes into contact with the substrate. After the mass of liquid comes into contact with the spot forming part of the object to be delivered, the pressurization of the liquid is stopped, and the dispensing vessel is left at rest until the movement of the liquid stops in a state where the mass of liquid is held between the delivery port and the object to be delivered. After that, the dispensing vessel is separated from the mass of liquid, and a spot is formed on the object to be delivered. <P>COPYRIGHT: (C)2008,JPO&INPIT

Description

  The present invention relates to a droplet forming substrate in which a large number of droplets (liquid mass) such as a liquid sample are formed on a substrate, and more particularly to a method for manufacturing a droplet forming substrate and a manufacturing apparatus for a droplet forming substrate.

  The droplet forming substrate is used as a biochip (microarray) such as a DNA chip, an RNA chip, a protein chip, or a sugar chain chip. A biochip is obtained by aligning and immobilizing hundreds to tens of thousands of biological materials and chemical substances on a substrate such as a glass slide of several square centimeters at high density. Biochips are used to search for substances that interact with substances on the chip and to obtain information from signal patterns of interacting substances. Biochips are expected in various fields such as therapeutic drug development tools, disease diagnosis, and health monitoring because they can process and analyze large amounts of information at once.

  The method for producing a biochip can be roughly classified into an optical lithographic method, a surface adhesion method, and an ink jet method.

  Among the dispensing methods, the surface adhesion method is a method in which the needle tip of a dispensing needle is immersed in a liquid containing a biological sample such as a DNA fragment and then the liquid is spotted by contacting the needle tip on a substrate. It is. The dispensing needle can be reused by washing with a washing solution or the like, and various types of biological samples can be spotted using the same dispensing needle. This method is advantageous in that the cost is low and it is easy to create a unique arrangement pattern. However, on the other hand, when the tip of the needle is first immersed in the liquid, the spot size varies between the spot of the liquid first formed and the spot after it due to excess liquid adhering around the tip of the dispensing needle. In some cases, the spot size of the liquid may vary depending on the contact state such as the contact time and contact angle between the needle tip and the substrate.

Under such circumstances, in order to solve the above problem, Japanese Patent No. 3436741 (Patent Document 1) discloses that after the dispensing needle is immersed in the liquid, excess liquid adhering to the needle tip is discarded. The method of removing by this is disclosed. Moreover, in JP 2004-325329 A (Patent Document 2), the distance between the dispensing pin and the dispensing object is kept constant by the positioning pin, the tip of the dispensing pin is not brought into contact with the surface of the dispensing object, A method is disclosed in which a droplet containing protein held at the tip of a dispensing pin is brought into contact with a dispensing object and dispensed in a non-contact manner by the surface tension of the droplet. Japanese Patent Laid-Open No. 2003-172744 (Patent Document 3) discloses a non-contact type trace liquid in which a trace liquid (biological sample) is dropped onto a measurement substrate on which a metal thin film is formed without damaging the metal thin film. A dripping method is disclosed.
Japanese Patent No. 3436741 JP 2004-325329 A JP 2003-172744 A

  However, in the method of Patent Document 1, since the dispensing needle directly contacts the substrate, there is a possibility that variation due to contact may occur. Further, in the method of Patent Document 2, since the liquid is collected by immersing the tip of the dispensing pin in the liquid, the spot size varies between the first and subsequent liquid spots due to excess liquid adhering around the tip of the dispensing pin. May occur. In addition, since the dispensing pin is moved in a state where the droplet is held, the droplet may drop at an unplanned place or timing.

  On the other hand, in the method of Patent Document 3, since the liquid is replenished from the tank to the dispensing needle by a pump, there is no problem that the above-described droplets are caused by dropping at an unplanned place or timing. In addition, by irradiating micron-order light and measuring the spread of the scattered light with a sensor, the size (shape) of the droplet is measured, and the amount of liquid sent to the tip of the needle is controlled. Quantification of However, in this method, since the droplet size is directly controlled by a pump or the like while measuring the droplet size with a sensor, it is necessary to control a very small amount of liquid. In order to control such a small amount of liquid, a highly accurate apparatus is required, and the price of the apparatus increases, so that the manufacturing cost of the droplet forming substrate also increases.

  Therefore, an object of the present invention is to provide a technique for producing a droplet formation substrate that can reduce variations in liquid spot size by a cheaper method. Another object of the present invention is to provide a dispensing technique capable of avoiding contamination that may be caused by dropping of a droplet at an unplanned place or timing.

  In order to achieve the above object, a method for manufacturing a droplet forming substrate according to the present invention includes a dispensing container that has a plurality of discharge ports at the bottom and contains a liquid, and a plurality of spot forming portions to which droplets are to be supplied. The predetermined gap in a range in which the distance between the discharge port and the spot forming portion does not exceed the holding limit length for holding the liquid mass formed at the discharge port. The process of setting the relative position of the dispensing container and the discharge target body, and pressurizing the liquid to generate the liquid mass at the discharge port, The process of growing the liquid mass until it comes into contact with the spot formation site of the discharge target body, the process of holding the liquid mass for a while between the discharge port and the spot formation region, and the dispensing container Separated from the object to be ejected, the liquid The containing the steps of the above droplets by transferring each on the spot formation region.

  In the above method, the liquid mass formed at the discharge port is held between the discharge port and the spot forming portion of the discharge target object until the liquid movement (change in the amount of liquid held) stops. By holding for a while, the liquid volume of the liquid mass at each spot forming site is adjusted to be the same. Since the physical conditions of each discharge port are the same, and the physical conditions of each spot formation site are also common, each droplet applied to the discharge target object side after separating the dispensing container from the discharge target object The amount will be the same. According to the above method, it is possible to manufacture a microarray having no variation in spot size by a cheaper method without requiring an expensive control device. Also, in the above method, the liquid mass held in the dispensing container is grown (hangs down) after determining the application position of the liquid droplet, so that the liquid droplet falls at an unplanned place or timing during the movement of the dispensing container. It is possible to avoid contamination caused by doing so.

  Preferably, each spot forming portion of the discharge target body is lyophilic, and the periphery of each spot forming portion is lyophobic. Thereby, the bottom face shape of the droplets adhering to each spot forming site can be made constant.

  Preferably, the periphery of each discharge port of the dispensing container is liquid repellent. Thereby, it is possible to avoid that the liquid mass is connected between adjacent discharge ports.

  Preferably, each spot forming portion of the discharge target body is lyophilic, the periphery of each spot forming portion is liquid repellent, and the periphery of each discharge port of the dispensing container is liquid repellent. Thereby, the shape of each liquid mass held between the discharge port and the spot forming region becomes the same, and the same amount of liquid droplets (liquid mass) can be applied to each spot forming region.

  Further, in the method for producing a droplet forming substrate of the present invention, the liquid is dispensed to the spot forming site on the discharge target body where the spot forming site is more lyophilic than at least the surrounding region of the spot forming site. A method of manufacturing a droplet forming substrate that forms a plurality of spots, wherein the tip of a discharge port provided at the bottom of a dispensing container holding a liquid is perpendicular to the spot forming portion of the discharge target object The process of setting the position of the dispensing container so that the distance in the direction becomes a predetermined gap that is equal to or less than the limit length in the falling direction of the liquid mass that can be held in the discharge port, and pressurizing the liquid. Forming a liquid mass at the discharge port and growing the liquid mass until it comes into contact with the substrate; and after the liquid mass comes into contact with the spot forming portion of the discharge target body, the pressurization of the liquid is stopped. , The discharge port and the discharge target The liquid container is held between the liquid container and the liquid container until the liquid movement stops, the process of allowing the liquid container to stand on the object to be ejected, Forming a spot of liquid on the discharge target body.

  In the above method, the liquid mass formed at the discharge port is held between the discharge port and the spot forming portion of the discharge target object until the liquid movement (change in the amount of liquid held) stops. By letting it stand, the amount of liquid forming the spot is adjusted. Therefore, it is possible to determine the balance between the force that draws the liquid on the discharge port side and the force that draws the liquid on the discharge target side without actively finely adjusting the liquid volume, and therefore an expensive control device is required. However, it is possible to manufacture a droplet forming substrate with no variation in spot size by a cheaper method. Further, in the above method, since the liquid mass held in the dispensing container is dropped, it is possible to avoid contamination caused by dropping of the droplet at an unplanned place or timing during the movement of the dispensing container. .

  Here, the object to be ejected is not particularly limited as long as it is a member that is generally used for producing a droplet forming substrate, and examples thereof include a substrate such as a glass substrate.

  According to another aspect of the present invention, a plurality of spots are formed by dispensing liquid to the spot forming site on the discharge target body, where the spot forming site is more lyophilic than at least the surrounding region of the spot forming site. And a tip of the discharge port of a dispensing container having a plurality of liquid storage chambers for storing liquid and a plurality of discharge ports respectively provided at the bottom of the liquid storage chamber And the dispensing container so that a vertical distance between the spot forming portion and the spot forming portion of the discharge target object is a predetermined gap that is equal to or less than a limit length in a dropping direction of the liquid mass that can be held in the discharge port. And the liquid is pressurized to form a liquid mass at the discharge port, and the liquid mass is grown until all of the liquid mass discharged from the discharge port contacts the object to be discharged. Process and all the above liquids Until the liquid is stopped in a state where the liquid mass is held between the discharge port and the discharge target object Droplet formation including a process of allowing the dispensing unit to stand on the object to be ejected and a process of separating the dispensing unit from the liquid mass to form a spot on the object to be ejected. A method for manufacturing a substrate is provided.

  In the above method, the liquid mass formed at the discharge port is held between the discharge port and the spot forming portion of the discharge target object until the liquid movement (change in the amount of liquid held) stops. By letting it stand, the amount of liquid forming the spot is adjusted. Therefore, it is possible to determine the balance between the force that draws the liquid on the discharge port side and the force that draws the liquid on the target side without separately finely adjusting the amount of liquid, so that an expensive control device is not required. Thus, it is possible to manufacture a droplet forming substrate with no variation in the spot size of the liquid by a cheaper method. In addition, according to the above method, even if the size of the liquid mass formed by the corresponding plurality of discharge ports varies due to the difference in the pressing force for pressing the liquid stored in each liquid storage chamber, the final In particular, since the liquid amount is adjusted by the balance of the above forces, it is possible to reduce the variation in the size of the formed spots. In the above method, since the liquid mass held in the dispensing container is moved (transferred), it is possible to avoid contamination caused by dropping of liquid droplets at an unplanned place or timing during the movement of the dispensing container. Become.

  It is preferable that the spot forming part of the discharge target body is lyophilic, and at least the periphery of the spot forming part is lyophobic. According to this, since at least the periphery of the spot formation site is liquid repellent, it is possible to more reliably form droplets in the spot formation site. Therefore, it is possible to more reliably reduce spot size variation.

  According to still another aspect of the present invention, a spot is formed by dispensing a liquid to the spot forming site on the discharge target object, the spot forming site having higher lyophilicity than at least the surrounding region of the spot forming site. A microarray manufacturing apparatus to be formed, which is accommodated in a plurality of liquid storage chambers for storing a liquid, the top of which is open, and the liquid storage chambers provided at the bottoms of the plurality of liquid storage chambers, respectively. A dispensing container comprising: a plurality of discharge ports for discharging liquid to the outside; and one or a plurality of lids that cover the openings of the plurality of liquid storage chambers to form one or a plurality of closed spaces , A pressurizing unit that pressurizes the closed space to form a liquid mass at the plurality of discharge ports, and a discharge target surface between a discharge port tip of the dispensing container and a discharge target object A substantially vertical distance is maintained at the discharge port. Moving means for moving the dispensing container in the substantially vertical direction so that a predetermined gap is equal to or less than a limit length in a dropping direction of the liquid mass that can be performed, and all the discharge ports formed in the plurality of discharge ports A detecting means for detecting that the liquid mass has contacted the object to be ejected, and a stop signal for stopping the pressurization to the pressurizing means in response to a detection signal sent from the detecting means, and the stop signal There is provided a droplet forming substrate manufacturing apparatus comprising: a control unit that transmits a movement signal to the moving unit so as to be separated from the discharge target object after a predetermined time after transmission.

  According to this, it is detected that the liquid mass formed in all the discharge ports has contacted the object to be discharged, the pressurization is stopped, and the dispensing container is separated from the object to be discharged after a predetermined time thereafter. Therefore, the liquid amount can be determined by a balance between the force for pulling the liquid on the discharge port side and the force for pulling the liquid on the discharge target side without separately finely adjusting the liquid amount. Therefore, since a highly accurate control mechanism is not required, a more inexpensive droplet forming substrate manufacturing apparatus can be provided. This also makes it possible to manufacture, for example, a microarray with no variation in spot size at a lower cost. In addition, even if there is a variation in the size of the liquid mass formed at the corresponding plurality of discharge ports due to the difference in the pressing force that presses the liquid stored in each liquid storage chamber, the above force Since the liquid amount is adjusted by the balance, it is possible to reduce the variation in the size of the formed spot. Further, in this apparatus, after positioning the dispensing container, the liquid mass is grown at the discharge port of the dispensing container and the liquid mass is moved (transferred) to the ejection target body side. It is possible to avoid contamination caused by dropping at an unplanned place or timing during movement.

  An apparatus for manufacturing a droplet forming substrate according to the present invention includes a dispensing container having a liquid storage chamber for storing a liquid, a plurality of discharge ports for discharging the liquid to the outside, and a discharge target to which the droplet is to be applied. A body, a moving means for setting the mutual position so that the dispensing container and the object to be ejected face each other, and pressurizing the liquid storage chamber to form a liquid mass at the plurality of ejection ports. A pressure unit, and a control unit that controls the moving unit and the pressurizing unit, and the control unit controls the moving unit so that the dispensing container and the discharge target body are vertically opposed to each other. The distance between the discharge port of the dispensing container and the discharge target is set to be a predetermined gap in a range that does not exceed the holding limit length of the liquid droplet that drops formed on the discharge port, By controlling the pressurizing means between the plurality of discharge ports of the dispensing container and the discharge target object By controlling the moving means after the formation of the lumps is separated and the dispensing container and the discharge target object, characterized in that.

  According to such a configuration, it is possible to simultaneously apply a plurality of substantially the same amount of droplets (sample solution, etc.) onto a target object (substrate, etc.) with a relatively simple configuration.

  Preferably, the apparatus further includes a detection unit that detects that one or a plurality of liquid masses formed at the plurality of discharge ports has contacted the discharge target object, and the control unit includes a plurality of the dispensing containers. After the detection unit detects the formation of a liquid mass between the discharge port and the discharge target body, the moving means is controlled to separate the dispensing container from the discharge target body. Thereby, the adhesion of the liquid mass is actually confirmed, and the dispensing container is separated, so that it is possible to ensure the work and shorten the work time.

  Preferably, a spot forming portion on which the droplet (liquid mass) of the discharge target body is to be applied is lyophilic, and at least a surrounding portion of the spot forming portion is lyophobic. Thereby, the bottom shape of the applied droplet (liquid mass) becomes constant.

  Preferably, the periphery of the discharge port of the dispensing container is liquid repellent. Thereby, it is possible to avoid the connection (contact) of the liquid mass between the adjacent discharge ports. Further, it is possible to set the arrangement interval of the discharge ports to be narrower than in the case where the liquid repellent treatment is not performed.

  Preferably, the control unit detects that the liquid mass has contacted the object to be ejected, and then passes a temporary time required for each liquid mass to be substantially the same amount, and then the dispensing container. And the discharge target object are separated from each other. Thereby, each droplet (liquid block) applied to the discharge target body becomes substantially the same amount.

  Preferably, the dispensing container includes a plurality of liquid storage chambers having discharge ports, the liquid storage chambers are grouped, and the pressurization is performed for each group. Thereby, the pressurization to the liquid is made uniform and the response of the growth of the liquid mass is made uniform.

  In another aspect of the microarray manufacturing apparatus of the present invention, the liquid is distributed to the spot forming site on the target to be ejected, where the spot forming site is more lyophilic than at least the surrounding region of the spot forming site. A microarray manufacturing apparatus for forming spots by pouring, wherein the liquid is provided at a plurality of liquid storage chambers for storing a liquid, the top of which is open, and at the bottom of each of the plurality of liquid storage chambers A dispensing container comprising a plurality of discharge ports for discharging the liquid stored in the storage chamber to the outside, and a lid that covers all the openings of the plurality of liquid storage chambers to form one closed space; A pressurizing unit that pressurizes the closed space through one or a plurality of pressurization ports provided in the lid and forms droplets at the plurality of discharge ports; and a discharge port of the dispensing container. Between the tip and the target The dispensing container is placed in the substantially vertical direction so that the distance in the substantially vertical direction with respect to the ejection target surface is a predetermined gap that is equal to or less than the limit length in the dropping direction of the liquid mass that can be held in the discharge port. A detecting means for detecting that all the liquid masses formed at the plurality of discharge ports are in contact with the discharge target object, and the addition according to a detection signal sent from the detecting means. Control means for transmitting a stop signal to stop the pressurization to the pressure means, and transmitting a movement signal to the movement means so as to be separated from the discharge target object after a predetermined time after the stop signal transmission.

  According to this, it is detected that the liquid mass formed in all the discharge ports has contacted the object to be discharged, the pressurization is stopped, and the dispensing container is separated from the object to be discharged after a predetermined time thereafter. Therefore, the liquid amount can be determined by a balance between the force for pulling the liquid on the discharge port side and the force for pulling the liquid on the discharge target side without actively finely adjusting the liquid amount (by active control). Further, since one closed space is formed between the lid and the liquid storage chamber, the liquid stored in the plurality of liquid storage chambers can be collectively pressurized only by pressing the closed space. It becomes possible. Therefore, a highly accurate control device is not required, and the structure of the dispensing container can be simplified. Further, in this apparatus, since the liquid volume of the liquid mass formed at all the discharge ports is adjusted by pressurizing one closed space, the liquid mass formation (growth) speed is likely to vary among the respective discharge ports. However, since the liquid amount is finally adjusted by balancing the above forces, it is possible to reduce the variation in the size of the formed spots. Therefore, a droplet forming substrate (more specifically, a microarray) having no variation in spot size can be manufactured at a lower cost. In addition, the device grows the liquid mass held at the discharge port after the dispensing container is positioned, thus avoiding contamination caused by dropping of liquid droplets at unscheduled locations or timing during the movement of the dispensing container. It becomes possible to do.

  It is preferable that the spot forming part of the discharge target body is lyophilic, and at least the periphery of the spot forming part is lyophobic. According to this, since at least the periphery of the spot formation site is liquid repellent, it is possible to more reliably form droplets in the spot formation site. Therefore, it is possible to more reliably reduce spot size variation.

  It is preferable that the shape of the lid body has a substantially reverse concave shape in cross section. According to this, it becomes possible to provide a closed space for pressurization on the lid side by making the shape of the lid substantially reverse concave in cross section, and pressurization on the side of the dispensing container including a plurality of liquid storage chambers Therefore, the outer wall of the dispensing container body can be lowered, and the liquid can be easily supplied to the liquid storage chamber.

  Moreover, you may provide the partition plate which has the some hole which changed the hole diameter according to the distance from a pressurization opening so that the opening of a cover body with a substantially reverse concave shape may be plugged. According to this, it is possible to equalize the force applied to each liquid storage chamber.

  The apparatus further includes (second) moving means for moving the dispensing container in a substantially horizontal direction with respect to the surface to be ejected so that the dispensing container is on a spot formation site on the object to be ejected. Is preferred. According to this, the dispensing container can be moved three-dimensionally to a desired position on the discharge target surface.

  The droplet forming substrate includes a microarray formed by spotting and solidifying a sample liquid of a biological substance or chemical substance on a substrate such as glass.

  Hereinafter, a technique for producing a droplet forming substrate of the present invention will be described with reference to the drawings. 1 and 2 are process diagrams for explaining a method of manufacturing a droplet forming substrate according to the present invention. In the embodiments described below, the manufacture of a microarray, which is an example of a droplet forming substrate, is described as an example.

  In the embodiment of the present invention, as shown in FIGS. 1A to 1C, a dispensing container 10 having a plurality of discharge ports (nozzles) 14 formed on the lower surface is used. In the dispensing container 10, a liquid storage chamber 12 is formed for each discharge port 14, and the same type or different types of sample solutions are stored in each liquid storage chamber 12. The dispensing container 10 is hermetically sealed, and is configured so that the pressure (air pressure) in the internal closed space can be adjusted from the outside. A water repellent process is performed around each discharge port 14, and a liquid mass 26 is formed at each discharge port 14 by external pressure. A discharge target body (substrate) 20 is disposed below the dispensing container 10. A lyophilic process area around which liquid repellent treatment is applied is formed on the discharge target object (substrate) 20 corresponding to each discharge port. By disposing the dispensing container 10 and the discharge target object (substrate) 20 in advance at a predetermined distance corresponding to the drooping amount (growth amount) of the liquid mass, a predetermined amount of the sample solution is a plurality of the discharge target objects 20. Are moved almost simultaneously to the discharge target area.

  First, as shown to Fig.1 (a), the sample solution discharged to the dispensing container main body 10a of the dispensing container 10 (refer FIG.1 (b)) is inject | poured. The dispensing container main body 10a includes a plurality of liquid storage chambers 12 capable of storing a liquid, and each of the liquid storage chambers 12 has a bottom wall (bottom portion) for discharging the stored liquid to the outside. A discharge port (nozzle hole) 14 is provided. The outermost outer wall of the dispensing container body 10a is provided one step higher than the partition wall (partition wall) separating the liquid storage chambers 12, and the inner peripheral edge thereof is cut out to form a step so that a lid, which will be described later, can be fitted. Forming. The lower surface 16 (hereinafter also referred to as a discharge port forming surface) of the dispensing container main body 10a in which the discharge ports 14 are formed is subjected to liquid repellent treatment entirely or partially, and the outer periphery of each discharge port 14 has liquid repellent treatment. Is given. By this liquid repellent treatment, the liquid masses 26 of the sample solution formed at the respective discharge ports 14 are not connected (mixed).

  Although simplified in FIG. 1A, a liquid passage is appropriately formed between the liquid storage chamber 12 and the discharge port 14 by using a technique of an ink jet head of a so-called ink jet printer, and a sample solution It is possible to suppress the self-weight drop and to adjust the balance of the discharge pressure between the discharge ports described later. In addition, when injecting the sample solution into each liquid storage chamber 12, the bottom of the dispensing container body 10 a may be covered with a lower cap (not shown) to prevent the sample solution from leaking from the discharge port 14.

  The liquid repellent treatment agent used for the liquid repellent treatment of the discharge port forming surface 16 is not particularly limited. For example, a xylene-based resin or a fluorine-containing polymer is used. As the xylene-based resin, for example, parylene resins such as parylene C and parylene N are used. Parylene resin can be coated by, for example, chemical vapor deposition. As the fluorine-containing polymer, an amorphous fluorine-containing polymer is particularly preferable. Specifically, fluorine-containing polymers such as polydiperfluoroalkyl fumarate, Teflon AF (trademark of DuPont), Cytop (trademark of Asahi Glass Co., Ltd.), alternating co-polymerization of diperfluoroalkyl fumarate and styrene Alternating copolymers of fluorine-containing ethylene and hydrocarbon-based ethylene, such as an alternating copolymer, an alternating copolymer of ethylene trifluorinated chloride and vinyl ether, an alternating copolymer of tetrafluoroethylene chloride and vinyl ester, or the like Or a derivative or fumarite (trademark of NOF Corporation) is preferable. Since these amorphous fluorine-containing polymers are selectively dissolved in a fluorine-based organic solvent, they can be dissolved in a solvent at an arbitrary concentration and coated by dipping coating or spray coating.

  Although it does not specifically limit as a material which comprises the dispensing container main body 10a, For example, ionomer, polyethylene, a polypropylene, poly- (4-methylpentene-1), an ethylene-propylene copolymer, ethylene-acetic acid Polyolefin such as vinyl copolymer (EVA), cyclic polyolefin, modified polyolefin, polyvinyl chloride, polyvinylidene chloride, polystyrene, polyamide, polyimide, polyamideimide, polycarbonate, acrylic resin (eg, polymethyl methacrylate), acrylonitrile-butadiene -Styrene copolymer (ABS resin), acrylonitrile-styrene copolymer (AS resin), butadiene-styrene copolymer, polyoxymethylene, polyvinyl alcohol (PVA), ethylene-vinyl alcohol copolymer Body (EVOH), polyethylene terephthalate (PET), polybutylene terephthalate (PBT), polyester such as polycyclohexane terephthalate (PCT), polyether, polyetherketone (PEK), polyetheretherketone (PEEK), polyetherimide, Polyacetal (POM), polyphenylene oxide, modified polyphenylene oxide, polysulfone, polyethersulfone, polyphenylene sulfide, polyarylate, aromatic polyester (liquid crystal polymer), polytetrafluoroethylene, polyvinylidene fluoride, other fluorine resins, epoxy resins, Phenol resins, urea resins, melamine resins, unsaturated polyesters, silicone resins, polyurethanes, etc., or copolymers based on these, Rend body, various kinds of resin materials such as polymer alloys, various glass materials, various kinds of metal or metal oxide materials, and the like.

  The sample solution used in the examples is not particularly limited as long as it is a solution containing biological sample components used to create a microarray. The biological sample is not particularly limited. For example, in addition to biomolecules such as proteins and nucleic acids, artificially synthesized oligonucleotides, polynucleotides, oligopeptides, polypeptides, PNA (peptide nucleic acid) And the like. Usually, such a biological sample is used by being dispersed in water.

  Next, as shown in FIG.1 (b), the whole upper opening of the dispensing container main body 10a is covered with the cover body (upper cap) 18, and the dispensing container main body 10a is sealed. The upper part of each liquid storage chamber 12 is covered with one lid 18, and each liquid storage chamber 12 shares one closed space 22. The closed space 22 is connected to a pressure adjusting means provided outside, which will be described later, via the tube 19 and functions as a pressure chamber for the stored sample solution. The dispensing container main body 10a is covered with the lid 18 and the pressure chamber is operated to remove the lower cap described above.

  The dispensing container 10 is disposed such that the lower surface thereof faces the upper surface of the flat plate-like target object 20. The distance (the height of the dispensing container 10) D between the dispensing container 10 and the object 20 to be ejected is set to a value determined in advance corresponding to the amount of dispensing (application) of the sample solution. In addition, horizontal alignment between each discharge port 14 of the dispensing container 10 and each spot formation site (scheduled position for forming a spot) on the discharge target 20 is also performed.

  Here, the distance D between the discharge port forming surface (lower surface) 16 of the dispensing container 10 and the discharge target surface (upper surface) of the discharge target body 20 is the falling direction of the liquid mass that can be held in the discharge port 14. Is determined to be a predetermined gap within the maximum length (the limit size at which the liquid mass does not fall). Further, the size of the liquid mass formed by the gap is set by changing the predetermined gap to various values within the limit range. The liquid mass formed at the discharge port 14 grows when the liquid is pushed out by the pressure applied to the closed space 22, and touches the discharge target surface 20 a subjected to the lyophilic treatment of the discharge target body 20, thereby causing the discharge port 14 side. To the object 20 to be ejected. Therefore, the transfer (application) liquid amount at the time of spot formation can be adjusted by setting the gap D. As will be described later, this predetermined gap is preferably equal to the radius of the discharge port 14 (see FIG. 4).

  The lid (upper cap) 18 is detachable and has a substantially concave cross section (U-shape). The lid (upper cap) 18 is fitted into the dispensing container main body 10a and is in close contact with the dispensing container main body 10a. One closed space 22 including a plurality of liquid storage chambers 12 is formed. In the subsequent step of FIG. 1C, the closed space 22 is pressurized by a pump (not shown) as a pressurizing means (pressure adjusting means) continuously provided through the tube 19, thereby It becomes possible to press all the liquids stored in the liquid storage chamber 12 at once. In addition, it is possible to prevent leakage of the sample solution from the discharge port 14 by reducing the pressure of the closed space 22 with the pressurizing means.

  The material constituting the lid 18 is not particularly limited. For example, natural rubber, isoprene rubber, butadiene rubber, styrene-butadiene rubber, nitrile rubber, chloroprene rubber, butyl rubber, acrylic rubber, ethylene-propylene rubber, Elastic materials such as hydrin rubber, urethane rubber, silicone rubber, and fluorine rubber can be used. Thus, by comprising the cover body 18 with an elastic material, it becomes possible to closely_contact | adhere to the upper end part of the dispensing container main body 10a, and the airtightness of the closed space 22 can be improved.

  The lid 18 may be formed of a hard member instead of an elastic material. In this case, the lid 18 or the vicinity of the junction between the lid 18 of the dispensing container body 10a and the dispensing container body 10a. Further, by providing a member that enhances adhesion such as a seal ring, the confidentiality of the closed space 22 can be enhanced.

  The lid 18 and the tube 19 are integrally formed in the present embodiment, but may be separate.

  As shown in FIG. 3, the discharge target 20 used in the present invention is formed such that the lyophilicity of the spot forming portion 20 a is relatively higher than the surrounding portions. The spot forming portion 20a is subjected to lyophilic treatment, and a region other than the spot forming portion (hereinafter also referred to as non-spot forming region) 20b is subjected to liquid repellent treatment. In this way, the spot formation site 20a is subjected to lyophilic treatment, and the non-spot formation region 20b is subjected to liquid repellent treatment, whereby the droplet spot of the transferred (applied) sample solution is more reliably formed in a desired shape (larger size). )).

  The method of lyophilic treatment and lyophobic treatment is not particularly limited, and methods generally used in the technical field of the present invention can be used. For example, as a patterning method of the spot forming portion 20a and the non-spot forming region 20b, there are methods such as drug application by ink jet method, film formation by photolithography, treatment by laser irradiation, electron beam irradiation, and the like.

  In addition, when the target 20 itself is lyophilic or lyophobic, one of the processes can be omitted. That is, for example, when the discharge target 20 itself is lyophilic, the non-spot forming region 20b may be subjected to a liquid repellent treatment. Further, when the discharge target 20 itself is lyophobic, the lyophilic process may be performed on the spot forming portion 20a.

  Further, the spot forming portion 20a may be a concave portion or a convex portion, preferably a concave portion. This makes it possible to define the spot size more reliably.

  Although it does not specifically limit as a board | substrate material which comprises the to-be-discharged target body 20, For example, glass, a film | membrane, a synthetic resin (polyethylene, polypropylene, polystyrene etc.) etc. are mentioned.

  Next, as shown in FIG.1 (c), the closed space 22 is pressurized via the tube 19 with the pump as a pressurizing means which is not shown in figure. Thereby, a liquid mass 26 is formed at each discharge port 14.

  Although a pump is not specifically limited, For example, a syringe pump, a tube pump, etc. are used suitably.

  In this embodiment, since there is one pressurizing port 24 for pressurizing the closed space 22, the growth rate of the liquid mass 26 formed in each discharge port 14 is slightly increased according to the distance from the pressurizing port 24. There is a difference. That is, as the distance from the pressurizing port 24 increases, it takes time to transmit the pressurizing force, so that the growth rate of the liquid mass 26 (the start time of formation of the liquid mass) becomes slower.

  Next, as shown in FIG. 2 (d), the liquid mass 26 is maintained until the closed space 22 is continuously pressurized until all the liquid masses 26 formed at the respective ejection ports 14 come into contact with the discharge target body 20. Grow. When all the liquid masses 26 come into contact with the discharge target body 20, the pressurization of the closed space 22 is stopped. A detection method as described later is used to detect whether or not all the liquid masses 26 are in contact with the discharge target body 20. In addition, it is good also as continuing a pressurization state for the predetermined time which each liquid lump determined by experiment etc. replaced with the detection of contact for contact.

  Thereafter, when the liquid mass 26 is held between the discharge port 14 and the spot forming portion 20a for a while (for a while), as shown in FIG. 2 (e), the self-weight of the liquid mass 26 and the liquid mass 26 are obtained. The liquid mass 26 is pulled downward by a force or the like acting when the liquid spreads to the spot forming site 20a, and the movement of the liquid stops when the force pulling the liquid mass 26 downward and the force pulling upward are balanced. In this way, since the liquid amount is adjusted by the balance between the force that pulls the liquid mass 26 downward and the force that pulls it upward, the liquid amount on all the spot forming portions 20a finally becomes substantially uniform (temporarily elapses). rear).

  Thereafter, as shown in FIG. 2 (f), the dispensing container 10 is gently pulled away from the substrate 20, so that the liquid mass 26 is cut off at the discharge port forming surface (nozzle surface) 16, and on the discharge target body 20. A spot will be formed.

  Next, the principle (gap setting) of the present invention will be described. FIG. 4 is an explanatory diagram for explaining the principle of the present invention.

As shown in the figure, for example, when the liquid mass 26 exiting from the discharge port (nozzle) having a radius r is substantially hemispherical, the distance (liquid mass height h) at which the liquid mass 26 contacts the flat substrate is defined as the gap D. And Assuming that the contact angle θ between the liquid and the inner wall of the nozzle is θ = 180 °, θ1 = 90 °, so that 2 × θ2 = θ1 is established from the so-called contact angle measurement θ / 2 method, and θ2 = 45 °. Therefore, the liquid mass height h = nozzle radius r = gap D. When the liquid discharged from the nozzle is made into a hemisphere (θ = 180 °), the pressure ΔP is expressed by ΔP = 2γ _L / r. Here, γ _L is the surface tension of the liquid and the nozzle. The liquid amount at this time is determined by (4/3) πr ³ × (1/2) which is the volume of the hemisphere, and the liquid weight is calculated by the density of the liquid.

  The liquid mass 26 in contact with the lyophilic surface of the discharge target body 20 moves to the discharge target body 20 side, and the liquid mass 26 is separated along the lower surface 16 on which the liquid repellent treatment is performed. Since each nozzle and each spot forming part are formed under the same conditions, the force with which the nozzle pulls the liquid and the force with which the spot forming part pulls the liquid are the same for each nozzle, cut from the nozzle surface, and formed on the flat substrate. The amount of spot liquid is the same for each nozzle.

  Next, an example of a microarray manufacturing apparatus used in the present invention will be described. FIG. 5 is a schematic diagram showing the overall configuration of the microarray manufacturing apparatus of this embodiment, and FIG. 6 is a block diagram for explaining a control system of the microarray manufacturing apparatus shown in FIG.

  As shown in FIGS. 5 and 6, the microarray manufacturing apparatus 1 applies a liquid (sample) by pressurizing or depressurizing the dispensing container 10 and the closed space 22 (see FIG. 2) in the dispensing container 10. Pump 106 as a pressurizing means for pushing out (solution) and preventing liquid leakage, a positioning mechanism (corresponding to a moving means) for relative positioning of the dispensing container 10 and the discharge target object 20, detection of positioning, etc. A control unit 300 for controlling the positioning mechanism and the pump 106, a storage unit 500 for storing control programs and data, an input unit 600 for inputting commands to the apparatus and displaying information.

  The pump 106 described above is configured, for example, of a syringe type (using a piston and a cylinder) or a diaphragm type, and a minute pressure can be set.

  The detection unit 400 detects that the liquid mass 26 formed in all the discharge ports 14 has come into contact with the discharge target body 20, and transmits a detection signal to the pressurization control unit 302. A conventionally known detection mechanism is used for the detection unit. Specifically, for example, the growth of a liquid mass formed at the discharge port 14 that is considered to be the slowest or the slowest is observed by an optical imaging means such as a CCD (Charged Coupled Device) camera, and the obtained image is displayed on a computer. It can be detected by processing. Alternatively, a laser light source and a light receiving sensor may be used to detect the change in the received light intensity when the liquid mass 26 crosses the laser light source generated from the laser light source.

  The positioning mechanism includes an x-axis direction moving unit 204 that moves the dispensing container 10 in the x-axis direction, and a y-axis direction movement in which the discharge target 20 is placed on a moving work table (stage) 210 and moved in the y-axis direction. The unit 206 includes a z-axis direction moving unit 202 that moves the dispensing container 10 in the z-axis direction.

  The control unit 300 includes a control computer, and controls a pressurization control unit 302 that controls the pump 106 according to a control program, an x-axis direction movement control unit 306 that controls the x-axis direction movement unit 204, and a y-axis direction movement unit 206. A y-axis direction control unit 308, a z-axis direction control unit 304 that controls the z-axis direction movement unit 206, and a pressurization control unit 302 that controls the pump 106 are configured. The control unit 300 controls the overall operation of the apparatus. The z-axis direction moving unit 202 corresponds to the first moving unit, and the x-axis direction moving unit 204 and the y-axis direction moving unit 206 correspond to the second moving unit.

  In the x-axis direction moving unit 204, an x-axis direction driving motor (for example, a stepping motor) is connected to a carrier (not shown) that carries the dispensing container 10 via a driving belt. The dispensing container 10 is configured to be driven in accordance with the supplied driving signal. When the x-axis direction moving unit 204 is driven, the carrier 108 moves in the x-axis direction.

  The y-axis direction moving unit 206 is configured so that the moving work table 210 on which the ejection target 20 is placed can be moved in the y-axis direction by a feed screw system driven by a y-axis direction driving motor (for example, a stepping motor). Has been.

  The z-axis direction moving unit 202 moves in the z-axis direction a fixed bar 208 that fixes a carrier that transports the dispensing container 10 by a feed screw system driven by a z-axis direction drive motor (for example, a stepping motor). It is configured to be possible.

  The pressurization control unit 302 controls the amount of liquid discharged from the dispensing container 10 (size of the liquid mass 26). Specifically, for example, the pressurization control unit 302 transmits a drive signal to the pump 106 after the discharge port forming surface 16 has been moved and fixed by the z-axis direction moving unit 202 for a predetermined distance. Further, a stop signal is transmitted to the pump 106 so as to stop pressurization in response to a detection signal transmitted by the detection unit 400 described later.

  As described above, in the storage unit 500, for example, a table created by experimentally measuring or calculating the relationship between the spot size and the length in the liquid drop direction (corresponding to the distance D), or A database such as a table created by experimentally measuring or calculating the relationship between the spot size and the liquid extrusion amount is constructed.

  In the configuration as described above, the control unit 300 responds to the relative positioning information in the xy directions such as the spot formation start position on the discharge target body 20 and the size of the discharge target body 20, which are input by the input unit 600. Then, a drive signal is sent to the x-axis direction moving unit 204 and the y-axis direction moving unit 206, and the dispensing container 10 is horizontally moved onto the spot formation site on the discharge target body 20, and the dispensing container 10 and the discharge target The body 20 is opposed.

  Further, the control unit 300 sets various kinds of process parameters such as the type of sample solution, the sample supply amount, the type / number of the target object 20 to be ejected, the number of spots, the spot size, or the planned amount of liquid to be ejected, which are input from the input unit 600. Accordingly, referring to information such as the surface tension between the sample solution and the used dispensing container 10 and the nozzle radius of the used dispensing container 10 from the database stored in the storage unit 500, the dispensing container 10 and the discharge target A gap D to be set between the object 20 and the pressure ΔP to be set to the pump 106 is calculated.

  The control unit 300 sends a drive signal to the z-axis direction moving unit 202 via the z-axis direction control unit 304 so as to move the dispensing container 10 by a predetermined distance in the vertical direction, and the nozzles of the dispensing container 10 and the discharge target The separation distance from the body 20 is set to be the gap D.

  Further, the control unit 300 pressurizes the closed space 22 of the dispensing container 10 by the pump 106 via the pressurization control unit 302, and causes the liquid mass 26 to hang down. The detection unit 400 detects that the liquid mass (or the liquid mass of the sample) formed at all the ejection ports is in contact with the object to be ejected, stops the pressurization, and sets the static pressure state. Thereafter, when a predetermined time elapses until the total liquid mass of each nozzle moves to the ejection target object 20 side, or the detection unit 400 has moved the entire liquid mass (or the sample liquid mass) to the ejection target object. Is detected, the control unit 300 sends a movement signal to the z-axis direction moving unit 202 so as to move the dispensing container 10 away from the discharge target body 20, and the dispensing container 10 is moved to a predetermined height position (or the original height Return to position. When the dispensing container 10 is separated, the closed space 22 can be set to a negative pressure to prevent liquid leakage from the discharge port 14.

  Note that when the detection unit 400 is used, there is an advantage that the operation can be ensured and the discharge cycle can be shortened, but the detection unit 400 is not essential. The predetermined time may be set to a time sufficient for the movement of the liquid held between the discharge port 14 of the dispensing container 10 and the discharge target body 20 to end (the liquid amount does not change). .

  The control unit 300 controls the positioning mechanism to repeat the application process of the sample solution to the substrate, and repeats the application process to all the discharge target objects 20.

  According to the present embodiment, the amount of liquid applied to the discharge target object can be set by setting the gap between the dispensing container (nozzle thereof) and the discharge target object. It is easy to apply substantially the same amount of liquid to the discharge target object.

  Further, it detects that the liquid mass formed in all the discharge ports is in contact with the object to be discharged, stops the pressurization, and separates the dispensing container from the object to be discharged after a predetermined time elapses. It is possible to move the entire liquid mass to the discharge target object without fine adjustment individually.

  Moreover, it becomes possible to pressurize the liquids stored in the plurality of liquid storage chambers at once by forming a single closed space between the lid and the liquid storage chamber and pressing the closed space. . Therefore, a highly accurate control device is not required, and the structure of the dispensing container can be simplified.

Further, in the microarray manufacturing apparatus of the present embodiment, the amount of liquid pushed out from all the discharge ports is adjusted by pressurizing one closed space, so that there is some variation in the liquid mass formation speed between the discharge ports. However, since each spot forming portion of each nozzle and the target to be ejected is formed under the same conditions, the balance of the force acting on the liquid mass at each nozzle is the same condition and the separation of each liquid mass is performed. Thus, it becomes possible to reduce the variation in the size of the formed spot. Therefore, a microarray with no variation in spot size can be manufactured at a lower cost.
Further, in the apparatus, the liquid mass is formed at the discharge port of the dispensing container after positioning the droplet application with the target object, and the held liquid mass is moved to the target object side. As in the case of using the droplet, it is possible to avoid the contamination caused by the droplet dropping at an unscheduled place or timing during the movement of the dispensing container.

  The present invention has been specifically described by taking an embodiment as an example. However, the present invention is not limited to the above embodiment, and the design may be changed as appropriate without departing from the scope or spirit of the present invention. Can be done.

  For example, in the above embodiment, the lid body 18 having a substantially concave cross section is used. However, as shown in FIG. 7, a substantially plate-like lid body 18 may be used. Further, the pressure port 24 is not limited to one, and a plurality of (two or more) pressure ports 24 may be provided as shown in FIG. Further, as shown in the figure, a partition wall may be provided in the plate-like lid 18 and a plurality of closed spaces 22 may be formed for each pressure port 24. According to this, when there are a large number of liquid storage chambers, the force applied to the liquid stored in each liquid storage chamber can be more equalized. Further, the pressure propagation time can be made more uniform. Moreover, you may provide the partition plate which has the some hole which changed the hole diameter according to the distance from a pressurization opening so that the opening of a cover body with a substantially concave cross section may be plugged up. According to this, it is possible to equalize the force applied to each liquid storage chamber.

  Moreover, in the said embodiment, although the discharge port 14 used the micro hole provided in the bottom part of the liquid storage chamber 12, it is not limited to this, For example, you may use a (hollow) dispensing needle.

FIG. 1 is a process diagram for explaining the microarray manufacturing method of the present invention. FIG. 2 is a process diagram for explaining the microarray manufacturing method of the present invention. FIG. 3 is a diagram for explaining a pattern of spot formation sites and non-spot formation regions of the discharge target object. FIG. 4 is an explanatory diagram for explaining the principle of the present invention. FIG. 5 is a schematic diagram showing the overall configuration of the microarray manufacturing apparatus of the present embodiment. FIG. 6 is a block diagram for explaining an example of the microarray manufacturing apparatus of the present embodiment. FIG. 7 is a view showing another example of a dispensing container. FIG. 8 is a view showing another example of a dispensing container.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Microarray manufacturing apparatus, 10 dispensing container, 10a dispensing container main body, 12 liquid storage chamber, 14 discharge port, 16 discharge port formation surface, 18 cover body, 19 tube, 20 to-be-discharged object body, 20a spot formation site, 20b Non-spot formation region, 22 closed space, 24 pressure port, 26 liquid mass, 106 pump, 108 carrier, 202 z-axis direction moving part, 204 x-axis direction moving part, 206 y-axis direction moving part, 208 fixed bar, 210 Moving work table (stage), 302 Pressure control unit, 304 x-axis direction movement control unit, 306 y-axis direction movement control unit, 308 z-axis direction movement control unit, 400 detection unit, 500 storage unit, 600 input unit

Claims (9)

A process of vertically arranging a dispensing container having a plurality of discharge ports at the bottom and containing a liquid and a target object having a plurality of spot forming portions to be supplied with droplets;
The dispensing container and the discharge target are set so that a distance between the discharge port and the spot forming portion is a predetermined gap in a range not exceeding a holding limit length for holding a liquid mass formed in the discharge port. The process of setting the position relative to the body,
Pressurizing the liquid to form the liquid mass at the discharge port, and growing the liquid mass until the liquid mass contacts each of the spot forming portions of the discharge target body;
A process of holding the liquid mass for a while between the discharge port and the spot forming region,
Separating the dispensing container from the object to be ejected, and transferring the liquid mass onto the spot formation region to form the droplets;
A method of manufacturing a droplet forming substrate including:   The method for manufacturing a droplet forming substrate according to claim 1, wherein the spot forming portion of the discharge target is lyophilic and the periphery of the spot forming portion is lyophobic.   The manufacturing method of the droplet formation board | substrate of Claim 1 or 2 made the periphery of the said discharge outlet of the said dispensing container liquid-repellent. A dispensing container having a liquid storage chamber for storing a liquid and a plurality of discharge ports for discharging the liquid to the outside;
A discharge target to which the liquid is to be applied;
Moving means for setting a mutual position so that the dispensing container and the discharge target object are opposed to each other;
Pressurizing means for pressurizing the liquid storage chamber to form a liquid mass at the plurality of discharge ports;
A controller for controlling the moving means and the pressurizing means;
The control unit controls the moving means so that the dispensing container and the discharge target body are vertically opposed to each other, and a distance between the discharge port of the dispensing container and the discharge target object is held in the discharge port. The liquid volume is set between the plurality of discharge ports of the dispensing container and the target object by controlling the pressurizing means to set a predetermined gap in a range not exceeding the retention limit length of the liquid volume to be discharged. The droplet forming substrate manufacturing apparatus is characterized in that after the liquid is formed, the moving means is controlled to separate the dispensing container and the discharge target object, and the liquid mass is transferred to the discharge target object. . Furthermore, a detection unit that detects that one or a plurality of liquid masses formed in the plurality of discharge ports has come into contact with the discharge target object,
The control unit controls the moving unit after detecting the formation of a liquid mass between a plurality of discharge ports of the dispensing container and the discharge target body by the detection unit, and controls the dispensing container and the discharge target. The manufacturing apparatus of the droplet formation board | substrate of Claim 4 which spaces apart a body.   The droplet formation according to claim 4 or 5, wherein a spot forming portion to which the liquid to be discharged is applied is lyophilic and at least a surrounding portion of the spot forming portion is lyophobic. Board manufacturing equipment.   The apparatus for manufacturing a droplet forming substrate according to claim 4, wherein the periphery of the discharge port of the dispensing container is liquid repellent.   The controller, after detecting that the liquid mass has contacted the object to be ejected, and after passing a temporary time required for each liquid mass to be substantially the same amount, The apparatus for manufacturing a droplet forming substrate according to claim 5, wherein the apparatus separates the object.   The droplet formation according to any one of claims 4 to 8, wherein the dispensing container includes a plurality of liquid storage chambers having discharge ports, the liquid storage chambers are grouped, and the pressurization is performed for each group. Board manufacturing equipment.

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