JP2004077490A - Micropipette and dispensing device - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a micropipette forming minute spots with high precision and rapidly and a productive dispensing apparatus forming minute spots by efficiently dispensing a few hundred to a few tens of thousand different samples at once using the micropipette. <P>SOLUTION: The micropipette comprises at least one base 10 in which an inlet 16 for injecting a sample from outside, a cavity 15 into which the sample is injected and which is filled therewith, and an outlet 12 for discharging the sample are formed. The base 10 forming the cavity 15 is made of ceramics. At least one wall of the base 10 is provided with a piezoelectric/electrostriction element 22. The sample moves in the cavity 15 in the form of a laminar flow. Through the actuation of the piezoelectric/electrostriction element 22, the volume in the cavity 15 is changed, and a fixed amount of the sample in the cavity 15 is discharged out of the outlet 12. <P>COPYRIGHT: (C)2004,JPO

Description

The present invention relates to a micropipette excellent in volume controllability of droplets and product productivity, which is suitably used for aligning and fixing small volumes of liquid droplets at high density, such as in the production of DNA chips, and a micropipette using the same. Note on the device.

In recent years, there has been remarkable progress in analyzing gene structures, and many gene structures including human genes have been revealed. For such gene structure analysis, a DNA chip in which thousands to 10,000 or more different types of DNA fragments are aligned and fixed as minute spots on a substrate such as a microscope slide glass has been used. .

As a method of forming minute spots in the production of this DNA chip, a QUIILL method, a pin & ring method, or a spring pin method is widely used, and even if any method is adopted, each minute spot is formed. It is important to keep the capacitance and shape variation low and keep the distance between each minute spot constant. On the other hand, there is great expectation for the development of a new method with good shape controllability of minute spots and excellent productivity for further densification.

Here, the QUIILL method is a method in which a sample is stored in a concave portion formed at a pin tip, and the pin tip is brought into contact with a substrate to transfer a sample in the concave portion onto the substrate to form a minute spot. However, there are problems such as durability or the like being deformed or damaged by contact with the substrate, and problems such as incomplete cleaning of the sample stored in the concave portion and cross contamination.

In the pin-and-ring method, after the sample solution in the microplate is reserved by the ring, the solution penetrates the inside of the reserved ring, captures the sample in the ring with the pin tip, and forms a spot on the substrate However, the number of rings that can be reserved at one time depends on the number of rings. Conventionally, the number of rings is about several, so it is necessary to form minute spots of thousands to tens of thousands of samples. Requires several hundred to several thousand washing and drying steps, and therefore the productivity is not necessarily high.

The spring pin method is a method in which a sample attached to a pin tip is transferred onto the substrate by pressing the pin tip onto the substrate to form a minute spot. Although this method softens the damage to the substrate and blows out the sample, basically, only one spotting can be performed with one reserve, which is inferior in productivity. Furthermore, these conventional methods for forming minute spots all transport the sample solution onto the substrate while exposing the sample solution to the atmosphere, so that the sample dries during transport and cannot be spotted, resulting in a very expensive sample. There is a problem that the use efficiency of the solution is poor. On the other hand, there is a study on spotting using a so-called ink jet method that has been put to practical use in printers.However, forming thousands to tens of thousands of samples in individual flow paths is a problem in terms of size and cost. In addition, in the ink jet method, it is necessary to previously fill the pump without bubbles before spotting, so that a large amount of a sample for purging is required and the use efficiency of the sample is extremely poor. In general, it is better for the liquid to move at a high speed in the flow path including the pump chamber to remove bubbles. Therefore, when the sample is stirred in the flow path, for example, when a delicate DNA solution is used as the sample. In some cases, DNA was damaged.

As a micropipette for forming a minute spot of a sample, for example, a micropipette using an inkjet method (see Patent Document 1) and a device that discharges a biological sample (see Patent Document 2) are disclosed.
JP-A-6-40030 JP-A-8-233710

However, the micropipette disclosed in Patent Literature 1 and Patent Literature 2 cannot always sufficiently satisfy the precision, speed, and efficiency in forming minute spots. The present invention has been made in view of the above-described problems of the related art, and an object of the present invention is to use a micropipette that enables high-precision and high-speed formation of minute spots and this micropipette. It is an object of the present invention to provide a dispensing apparatus excellent in productivity capable of efficiently dispensing hundreds to tens of thousands of different samples at one time to form minute spots.

That is, according to the present invention, an inlet for injecting a sample from the outside, a cavity into which the sample is injected and filled, and an outlet for discharging the sample are formed in at least one or more substrates. A micropipette, wherein the substrate forming the cavity is made of ceramics, a piezoelectric / electrostrictive element is provided on at least one wall surface of the substrate, and the sample is configured to move in a laminar flow in the cavity; A micropipette is provided in which the volume in the cavity is changed by driving the piezoelectric / electrostrictive element, and a certain amount of sample in the cavity is discharged from the discharge port.

In the micropipette of the present invention, by adopting such a configuration, a minute amount of liquid is discharged from the discharge port corresponding to each drive of the piezoelectric / electrostrictive element, and the volume thereof is small and constant without variation. By using the piezoelectric / electrostrictive element, the driving cycle can be adapted to a high frequency, and the time required for ejection can be shortened. In addition, since the sample moves in the closed space until the ejection after the sample injection, the sample does not dry on the way. Furthermore, since the entire substrate can be formed small and compact, the flow path through which the sample moves can be shortened, and the deterioration of the use efficiency due to the sample adhering to the flow path wall can be reduced.

In the micropipette of the present invention, the cavity is previously filled with a replacement solution such as a buffer solution or a physiological saline solution, and then the sample is injected from the injection port into the cavity while performing laminar flow replacement. It is preferable to drive the element to discharge the sample in the cavity from the discharge port. The end point of the laminar flow replacement completion may be controlled in advance by determining the moving speed and volume of the sample and the replacement time, but it is more preferable to grasp the end point by detecting a change in the fluid property in the cavity. . The sample may be laminar-flow-replaced into the cavity from the injection port while driving the piezoelectric / electrostrictive element. After reliably replacing the inside of the cavity with an inexpensive replacement solution in advance, laminar flow replacement of the expensive sample can completely prevent the occurrence of ejection failure and efficiently discharge the expensive sample. Further, in the micropipette of the present invention, the cavity is previously filled with a replacement solution such as a buffer solution or physiological saline, and then the sample is injected while being replaced from the injection port into the cavity, and the end point of the completion of replacement is determined. After detecting the change in the fluid characteristics in the cavity, it is preferable to drive the piezoelectric / electrostrictive element to discharge the sample in the cavity from the discharge port. By grasping the completion of replacement by detecting a change in the fluid characteristics in the cavity, even if the sample and the replacement liquid mix slightly in the flow path, it is possible to distinguish between the mixed part and the unmixed part. Since it can be easily and accurately determined, the amount of the sample that must be mixed with the replacement liquid and purged can be reduced, and the usage efficiency of the sample can be increased.

Further, it is preferable that the change in the fluid property in the cavity is grasped by applying a voltage for exciting vibration to the piezoelectric / electrostrictive element and detecting a change in an electric constant accompanying the vibration. By doing so, there is no need to install a special detection element or the like, and low-cost, high-precision detection can be performed.

In the micropipette of the present invention, a sample inlet, a cavity, a sample outlet, and a plurality of piezoelectric / electrostrictive elements are formed in a single substrate, respectively. A plurality of units each having one sample injection port, cavity, sample discharge port, and the piezoelectric / electrostrictive element are fixed to a fixing jig in a substrate. And at least two types of portions, namely, a combination of a piezoelectric / electrostrictive element, and a sample injection port and a sample discharge port, are formed separately on at least two types of bases, and are joined to each other. At least one cavity and a piezoelectric / electrostrictive element are formed in each of the substrates, and at least one of the substrates is formed with at least one of a sample injection port and a sample discharge port. Is formed joining the units to one base, it is preferred that more than one of the units is integrally fixed.

Since each part is formed in a plurality of places in one substrate, the whole body is compact, the discharge ports can be arranged with high precision and high density, and plural kinds of samples can be discharged simultaneously. it can. In addition, a structure in which a plurality of units each of which is formed in one body is fixed in one body to form the entire body as a whole facilitates the manufacture of each body and improves the yield. Furthermore, by joining at least two or more substrates formed with each part to form the whole, the range of material selection of the substrate is expanded, and it becomes possible to select the most suitable material for each part, while the element It is possible to improve the yield, achieve high-precision and high-density arrangement of discharge ports, and simultaneously discharge a plurality of types of samples.

Further, the base is a flat plate, and the discharge port for the sample is formed on the side surface or the main plane of the base, or the base is flat and the discharge port for the sample is formed on one main plane of the base. It is preferable that the sample inlet is formed on the other main plane. By forming the substrate in a flat plate shape, the substrate can be manufactured by laminating green sheets and the like as described later, and the whole becomes thin and compact. If the discharge port is formed on the main plane of the base, the substrate can be set in parallel with the flat plate on which the discharge port is formed, and the discharge distance of the droplet can be easily made constant. The shape becomes stable. In addition, when the discharge ports are formed on the side surfaces of the base, the plate-like bases are vertically arranged, so that the density of the discharge ports can be easily increased. Further, by forming the injection port and the discharge port on different main planes of the substrate, the length of the flow path from the injection port to the discharge port is almost the same as the thickness of the flat plate, and the flow path of the sample liquid is reduced. This has the advantages of being short and simple, reducing problems such as bubbles being caught in the middle of the flow path and causing ejection failure, and improving the sample use efficiency.

Furthermore, a mode in which at least two or more sample inlets are connected to one cavity may be employed. With this configuration, the cavity can be filled with certainty by adjusting the timing of the sample or the replacement liquid from the plurality of injection ports, and sucking and pushing out.

In the micropipette of the present invention, the base on which the cavity and the piezoelectric / electrostrictive element are formed is preferably made of zirconia ceramics, or all the bases are preferably made of zirconia ceramics. It is preferable that it is produced using a sheet lamination firing method. Zirconia, especially stabilized zirconia and partially stabilized zirconia, have high mechanical strength even in the form of a thin plate, high toughness, durability in acid / alkali solutions, and reactivity with piezoelectric films and electrode materials. Suitable for small size. The base on which at least one of the injection port and the discharge port is formed may be made of a metal or resin excellent in its moldability and cost.

Note that the piezoelectric / electrostrictive film in the piezoelectric / electrostrictive element has an electromechanical mechanical coupling coefficient and a piezoelectric constant, which are mainly composed of components composed of lead zirconate, lead titanate, and lead magnesium niobate. This is preferred because the reactivity with the substrate (zirconia ceramics) during sintering of the film is small and a stable composition can be obtained.

According to the present invention, an injection port for injecting a sample from outside, a cavity filled with the sample, and a discharge port for discharging the sample are formed in one or more substrates. A dispensing apparatus using a plurality of micropipettes, wherein the substrate forming ceramics is formed of ceramics, a piezoelectric / electrostrictive element is provided on at least one wall surface of the substrate, and the sample is moved in a laminar flow in the cavity. Wherein the discharge ports are arranged vertically and horizontally, and different types of liquid samples are respectively discharged from the discharge ports. Furthermore, according to the present invention, an inlet for injecting a sample from the outside, a cavity into which the sample is injected and filled, and an outlet for discharging the sample are formed in at least one or more substrates. The substrate forming the cavity is made of ceramics, a piezoelectric / electrostrictive element is provided on at least one wall surface of the substrate, a replacement liquid is filled in the cavity in advance, and then a sample is injected from the injection port into the cavity. After the completion of the replacement of the sample in the cavity is detected by detecting the change in the fluid property in the cavity, the volume in the cavity is changed by driving the piezoelectric / electrostrictive element. A dispenser using a plurality of micropipette for discharging a certain amount of sample in the cavity from the discharge port, wherein the discharge port is Are aligned horizontally, the dispensing device is provided, characterized in that different kinds of liquid samples from the discharge port is discharged. According to these dispensing apparatuses, by using a plurality of micropipettes, many types of samples can be simultaneously supplied at a time, and a pipette partially defective can be easily replaced. Further, since the ejection ports are arranged vertically and horizontally, it is preferably used when a minute spot which is two-dimensionally arranged and fixed, such as a DNA chip, is required.

In this dispensing device, in order to increase the use efficiency of the sample, a cartridge filled with different types of liquid samples is attached to each of the sample inlets, and a mechanism for discharging different liquid samples from the outlet is provided. Preferably, an aqueous solvent or an organic solvent is provided at each of the sample injection ports in order to discharge thousands to tens of thousands of types of DNA fragments and the like into fine spots without contamination and with high purity. It is preferable to provide a mechanism for attaching the filled cartridge and cleaning a space from the inlet to the outlet formed in the base. Further, in this dispensing apparatus, it is preferable that an anisotropic flying-drop shielding plate made of a thin plate having a hole having the same central axis as the discharge port is provided outside the discharge port. By doing so, even if the ejection direction of the ejected droplets is bent, the droplets do not reach the substrate, thereby preventing a spotting position shift and a defect of mixing with an adjacent spot.

As described above, according to the micropipette of the present invention, a minute spot can be formed with high accuracy and at high speed. According to the dispensing apparatus using this micropipette, it is possible to efficiently dispense hundreds to tens of thousands of different samples at one time to form minute spots, thereby dramatically improving productivity. I do.

A basic structure of a micropipette according to the present invention is that at least one or more substrates are provided with a sample inlet, a cavity filled with the sample, and a sample outlet, and at least one of the bases forming the cavity is formed. It has a piezoelectric element on the wall. The micropipette is preferably configured so that the sample moves in a laminar flow in the cavity. The micropipette thus configured changes the volume in the cavity by driving the piezoelectric / electrostrictive element, and discharges a certain amount of the sample in the cavity from the discharge port, thereby forming a minute spot such as a DNA chip. It can be efficiently formed with high accuracy and at high speed.

Hereinafter, the present invention will be described in detail with reference to examples shown in the drawings, but the present invention is not limited to these examples. FIG. 2 shows an example of the micropipette of the present invention. In FIG. 2, the nozzle portion 11 is formed by forming a thin plate-shaped nozzle plate 13 provided with a discharge port 12 having at least one or more nozzle holes with a green sheet of zirconia ceramics, while the pump portion 21 has at least A spacer plate 25 on which one or more windows 28 are formed, and a closing plate 23 that is overlaid on one side of the spacer plate 25 and covers the windows 28 are also formed of green sheets of zirconia ceramic, respectively. The entire body is laminated and integrally fired to form the base 10. The closing plate 23 is provided with a sample injection port 16, which is connected to the introduction hole 14 connected to the window 28 formed in the spacer plate 25 and the communication path 17. On the outer surface of the closing plate 23, a piezoelectric / electrostrictive element 22 including a lower electrode 31, a piezoelectric / electrostrictive layer 32, and an upper electrode 33 is formed.

According to the micropipette having the above configuration, when an electric field is generated between the upper electrode 33 and the lower electrode 31, the piezoelectric / electrostrictive layer 32 is deformed, and the cavity formed by covering the window 28 is formed. As the volume of the pressurizing chamber (15) decreases, the sample (liquid containing DNA fragments) filled in the cavity (15) is discharged at a predetermined speed from the discharge port (12) communicating with the cavity (15). A DNA chip or the like which is aligned and fixed as a minute spot on a substrate can be manufactured. A device structure of a so-called ink jet system as shown in FIG. 2 is described in, for example, JP-A-6-40030, which can be referred to.

In the micropipette having the above-described configuration, the shape and the flow path dimensions are such that a liquid sample containing a DNA fragment or the like moves in a laminar flow in the cavity (pressurizing chamber) 15.

An example of a specific cavity will be described with reference to FIG. The cavity 3 has a long shape as shown in FIG. 1 and has an inlet 1 or an inlet 4 for introducing a sample at one end, and an outlet 2 at the other end. With such a shape, the cavity 3 is formed as a part of a flow path from the inlet 1 to the outlet 2, from the inlet 1 or from the inlet 1 through the communication path 5 and the inlet 4 to the inside of the cavity 3. Can be guided to the discharge port 2 without disturbing the flow of the sample moving to the discharge port 2. The specific dimensions of the cavity 3 vary depending on the type of the sample, the size of the droplet to be prepared, and the formation density. For example, a DNA fragment having a molecular number of about 1 to 10000 at a concentration of 1 μg / μl in a × 3 SSC buffer ( A sample dispersed in an aqueous solution of 0.45 M sodium chloride (0.045 M sodium citrate (pH 7.0)) and a microchip for manufacturing a DNA chip or the like that requires spotting with a droplet diameter of several hundred microns at a pitch of several hundred microns. In the case of a pipette, the cavity length (L) is preferably 1 to 5 mm, the cavity width (W) is preferably 0.1 to 1 mm, and the cavity depth (D) is preferably 0.1 to 0.5 mm. The inner wall of the cavity is preferably smooth so that there are no projections that disturb the flow, and the material is preferably made of ceramics having a good affinity for the sample solution.

In the micropipette of the present invention, preferably, the cavity is previously filled with a replacement solution such as a buffer solution or physiological saline, and then the sample is injected from the injection port into the cavity while undergoing laminar flow replacement. / Driving the electrostrictive element. In this case, it is preferable to grasp the completion of the laminar flow replacement of the sample in the cavity by detecting a change in the fluid characteristic in the cavity. The replacement of the sample with the replacement liquid in the cavity is preferably performed by laminar flow.However, when the type of the sample is changed, the liquid moving speed is extremely high, or in the case where the sample is in the cavity near the introduction hole, The flow need not necessarily be laminar. In this case, the purge amount of the sample increases due to the mixing of the sample and the replacement liquid. However, the increase in the purge amount can be minimized by judging the completion of the replacement by detecting a change in the fluid characteristic in the cavity. Here, a change in the fluid characteristic in the cavity is grasped by applying a voltage for exciting vibration to the piezoelectric / electrostrictive element and detecting a change in an electrical constant accompanying the vibration. The detection of such a change in the fluid characteristic is described in, for example, Japanese Patent Application Laid-Open No. 8-201265, and the contents thereof can be referred to.

Specifically, for a given piezoelectric / electrostrictive element, at predetermined intervals, an electrical connection from a power supply for ejection drive is disconnected by a relay, and at the same time, means for measuring a resonance frequency is connected by a relay, The impedance or resonance frequency at that time is electrically measured. This makes it possible to grasp whether the viscosity, specific gravity, etc. of the liquid are the target sample (liquid containing DNA fragments and the like). That is, according to the micropipette of the present invention, since the micropipette itself functions as a sensor, the configuration of the micropipette can be simplified.

Next, in the micropipette of the present invention, a replacement solution such as a buffer solution or physiological saline is injected into the cavity from the injection port while discharging the sample, and similarly, the sample remaining in the cavity by laminar flow replacement is removed. Discharge completely and be ready for the next sample injection. In this case, whether or not the sample remains in the cavity (whether or not the sample can be ejected) can also be detected by detecting a change in the fluid characteristic in the cavity. As described above, when the micropipette of the present invention is used, the purge amount of the sample not used by the laminar flow replacement or replacement completion detecting mechanism can be extremely reduced, and the use efficiency of the sample can be improved.

3 (a) (b) to 9 (a) and 9 (b) show other examples of the micropipette of the present invention. 3 (a) and 3 (b), a plurality of sample injection ports 16, cavities 15, sample discharge ports 12, and piezoelectric / electrostrictive elements 22 are formed in one substrate 40, respectively. The upper electrode 33 and the lower electrode 31 of the piezoelectric / electrostrictive element 22 are drawn out at once. In this case, different types of samples can be simultaneously discharged, and a DNA chip or the like can be efficiently manufactured with high productivity, which is preferable.

The micropipette shown in FIGS. 4A and 4B is a unit in which one sample injection port 16, one cavity 15, one sample discharge port 12, and one piezoelectric / electrostrictive element 22 are formed in one substrate. 4 (c) and 4 (d) show an embodiment in which a plurality of fixing jigs 35 (collectively referred to as holding jigs 18, positioning pins 19 and fixing plates 20 described later) are fixed. Each unit is fixed to a fixed plate 20 by a holding jig 18 for holding a tube (communication passage) 17 for supplying a sample to a sample inlet 16 and a positioning pin 19. 4 (a) and 4 (b), the fixing is performed by pressing both ends of the jig 18 to the fixing plate 20 with the screws 35A. Or an adhesive or the like.

In FIGS. 3A and 3B and FIGS. 4A to 4D, the base 40 on which the sample inlet 16, the cavity 15, and the sample outlet 12 are formed is made of ceramics. Stabilized zirconia, partially stabilized zirconia, alumina, magnesia, silicon nitride, or the like can be used. Among them, stabilized / partially stabilized zirconia is most preferably employed because of its high mechanical strength, high toughness, and low reactivity with piezoelectric films and electrode materials even in thin plates. When using stabilized / partially stabilized zirconia as a material for the base 40 or the like, at least a portion where the piezoelectric / electrostrictive element 22 is formed contains an additive such as alumina or titania. Is preferred. The piezoelectric / electrostrictive layer of the piezoelectric / electrostrictive element 22 is made of piezoelectric ceramics such as lead zirconate, lead titanate, lead magnesium niobate, lead magnesium tantalate, nickel nickel niobate, lead zinc niobate, Composite manganese containing lead manganese niobate, lead antimony stannate, lead manganese tungstate, lead cobalt niobate, barium titanate and the like or a component obtained by combining any of these can be used. A material mainly containing a component composed of lead zirconate, lead titanate and lead magnesium niobate is preferably used. This is because such a material has a high electromechanical coupling coefficient and a piezoelectric constant, and has low reactivity with the sensor substrate material during sintering of the piezoelectric film, so that a material having a predetermined composition can be stably formed. Based on what you can do.

Further, the above piezoelectric ceramics, lanthanum, calcium, strontium, molybdenum, tungsten, barium, niobium, zinc, nickel, manganese, cerium, cadmium, chromium, cobalt, antimony, iron, yttrium, tantalum, lithium, bismuth, tin, etc. A ceramic to which an oxide, a combination of any of these, or another compound is appropriately added may be used. For example, it is also preferable to use ceramics containing lead zirconate, lead titanate and lead magnesium niobate as main components, and containing lanthanum and strontium.

On the other hand, the upper electrode and the lower electrode in the piezoelectric / electrostrictive element are preferably made of a conductive metal which is solid at room temperature and includes, for example, aluminum, titanium, chromium, iron, cobalt, nickel, copper, Single metals such as zinc, niobium, molybdenum, ruthenium, palladium, rhodium, silver, tin, tantalum, tungsten, iridium, platinum, gold, and lead, or alloys combining any of these are used. Alternatively, a cermet material in which the same material as the detection plate is dispersed may be used. These bases, piezoelectric / electrostrictive elements, and electrode materials are commonly used in the present invention.

FIGS. 5A and 5B show a base body 40 in which one cavity 15, one piezoelectric / electrostrictive element 22, and one introduction hole 14 are formed, and one injection port 16 and one communication path 17 in two places. This is an embodiment of a micropipette in which a base 39 having a plurality of discharge ports 12 formed separately from a base 39 formed and then joined together by an adhesive 34 to be integrated. The base 40 is made of partially stabilized zirconia, the base 39 is made of stainless steel, and the base 38 is made of polyimide resin. The joining with each other may be performed mechanically, but joining with an adhesive or heat diffusion is preferable from the viewpoint of maintaining the sealing property of the flow path. The adhesive to be used is appropriately selected depending on the material of the substrate, the combination of the coefficient of thermal expansion, etc., and the sample solution resistance, but is based on vinyl, acrylic, phenol, polyamide, resorcinol, urea, melanin, Polyester-based, epoxy-based, furan-based, polyurethane-based, silicone-based, rubber-based, polyimide-based, and polyolefin-based adhesives are suitable.Especially, from the viewpoint of adhesive strength and durability, epoxy-based and polyimide-based adhesives are preferred. is there. Further, each of the adhesives may be a mixture of minute beads such as glass to keep the thickness of the adhesive constant.

FIGS. 6 (a) and 6 (b) show still another example of the micropipette of the present invention. This micropipette is what is called an edge type. , The cavity 15, the sample discharge port 12, and the piezoelectric / electrostrictive element 22 are formed at a plurality of locations, respectively. In this micropipette, the sample outlet 12 is formed on the side surface of the base 40, and the sample injected from the ordinary pipette 45 into the sample inlet 16 passes through the communication passage 17 in the base 40 and the cavity 15. The volume inside the cavity 15 is changed by driving the piezoelectric / electrostrictive element 22, and a certain amount of the sample filled in the cavity 15 is discharged from the discharge port 12.

FIGS. 7 (a) and 7 (b) show still another example of the micropipette of the present invention, which is the same as FIGS. 3 (a) (b) to 5 (a) (b). 6 (a) and 6 (b), a sample injection port 16, a cavity 15, a sample discharge port 12, and a piezoelectric / electrostrictive element 22 are provided in a single base 40 as in FIGS. Are formed at a plurality of locations, respectively. In this micropipette, the sample outlet 12 is formed on the main plane of the base 40. The cavity 15 and the sample inlet 16 are connected by an introduction hole 14 and a communication path 17.

FIGS. 8A and 8B show that the base 40 is flat, the sample outlet 12 is formed on one main plane of the base, and the sample inlet 16 is formed on the other main plane. This is an example. The piezoelectric / electrostrictive element 22 is formed in the same main plane as the discharge port. FIGS. 9A and 9B show an embodiment in which two sample injection ports 16 are connected to one cavity 15. The piezoelectric / electrostrictive element 22 is formed in the same main plane as the sample inlet 16, and the sample outlet 12 is formed in the other main plane.

Next, a dispensing apparatus using the above-described micropipette will be described. FIG. 10 shows an example of the dispensing device 55. The dispensing device 55 shown in FIG. 10 includes a plurality of micropipettes 50 (50a, 50b, 50c) each having a sample inlet 52 and a sample outlet 51 shown in FIGS. It is configured to stand upright. That is, each of the micropipette 50a, 50b, 50c has the sample injection port 52a, 52b, 52c on the upper side, the sample discharge port 51a, 51b, 51c on the lower side, and the sample discharge port 51a, 51b, 51c. Liquid samples of different types are discharged from the sample discharge ports 51a, 51b, and 51c, respectively, which are arranged vertically and horizontally. Below the sample outlets 51a, 51b, and 51c, an anisotropic flight shielding plate 53 made of a thin plate having a hole having the same central axis as the sample outlet is provided.

In the dispensing device 55 having such a configuration, as shown in FIG. 12, cartridges 60 filled with liquid samples of different types are attached to the sample inlets 52a, 52b, and 52c, respectively. It is preferable that a mechanism for discharging different liquid samples from the discharge ports 51a, 51b, 51c be provided in that the sample can be discharged efficiently. Further, it is difficult to attach a cartridge filled with a physiological saline solution or an organic solvent to each of the sample inlets, and to provide a mechanism for washing a space from the inlet to the outlet formed in the substrate for several thousand times. It is desirable to discharge various types of DNA fragments from as small as tens to tens of thousands to fine spots without contamination and with high purity. In addition, the method of injecting a sample or the like from the cartridge into each of the sample injection ports is such that, after setting the cartridge in the injection port, the bottom of the cartridge is opened with a needle or the like, or a needle or the like is formed near the injection port in advance. Alternatively, it may be opened simultaneously with the setting. Note that a mechanism may be added in which gas or the like is pressure-fed after opening and the sample or the like is forcibly pushed out.

Next, an example of a method for manufacturing a DNA chip using the dispensing device 55 according to the present invention will be described. After setting a cartridge containing a buffer solution as a replacement solution in advance, fill the cavity in each micropipette with the buffer solution, further set a cartridge containing a DNA fragment sample at an inlet, and use a needle or the like to set the bottom of the cartridge with a needle or the like. Is opened, and the sample is injected into the injection port. Thereafter, the piezoelectric / electrostrictive element is driven to discharge laminar flow of the sample in the cavity while discharging the prefilled buffer solution from the discharge port.

The end point of the replacement is detected by switching the relay so that the piezoelectric / electrostrictive element acts as a sensor for detecting the viscosity and specific gravity of the liquid in the cavity. After the replacement, the DNA chip is manufactured by driving the piezoelectric / electrostrictive element under the driving conditions corresponding to the amount of droplet corresponding to the required spot diameter and repeating spotting. Usually, one to several hundred drops are discharged from a micropipette to form one spot. When the amount of the sample in the injection port is reduced, the sample can be used up without remaining in the micropipette by adding a buffer solution to prevent air bubbles from entering the flow path and continuing ejection. Completion of the replacement of the sample with the replacement liquid (end of discharge of the sample) is similarly performed by detecting the viscosity and specific gravity of the liquid using a piezoelectric / electrostrictive element. It is also preferable to dry the solvent while forming microdroplets on the substrate using a sample solution whose concentration has been reduced in advance. By performing such a method, the amount of the sample remaining in the flow channel can be further reduced, and the use efficiency of the sample is improved. Further, it is preferable to use the replacement liquid to be used, and the sample itself, from which the dissolved gas in the solution has been removed through a deaeration operation in advance. By using such a solution, when filling the flow channel with the solution, even if bubbles are caught in the middle of the flow channel and the filling is incomplete, the bubbles can be dissolved in the solution to avoid problems, and the discharge can be prevented. It is also possible to prevent bubbles from being generated in the fluid in the middle of the process and causing a discharge failure.

INDUSTRIAL APPLICABILITY The micropipette and dispensing method of the present invention are effectively used in the fields of research, drug discovery, diagnosis, and medical care, for example, for analyzing gene structure, detecting gene expression, and studying gene function.

It is explanatory drawing which shows an example of a cavity. FIG. 2 is a cross-sectional view illustrating an example of the micropipette of the present invention. FIGS. 3A and 3B show another example of the micropipette of the present invention, wherein FIG. 3A is a plan view and FIG. 4 (a) is a plan view, FIG. 4 (b) is a side view, FIG. 4 (c) is an enlarged plan view of each unit, and FIG. 4 (d) shows another example of the micropipette of the present invention. FIG. 4 is a sectional view of FIG. FIGS. 5A and 5B show still another example of the micropipette of the present invention. FIG. 5A is a plan view, and FIG. FIGS. 6A and 6B show still another example of the micropipette of the present invention. FIG. 6A is a plan view, and FIG. FIGS. 7A and 7B show still another example of the micropipette of the present invention, wherein FIG. 7A is a plan view and FIG. 7B is a cross-sectional view taken along the line DD of FIG. 7A. FIGS. 8A and 8B show still another example of the micropipette of the present invention, in which FIG. 8A is a plan view and FIG. FIGS. 9A and 9B show still another example of the micropipette of the present invention, in which FIG. 9A is a plan view and FIG. 9B is a sectional view taken along line FF of FIG. 9A. It is a perspective view showing an example of a dispensing device. 11A and 11B show a micropipette used in the dispensing apparatus of FIG. 10, FIG. 11A is a plan view, and FIG. 11B is a GG cross-sectional view of FIG. It is a perspective view showing the situation where a cartridge is attached to a pipetting device.

Explanation of reference numerals

DESCRIPTION OF SYMBOLS 1 ... Injection port, 2 ... Discharge port, 3 ... Cavity, 4 ... Inlet, 5 ... Communication path, 10 ... Base, 11 ... Nozzle part, 12 ... Discharge port, 13 ... Nozzle plate, 14 ... Introduction hole, 15 ... Cavity, 16: sample inlet, 17: communication path, 18: holding jig, 19: positioning pin, 20: fixing plate, 21: pump unit, 22: piezoelectric / electrostrictive element, 23: closing plate, 25: spacer Plate, 28: Window, 31: Lower electrode, 32: Piezoelectric / electrostrictive layer, 33: Upper electrode, 34: Adhesive, 35: Fixing jig, 38: Base, 39: Base, 40: Base, 50, 50a, 50b, 50c: micropipette, 51, 51a, 51b, 51c: sample outlet, 52, 52a, 52b, 52c: sample inlet, 53: anisotropic flight shield plate, 55: dispenser, 60: cartridge .

Claims (27)

An inlet for injecting a sample from the outside, a cavity into which the sample is injected and filled, and an outlet for discharging the sample, formed in at least one or more substrates, and the substrate forming the cavity Is a micropipette comprising ceramics, comprising a piezoelectric / electrostrictive element on at least one wall surface of the base, and configured so that the sample moves in a laminar flow in the cavity,
A micropipette wherein a volume in the cavity is changed by driving the piezoelectric / electrostrictive element, and a certain amount of sample in the cavity is discharged from the discharge port. After filling the cavity with the replacement liquid in advance, and then injecting the sample from the injection port into the cavity while performing laminar flow replacement, the piezoelectric / electrostrictive element is driven and a certain amount of the sample in the cavity is removed. The micropipette according to claim 1, wherein the micropipette is discharged from the discharge port. After filling the cavity with a replacement liquid in advance, and injecting a sample into the cavity by laminar displacement from the injection port while driving the piezoelectric / electrostrictive element, driving the piezoelectric / electrostrictive element, 2. The micropipette according to claim 1, wherein a certain amount of the sample in the cavity is discharged from the discharge port. 4. The micropipette according to claim 2, wherein completion of laminar flow replacement of the sample in the cavity is grasped by detecting a change in fluid characteristics in the cavity. An inlet for injecting a sample from the outside, a cavity into which the sample is injected and filled, and an outlet for discharging the sample, formed in at least one or more substrates, and the substrate forming the cavity Is formed of ceramics, a piezoelectric / electrostrictive element is provided on at least one wall surface of the base, the volume of the cavity is changed by driving the piezoelectric / electrostrictive element, and a certain amount of sample in the cavity is discharged to the discharge port. A micropipette ejected from
Filling the cavity with a replacement liquid in advance, then injecting a sample from the injection port while replacing the cavity into the cavity, and detecting the completion of the replacement of the sample in the cavity and detecting a change in the fluid characteristic in the cavity. A micropipette, characterized in that the piezoelectric / electrostrictive element is driven after grasping by the above method, and a certain amount of sample in the cavity is discharged from the discharge port. 5. The method according to claim 4, wherein a change in fluid characteristics in the cavity is grasped by applying a voltage for exciting vibration to the piezoelectric / electrostrictive element and detecting a change in an electric constant caused by the vibration. Or the micropipette according to 5. The said inlet, the said cavity, the said discharge opening, and the said piezoelectric / electrostrictive element are each formed in two or more places in one said base material, The Claim 1 characterized by the above-mentioned. The micropipette described in 1. A plurality of units each having one of the injection port, the cavity, the discharge port, and the piezoelectric / electrostrictive element formed in one base are fixed to a fixing jig. The micropipette according to any one of claims 1 to 6. The combination of the cavity and the piezoelectric / electrostrictive element, and the three types of portions of the injection port and the discharge port are formed separately in at least two or more types of bases, and are joined to each other. The micropipette according to any one of claims 1 to 6. At least the cavity and the piezoelectric / electrostrictive element are formed in one base, and at least one of the bases is formed with at least one of the injection port and the discharge port. The micropipette according to any one of claims 1 to 6, wherein a unit joined to the plurality of substrates is formed, and at least one of the units is fixedly integrated. The micropipette according to any one of claims 1 to 10, wherein the base is flat, and the discharge port is formed on a side surface or a main plane of the base. The substrate according to any one of claims 1 to 10, wherein the base is flat, the discharge port is formed on one main plane of the base, and the injection port is formed on the other main plane. The micropipette according to claim 1 or 2. The micropipette according to any one of claims 1 to 12, wherein at least two or more inlets are connected to one cavity. 14. The micropipette according to claim 1, wherein at least the substrate on which the cavity and the piezoelectric / electrostrictive element are formed is made of zirconia ceramics. The micropipette according to any one of claims 1 to 14, wherein the substrate is made of zirconia ceramics. The micropipette according to any one of claims 1 to 15, wherein the substrate is manufactured using a green sheet laminating and firing method. The micropipette according to any one of claims 1 to 13, wherein the substrate on which at least one of the inlet and the outlet is formed is made of metal or resin. 18. The piezoelectric / electrostrictive film in the piezoelectric / electrostrictive element has a main component of a component composed of lead zirconate, lead titanate, and lead magnesium niobate, according to any one of claims 1 to 17, wherein Micropipette as described. An injection port for injecting a sample from outside, a cavity filled with the sample, and a discharge port for discharging the sample are formed in at least one or more substrates, and at least one of the substrates forming the cavity is formed. A dispensing apparatus using a plurality of micropipettes each including a piezoelectric / electrostrictive element on one wall surface and configured to move a sample in a laminar flow in the cavity,
A dispensing apparatus wherein the discharge ports are arranged vertically and horizontally, and different types of liquid samples are discharged from the discharge ports. After filling the plurality of cavities with the replacement liquid in advance, and then injecting different types of samples from the injection port into the plurality of cavities while performing laminar flow replacement, by driving the piezoelectric / electrostrictive element, 20. The dispensing apparatus according to claim 19, wherein different types of samples in the plurality of cavities are discharged from the discharge ports. The plurality of cavities are filled with a replacement liquid in advance, and different types of samples are injected into the plurality of cavities by laminar displacement from the injection ports while driving the piezoelectric / electrostrictive elements. 20. The dispensing apparatus according to claim 19, wherein the electrostrictive element is driven to discharge different types of samples in the plurality of cavities from the discharge ports. 22. The dispensing device according to claim 20, wherein completion of laminar flow replacement of the sample in the plurality of cavities is grasped by detecting a change in fluid characteristics in the cavities. An inlet for injecting a sample from the outside, a cavity into which the sample is injected and filled, and an outlet for discharging the sample, formed in at least one or more substrates, and the substrate forming the cavity Is made of ceramics, a piezoelectric / electrostrictive element is provided on at least one wall surface of the substrate, a replacement liquid is filled in the cavity in advance, and a sample is injected from the injection port while being replaced in the cavity. After the completion of the replacement of the sample in the cavity is grasped by detecting the change in the fluid characteristic in the cavity, the volume in the cavity is changed by driving the piezoelectric / electrostrictive element, and a certain amount of the cavity in the cavity is changed. A dispensing apparatus using a plurality of micropipettes for discharging a sample from the discharge port,
A dispensing apparatus wherein the discharge ports are arranged vertically and horizontally, and different types of liquid samples are discharged from the discharge ports. 23. A fluid characteristic in the plurality of cavities is grasped by applying a voltage for exciting vibration to the piezoelectric / electrostrictive element and detecting a change in an electric constant caused by the vibration. Or the dispensing device according to 23. 25. A mechanism according to claim 19, further comprising: attaching a cartridge separately filled with different types of liquid samples to each of said inlets, and discharging different liquid samples from said outlets. The dispensing device according to claim 1. A cartridge filled with an aqueous solvent or an organic solvent is attached to each of the inlets, and a mechanism for washing a space from the inlet to the outlet formed in the base is provided. The dispensing device according to any one of claims 19 to 25. 27. An anisotropic flying drop shielding plate made of a thin plate with a hole having the same central axis as the ejection port is provided outside the ejection port. Note device.

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