Abstract:
A small droplet discharging device is provided capable of producing high-density arrays. The droplet discharging device comprises a first substrate comprising a plurality of reservoir chambers for holding a liquid, a second substrate comprising a plurality of discharge units comprising supply openings for receiving a supply of a liquid stored in the reservoir chambers, pressurizing chambers for applying a pressure to the liquid supplied from the supply openings, and discharge openings for discharging the liquid pressurized in the pressurizing chambers to the outside, and a third substrate sandwiched between the first substrate and the second substrate and comprising channels for connecting the plurality of reservoir chambers with the plurality of supply openings corresponding thereto, wherein the plurality of supply openings provided in the second substrate are arranged such that relative positions thereof have a zigzag disposition.

Description:
RELATED APPLICATIONS 
     This application claims priority to Japanese Patent Application No. 2003-382891 filed Nov. 12, 2003 which is hereby expressly incorporated by reference herein in its entirety. 
     BACKGROUND OF THE INVENTION 
     The present invention relates to a droplet discharging device for discharging liquid drops of a very small amount, an apparatus for manufacturing a microarray and a method for manufacturing a microarray that use this droplet discharging device. 
     In recent years, decoding the base sequence of DNA and conducting functional analysis of genetic information became an important task, and DNA microarrays have been used as a technique for monitoring the gene expression patterns and screening new genes. With the array, the gene expression quantity is evaluated by preparing a probe DNA, high-density spotting on a substrate such as a slide glass, hybridizing a target DNA having a base sequence complementary with the probe DNA, of the fluorescent labeled target DNA, and observing the fluorescent pattern. 
     Further, a protein chip in which a protein is applied with a high density onto a substrate by employing the above-described technology has also been developed and used for expression analysis of proteins or analysis of protein interaction. 
     In order to manufacture such a microarray, a large number of probes have to be placed with a high density on a substrate. A method employing a droplet discharging device, for example, by using an ink-jet system, is such a method for placing a large number of probes on a substrate with a high density. 
     For example, Japanese Patent Application Laid-open No. 2002-286735 discloses a droplet discharging device comprising liquid supply openings arranged in the form of a matrix and connected with respective channels to a plurality of reservoir chambers arranged in the form of a matrix, wherein the arrangement pitch of the reservoir chambers is larger than the arrangement pitch of nozzle holes. With this droplet discharging device, the problem of increasing the degree of freedom in designing the nozzle spacing and the disposition spacing of the corresponding reservoir chambers was resolved by employing a multilayer configuration of channel substrates forming channels connecting the nozzles and the reservoir chambers. 
     However, because the channels in such a droplet discharging device have a multilayer structure, there is still space for improvement from the standpoint of device miniaturization. 
     SUMMARY 
     Accordingly, it is an object of the present invention to provide a small droplet discharging device capable of producing high-density arrays. 
     The present invention provides a droplet discharging device comprising a first substrate comprising a plurality of reservoir chambers (liquid reservoir chambers) for holding a liquid, a second substrate comprising a plurality of discharge units comprising supply openings for receiving a supply of the liquid stored in the reservoir chambers, pressurizing chambers for applying a pressure to the liquid supplied from the supply openings, and discharge openings for discharging the liquid pressurized in the pressurizing chambers to the outside, and a third substrate sandwiched between the first substrate and the second substrate and comprising channels for connecting the plurality of reservoir chambers with the plurality of supply openings corresponding thereto, wherein the plurality of supply openings provided in the second substrate are arranged such that relative positions thereof have a zigzag disposition. 
     With such a configuration, because the supply openings have a zigzag disposition, there is a high degree of freedom in selecting the width (channel width) of a channel itself, even if the prescribed width between the channels (wall thickness) is ensured. Therefore, excellent sealing ability between the channels is attained and the increase in channel resistance caused by restricting the channel width can be prevented. Further, because the supply openings have such a disposition, the degree of freedom in selecting the disposition of channels and reservoir chambers is increased and the droplet discharging device can be miniaturized. 
     It is preferred that the channels connected to the plurality of supply openings disposed in a zigzag fashion be formed alternately on both sides of the arrangement of the supply openings. With such a configuration, dispersing the channels makes it possible to dispose dispersedly the reservoir chambers connected thereto. Therefore, the droplet discharging device can be further miniaturized. 
     It is preferred that all the channels formed in the third substrate have an almost the same length. As a result, the difference in channel resistance caused by the spread in channel length can be reduced. Therefore, spread of discharge characteristics between the discharge openings (nozzles) can be prevented. 
     It is preferred that the second substrate comprise an electrode substrate having electrodes on the surface, a pressurizing chamber substrate disposed via a small gap opposite the electrode substrate and comprising pressurizing chambers with a pressure inside thereof adjusted by the displacement of diaphragms oscillating under the effect of an electrostatic force induced by the electrodes, and a nozzle substrate disposed on the opposite side of the pressurizing chamber substrate and having nozzle holes for discharging the liquid filling the pressurizing chambers to the outside. With such an electrostatic drive system, no heat is generated. Therefore, even when a solution containing a biological sample is used as the liquid for ejection, the effect of heat on the biological sample is prevented. 
     The drive system of the droplet discharging device is not limited to electrostatic drive and may be a piezoelectric drive system that generates no heat. 
     It is preferred that one of the first substrate and the third substrate be composed of glass, the other of the first substrate and the third substrate be composed of silicon, and the first substrate and the third substrate be bonded by anodic bonding. If the glass substrate and silicon substrate are thus used the main structural members, then it is possible to employ a lithographic technique that has been used in a semiconductor fabrication process or the like. Therefore, such substrates can be easily designed and processed. Further, anodic bonding uses no other elements such as adhesives during bonding. Therefore, the effect of other elements, such as contamination of the liquid by the admixture of the adhesive components, can be prevented. 
     Some of the channels may be formed in the first substrate. As a result, the degree of freedom in designing the channels is increased. 
     It is preferred that the channels formed in the first substrate and/or third substrate be formed by using a photolithography technology. With such a technology, fine channels can be easily processed. Furthermore, parameters can be changed by merely changing the pattern of the photomask, which is convenient from the standpoint of design changes. 
     The second aspect of the present invention is an apparatus for manufacturing a microarray comprising the above-described droplet discharging device and positioning means for adjusting relative positions of the droplet discharging device and the substrate for fixing the liquid drops discharged from the droplet discharging device. 
     With such a configuration, because the aforementioned miniature droplet discharging device is provided, the operability can be improved and a highly accurate microarray can be provided at a low cost. 
     The method for manufacturing a microarray in accordance with the present invention advantageously uses the discharging device for spotting various proteins on a chip for the fabrication of protein chips especially suitable for medical diagnostics. 
     The third aspect of the present invention is a method for manufacturing a microarray by which liquid drops are discharged onto a substrate and a microarray is manufactured by using the aforementioned droplet discharging device. 
     With such a configuration, because the aforementioned miniature droplet discharging device is provided, the operability can be improved and a highly accurate microarray can be provided at a low cost. 
    
    
     
       DESCRIPTION OF DRAWINGS 
         FIG. 1  is a plan view illustrating the droplet discharging device of the present embodiment. 
         FIG. 2A  is a cross-sectional view along the A–A′ line in  FIG. 1 , and  FIG. 2B  is a cross-sectional view along the B–B′ line in  FIG. 1 . 
         FIG. 3  illustrates the mutual arrangement of supply openings and nozzle holes in the head portion. 
         FIG. 4  illustrates the device for manufacturing a microarray of the present embodiment. 
         FIG. 5  illustrates the droplet discharging device of a comparative example. 
     
    
    
     DETAILED DESCRIPTION 
     An embodiment of the present invention will be described below with reference to the drawings. 
       FIG. 4  illustrates the apparatus for manufacturing a microarray of the present embodiment. 
     An apparatus  500  for the manufacture of a microarray of the present embodiment is designed for the manufacture of a microarray in which a plurality of spots are disposed on a substrate  10  for microarray production and is composed of a table  510 , an Y-direction drive shaft  520 , a droplet discharging device  100 , an X-direction drive shaft  530 , a drive unit  540 , and a control computer  600  as control means. The position control is conducted with the table  510 , Y-direction drive shaft  520 , X-direction drive shaft  530 , drive unit  540 , and control computer  600  (position control means). 
     The table  510  serves for carrying the substrate  10  constituting the microarray. The table  510  can carry a plurality of substrates  10  and is so composed that it can fix each substrate  10 , for example, by vacuum chucking. 
     The Y-direction drive shaft  520  can freely move the table  510  along the Y direction shown in the figure. The Y-direction drive shaft  520  is connected to a drive motor (not shown in the figure) contained in the drive unit  540  and moves the table  510  by receiving the drive force from the drive motor. 
     The droplet discharging device  100  discharges a biological sample solution toward the substrate  10  based on a drive signal supplied from the control computer  600 . The nozzle plane for discharging the solution is disposed on the X-direction drive shaft  530  so as to face the table  510 . For example, a DNA or a protein is used as a biological sample contained in the biological sample solution. The configuration of the droplet discharging device  100  will be described hereinbelow in greater detail. 
     The X-direction drive shaft  530  serves to move freely the droplet discharging device  100  along the X direction shown in the figure. The X-direction drive shaft  530  is connected to a drive motor (not shown in the figure) contained in the drive unit  540  and moves the droplet discharging device  100  by receiving the drive force from the drive motor. 
     The drive unit  540  comprises motors or other drive mechanisms for driving the Y-direction drive shaft  520  and X-direction drive shaft  530 . Those motors or mechanisms operate based on the drive signals supplied from the control computer  600 , thereby controlling the relative positions of the droplet discharging device  100  and table  510  carrying the substrate  10 . 
     The control computer  600  is disposed inside the housing of the drive unit  540 , controls the operation (discharge timing of the solution, discharge frequency, and the like) of the droplet discharging device  100 , and controls the operation of the drive unit  540 . 
     The droplet discharging device of the present embodiment will be described hereinbelow with reference to  FIG. 1  and  FIG. 2 . 
       FIG. 1  is a plan view illustrating the droplet discharging device of the present embodiment.  FIG. 2A  is a cross-sectional view along the A–A′ line in  FIG. 1 .  FIG. 2B  is a cross-sectional view along the B–B′ line in  FIG. 1 . 
     The droplet discharging device  100  has a first substrate  110  comprising a plurality of liquid reservoir chambers (reservoir chambers)  111 , a second substrate  150  comprising a plurality of pressurizing chambers  131  for applying a pressure to the liquid supplied from the liquid reservoir chambers  111 , and a third substrate  160  comprising a plurality of channels  161  for connecting the liquid reservoir chambers  111  to corresponding pressurizing chambers  131 . 
     The second substrate (head portion)  150  comprises an electrode substrate  120 , a pressurizing chamber substrate  130 , and a nozzle substrate  140 . 
     The electrode substrate  120  comprises a plurality of recesses  123  in the surface facing the pressuring chamber substrate  130 , and individual electrodes  122  are formed at the bottom surface of each recess  123 . Further, supply openings  121  for introducing the liquid accommodated in the liquid reservoir chambers  111  to the pressurizing chambers  131  are formed in the electrode substrate  120 . 
     The supply openings  121  are disposed in a zigzag fashion, when the second substrate  150  is viewed from the upper surface. 
       FIG. 3  shows mutual arrangement of the nozzle holes  141  and supply openings  121  in the second substrate.  FIG. 3A  is a plan view and  FIG. 3B  is a cross-sectional view along the C–C′ line in  FIG. 3A . As shown in  FIG. 3A , in order to miniaturize the head, the supply openings  121  are disposed with a spacing almost equal to that of the nozzle holes  141 . Furthermore, a plurality of supply openings  121  are disposed in a zigzag fashion so as to be displaced with respect to each other. 
     The pressurizing chamber substrate  130  comprises pressurizing chambers  131  for applying a force for pressing liquid drops to the outside. The bottom portion of the pressurizing chambers  131  are in the form of thin plates (diaphragms). If a voltage is applied from an external power source (not shown in the figure) to the common electrode (not shown in the figure) formed on the pressurizing chamber substrate  130  and an electrode  122  formed in the electrode substrate  120 , then the bottom portion is pulled by an electrostatic force to the electrode substrate  120 . If the voltage is thereafter switched off, the bottom portion returns to the original position. At this time, the pressure of the pressurizing chamber  131  temporarily increases and a liquid drop is thereby pushed to the outside. 
     The nozzle substrate  140  has discharge openings (nozzle holes)  141  for discharging the liquid pushed out of the pressurizing chambers  131  to the outside. 
     Such electrode substrate  120 , pressurizing chamber substrate  130 , and nozzle substrate  140  are composed, for example, of glass or silicon. 
     The third substrate  160  and the first substrate  110  are stacked on the second substrate  150  of the above-described configuration. 
     A plurality of liquid reservoir chambers  111  for holding (accommodating) a liquid are formed in the first substrate (reservoir chamber substrate)  110 . The liquid reservoir chambers  111  are so disposed as to be positioned alternately on both sides of the supply openings  121  for the liquid in the second substrate  150 , those openings being arranged in a zigzag fashion as shown in  FIG. 1 . Thus, because the liquid reservoir chambers  111  are disposed dispersedly with respect to the arrangement of supply openings  121 , the space utilization efficiency is increased by comparison with the configuration in which the liquid reservoir chambers  111  are disposed in a row and the droplet discharging device  100  can be miniaturized. 
     The third substrate (channel substrate)  160  is disposed between the first substrate  110  and second substrate  150 . The fine channels  161   a  extending in the plane direction and connecting the liquid reservoir chambers  111  and the supply openings  121  are formed in the surface of the third substrate  150  that faces the first substrate  110 . The channels  161   a  descend vertically above the supply openings  121  and are connected to the supply openings  121 . For example, a silicon substrate can be used as the third substrate  160 , and fine channels  161   a  are formed, for example, by using photolithography. 
     It is preferred that the first substrate and third substrate be bonded by anodic bonding, but this condition is not limiting. If they are thus bonded by anodic bonding, it is not necessary to introduce an adhesive or the like therebetween and the effect produced on biological samples is small. However, it does not mean that bonding with an adhesive is excluded from the scope of the present invention. Thus, the substrates may be bonded with an adhesive. In this case, it is preferred that an adhesive be selected which produces little effect on biological sample. When anodic bonding is employed, a glass substrate, more specifically, borosilicate glass substrate is used as the first substrate. 
       FIG. 1  shows a mutual arrangement of the first substrate  110  (or third substrate  160 ) and second substrate  150 , in particular, a mutual arrangement of the supply openings  121 , channels  161   a , and liquid reservoir chambers  111 . When the channels  161   a  are formed by laminating together the substrates in which fine grooves were formed, as in the droplet discharging device  100  of the present embodiment, the prescribed wall thickness (δ) has to be ensured in order to provide for sealing between the adjacent channels  161   a . However, when the channels are arranged in the same direction, if the necessary wall thickness (δ) is ensured, the channel width (W) decreases and channel resistance increases. In the present embodiment, the prescribed wall thickness δ 1  between the channels can be provided, without reducing the width W 1  of the channels  161   a , by disposing the supply openings  121  in a zigzag fashion and forming the channels  161   a  alternately on both sides of the arrangement of the supply openings  121 . Furthermore, disposing the supply openings  121  in a zigzag fashion makes it possible to disperse the liquid reservoir chambers  111  on both sides of the arrangement of the supply openings  121  and increases the degree of freedom in designing the disposition of the liquid reservoir chambers  111  and channel width. Further, because the distance of the channels from the liquid reservoir chambers  111  to supply openings  121  can be formed almost the same for all the channels, the spread of discharge characteristic between the nozzles caused by spread in the channel length can be prevented. 
     With the present embodiment, even if the prescribed value of the width (thickness of the wall portion) δ 1  between the channels  161   a  is ensured by disposing the supply openings  121  in a zigzag fashion, because the degree of freedom in selecting the width (channel width) W 1  of the channels  161   a  themselves is high, excellent sealing between the channels is obtained and the increase in channel resistance caused by restricting the channel width can be prevented. Moreover, since the length of all the channels is the same, spread of discharge characteristic between the nozzles can be prevented. Further, disposing the channels  161   a  and liquid reservoir chambers  111  alternately on both side of the arrangement of the supply openings provides for dispersed disposition of the channels  161   a  and liquid reservoir chambers thereby enabling the miniaturization of the droplet discharging device  100 . Therefore, an inexpensive droplet discharging device with good operability can be provided. 
     Further, if a microarray is produced by using such a droplet discharging device, the microarray with good accuracy can be provided at a low cost. 
     In the present embodiment, a glass substrate was used as the first substrate and the silicon substrate was used as the third substrate, but such a configuration is not limiting and a silicon substrate may be used as the first substrate and a glass substrate may be used as the third substrate. Further, this combination of the materials is also not limiting. 
     Further, in the above-described embodiment, an example was considered in which the channels were formed on the third substrate  160 , but such a configuration is not limiting and some of the channels may be formed on the first substrate  110 . More specifically, only the channels  161   b  in the vertical direction which are connected to the supply openings  121  may be formed in the third substrate  160 , and the channels  161   a  in the horizontal direction may be formed in the rear surface (the surface facing the third substrate  160 ) of the first substrate  110 . In this case, it is preferred that the first substrate  110  be a silicon substrate, because fine channels can be formed with good accuracy. Further, when the first substrate  110  and the third substrate  160  are bonded by anodic bonding, a glass substrate may be used as the third substrate  160 . 
       FIG. 5  shows a droplet discharging device as a comparative example for explaining the effect of the present invention. 
       FIG. 5A  illustrates a head portion as a comparative example in which the supply openings are disposed in a linear fashion.  FIG. 5B  illustrates a droplet discharging device as a comparative example for explaining the mutual arrangement of supply openings and liquid reservoir portions. 
     When a plurality of supply openings  121  are disposed in a linear fashion according to the spacing between the nozzle holes, as shown in  FIGS. 5A , B, if the prescribed wall thickness δ 2  between the channels  161   a  is ensured, the channel width W 2  of the channels  161   a  is restricted. Therefore, the reduction in the channel width W 2  increases the channel resistance and the discharge ability of liquid drops with a high viscosity is decreased. The resultant problem is that a limitation is placed on the types of liquid that can be discharged. Further, as shown in  FIG. 5B , when all the liquid reservoir chambers  111  are disposed on one side of the arrangement of the supply openings  121 , the outer shape of the droplet discharging device increases in size, operability of the device is degraded, and cost thereof is increased. 
     Another problem is that because of the spread of the channel lengths, uniform discharge characteristic cannot be obtained for all the nozzles due to the difference in the channel resistance caused by the difference in the channel length. 
     The droplet discharging device in accordance with the present invention resolves the aforementioned problems.