JP4880540B2 - Sample deposition equipment - Google Patents

Description

  The present invention relates to a sample deposition method and a sample deposition apparatus that can deposit a small amount of sample to be subjected to biochemical inspection on a substrate, and more particularly, to use different types of samples on the same substrate without using a mask member. The present invention relates to a sample deposition method and a sample deposition apparatus capable of depositing a large number.

  In recent years, what are called microarrays, DNA chips, protein chips, organic compound chips and the like (hereinafter referred to as “microarrays”) have attracted attention in the fields of DNA analysis and genome drug discovery. In this microarray, a small amount of samples (proteins, etc.) having different types and characteristics are arranged in a matrix on a substrate as large as a slide glass for an optical microscope. Therefore, it is possible to detect a sample that reacts with the chemical by applying some chemical to the entire microarray. As described above, the microarray can perform a large amount of tests at a time and confirm the results at a glance.

  In particular, when manufacturing microarrays for DNA, etc., it is necessary to arrange many types of samples on a single substrate, so the area occupied by one sample (part of DNA) on the substrate is on the order of microns. Sometimes it becomes.

  When manufacturing the microarray, that is, in order to deposit and arrange a plurality of types of samples on a substrate, it is necessary to deposit only a target type of sample in a predetermined minute region.

As shown in FIG. 9A, the conventional microarray manufacturing apparatus sprays the sample 1 charged by the spraying means, applies a potential to the mask 2 having minute holes, and passes through the holes of the mask 2. By depositing the sample on the substrate 3, the target sample 1 is deposited only in a minute region on the substrate. Then, by changing the type of the sample 1 to be sprayed and changing the relative position between the mask 2 and the substrate 3, different types of samples 1 are arranged in a matrix (see Patent Document 1).
JP 2001-281252 A

  However, as in the conventional microarray manufacturing apparatus, the sample 2 is deposited on the substrate 3 by passing the sample 1 through the holes of the charged mask 2 (or naturally charged) using the mask 2 having minute holes. Then, as shown in FIG. 9B, the peripheral edge of the deposited sample 1 may rise. This is presumably because the sample 1 that has reached the outside of the micropores of the mask 2 repels the charged mask 2 and gathers in the micropores, and passes through the micropores as it is and is deposited.

  As described above, a microarray in which a sample that is not uniformly deposited is arranged may not exhibit an appropriate reaction when subjected to some test, and the performance as a microarray cannot be guaranteed.

  The present invention has been made in view of the above problems, and an object of the present invention is to provide a sample deposition method and a sample deposition apparatus capable of uniformly depositing a sample on a minute region on a substrate without using a mask.

  In order to achieve the above object, a sample deposition method according to the present invention is a sample deposition method including a plurality of electrodes and depositing a sample on a predetermined region on a substrate made of an insulator, and spraying the sample A spraying step, a charging step for charging the sample, and a reverse polarity potential application step for applying a potential having a polarity opposite to the polarity of the charged sample to an electrode corresponding to a predetermined region on the substrate; And a repelling potential applying step of applying an alternating potential to the electrode adjacent to the electrode corresponding to a predetermined region, the polarity being the same as the polarity of the charged sample.

  Further, a sample deposition apparatus according to the present invention is a sample deposition apparatus that includes a plurality of electrodes and deposits a sample in a region corresponding to the electrode of a substrate made of an insulator, and spraying means for spraying the sample; A charge applying means for charging the sample; a first power supply for applying a potential having a polarity opposite to the polarity of the charged sample; and a DC potential having the same polarity as the polarity of the charged sample, or an alternating current It is characterized by comprising: a second power source for applying a potential; and selective connection means for selecting whether to connect each of the electrodes and the first power source or to the second power source.

  This makes it possible to deposit the sample uniformly in a predetermined region on the substrate without using a mask.

  Furthermore, an induction member that guides the sprayed sample in a predetermined direction and a third power source that applies a potential having the same polarity as the polarity of the charged sample or a ground potential to the induction member. preferable.

  As a result, the sprayed sample can be guided in a predetermined direction, and the deposition amount on the substrate with respect to the spray amount of the sample can be improved.

  The guide member preferably includes a guide surface having a constant distance from a region where the sample is deposited.

  This makes it possible to guide the sprayed sample toward a predetermined area on the substrate.

  Further, an annular converging member that guides the sprayed sample to converge within a predetermined range, and a fourth power source that applies a potential having the same polarity as the polarity of the charged sample or a ground potential to the converging member It is preferable to comprise.

  Thereby, the sprayed sample can be converged on a minute region, and the sample can be deposited on the minute region on the substrate. Therefore, it is possible to manufacture a substrate on which many samples are deposited.

  Furthermore, it is preferable to include an adsorption member that adsorbs the sprayed sample and an adsorption power source that applies a potential having a polarity opposite to the polarity of the charged sample.

  As a result, when different samples are continuously deposited by the same sample deposition apparatus, the spray space of the sample can be cleaned at the transition of the sample, and contamination of the sample to be deposited next can be prevented. Become.

  According to the present invention, a sample can be uniformly deposited in a predetermined region on a substrate.

  Next, a sample deposition apparatus according to the present invention will be described with reference to the drawings.

  FIG. 1 is a sectional view schematically showing a sample deposition apparatus, partially broken away.

  As shown in the figure, the sample deposition apparatus 100 is an apparatus for spraying a sample and depositing the sprayed sample only on a predetermined region. The capillary 101 as the spraying means, the charge applying means 110, and the first power source 102, a second power source 103, a connection selection means (not shown), a guide member 104, a third power source 105, a converging member 106, a fourth power source 107, a chamber 108, a fifth power source 109, , A pump 131, a pressure adjusting means 132, an on-off valve 133, and an XY stage 135. A sample chip 200 is placed on the XY stage 135.

  The capillary 101 is a device capable of spraying a sample stored therein, and has a nozzle portion provided with a fine hole for spraying at a tapered tip, and a cylinder that stores the sample and communicates with the nozzle portion. The storage part of a shape is provided. The capillary 101 is detachably attached to an L-shaped attachment member that hangs down from the ceiling of the chamber 108. The capillary 101 is preferably arranged so that the distance between the tip of the capillary 101 and the sample chip 200 placed on the XY stage 135 is in the range of 10 mm to 500 mm. If the distance between the sample chip 200 and the capillary 101 is less than 10 mm, the sample sprayed from the capillary 101 cannot be completely converged and it is difficult to deposit the sample in a predetermined region. On the other hand, if the distance is longer than 500 mm, the voltage necessary for converging the sprayed sample becomes very large, and abnormal discharge or the like occurs. Since the equipment itself becomes larger, the cost increases. These make it difficult to construct realistic equipment. The preferred range of the distance is 100 mm or more and 300 mm or less. The capillary 101 can be easily replaced with a capillary or the like in which other types of samples are stored by a robot (not shown) or manually.

  The charge applying unit 110 is a device for applying a charge to the sample. In this embodiment, the charge applying unit 110 applies a charge to the sample stored in the capillary 101. The charge applying unit 110 includes a rod electrode 111 and a sixth power source 112.

  The rod electrode 111 is a rod-shaped metal electrode that is in direct contact with the sample and directly charges the sample, and is immersed in the sample inside the capillary 101.

  The sixth power source 112 is a DC power source for supplying a charge to the sample. In the present embodiment, the sixth power source 112 supplies a negative charge to the sample, that is, charges the sample negatively. The sixth power source 112 has a function of maintaining the rod electrode 111 at the ground potential (0 V) and continuing to supply charges to the rod electrode 111.

  The first power source 102 is a DC power source that applies a potential of a polarity opposite to the polarity of the charged sample, in the case of the present embodiment, a positive polarity to a plurality of electrodes included in the sample chip 200. The connection selection means determines which electrode of the sample chip 200 the first power supply 102 applies a potential to. The connection selection means will be described later.

  The second power source 103 is an AC power source that applies an AC potential to an electrode other than the electrode connected to the first power source 102 (for example, an electrode adjacent to the electrode connected to the first electrode).

  Note that the second power supply 103 may be a DC power supply that applies a potential having the same polarity as that of the charged sample, and in the case of this embodiment, a negative polarity.

  The guide member 104 applies a potential of the same polarity (negative in this embodiment) as the polarity of the sample sprayed from the capillary 101 or a ground potential, thereby applying an electric field to the sample that is sprayed and flies randomly. This is a member that is guided in a predetermined direction by the formation of. The guide member 104 has a shape of a part of a hollow true sphere, and is attached to the chamber 108 in a state where the so-called bowl is turned down. A circular hole is provided in the upper end portion of the guide member 104, and the capillary 101 is disposed at the center of the hole. In other words, the guide member 104 is disposed so as to surround the capillary 101 disposed at the center of the hole. The inner surface of the guide member 104 has a shape that is constant in distance from a region where a sample is deposited, in this embodiment, a predetermined region of the sample chip 200 placed on the XY stage 135, and the predetermined member. It is arrange | positioned in the position where distance with this area becomes constant. The inner surface of the guiding member 104 functions as a guiding surface on which electric lines of force for guiding the sample are generated. Further, the chamber 108 and the guide member 104 are in an insulated state.

  The presence of the guide member 104 can improve the amount of the deposited sample with respect to the amount of the sprayed sample.

  The third power source 105 is a power source that applies a potential having the same polarity as that of the charged sample, in the case of this embodiment, a negative polarity to the induction member 104. The third power source 105 also has a function of maintaining the induction member 104 at the ground potential (0 V).

  The converging member 106 is a member for further converging the sample induced by the induction member 104 by applying a potential having the same polarity as the polarity of the sample sprayed from the capillary 101 or a ground potential to form an electric field. . The converging member 106 has an annular shape, and is disposed so as to surround an inverted conical shape (indicated by a one-dot chain line in the figure) connecting a region where the sample is deposited and a lower end edge of the guiding member 104. The converging member 106 is attached to a column that is erected on the floor surface of the chamber 108, and is insulated from the chamber 108. In this embodiment, the cross section orthogonal to the circumferential direction of the converging member 106 is a rectangle, but an arbitrary shape such as a circle or an ellipse can be selected. Further, in the present embodiment, the converging member 106 is disposed outside the one-dot chain line in the figure, but may be disposed so as to cover the one-dot chain line or inside the one-dot chain line. In the figure, the alternate long and short dash line is a virtual line.

  The presence of the converging member 106 makes it possible to adjust the size of the region on which the sample is deposited. Therefore, when the size of an electrode (described later) of the sample chip 200 and the region of the sample to be converged by the converging member 106 are substantially matched, the uniformity when the sample is deposited can be further improved. The deposition region is adjusted by adjusting three parameters: the distance between the sample chip 200 and the converging member 106, the diameter of the converging member 106, and the potential applied to the converging member 106.

  The fourth power source 107 is a DC power source that applies a potential having the same polarity as that of the charged sample, in the present embodiment, a negative polarity to the converging member 106. Note that the fourth power source 107 also has a function of maintaining the converging member 106 at the ground potential (0 V).

  The chamber 108 is a box for avoiding dust and the like from being mixed into the sample, and at least the inner surface is made of a conductive material. In the state where the sample is deposited, the chamber 108 is applied with a potential having the same polarity as the sample or a ground potential, and contributes to the formation of an electric field necessary for depositing the sample. Further, the chamber 108 is applied with a potential having a polarity opposite to that of the sample in a state where the deposition of the sample is completed, and adsorbs the sample remaining in the space in the chamber 108 to increase the cleanliness in the chamber 108. That is, the chamber 108 functions as an adsorption member. Further, when various members are charged and there is a concern about the occurrence of abnormal discharge such as corona discharge, an AC voltage is applied to the chamber 108 and the various members are neutralized.

  The fifth power source 109 is a power source that can switch the polarity and apply a DC voltage to the chamber 108 or an AC voltage to the chamber 108 as described above. In addition, when the chamber 108 functions as an adsorption member, the fifth power source 109 functions as an adsorption power source. The fifth power source 109 also has a function of maintaining the chamber 108 at the ground potential (0 V) as described above.

  The pump 131 is a pump that supplies a pressure necessary for spraying a sample from the capillary 101 as compressed air.

  The pressure adjusting means 132 is a throttle valve that adjusts the pressure of the compressed air supplied from the pump 131 to obtain an optimum pressure for spraying the sample.

  The on-off valve 133 is a valve that determines whether or not to supply compressed air to the capillary 101. The on-off valve 133 and the pressure adjusting means 132 are controlled by a control device (not shown).

  The XY stage 135 is a table on which the sample chip 200 is placed and can move the sample chip 200 in the horizontal direction.

  2A and 2B are diagrams conceptually showing the sample chip 200, where FIG. 2A is a top view and FIG. 2B is a front view.

  As shown in the figure, the sample chip 200 is a member on which a plurality of samples are deposited, and includes a substrate 201, an electrode 202, a terminal 203, and a connection portion 204.

  The substrate 201 is a member that becomes a base of the sample chip 200, and is made of transparent glass having insulating properties.

  The electrode 202 is a member that defines a region where a sample is deposited, and is made of a transparent material having conductivity (for example, ITO (Indium Tin Oxide)). The electrode 202 is provided on the surface of the substrate 201.

  As described above, if the substrate 201 and the electrode 202 are transparent, the deposited sample can be observed with a transmission optical microscope.

  The terminal 203 is a terminal for connecting to the connection selection means, and is made of ITO like the electrode 202.

  The connection portion 204 is a portion that electrically connects the electrode 202 and the terminal 203 and is made of ITO.

  Note that although the substrate 201 and the electrode 202 are made of a transparent material in this embodiment mode, they are not particularly limited. The material which comprises the sample chip 200 does not ask | require the sample deposition apparatus 100 concerning this invention.

  FIG. 3 is a front view conceptually showing the sample chip and the connection selection means, where (a) is an external view and (b) is a circuit diagram.

  As shown in the figure, the terminal 203 and the connection selection means 120 of the sample chip 200 are detachable, and the sample chip 200 and the new sample chip 200 are separated by separating the sample chip 200 and the connection selection means 120. It is possible to exchange.

  The connection selection unit 120 is a device that includes a switch 121 that can select whether an electrode to be connected is connected to the first power source 102 or the second power source 103. The connection selection means 120 includes a number of connectors protruding downward corresponding to the number of terminals 203 of the sample chip 200. Further, the connection selection means 120 can be separated from and connected to the sample chip 200, and in a state where the connection selection means 120 and the sample chip 200 are in contact, each connector 123 and the terminal 203 are electrically connected. That is, the connection selection unit 120 and the electrode 202 are connected.

  The switch 121 is provided corresponding to each connector 123, and connects the connected connector 123 and the first power supply 102, or connects the connector 123 and the second power supply 103. It is possible to select.

  The connection selection means 120 is connected to a control device (not shown), receives a command from the control device, sets the switch 121, and selects the power source connected to the connector 123 by the control device. It has become.

  Next, a sample deposition method using the sample deposition apparatus 100 will be described.

  FIG. 4 is a conceptual diagram showing each stage of the sample deposition state at a time with different magnifications.

  First, the capillary 101 filled with a predetermined sample is attached to the sample deposition apparatus 100.

  Next, the sixth power source 112 is driven to charge the sample negatively through the rod electrode 111 (charging step).

  In addition, an electric field is formed by applying a potential to each part of the sample deposition apparatus 100 and the electrode 202 of the sample chip 200. Specifically, a positive potential having a polarity opposite to the polarity of the charged sample is applied to the electrode 202 corresponding to a predetermined region of the sample chip 200 (the leftmost electrode of the sample chip 200 shown in the figure). Is applied (reverse polarity potential application step). The application of the potential is performed via the connection selection unit 120 using the first power source 102. The potential to be applied is selected from the range of 20 V or more and 3 KV or less when the ground potential is 0 V. When the voltage is less than 20 V, the ability of the electrode 202 to attract the charged sample is remarkably reduced, and the sample is not deposited in a predetermined region. This is because if it is higher than 3 KV, the probability of occurrence of abnormal discharge increases. Note that the potential applied to the electrodes 202 depends on the potential applied to the adjacent electrodes 202 and the distance between the electrodes.

  Further, an alternating potential is applied to the electrode 202 adjacent to the electrode 202 corresponding to the region of the sample chip 200 and all the electrodes 202 on the right side of the adjacent electrode 202 (repellent potential applying step). The application of the potential is performed via the connection selection unit 120 using the second power source 103. By applying an alternating potential to the electrode 202 existing in the region where the sample is not deposited, it is possible to avoid the deposition of the sample. This can be considered that the average value of the AC potential (potential 0 V) is applied to the electrode 202 in consideration of the moving speed of the sample in the space and the frequency of the AC potential. Therefore, compared to the electrode 202 to which a positive potential is applied, the potential of the electrode 202 to which an AC potential is applied is closer to the polarity of the sample, and it is considered that deposition of the sample can be avoided. Moreover, the effect which can suppress discharge with another electrode can be enjoyed by applying an alternating potential. Note that, when the electrode 202 other than the electrode 202 to which a positive potential is applied is grounded, a positive charge appears on the surface of the electrode 202 when the negative sample approaches the electrode 202, so that the sample repelling effect is poor and the discharge is performed. It is also difficult to deter.

  The AC voltage applied by the second power source to the electrode 202 is preferably in the range of 5 V to 1 KV in terms of effective value. If it is less than 5 V, the effect of repelling the sample cannot be expected. If it is higher than 1 KV, improvement in the effect of avoiding the sample cannot be expected, and there is a high possibility that dielectric breakdown will occur. The frequency is preferably 20 Hz or more and 50 KHz or less. If it is less than 20 Hz, it cannot be expected to completely avoid sample adhesion. If the frequency is higher than 50 KHz, the improvement in the repelling effect is hardly desired, and noise is generated, and there is a concern about adverse effects on others.

  Note that the potential applied by the second power source 103 may be a potential having the same polarity as the sample, that is, a negative DC potential.

  Further, a negative potential is applied to the induction member 104 using the third power source. A negative potential is applied to the converging member 106 using the fourth power source 107. Further, a negative DC potential is applied to the chamber 108 using the fifth power source.

  After forming an electric field in the sample deposition apparatus 100 as described above, compressed air is sent to the capillary 101 to spray the sample (spraying step).

  As shown in the upper part of the figure, the sprayed sample is scattered in a random manner as charged small particles, but is directed toward a predetermined region of the sample chip 200 by the electric field (electric field lines) formed by the induction member 104. Be guided to.

  The induced grain of the sample is further converged by the converging member 106 (the middle stage in the figure).

  Finally, the sample is attracted to the electrode 202 to which a reverse polarity voltage is applied, and is deposited in a predetermined region of the sample chip 200 (the lower part of the figure).

  Here, the distance between the converging member 106 and the sample chip 200 is sufficiently longer than the distance between the conventional sample chip and the mask. Therefore, the sample converged by the converging member 106 takes a considerable time to reach the sample chip 200. At this time, the sample flies evenly and deposits uniformly on the sample chip 200.

  By employing the above apparatus and method, it is possible to deposit a sample uniformly in a predetermined region on the sample chip 200.

  Next, the capillary 101 is filled with another type of sample, and the sample chip 200 is moved by the XY stage 135 so that the region where the next sample is deposited comes to the center of the converging member 106.

  By repeating the steps as described above, a plurality of different types of samples are deposited on the sample chip 200.

  In this embodiment, the guiding member 104 and the converging member 106 are used. However, the present invention does not necessarily require the guiding member 104 and the converging member 106. In other words, a charged sample is sprayed from the capillary 101, a potential opposite to that of the charged sample is applied to the electrode corresponding to a predetermined region of the sample chip 200, and an alternating current or a potential of the same polarity is applied to the other electrodes. As a result, the sample can be uniformly deposited in a predetermined region on the sample chip 200. The guiding member 104 and the converging member 106 are for enhancing the above-described effect, and exhibit the effect particularly when the predetermined region is very small.

  As shown in FIG. 5, when the electrode 202 provided in the sample chip 200 is rectangular, as shown in FIG. 6, if the guide member 104 and the converging member 106 are configured in a rectangle corresponding to the shape of the electrode 202, the electrode The sample can be deposited in a shape corresponding to the shape of 202.

  In addition, as shown in FIG. 7, if a multi-capillary including a plurality of capillaries 101 is used, labor for replacing the capillaries is reduced. Note that the capillary 101 from which the sample is sprayed can be executed by selecting the on-off valve 133. The sample can be charged by forming the capillary 101 with a conductor and applying a potential to the capillary 101 itself.

  Moreover, it is preferable to set the sample deposition apparatus 100 to the following potential state.

  That is, a potential selected from 20 V or more and 3 KV or less is applied to the electrode 202 of the sample chip 200. In this case, the polarity (+, −) may be any, but here, it is assumed that the sex polarity (+) is applied. The converging member 106 is grounded. The guide member 104 is grounded. Capillary 101 is grounded. Note that with the grounding, the capillary 101 or the like may be directly connected to the ground. Alternatively, the capillary 101 or the like may be connected to the ground via a power source to maintain the capillary 101 or the like at 0 V (ground potential).

  As a result, the electrode 202 of the sample chip 200 has a positive polarity, and the other members have 0 V (ground potential). Similarly, the capillary 101 has a potential of 0 V, but is induced by the positive polarity of the electrode 202 of the sample chip 200, and a negative charge is induced in the capillary 101.

  Therefore, the sample discharged from the capillary 101 is negatively charged and flies toward the electrode 202 of the sample chip 200.

  In such a potential state, many parts of the sample deposition apparatus 100 are at ground potential, so that abnormal discharge is unlikely to occur and the sample can be safely deposited in a stable state.

  Further, as shown in FIG. 8, a sample deposition apparatus 100 may be arranged in series, and an in-line deposition apparatus including a loader 211 and an unloader 212 may be configured. Thereby, it is possible to improve the throughput for manufacturing the sample chip 200. The sample deposition apparatus 100 may be arranged not only in a straight line but also in a circle.

  In the above embodiment, the ground potential is set to 0 V, and the polarity is determined based on the ground potential. However, the present invention is not limited to this. For example, the polarity may be determined based on a potential biased by a predetermined voltage from the ground potential.

  The present invention can be used for manufacturing a microarray.

It is sectional drawing which shows a sample deposition apparatus roughly and is shown partly broken. It is a figure which shows a sample chip | tip conceptually, (a) is a top view, (b) is a front view. It is a front view which shows a sample chip | tip and a connection selection means notionally, (a) is an external view, (b) is a circuit diagram. It is the conceptual diagram which showed each step of the deposition state of the sample at once, changing the expansion rate. It is a figure which shows the other kind of sample chip | tip conceptually, (a) is a top view, (b) is a front view. It is a perspective view which shows other embodiment of a sample deposition apparatus. It is sectional drawing which shows the other embodiment of a sample deposition apparatus roughly, and is shown partly broken. It is a front view which shows notionally the in-line type sample deposition apparatus. It is a conceptual diagram which shows the deposition state of the sample using the conventional mask.

Explanation of symbols

DESCRIPTION OF SYMBOLS 100 Sample deposition apparatus 101 Capillary 102 1st power supply 103 2nd power supply 104 Guidance member 105 3rd power supply 106 Convergence member 107 4th power supply 108 Chamber 109 5th power supply 110 Charge provision means 111 Rod electrode 112 6th power supply 120 Connection selection means 121 Switch 123 Connector 131 Pump 132 Pressure adjusting means 133 On-off valve 135 XY stage 200 Sample chip 201 Substrate 202 Electrode 203 Terminal 204 Connection section 211 Loader 212 Unloader

Claims (1)

A sample deposition apparatus comprising a plurality of electrodes and depositing a sample in a region corresponding to the electrodes of a substrate made of an insulator,
Spraying means for spraying the sample;
Charge applying means for charging the sample;
A first power source for applying a potential having a polarity opposite to that of the charged sample;
A second power source for applying a DC potential having the same polarity as that of the charged sample or an AC potential;
A selective connection means for selecting whether to connect each of the electrodes and the first power source or to the second power source ;
A guiding member for guiding the sprayed sample in a predetermined direction;
A third power source for applying a potential having the same polarity as the polarity of the charged sample or a ground potential to the induction member ;
The guide member includes a sample deposition device a distance between the area where the sample is deposited Ru provided with a guiding surface becomes constant.

Images