Patent Application
20100062446
1. Field of the Invention
The present invention relates to a reactor plate suitable for use in various assays and analyses such as biological and biochemical assays and general chemical analyses in the fields of medical care and chemistry, and a reaction processing method for processing such a reactor plate.
2. Description of the Related Art
As small reactors for use in biochemical assays or general chemical analyses, micro multi-chamber devices are used. Examples of such devices include micro well reactor plates such as a microtiter plate constituted from a plate-shaped substrate having a plurality of wells formed in the surface thereof (see, for example, Patent Document 1), and the like.
Further, as a structure for dispensing a small amount of liquid which can quantitatively treat a small amount of liquid, a structure having a first channel, a second channel, a third channel which is in communication with the first channel through an opening provided in the channel wall of the first channel, and a fourth channel which is in communication with the second channel through an opening provided in the channel wall of the second channel, connects one end of the third channel to the second channel, and has relatively lower capillary attraction than the third channel is developed (see, for example, Patent Documents 2, 3). When such a structure for dispensing a small amount of liquid is used, a liquid introduced into the first channel is drawn into the third channel, and then the liquid remaining in the first channel is removed. As a result, the liquid having a volume corresponding to the capacity of the third channel is dispensed into the second channel.
When a conventional micro well reactor plate is used, the top surface of the reactor plate is open to the atmosphere. Therefore, there is a possibility that foreign matter will enter a sample from outside, or otherwise, a reaction product will pollute an outside environment.
Further, in the structure for dispensing a small amount of liquid disclosed in Patent Documents 2 and 3, each of the first and second channels has a port for introducing a liquid at each end thereof. However, these ports are open to the atmosphere and, therefore, there is a possibility that a reaction product will leak through the ports and then pollute an outside environment.
Therefore, it is an object of the present invention to provide a reactor plate which can prevent the entry of foreign matter from outside and the pollution of an outside environment, and a reaction processing method using such a reactor plate.
The present invention is directed to a reactor plate including a sealed reaction well, a reaction well channel connected to the reaction well, a sealed well provided separately from the reaction well, a sealed well channel connected to the sealed well, a syringe for sending a liquid, a syringe channel connected to the syringe, and a rotary switching valve for connecting the syringe channel to the reaction well channel or the sealed well channel, wherein the sealed well channel is openably sealed with the rotary switching valve at an end thereof not connected to the sealed well.
In the reactor plate according to the present invention, since the reaction well and the sealed well are sealed, it is possible to prevent the entry of foreign matter from the outside of the reactor plate and the pollution of an outside environment with a liquid.
Further, since the sealed well channel is openably sealed with the rotary switching valve at an end thereof not connected to the sealed well, it is possible, even when the sealed well previously contains a liquid such as a liquid reagent or dilution water, to prevent the entry of foreign matter into the sealed well from the outside and the pollution of an environment outside the rector plate with the liquid contained in the sealed well.
In a case where the reactor plate according to the present invention is intended to be used for measuring a gene-containing sample, a sample previously subjected to gene amplification reaction may be introduced into the reactor plate, or a gene amplification reagent may be previously contained in the reaction well or the reactor plate may be designed to allow a gene amplification reagent to be dispensed into the reaction well so that gene amplification reaction can be carried out in the reaction well of the reactor plate.
Examples of the gene amplification reaction include PCR method and LAMP method. As PCR method for amplifying DNA, a method is proposed for directly subjecting a sample such as blood to PCR reaction without pretreating the sample. More specifically, this method is a nucleic acid synthesis method for amplifying a target gene contained in a gene-containing sample by adding a gene-containing body contained in the gene-containing sample or the gene-containing sample itself and then adjusting the pH of the thus obtained reaction mixture to 8.5 to 9.5 (25° C.) (see Patent Document 4).
Further, conventional micro devices having a rotary switching valve thereon are disclosed in, for example, Patent Documents 5 to 7.
In the reactor plate according to the present invention, the rotary switching valve may have a sealing plate made of an elastic material and having a first through hole to be connected to the syringe channel and a second through hole to be connected to the reaction well channel and the sealed well channel.
In this case, a surface of the sealing plate opposed to the syringe channel, the reaction well channel, and the sealed well channel may be covered with a fluorine resin layer formed thereon or a fluorine resin member placed thereon and having a through hole provided at a position corresponding to the position of the first through hole and a through hole provided at a position corresponding to the position of the second through hole. Examples of a fluorine resin for forming the fluorine resin layer or the fluorine resin member include PTFE (polytetrafluoroethylene) and PCTFE (polychlorotrifluoroethylene). However, the fluorine resin is not limited to PTFE and PCTFE, and any other fluorine resins may be used.
An example of the sealed well includes a sample well for containing a sample liquid.
For example, the sample well may be hermetically sealed with an elastic member which allows a dispensing device having a sharp tip to pass through to form a through hole and which also allows the through hole to be closed by pulling out the dispensing device due to its elasticity.
Further, the sample well may previously contain a liquid for pretreating a sample or a reagent.
The reactor plate according to the present invention may further include one or more reagent wells, each of which is constituted from the sealed well, other than the sample well. The reagent well previously contains a reagent to be used for the reaction of a sample liquid and is sealed with a film, or has an openable and closable cap so that the reagent can be injected thereinto. An example of the film for sealing the reagent well to prevent the leakage of the reagent includes one through which a dispensing device having a sharp tip can pass.
In a case where the reactor plate according to the present invention is intended to be used for gene analysis, the reactor plate preferably includes a gene amplification well which is constituted from the sealed well and used for carrying out gene amplification reaction. The gene amplification well preferably has a shape suitable for controlling a temperature according to a predetermined temperature cycle. It is to be noted that gene amplification can also be carried out in the reaction well.
The rotary valve may have a port to be connected to the syringe at the center of rotation. In this case, the syringe may be placed on the rotary valve.
The reaction well can be used for carrying out at least any one of color reaction, enzymatic reaction, fluorescence reaction, chemiluminescence reaction, and bioluminescence reaction.
The reaction well may be made of an optically-transparent material so that optical measurement can be carried out from the bottom of the reaction well or from above the reaction well.
In a case where a liquid to be introduced into the reaction well channel contains a gene, the reaction well may contain a probe which reacts with the gene.
Further, the probe may be fluorescently-labeled.
The reactor plate according to the present invention may further include a reaction well air vent channel connected to the reaction well. As a specific example of the channel configuration of the reactor plate according to the present invention, the reaction well channel which is constituted from a groove formed in the contact surface between two members bonded together, or from the groove and a through hole formed in both or one of the members and which includes a main channel connected to the syringe channel, a metering channel branched off the main channel and having a predetermined capacity, and an injection channel whose one end is connected to the metering channel and whose other end is connected to the reaction well can be mentioned. In this case, the main channel and the reaction well air vent channel can be hermetically sealed, the injection channel is formed narrower than the metering channel, and does not allow the passage of a liquid at a liquid introduction pressure applied to introduce the liquid into the main channel and the metering channel and at a purge pressure applied to purge the liquid from the main channel but allows the passage of the liquid at a pressure higher than the liquid introduction pressure and the purge pressure.
The present invention is also directed to a reaction processing method using the reactor plate according to the present invention having the channel configuration described above as an example of a channel configuration, the method including filling the main channel and the metering channel with a liquid at the introduction pressure, purging the liquid from the main channel by flowing a gas through the main channel while allowing the liquid to remain in the metering channel, and injecting the liquid contained in the metering channel into the reaction well through the injection channel by creating a positive pressure higher than the introduction pressure in the main channel, or by creating a negative pressure in the reaction well, or by creating a positive pressure higher than the introduction pressure in the main channel and creating a negative pressure in the reaction well.
Here, the phrase “the injection channel is formed narrower than the metering channel” means that in a case where the injection channel is constituted from a plurality of channels, each of the channels constituting the injection channel is formed narrower than the metering channel.
In the above-described channel configuration, since the main channel and the reaction well air vent channel can be hermetically sealed, it is possible to prevent the entry of foreign matter from the outside of the reactor plate and the pollution of an environment outside the reactor plate with the liquid.
In the channel configuration described above as an example of a channel configuration, the contact angle of the injection channel with a water drop is, for example, 90° or larger, and the area of an interface between the injection channel and the metering channel is, for example, 1 to 10,000,000 μm2 (square micrometers). It is to be noted that in a case where the injection channel is constituted from a plurality of channels, the phrase “the area of an interface between the injection channel and the metering channel” means the area of an interface between each of the channels constituting the injection channel and the metering channel.
The reactor plate according to the present invention may include the plurality of reaction wells. In this case, the metering channel and the injection channel may be provided for each of the reaction wells, and the plurality of metering channels may be connected to the main channel.
The reactor plate according to the present invention may further include a projecting portion which projects from a top inner surface of the reaction well. In this case, the other end of the injection channel is located at the tip of the projecting portion. The projecting portion includes one having a proximal end and a distal end narrower than the proximal end.
As described above, the reactor plate according to the present invention includes a sealed reaction well, a reaction well channel connected to the reaction well, a sealed well provided separately from the reaction well, a sealed well channel connected to the sealed well, a syringe for sending a liquid, a syringe channel connected to the syringe, and a rotary switching valve for connecting the syringe channel to the reaction well channel or the sealed well channel. Therefore, it is possible to prevent the entry of foreign matter from the outside of the reactor plate and the pollution of an environment outside the reactor plate with a liquid contained in the reactor plate.
Further, the sealed well channel is openably sealed with the rotary switching valve at an end thereof not connected to the sealed well. This makes it possible, even when the sealed well previously contains a liquid such as a liquid reagent or dilution water, to prevent the entry of foreign matter into the sealed well from the outside and the pollution of an environment outside the rector plate with the liquid contained in the sealed well.
In a case where the reactor plate according to the present invention is intended to be used for measuring a gene-containing sample, the sample injected into the reactor plate and then introduced into the reaction well can be processed in a closed system. Therefore, it is possible to prevent the pollution of an environment outside the reactor plate and the contamination of the sample with foreign matter coming from outside the reactor plate.
In the reactor plate according to the present invention, the rotary switching valve may have a sealing plate made of an elastic material and having a first through hole to be connected to the syringe channel and a second through hole to be connected to the reaction well channel and the sealed well channel. This makes it possible to reliably seal the sealed well channel due to the elasticity of the sealing plate.
Further, a surface of the sealing plate opposed to the syringe channel, the reaction well channel, and the sealed well channel may be covered with a fluorine resin layer formed thereon or a fluorine resin member placed thereon and having a through hole provided at a position corresponding to the position of the first through hole and a through hole provided at a position corresponding to the position of the second through hole. This makes it possible to more reliably seal the sealed well channel due to the elasticity of the sealing plate and hermeticity achieved by the surface covered with a fluorine resin. The fluorine resin layer or the fluorine resin member is effective as a member for sealing the rotary switching valve because a fluorine resin has a small coefficient of friction. Further, the fluorine resin layer or the fluorine resin member can hermetically seal the reaction well channel and the sealed well channel because a fluorine resin is highly airtight. Therefore, it is possible to prevent a liquid contained in the reaction well or the sealed well from being vaporized. This makes it possible to store the unused reactor plate for a long period of time.
For example, by providing a sample well for containing a sample liquid as the sealed well, it is possible to eliminate the necessity to separately prepare a well for containing a sample.
Further, the sample well may be hermetically sealed with an elastic member which allows a dispensing device having a sharp tip to pass through to form a through hole and which also allows the through hole to be closed by pulling out the dispensing device due to its elasticity. This makes it possible to inject a sample liquid into the sample well sealed with the elastic member and to prevent the sample liquid injected into the sample well from leaking out of the sample well.
Further, the sample well may previously contain a liquid for pretreating a sample or a reagent. This makes it possible to eliminate the necessity to dispense a liquid for pretreating a sample or a reagent into the sample well.
The reactor plate according to the present invention may further include one or more reagent wells, each of which is constituted from the sealed well, other than the sample well. By allowing the reagent well to previously contain a reagent to be used for the reaction of a sample liquid and sealing it with a film or by allowing the reagent well to have an openable and closable cap so that the reagent can be injected thereinto, it is possible to eliminate the necessity to separately prepare a well for containing the reagent.
The reactor plate according to the present invention may further include a gene amplification well which is constituted from the sealed well and used for carrying out gene amplification reaction. By providing such a gene amplification well, it is possible to amplify a target gene in the reactor plate by gene amplification reaction such as PCR method or LAMP method even when a sample liquid contains only a very small amount of the target gene, thereby increasing analytical precision.
By providing a port to be connected to the syringe at the center of rotation of the rotary valve, it is possible to simplify a channel configuration.
Further, by providing a port to be connected to the syringe at the center of rotation of the rotary valve and placing the syringe on the rotary valve, it is possible to shorten or eliminate a channel between the port and the syringe, thereby simplifying the structure of the reactor plate. In addition, it is also possible to effectively utilize a region on the switching valve, thereby making it possible to make the planar size of the reactor plate smaller as compared to a case where the syringe is placed in a region other than the region on the switching valve.
In a case where the reactor plate according to the present invention is intended to be used for measuring a gene-containing sample, the reactor plate may be designed to allow gene amplification reaction to be carried out in the reaction well. This makes it possible to eliminate the necessity to prepare a sample which has been subjected to gene amplification reaction outside the reactor plate.
The reaction well may be made of an optically-transparent material so that optical measurement can be carried out from the bottom of the reaction well or from above the reaction well. This makes it possible to optically measure a liquid contained in the reaction well without transferring the liquid into another well.
In a case where a liquid to be introduced into the reaction well channel contains a gene, the reaction well may contain a probe which reacts with the gene. This makes it possible to detect a gene having a base sequence corresponding to the probe in the reaction well.
The reactor plate according to the present invention may further include a reaction well, a reaction well channel connected to the reaction well, and a reaction well air vent channel connected to the reaction well. As an example of the channel configuration of the reactor plate according to the present invention, the reaction well channel which is constituted from a groove formed in the contact surface between two members bonded together, or from the groove and a through hole formed in both or one of the members and which includes a main channel connected to the syringe channel, a metering channel branched off the main channel and having a predetermined capacity, and an injection channel whose one end is connected to the metering channel and whose other end is connected to the reaction well can be mentioned. In this case, the main channel and the reaction well air vent channel can be hermetically sealed, and the injection channel is formed narrower than the metering channel, and does not allow the passage of a liquid at a liquid introduction pressure applied to introduce the liquid into the main channel and the metering channel and at a purge pressure applied to purge the liquid from the main channel but allows the passage of the liquid at a pressure higher than the liquid introduction pressure and the purge pressure. By carrying out the reaction processing method according to the present invention with the reactor plate according to the present invention of the above channel configuration, it is possible to prevent the entry of foreign matter from the outside of the reactor plate and the pollution of an outside environment with a liquid.
Further, by providing the reaction well air vent channel connected to the reaction well, it is possible to move a gas between the reaction well and the reaction well air vent channel when a liquid is injected into the reaction well through the injection channel, thereby making it possible to smoothly inject a liquid into the reaction well. The reaction well air vent channel can also be used to suck a gas contained in the reaction well to decompress the reaction well to inject a liquid into the reaction well.
In the channel configuration described above as an example of a channel configuration, the contact angle of the metering channel and the injection channel with a water droplet is preferably 90° or larger, and the area of an interface between the injection channel and the metering channel is preferably 1 to 10,000,000 μm2. This makes it difficult for a liquid to enter the injection channel when the liquid is introduced into the main channel and the metering channel, thereby making it possible to increase an introduction pressure applied to introduce a liquid into the main channel and the metering channel.
The reactor plate according to the present invention may include a plurality of the reaction wells. In this case, by providing the metering channel and the injection channel for each of the reaction wells and connecting the plurality of metering channels to the main channel, it is possible to introduce a liquid into the plurality of metering channels one after another and then simultaneously inject the liquid into the plurality of reaction wells through the injection channels.
Further, a projecting portion may be provided so as to project from a top inner surface of the reaction well. In this case, the other end of the injection channel is located at the tip of the projecting portion. By allowing the projecting portion to have a proximal end and a distal end narrower than the proximal end, a liquid to be injected into the reaction well through the injection channel can be easily dropped into the reaction well.
A reactor plate 1 includes a plurality of reaction wells 5 each having an opening in one surface of a well base 3. In the reactor plate 1 according to this embodiment of the present invention, the reaction wells 5 are arranged in an array of 6 rows and 6 columns in a staggered format. In each of the reaction wells 5, a reagent 7 and a wax 9 are contained.
The material of the well base 3 including the reaction wells 5 is not particularly limited. However, in a case where the reactor plate 1 is intended to be disposable, the material of the well base 3 is preferably a cheaply-available material. Preferred examples of such a material include resin materials such as polypropylene and polycarbonate. In a case where the reactor plate 1 is intended to be used to detect a substance in the reaction well 5 by absorbance, fluorescence, chemiluminescence, or bioluminescence, the container base 3 is preferably made of an optically-transparent resin so that optical detection can be carried out from the bottom of the reaction well 5. Particularly, in a case where the reactor plate 1 is intended to be used for fluorescence detection, the container base 3 is preferably made of a low self-fluorescent (i.e., fluorescence emitted from a material itself is weak) and optically-transparent resin, such as polycarbonate. The thickness of the well base 3 is in a range of 0.2 to 4.0 mm (millimeters), preferably in a range of 1.0 to 2.0 mm. From the viewpoint of low self-fluorescence, the thickness of the well base 3 for fluorescence detection is preferably small.
Referring to
The main channel 13 is constituted from one channel, and is therefore bent so as to run in the vicinity of all the reaction wells 5. One end of the main channel 13 is connected to a channel 13a constituted from a through hole provided in the well base 3. The channel 13a is connected to a port of a switching valve 63 (which will be described later). The other end of the main channel 13 is connected to the liquid drain space 29 provided in the well base 3. The main channel 13 is constituted from a groove having a depth of, for example, 400 μm (micrometers) and a width of, for example, 500 μm. It is noted that a part of the main channel 13 having a predetermined length (e.g., 250 μm) and located downstream of a position, to which the metering channel 15 is connected, has a width smaller than that of the other part of the main channel 13, and the width of such a part is, for example, 250 μm.
The metering channel 15 branches off the main channel 13, and is provided for each of the reaction wells 5. The end of the metering channel 15 on the opposite side of the main channel 13 is located in the vicinity of the reaction well 5. The depth of a groove constituting the metering channel 15 is, for example, 400 μm. The metering channel 15 is designed to have a predetermined internal capacity of, for example, 2.5 μL (microliters). A part of the metering channel 15 connected to the main channel 13 has a width larger than that of the above-described narrow part of the main channel 13 (e.g., 500 μm). Therefore, at a position where the metering channel 15 branches off the main channel 13, the resistance to the flow of a liquid coming from one end of the main channel 13 is larger in the main channel 13 than in the metering channel 15. For this reason, the liquid coming from one end of the main channel 13 first flows into the metering channel 15 to fill the metering channel 15, and then flows downstream through the narrow part of the main channel 13.
The injection channel 17 is also provided for each of the reaction wells 5. One end of the injection channel 17 is connected to the metering channel 15, and the other end of the injection channel 17 is connected to the recess 27 located above the reaction well 5 so as to be led to the space above the reaction well 5. The injection channel 17 is designed to have a size allowing the liquid-tightness of the reaction well 5 to be maintained in a state where there is no difference between the pressure in the reaction well 5 and the pressure in the injection channel 17. According to the present embodiment, the injection channel 17 is constituted from a plurality of grooves, and each groove has a depth of, for example, 10 μm and a width of, for example, 20 and the pitch between the adjacent grooves is, for example, 20 μm, and the thirteen grooves are provided in a region having a width of 500 μm. In this case, the area of an interface between the groove constituting the injection channel 17 and the metering channel 15, that is, the cross-sectional area of the groove constituting the injection channel 17 is 200 μm. The recess 27 has a depth of, for example, 400 μm, and has a circular planar shape smaller than that of the reaction well 5.
The reaction well air vent channel 19 is provided for each of the reaction wells 5. One end of the reaction well air vent channel 19 is connected to the recess 27, which is located above the reaction well 5, at a position different from the position, to which the injection channel 17 is connected, so as to be located above the reaction well 5. The reaction well air vent channel 19 is designed to have a size allowing the liquid-tightness of the reaction well 5 to be maintained in a state where there is no difference between the pressure in the reaction well 5 and the pressure in the reaction well air vent channel 19. The other end of the reaction well air vent channel 19 is connected to the reaction well air vent channel 21. According to the present embodiment, the reaction well air vent channel 19 is constituted from a plurality of grooves, and each groove has a depth of, for example, 10 μm and a width of, for example, 20 μm, and the pitch between the adjacent grooves is, for example, 20 μm, and the thirteen grooves are provided in a region having a width of 500 μm.
The reactor plate according to the present embodiment has a plurality of the reaction well air vent channels 21. To each of the reaction well air vent channels 21, the plurality of reaction well air vent channels 19 are connected. These reaction well air vent channels 21 are provided to connect the reaction well air vent channels 19 to the air drain space 31 provided in the well base 3. Each of the reaction well air vent channels 21 is constituted from a groove having a depth of, for example, 400 μm and a width of, for example, 500 μm.
The drain space air vent channel 23 is provided to connect the liquid drain space 29 to a port of the switching valve 63 (which will be described later). One end of the drain space air vent channel 23 is located above the liquid drain space 29. The other end of the drain space air vent channel 23 is connected to a channel 23a constituted from a through hole provided in the well base 3. The channel 23a is connected to a port of the switching valve 63 (which will be described later). The drain space air vent channel 23 is constituted from a groove having a depth of, for example, 400 μm and a width of, for example, 500 μm.
The drain space air vent channel 25 is provided to connect the air drain space 31 to a port of the switching valve 63 (which will be described later). One end of the drain space air vent channel 25 is located above the air drain space 31. The other end of the drain space air vent channel 25 is connected to a channel 25a constituted from a through hole provided in the well base 3. The channel 25a is connected to a port of the switching valve 63 (which will be described later). The drain space air vent channel 25 is constituted from a groove having a depth of, for example, 400 μm and a width of, for example, 500 μm
On the channel base 11, a channel cover 33 (not shown in
Referring to
In the well base 3, a sample channel 35a constituted from a through hole extending from the bottom of the sample well 35 to the back surface of the well base 3 and a sample well air vent channel 35b constituted from a through hole extending from the top surface to the back surface of the well base 3 are provided in the vicinity of the sample well 35. On the well base 3, a projecting portion 35c is provided so as to surround an opening of the sample well 35. In the projecting portion 35c, a sample well air vent channel 35d constituted from a through hole is provided so as to be located above the sample well air vent channel 35b. In the surface of the projecting portion 35c, a sample well air vent channel 35e which allows the sample well 35 to communicate with the sample well air vent channel 35d is provided.
The sample well air vent channel 35e is constituted from one or more narrow holes, and each narrow hole has a width of, for example, 5 to 200 μm and a depth of, for example, 5 to 200 μm. The sample well air vent channel 35e is provided to maintain the liquid-tightness of the sample well 35 in a state where there is no difference between the pressure in the sample well 35 and the pressure in the sample well air vent channel 35d. On the projecting portion 35c, a septum 41 as an elastic member to cover the sample well 35 and the air vent channel 35d is provided. The septum 41 is made of an elastic material such as silicone rubber or PDMS. Therefore, a dispensing device having a sharp tip can pass through the septum 41 to form a through hole, but the through hole can be closed by pulling the dispensing device out of the septum 41 due to its elasticity. On the septum 41, a septum stopper 43 for fixing the septum 41 is provided. The septum stopper 43 has an opening located above the sample well 35. According to the present embodiment, a reagent 45 is previously contained in the sample well 35.
As shown in
The reagent well air vent channel 37e is constituted from one or more narrow holes, and each narrow hole has a width of, for example, 5 to 200 μm and a depth of, for example, 5 to 200 μm. The reagent well air vent channel 37e is provided to maintain the liquid-tightness of the reagent well 37 in a state where there is no difference between the pressure in the reagent well 37 and the pressure in the reagent well air vent channel 37d. On the projecting portion 37c, a film 47 made of, for example, aluminum to cover the reagent well 37 and the air vent channel 37d is provided. In the reagent well 37, dilution water 49 is contained.
As shown in
Referring to
In the well base 3, the bellows 53 is also provided at a position other than the positions of a region where the reaction wells 5 are arranged, the drain spaces 29 and 31, the wells 35, 37, and 39, and the syringe 51. The bellows 53 has a sealed internal space, and the internal capacity of the bellows 53 is passively variable by extraction and contraction. The bellows 53 is placed in, for example, a through hole 53a provided in the well base 3.
A well bottom 55 is attached to the back surface of the well base 3 at a position other than the position of a region where the reaction wells 5 are arranged. In the well bottom 55, an air vent channel 53b is provided at a position allowing the air vent channel 53b to communicate with the bellows 53. The bellows 53 is connected to the well bottom 55 so as to be in close contact with the surface of the well bottom 55. The well bottom 55 is provided to guide the channels 13a, 23a, 25a, 35a, 35b, 37a, 37b, 39a, 39b, 51c, and 53b to predetermined port positions.
On the surface of the reaction well bottom 55 located on the opposite side from the well base 3, the rotary switching valve 63 is provided. The switching valve 63 is constituted from disk-shaped sealing plate 57, rotor upper 59, and rotor base 61. The switching valve 63 is attached to the well bottom 55 by means of a lock 65.
The sealing plate 57 is made of an elastic material such as a silicone rubber or PDMS. The surface of the sealing plate 57 is covered with a layer made of, for example, PTFE (not shown). The sealing plate 57 has a through hole (second through hole) 57a, a through groove 57b, and a through hole (first through hole) 57c. The through hole 57a is provided in the vicinity of the peripheral portion of the sealing plate 57, and is connected to any one of the channels 13a, 35a, 37a, and 39a. The through groove 57b is provided inside the through hole 57a and on a circle concentric with the sealing plate 57, and is connected to at least two of the channels 23a, 25a, 35b, 37b, 39b, and 53b. The through hole 57c is provided at the center of the sealing plate 57, and is connected to the syringe channel 51c.
The rotor upper 59 has a through hole 59a, a groove 59b, and a through hole 59c. The through hole 59a is provided at a position corresponding to the through hole 57a provided in the sealing plate 57. The groove 59b is provided in the surface of the rotor upper 59 so as to correspond to the through groove 57b provided in the sealing plate 57. The through hole 59c is provided at the center of the rotor upper 59.
The rotor base 61 has a groove 61a. The groove 61a is provided in the surface of the rotor base 61 to connect the through hole 59a provided in the peripheral portion of the rotor upper 59 and the through hole 59c provided at the center of the rotor upper 59 to each other.
By rotating the switching valve 63, the syringe channel 51c is connected to any one of the channels 13a, 35a, 37a, and 39a, and at the same time, the air vent channel 53b is also connected to at least any one of the channels 23a, 25a, 35b, 37b, and 39b.
The switching valve 63 shown in
As described above, the injection channel 17 provided in the reactor plate 1 is designed so that the liquid-tightness of the reaction well 5 is maintained in a state where there is no difference between the pressure in the injection channel 17 and the pressure in the reaction well 5. The reaction well air vent channel 19 is also designed so that the liquid-tightness of the reaction well 5 is maintained in a state where there is no difference between the pressure in the reaction well 5 and the pressure in the reaction well air vent channel 19. The main channel 13 constituting the reaction well channel, the liquid drain space 29 connected to the main channel 13, and the drain space air vent channel 23 can be hermetically sealed by switching of the switching valve 63. The wells 35, 37, and 39 are sealed with the septum 41 or the film 47. The channels 35a, 35b, 37a, 37b, 39a, and 39b connected to the wells 35, 37, and 39, respectively, can be hermetically sealed by switching the switching valve 63. One end of the air vent channel 53b is connected to the bellows 53 and, therefore, the air vent channel 53b is hermetically sealed. As described above, the wells and channels in the reactor plate 1 constitute a closed system. It is noted that even in a case where the reactor plate 1 does not have the bellows 53 and the air vent channel 53b is connected to the atmosphere outside the reactor plate 1, the air vent channel 53b can be cut off from the wells and the channels other than the air vent channel 53b provided in the reactor plate 1 by switching of the switching valve 63 and, therefore, the wells for containing a liquid and the channels for flowing a liquid can be hermetically sealed.
Although the reagent 45 is previously contained in the sample well 35 and the dilution water 49 is previously contained in the reagent well 37 of the reactor plate 1, it is possible to prevent the entry of foreign matter from the outside of the wells 35, 37 and the pollution of an environment outside the wells 35, 37 with the liquids 45, 49 contained in the wells 35, 37 even when the wells 35, 37 previously contain the liquids 45, 49 because the sample well channel 35a connected to the sample well 35 and the reagent well channel 37a connected to the reagent well 35 are sealed with the switching valve 63. Further, since the surface of the sealing plate 57 is covered with the PTFE layer having high airtightness, it is possible to prevent liquids 45, 49 contained in the wells 35, 37 from being vaporized and to store the unused reactor plate 1 for a long period of time.
The reaction processing apparatus includes a temperature control system for controlling the temperature of the reaction wells 5, a syringe driving unit 69 for driving the syringe 51, and a switching valve driving unit 71 for switching the switching valve 63.
A dispensing device having a sharp tip (not shown) is prepared, and the dispensing device is passed through the septum 41 provided on the sample well 35 to dispense, for example, 5 μL of a sample liquid into the sample well 35. After the completion of the dispensing of the sample liquid, the dispensing device is pulled out of the septum 41. By pulling the dispensing device out of the septum 41, a through hole formed in the septum 41 is closed due to the elasticity of the septum 41.
The syringe driving unit 69 is connected to the plunger 51b of the syringe 51, and the switching valve driving unit 71 is connected to the switching valve 63.
As shown in
The syringe 51 is allowed to slide to mix the sample liquid and the reagent 45 contained in the sample well 35. Then, for example, only 10 μL of the liquid mixture contained in the sample well 35 is sucked into the channel in the switching valve 63, the syringe channel 51c, and the syringe 51. At this time, the bellows 53 expands and contracts with changes in the volume of a gas contained in the sample well 35 because the sample well 35 is connected to the bellows 53 through the air vent channels 35e, 35d, and 35b, the switching valve 63, and the air vent channel 53b.
As shown in
As shown in
As shown in
As shown in
As shown in
As shown in
The switching valve 63 is returned to its initial state shown in
Alternatively, the wax 9 may be melted before the injection of the diluted mixture into the reaction wells 5 by heating the reaction wells 5 by the temperature control system 67 so that the diluted mixture is injected into the reaction wells 5 containing the melted wax 9. In this case, the diluted mixture injected into each of the reaction wells 5 immediately sinks below the wax 9, and is then mixed with the reagent 7 so that a reaction occurs. Even when the switching valve 63 is in the connection state shown in
As described above, by using the reactor plate 1, it is possible to perform reaction processing in a closed system. In addition, it is also possible to maintain the hermeticity of the reactor plate 1 before and after reaction processing.
According to the present embodiment, grooves for forming the channels 13, 15, 17, 19, 21, and 23 are provided in the channel base 11, but the present invention is not limited to this embodiment. For example, grooves for forming all or part of these channels may be provided in the surface of the well base 3.
On the well base 3, a channel spacer 73 is provided to cover a region where the reaction wells 5 are arranged. On the channel spacer 73, the channel base 11 and the channel cover 33 are further provided in this order. The channel spacer 73 is made of, for example, PDMS or silicone rubber. The thickness of the channel spacer 73 is, for example, from 0.5 to 5.0 mm. The channel spacer 73 has a projecting portion 75 projecting into each of the reaction wells 5. The projecting portion 75 is substantially trapezoidal in cross section. For example, the proximal end of the projecting portion 75 has a width of 1.0 to 2.8 mm, and the distal end of the projecting portion 75 has a width of 0.2 to 0.5 mm. That is, the distal end of the projecting portion 75 is narrower than the proximal end of the projecting portion 75. Further, the projecting portion 75 has a super-water-repellent surface. In this regard, it is noted that it is not always necessary to subject the surface of the projecting portion 75 to water-repellent treatment.
Further, in the channel spacer 73, an injection channel 77 is provided at a position corresponding to each of the projecting portions 75. The injection channel 77 is constituted from a through hole extending from the distal end of the projecting portion 75 to the surface of the channel spacer 73 where the projecting portion 75 is not provided. The injection channel 77 has an inner diameter of, for example, 500 μm. The opening of the injection channel 77 provided on the channel base 11 side is connected to the injection channel 17 provided in the channel base 11. It is noted that the reactor plate according to another embodiment of the present invention is different from the reactor plate described above with reference to
Further the channel spacer 73 has a reaction well air vent channel 79 constituted from a through hole. The reaction well air vent channel 79 is provided to allow the reaction well 5 to communicate with the reaction well air vent channel 19 provided in the channel base 11.
Although not shown in
According to the embodiment of the present invention shown in
Further, by placing the tip of the projecting portion 75 in the vicinity of the side wall of the reaction well 5 so that when a liquid passes through the injection channel 77 and is then discharged from the tip of the projecting portion 75, a droplet of the liquid formed at the tip of the projecting portion 75 can come into contact with the side wall of the reaction well 5, it is possible to inject the liquid into the reaction well 5 along the side wall of the reaction well 5, thereby making it possible to more reliably inject the liquid into the reaction well 5. However, the projecting portion 75 may be provided at a position which does not allow a droplet formed at the tip of the projecting portion 75 to be brought into contact with the side wall of the reaction well 5.
The reactor plate according to another embodiment of the present invention shown in
The reactor plate according to another embodiment of the present invention shown in
By providing the linear projecting portion 85 in such a manner that a space is left between the tip of the linear projecting portion 85 and the top surface of the reaction well 5 and between the tip of the linear projecting portion 85 and the side wall of the reaction well 5, it is possible to prevent a liquid contained in the reaction well 5 from reaching the top surface of the reaction well 5 through the side wall of the reaction well 5. The linear projecting portion 85 becomes particularly effective by subjecting the surface of at least the tip of the linear projecting portion 85 to water-repellent treatment.
The stepped portion 83 and the linear projecting portion 85 shown in
In each of these various embodiments described above with reference to
Although the present invention has been described above with reference to the various embodiments, the present invention is not limited to these embodiments. The shape, material, position, number, and size of each component and the channel configuration of the reactor plate in the above description are merely examples, and various changes can be made without departing from the scope of the present invention defined in claims.
For example, the bellows 53 connected to the air vent channel 53b may have another structure as long as it is a variable capacity member whose internal capacity is passively variable. Examples of such a bellows 53 having another structure include a bag-shaped one made of a flexible material and a syringe-shaped one.
The reactor plate according to the present invention does not always need to have a variable capacity member such as a bellows 53. Further, in a case where a liquid such as a reagent is not previously contained in the well 35, 37, or 39, the air vent channel thereof does not always need to partially have the channel 35e, 37e, or 39e constituted from a narrow hole.
In each of the above embodiments, the air vent channels 35b, 37b, and 39b, which communicate with the wells 35, 37, and 39 provided as sealed wells, are connected to the air vent channel 53b through the switching valve 63, but may be directly connected to the outside of the reactor plate or a variable capacity part such as a bellows 53. Each of the wells 35, 37, and 39 may be sealed by using an openable and closable cap.
In each of the above embodiments, the well base 3 is constituted from one component, but may be constituted from two or more components.
The reagent contained in the reaction well 5 may be a dry reagent.
It is noted that the sample well 35 and the reaction well 5 do not always need to previously contain a reagent.
In each of the above embodiments, the reagent well 37 contains dilution water 49, but may contain a reagent instead of the dilution water 49.
The well base 3 may further have a gene amplification well for carrying out gene amplification reaction. For example, the empty reagent well 37 may be used as a gene amplification well.
By previously placing a reagent for gene amplification reaction in the reaction well 5, it is possible to carry out gene amplification reaction in the reaction well 5.
In a case where a liquid to be introduced into the main channel 13 contains a gene, a probe which reacts with the gene may be previously placed in the reaction well 5.
In each of the above embodiments, the syringe 51 is placed on the switching valve 63. However, the position of the syringe 51 is not limited to a position on the switching valve 63, and the syringe 51 may be placed at any position.
Further, the reactor plate according to the present invention does not always need to have the syringe 51, and a syringe provided outside the reactor plate may be used to discharge and suck a liquid or a gas.
Further, in each of the embodiments described above, only one rotary switching valve 63 is provided as a switching valve. However, the reactor plate according to the present invention may have a plurality of switching valves.
Further, in each of the embodiments described above, the surface of the sealing plate 57 is covered with the PTFE layer formed thereon. However, as shown in
As in the case of the PTFE layer formed on the surface of the sealing plate 57, the PTFE member 87 placed on the surface of the sealing plate 57 can reliably seal the ends of the channels. Therefore, it is possible to prevent a liquid contained in the reaction well and/or the sealed well from being vaporized. This makes it possible to store the unused reactor plate 1 for a long period of time.
The layer formed on the surface of the sealing plate 57 or the member placed on the surface of the sealing plate 57 may be made of a fluorine resin other than PTFE (e.g., PCTFE).
The sealing plate 57 does not always need to have the PTFE layer formed on the surface thereof.
Further, the sealing plate 57 may be made of a material other than the elastic material such as a fluorine resin (e.g., PTFE or PCTFE).
In each of the above embodiments, a liquid filling the metering channel 15 is injected into the reaction well 5 through the injection channel 17 by applying a pressure to the inside of the main channel 13 after air purge, but the reaction processing method according to the present invention is not limited to such a method. For example, a liquid filling the metering channel 15 may be injected into the reaction well 5 through the injection channel 17 by creating a negative pressure in the reaction well air vent channel 21 and then in the reaction well 5. In this case, it is necessary to change the channel configuration of the reactor plate so that a negative pressure can be created in the reaction well air vent channel 21 by using the syringe 51. Alternatively, another syringe may be additionally prepared. In this case, a positive pressure is created in the main channel 13 and a negative pressure is created in the reaction well 5 to inject the liquid into the reaction well 5.
In each of the above embodiments, one main channel 13 is provided, and all the metering channels 15 are connected to the main channel 13. However, the channel configuration of the reactor plate according to the present invention is not limited thereto. For example, a plurality of main channels may be provided. In this case, one or more metering channels may be connected to each of the main channels.
In the above specific example of the channels in the reactor plate according to the present invention, the main channel can be hermetically sealed. In this regard, the main channel may be hermetically sealed by, for example, allowing both ends of the main channel to be openable and closable. The phrase “allowing both ends of the main channel to be openable and closable” includes a case where each end of the main channel is connected to another space, and the end of the space located on the opposite side from the main channel is openable and closable. In the case of each of the above embodiments, such ‘another space’ corresponds to, for example, the channel 13a, the liquid drain space 29, the drain space air vent channel 23, or the channel 23a.
In the above specific example of the channels in the reactor plate according to the present invention, the reaction well air vent channel can be hermetically sealed. In this regard, the reaction well air vent channel may be hermetically sealed by, for example, allowing the end of the reaction well air vent channel located on the opposite side from the reaction well to be openable and closable. The phrase “allowing the end of the reaction well air vent channel located on the opposite side from the reaction well to be openable and closable” includes a case where the end of the reaction well air vent channel located on the opposite side from the reaction well is connected to another space, and the end of the space located on the opposite side from the reaction well air vent channel is openable and closable. In the case of each of the above embodiments, such another space' corresponds to, for example, the air drain space 31, the drain space air vent channel 25, or the channel 25a.
In the case of such an aspect, a liquid is introduced into the main channel and the metering channels, and next, the liquid is purged from the main channel, and further, the liquid remaining in the metering channels is injected into the reaction wells, and thereafter, both ends of the main channel and one end of the reaction well air vent channel located on the opposite side from the reaction well are closed to hermetically seal the main channel and the reaction well air vent channel.
The present invention can be applied to measurements of various chemical and biochemical reactions.
| Number | Date | Country | Kind |
|---|---|---|---|
| 2007-106507 | Apr 2007 | JP | national |
| Filing Document | Filing Date | Country | Kind | 371c Date |
|---|---|---|---|---|
| PCT/JP2008/053642 | 2/29/2008 | WO | 00 | 10/12/2009 |
