JP2007040969A - Biochemical reaction cassette - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a biochemical reaction cassette having a structure for uniforming the flow of a liquid in a reaction chamber by adding thereto a simple structure. <P>SOLUTION: This biochemical reaction cassette is one for target nucleic acid detection, having: a probe fixing area for target nucleic acid detection, and having, in the probe fixing area; a reaction chamber for putting a specimen to reaction; an inlet port for inletting the specimen into the reaction chamber; and a discharge port for discharging the specimen out of the reaction chamber. This reaction cassette is equipped with a fluid resistance part for reducing a flow-path cross-section area in a flow path comprising the inlet port, the reaction chamber, and the discharge port, and is characterized in that the flow of the fluid in the reaction chamber is controlled by the resistance part. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  The present invention includes a probe carrier such as a DNA microarray that can be suitably used when a test for the presence or absence of a gene derived from a pathogenic bacterium in a specimen such as blood is performed and used as a material for determining the health condition of a test subject. It relates to a biochemical reaction cassette. More specifically, the present invention relates to the structure of a biochemical reaction cassette for making the flow velocity of the liquid flowing in the reaction chamber at least uniform.

  Many methods using a hybridization reaction using a probe carrier typified by a DNA microarray have been proposed as methods for rapidly and accurately analyzing the base sequence of a nucleic acid and detecting a target nucleic acid in a nucleic acid sample. A DNA microarray is a probe having a base sequence complementary to a target nucleic acid fixed on a solid phase such as a bead or a glass plate at a high density. The detection of a target nucleic acid using this probe is generally as follows. It has a process.

As a first step, a target nucleic acid is amplified by an amplification method typified by a PCR method. Specifically, first, first and second primers are added to a nucleic acid sample, and a temperature cycle is applied. The first primer specifically binds to a part of the target nucleic acid, and the second primer specifically binds to a part of the nucleic acid complementary to the target nucleic acid. When the double-stranded nucleic acid containing the target nucleic acid is bound to the first and second primers, the double-stranded nucleic acid containing the target nucleic acid is amplified by the extension reaction. After the double-stranded nucleic acid sufficiently containing the target nucleic acid is amplified, a third primer is added to the nucleic acid sample and subjected to a temperature cycle. The third primer is labeled with an enzyme, a fluorescent substance, a luminescent substance, or the like, and specifically binds to a part of the nucleic acid complementary to the target nucleic acid. When a nucleic acid complementary to the target nucleic acid and the third primer are bound, the target nucleic acid labeled with an enzyme, a fluorescent substance, a luminescent substance or the like is amplified by an extension reaction. As a result, a labeled target nucleic acid is produced when the target nucleic acid is contained in the nucleic acid sample, and a labeled target nucleic acid is not produced when the target nucleic acid is not contained in the nucleic acid sample.

  As a second step, the nucleic acid sample is brought into contact with a DNA microarray and subjected to a hybridization reaction with a probe of the DNA microarray. When there is a target nucleic acid complementary to the probe, the probe and the target nucleic acid form a hybrid.

  As a third step, the target nucleic acid is detected. Whether the probe and the target nucleic acid form a hybrid can be detected by a labeling substance of the target nucleic acid, thereby confirming the presence or absence of a specific base sequence.

  DNA microarrays utilizing this hybridization reaction are expected to be applied to medical diagnosis for identifying pathogenic bacteria and genetic diagnosis for examining patient constitutions. However, the nucleic acid amplification, hybridization, and detection steps are often performed by individual devices, and the work is complicated, requiring a considerable amount of time for diagnosis. In particular, in a configuration in which a hybridization reaction is performed on a slide glass, the probe fixing surface is exposed, and there is a possibility that the probe may be lost or contaminated when a finger touches the slide glass. Must be done carefully. In order to solve these problems, several biochemical reaction cassette structures have been proposed in which a DNA microarray is provided in a reaction chamber, a hybridization reaction is performed in the reaction chamber, and detection is possible thereafter.

  Japanese National Publication No. 10-505410 discloses a structure and a manufacturing method for forming a cavity. Japanese Patent Application Laid-Open Nos. 2003-302399 and 2004-093558 disclose a chamber structure for preventing bubbles from remaining during the initial filling of the liquid. Furthermore, Japanese Patent Laid-Open No. 2002-243748 discloses a structure for forming a uniform spread and flow of liquid.

In such a biochemical reaction cassette structure, the volume of the reaction chamber is as small as about several tens of μL, the height of the reaction chamber is low, and the reaction chamber has a planar space. As a result, the amount of reagents to be used is small, and a laminar flow is generated in the reaction chamber. Furthermore, in order to increase the efficiency of the hybridization reaction between the probe on the solid phase and the target nucleic acid, the liquid is preferably stirred in the reaction chamber. As the simplest method, there is a method in which the liquid in the reaction chamber is swung by pushing and pulling the liquid at the inlet.
Japanese National Patent Publication No. 10-505410 JP 2003-302399 A JP 2004-093558 JP JP 2002-243748 A

  An example of a biochemical reaction cassette is shown in FIGS. This biochemical reaction cassette has a substrate 111 and a housing 112. It is assumed that the reaction chamber 103 of the biochemical reaction cassette 110 is filled with a liquid. When the liquid is further fed from the inlet 106, the flow velocity 122 near the center becomes faster when the flow velocity 122 near the center of the reaction chamber 103 is compared with the flow velocity 121 and 123 near the end. Therefore, when the liquid in the reaction chamber 103 is swung by pushing and pulling the liquid at the inlet 106 or the outlet 107, the frequency of contact between the probe on the solid phase and the target nucleic acid is near the center and near the end of the reaction chamber 103. Then it will be different. In addition, after the hybridization reaction is completed, a washing solution is flowed into the reaction chamber 103 in order to remove unreacted nucleic acids. Also at this time, the flow rate 122 near the center of the reaction chamber 103 is different from the flow rates 121 and 123 near the ends, so that the ratio of the unreacted nucleic acid to be removed and the target nucleic acid reacted with the probe on the solid phase are peeled off. Probabilities are different. As a result, the brightness at the time of detection varies depending on the position of the probe, which may adversely affect the diagnosis result.

  In the configuration of Japanese Patent Publication No. 10-505410, a laminar flow is generated in the cavity, but the difference in flow velocity between the center and the end of the cavity cannot be resolved. In addition, in the configurations of Japanese Patent Application Laid-Open Nos. 2003-302399 and 2004-093558, the liquid spreads uniformly in the chamber at the initial filling of the liquid, but the flow rate in the chamber is increased when the liquid is filled in the chamber. There is no effect of homogenization. Furthermore, the configuration of Japanese Patent Laid-Open No. 2002-243748 has a complicated structure and has a limit in reducing the cost for manufacturing the cassette.

  The objective of this invention is providing the biochemical reaction cassette which has the structure which can equalize the flow of the liquid in a reaction chamber by addition of a simple structure.

The biochemical reaction cassette of the present invention has a fixed region of a probe for detecting a target nucleic acid, a reaction chamber for causing a sample to react with the probe fixed region, and an inlet for injecting the sample into the reaction chamber; A biochemical reaction cassette for detecting a target nucleic acid having a discharge port for discharging a sample from the reaction chamber,
A flow path configured to include the inlet, the reaction chamber, and the discharge port includes a fluid resistance portion that reduces a cross-sectional area of the flow path, and the fluid resistance portion allows a fluid flow in the reaction chamber to flow. A biochemical reaction cassette characterized by being controlled.

The biochemical reaction device of the present invention includes a probe fixing region for detecting a target nucleic acid, a reaction chamber for reacting a sample in the probe fixing region, and an inlet for injecting the sample into the reaction chamber. A biochemical reaction apparatus for detecting a target nucleic acid having a discharge port for discharging a sample from the reaction chamber,
A flow path configured to include the inlet, the reaction chamber, and the discharge port includes a fluid resistance portion that reduces a cross-sectional area of the flow path, and the fluid resistance portion allows a fluid flow in the reaction chamber to flow. It is a biochemical reactor characterized by being controlled.

  Another aspect of the biochemical reaction cassette of the present invention includes a reaction chamber having a reaction site for performing a biochemical reaction, an inlet for injecting a sample into the reaction chamber, and between the inlet and the reaction chamber. And a buffer chamber for controlling a flow rate of the sample supplied to the reaction chamber.

  Another aspect of the biochemical reaction device of the present invention includes a reaction chamber having a reaction site for performing a biochemical reaction, an inlet for injecting a sample into the reaction chamber, and between the inlet and the reaction chamber. And a buffer chamber for controlling a flow rate of the sample supplied to the reaction chamber.

  According to the present invention, the member for reducing the cross-sectional area of the flow path is disposed in the flow path having the inlet, the reaction chamber, and the outlet, and the buffer chamber is provided. The flow rate is controlled, and the flow rate in the reaction chamber can be made uniform. The fluid resistance member can be configured by a slot portion whose ceiling portion is lower than the reaction chamber, a protrusion from the ceiling portion, a columnar member, a partition member having a large number of communication holes, and the like.

  Embodiments according to the present invention will be described below with reference to the drawings.

Example 1
FIG. 1 is a perspective view showing the structure when the biochemical reaction apparatus according to the first embodiment of the present invention is a cassette type. FIGS. 2A and 2B are a plan view and a cross-sectional view showing the structure of the biochemical reaction cassette according to the first embodiment of the present invention. FIG. 3 is a plan view showing the state of the liquid flowing in the biochemical reaction cassette according to the first embodiment of the present invention.

  First, the structure of the cassette will be described. The cassette 10 has a configuration in which a glass substrate 11 and a housing 12 made of polycarbonate are joined. In addition, the joining form of the housing | casing part with respect to a glass substrate is not limited to the illustrated example, A various form can be taken. The material of the housing 12 is not limited to polycarbonate, but may be plastic other than polycarbonate, glass, rubber, silicon, and a composite material composed of at least two of these. On the joint surface between the glass substrate 11 and the housing 12, a recess having a predetermined cross-sectional shape is provided in the housing 12, and the first buffer chamber 1 and the first slot portion 2 are provided between the glass substrate 11 and the housing 12. The reaction chamber 3, the second slot portion 4, and the second buffer chamber 5 are formed. The bottom surface of each space constituting these portions formed between the glass substrate 11 and the housing 12 is formed of a part of the surface of the glass substrate 11. Here, since each space which becomes a buffer chamber, a slot part, and a reaction chamber is provided in the housing | casing 12, the bottom face of each space has comprised the same plane. However, a configuration in which part or all of the buffer chamber, the slot portion, and the reaction chamber are provided on the glass substrate 11 and the bottom surfaces of the spaces do not form the same plane may be employed.

  The ceiling portions 2a and 4a of the slot portions made of the protruding members are lower than the ceiling portions of the buffer chamber and the reaction chamber, and the upper portions of the slot portions form the partition portions of the buffer chamber and the reaction chamber.

  A probe fixing region 13 is provided on a part of the surface of the glass substrate 11 on the bottom surface of the reaction chamber 3. When the target nucleic acid is contained in the liquid filled in the reaction chamber 3, The probe in the probe fixing region 13 reacts. The combination of the target nucleic acid and the probe can be selected according to the detection purpose such as when both of them are DNA.

  The liquid is injected into the first buffer chamber 1 from the injection port 6, passes through the first slot portion 2, the reaction chamber 3, the second slot portion 4 and the second buffer chamber 5 in this order, and is connected to the second buffer chamber 5. It is discharged out of the cassette 10 through the discharge port 7. That is, a liquid flow path is formed from these portions.

  When the dimensions of each space are expressed by coordinates X (width) × Y (length) × Z (height: distance from each bottom surface to the ceiling) in FIG. 1, the first buffer chamber 1 is 10 × 2 ×. 0.5 mm, the first slot 2 is 10 × 1 × 0.1 mm, and the reaction chamber 3 is 10 × 10 × 0.5 mm. Further, the second slot portion 4 is 10 × 1 × 0.1 mm, and the second buffer chamber 5 is 10 × 2 × 0.5 mm. However, each space is not limited to this size, and the heights of the ceilings of the first slot portion 2 and the second slot portion 4 are the same as those of the first buffer chamber 1, the reaction chamber 3, and the second buffer chamber 5. The height may be lower than the height and can provide a desired slot function.

  Further, the ceiling portion of the reaction chamber 3 is formed as a flat surface having a constant height with respect to the bottom surface (the height with respect to the bottom surface is constant throughout the reaction chamber). It can be appropriately changed as necessary. Moreover, the shape of the ceiling part of the slot parts 2 and 4 is not one plane (the height with respect to the bottom face is constant over the entire slot part), and can be changed as needed. However, considering the simplification of the structure of the cassette and the simplification of the manufacturing process, the illustrated structure is one of preferred embodiments.

  On the other hand, the heights of the ceiling portions 1a, 3a, and 5a of the first buffer chamber 1, the reaction chamber 3, and the second buffer chamber 5 need not necessarily match within the range in which the intended function of each portion can be obtained. Absent. Further, the heights of the ceiling portions 2a and 4a of the first slot portion 2 and the second slot portion 4 do not necessarily need to match.

  In the illustrated example, the width (length in the X-axis direction in FIG. 1) of each buffer chamber, each slot portion, and the reaction chamber with respect to the flow path direction is the same, but these widths are not the same between the portions. Not necessary. However, it is preferable to match the widths of these portions from the standpoint of not complicating the manufacturing process and obtaining a uniform flow rate more effectively in the reaction chamber.

  On the other hand, as shown in the drawing, the height of the ceiling of the buffer chamber is constant throughout the buffer chamber, and the height of the ceiling of the reaction chamber is also preferably constant throughout the reaction chamber in order to obtain a uniform flow. . The same applies to Example 2 described later.

  Next, a method for detecting a target nucleic acid using a cassette will be described. First, a nucleic acid sample is prepared, and the target nucleic acid is amplified according to the method described above as necessary. When the target nucleic acid is present in the nucleic acid sample, a target nucleic acid labeled with a fluorescent substance is generated in the amplification step. Here, the labeling substance is a fluorescent substance, but it may be a luminescent substance or an enzyme. The solution of the nucleic acid sample is injected into the cassette 10 from the injection port 6 using a liquid injection means (not shown). When the solution is filled in the first buffer chamber 1, the first slot portion 2, the reaction chamber 3, the second slot portion 4 and the second buffer chamber 5, the solution is heated and the target nucleic acid in the solution and the probe fixing region 13 are over. The hybridization reaction with the probe proceeds. At this time, the solution is reciprocated in the reaction chamber 3 and stirred in order to increase the frequency with which the target nucleic acid in the solution under temperature conditions necessary for the hybridization reaction contacts the probe on the probe fixing region 13. At this time, the first buffer chamber 1, the first slot portion 2, the reaction chamber 3, the second slot portion 4 and the second buffer chamber 5 are always filled with the solution.

  When a nucleic acid sample solution is fed from the inlet 6 side for stirring, a flow as shown in FIG. If there is no resistance in the passage of the liquid, the solution flows in a substantially straight line from the inlet 6 to the outlet 7, but the first slot 2 becomes a resistance, so that the solution spreads over the entire first buffer chamber 1. Streams 21, 22 and 23 are generated. As a result, the pressure of the entire first buffer chamber 1 rises, and an equal pressure is applied to the first slot portion 2. Therefore, the solution pushed out from the first slot portion 2 has a uniform flow rate 24, 25 and 26 in the reaction chamber 3. When the solution of the nucleic acid sample necessary for stirring is fed from the inlet 6, the solution of the nucleic acid sample is then fed from the outlet 7 side. Based on the same principle as when the solution is fed from the inlet 6 side, as shown in FIG. 3 (b), uniform flow rates 34, 35 in the reaction chamber 3 according to the solution flows 21, 22, 23, etc. And 36 etc. occur. After the amount of nucleic acid sample solution necessary for stirring is fed from the discharge port 7, the solution of the nucleic acid sample is again fed from the injection port 6. After that, the liquid feeding from the discharge port 7 and the liquid feeding from the injection port 6 are repeated, and the solution in the reaction chamber 3 is stirred. Since a uniform flow rate is generated in the reaction chamber 3, the probe present at any position in the probe fixing region 13 has the same frequency of contact with the target nucleic acid in the nucleic acid sample, and the hybridization reaction depends on the position of the probe fixing region 13. No difference in the degree of progress.

  If the nucleic acid sample is still filled in the reaction chamber 3 or if the nucleic acid sample remains attached to the wall surface of the reaction chamber 3, the background at the time of detection is lowered, so these need to be washed away. During cleaning, the cleaning solution continues to flow from the inlet 6 for a certain period of time, but at this time, uniform flow rates 24, 25, 26, etc. are generated in the reaction chamber 3 as shown in FIG. The principle that the flow rate of the washing solution becomes uniform is the same principle that the flow rate of the solution of the nucleic acid sample fed during stirring becomes uniform. Since the flow rate of the cleaning liquid becomes uniform, the degree to which the nucleic acid sample attached to the wall surface of the reaction chamber 3 is washed out becomes constant regardless of the position of the reaction chamber 3. Furthermore, the target nucleic acid bound to the probe may be peeled off by the flow of the washing solution. However, even when a part of the target nucleic acid is peeled off from the probe fixing region, the flow rate of the washing liquid becomes uniform, so that the probability that the target nucleic acid is peeled off is constant regardless of the position of the probe fixing region 13. Therefore, it is possible to reduce variation in fluorescence luminance when the presence or absence of the target nucleic acid labeled with a fluorescent substance is detected by an optical system (not shown) after washing.

  As explained above, since the flow rate of the solution of the nucleic acid sample and the washing solution flowing in the reaction chamber 3 becomes uniform, the binding ratio between the probe and the target nucleic acid becomes constant regardless of the position, and the detection accuracy is improved. Can be improved.

(Example 2)
FIG. 4 is a perspective view showing the structure of a biochemical reaction cassette according to the second embodiment of the present invention. FIGS. 5A and 5B are a plan view and a cross-sectional view showing the structure of the biochemical reaction cassette according to the second embodiment of the present invention. FIG. 6 is a plan view showing the state of the liquid flowing in the biochemical reaction cassette according to the second embodiment of the present invention.

  First, the structure of the cassette will be described. The cassette 60 has a configuration in which a glass substrate 61 and a housing 62 are joined. On the joint surface between the glass substrate 61 and the housing 62, a recess having a predetermined cross-sectional shape is provided in the housing 62, and the buffer chamber 51, the slot portion 52, and the reaction chamber 53 are provided between the glass substrate 61 and the housing 62. And the taper part 54 is formed. The bottom surface of each space constituting these portions formed between the glass substrate 61 and the casing 62 is formed of a part of the surface of the glass substrate 61. Here, since each space which becomes a buffer chamber, a slot part, and a reaction chamber is provided in the housing | casing 62, the bottom face of each space has comprised the same plane. However, a configuration in which part or all of the buffer chamber, the slot portion, and the reaction chamber are provided on the glass substrate 61 and the bottom surfaces of the spaces do not form the same plane may be employed.

  The ceiling portion of the slot portion 52 is lower than the ceiling portions of the buffer chamber 51 and the reaction chamber 53, and the upper portion of the slot portion 52 forms a partition portion for the buffer chamber 51 and the reaction chamber 53.

  A probe fixing region 63 is provided on the surface of the substrate 61 which is the wall surface of the reaction chamber 53, and the target nucleic acid contained in the solution filled in the reaction chamber 53 reacts with the probe in the probe fixing region 63. ing. The liquid is injected into the buffer chamber 51 through the injection port 56, passes through the slot portion 52 and the reaction chamber 53, and is discharged out of the cassette 60 through the discharge port 57 connected to the reaction chamber 53. When the dimensions of each space are expressed by the coordinates X (width) × Y (length) × Z (height) in FIG. 4 as in the first embodiment, the buffer chamber 51 is 10 × 2 × 0.5 mm, slot. The part 52 is 10 × 1 × 0.1 mm, and the reaction chamber 53 is 10 × 13 × 0.5 mm. Further, a tapered portion 54 having a 45 ° inclination with respect to the Y direction is provided on the side wall surface of the reaction chamber 53. However, each space is not limited to this dimension, and the height of the slot portion 52 is lower than the height of the buffer chamber 51 and the reaction chamber 53 and may be within a range in which the intended slot function can be obtained. . In addition, the heights of the buffer chamber 51 and the reaction chamber 53 do not need to match within a range in which the intended function of each part can be obtained.

  In the illustrated example, the portions other than the tapered portion of the reaction chamber, the buffer chamber, and the slot portion are formed to have the same width in the flow direction, but these widths may not be the same between the portions. However, it is preferable to match the widths of these portions from the standpoint of not complicating the manufacturing process and obtaining a uniform flow rate more effectively in the reaction chamber.

  Next, a method for detecting a target nucleic acid using a cassette will be described. First, a nucleic acid sample is prepared, and the target nucleic acid is amplified according to the method described above as necessary. When the target nucleic acid is present in the nucleic acid sample, a target nucleic acid labeled with a fluorescent substance is generated in the amplification step. Here, the labeling substance is a fluorescent substance, but it may be a luminescent substance or an enzyme. The nucleic acid sample solution is injected into the cassette 60 from the injection port 56 using liquid injection means (not shown). When the solution is filled in the buffer chamber 51, the slot portion 52, and the reaction chamber 53, the solution is heated, and the hybridization reaction between the target nucleic acid in the solution and the probe on the probe fixing region 63 proceeds. At this time, in order to increase the frequency with which the target nucleic acid in the solution under the temperature condition necessary for the hybridization reaction contacts the probe on the probe fixing region 63, the solution is reciprocated in the reaction chamber 53 and stirred. At this time, the buffer chamber 51, the slot 52 and the reaction chamber 53 are always filled with the solution. When a solution of a nucleic acid sample is fed from the inlet 56 side for stirring, a flow as shown in FIG. If there is no resistance in the passage of the liquid, the solution flows in a substantially straight line from the inlet 56 to the outlet 57. However, since the slot 52 becomes a resistance, the solution flows 71, 72 and 73 or the like occurs. As a result, the pressure in the entire buffer chamber 51 increases, and an equal pressure is applied to the slot portion 52. Therefore, the solution pushed out from the slot 52 has uniform flow rates 74, 75, and 76 in the reaction chamber 53. When a solution of the nucleic acid sample necessary for stirring is fed from the inlet 56, the solution of the nucleic acid sample is fed from the outlet 57 side this time. When the solution is fed from the discharge port 57 side, there is no resistance due to the slot portion, and as a result, differences in the flow rates 81, 82, and 83 in the reaction chamber 53 occur as shown in FIG. Further, flows 84, 85, 86 and the like are generated in front of the slot portion 52.

  After the amount of the nucleic acid sample solution necessary for stirring is fed from the outlet 57, the solution of the nucleic acid sample is again fed from the inlet 56. After this, liquid feeding from the discharge port 57 and liquid feeding from the injection port 56 are repeated, and the solution in the reaction chamber 53 is stirred. Since the state of the flow varies depending on the solution feeding direction, the stirring efficiency in the reaction chamber 53 is improved. Thereby, the concentration distribution of the target nucleic acid in the solution filled in the reaction chamber 53 is always constant regardless of the position, and the difference in the degree of progress of the hybridization reaction depending on the position of the probe fixing region 63 is eliminated.

  If the nucleic acid sample is still filled in the reaction chamber 53 or if the nucleic acid sample remains attached to the wall surface of the reaction chamber 53, the background at the time of detection is lowered, so these need to be washed away. During cleaning, the cleaning liquid continues to flow from the inlet 56 for a certain period of time, but at this time, uniform flow rates 74, 75, 76, etc. are generated in the reaction chamber 53 as shown in FIG. The principle that the flow rate of the washing solution becomes uniform is the same principle that the flow rate of the solution of the nucleic acid sample fed from the inlet 56 side during stirring becomes uniform. Since the flow rate of the cleaning liquid becomes uniform, the degree to which the nucleic acid sample attached to the wall surface of the reaction chamber 53 is washed out becomes constant regardless of the position of the reaction chamber 53. Furthermore, the target nucleic acid bound to the probe may be peeled off by the flow of the washing solution. However, even when a part of the target nucleic acid is peeled off from the probe fixing region, the flow rate of the cleaning liquid becomes uniform, so the probability that the target nucleic acid is peeled off is constant regardless of the position of the probe fixing region 63. Therefore, it is possible to reduce variation in fluorescence luminance when the presence or absence of the target nucleic acid labeled with a fluorescent substance is detected by an optical system (not shown) after washing.

  In Example 2, since the flow state varies depending on the liquid feeding direction in the reaction chamber 53, the stirring efficiency of the target nucleic acid in the reaction chamber 53 is improved, and the progress of the hybridization reaction is constant regardless of the position. It becomes. In addition, since the flow rate of the washing liquid is uniform, the degree to which the target nucleic acid is washed out is constant regardless of the position, so that the detection accuracy can be improved.

  As described above, as described in Examples 1 and 2, by providing a buffer chamber via the slot at least upstream of the reaction chamber, when the liquid flows from the inlet to the outlet, Variation in flow rate is suppressed. That is, the liquid sample can be uniformly supplied to the probe region. That is, when the liquid is going to flow from the inlet to the outlet while the buffer chamber, the slot portion, and the reaction chamber are filled with the liquid, the slot portion becomes a resistance, and the liquid supplied to the buffer chamber is spread over the entire buffer chamber. It tries to flow to spread. Thereby, the pressure in the buffer chamber rises as a whole, and the liquid is pushed out from the slot portion to the reaction chamber. At this time, the force with which the pressure in the buffer chamber pushes the liquid out of the slot portion is uniform in the width direction of the slot portion, so that the flow rate in the reaction chamber is made uniform.

  On the other hand, in the configuration in which the buffer chamber is provided on the downstream side via the slot portion in addition to the upstream side of the reaction chamber, the flow rate in the reaction chamber becomes uniform even if the liquid is swung for stirring.

  Furthermore, in the configuration in which the buffer chamber is provided only through the slot portion on the upstream side of the reaction chamber (the configuration in which the buffer chamber and the slot portion are not provided on the downstream side), the liquid is allowed to flow from the inlet for the reason described above. Sometimes the flow rate in the reaction chamber is uniform. Furthermore, when the liquid is allowed to flow from the discharge port, a difference in flow velocity occurs in the width direction in the reaction chamber. Therefore, when the liquid is oscillated for stirring, since the state is different between the forward direction and the reverse direction of the flow, the stirring efficiency in the reaction chamber can be improved.

  With the configuration as described above, it is possible to provide a biochemical reaction cassette that has a volume that can secure a necessary liquid in the buffer chamber and the reaction chamber, and that has improved flow rate uniformity and stirring efficiency by the effect of the slot portion. .

  In addition, the slot part in said Example 1 and 2 is comprised as a flow path with a ceiling part lower than a buffer chamber and a reaction chamber. However, it is also possible to form a partition portion having a slot function by providing projections extending from the ceiling portion to the bottom surface in a direction toward the bottom surface so as to extend to a position having a predetermined distance from the bottom surface.

(Example 3)
FIG. 7 is a perspective view showing the structure of a biochemical reaction cassette according to the third embodiment of the present invention. FIGS. 8A and 8B are a plan view and a cross-sectional view showing the structure of a biochemical reaction cassette according to the third embodiment of the present invention.

  The structure of the cassette is such that the first slot portion 2 in the first embodiment is a first columnar member 14 and the second slot portion 4 is a second columnar member 15. The liquid passes through the interval formed by the first columnar members 14 and the interval formed by the second columnar members 15. Other structures are the same as those in the first embodiment.

  The cassette according to this embodiment is manufactured by integrally molding the casing 12. However, the manufacturing method is not limited to this, and the first columnar member 14 and the second columnar member 17 may be bonded and fixed to the casing 112 shown in FIGS. 11 and 12.

  With the configuration as described above, the first columnar member 14 and the second columnar member 15 reduce the flow path cross-sectional area, and exhibit the same effects as the first slot portion 2 and the second slot portion 4. That is, since the nucleic acid sample solution flowing in the reaction chamber 3 and the flow rate of the washing liquid become uniform, the binding ratio between the probe and the target nucleic acid becomes constant regardless of the position, and the detection accuracy can be improved. .

Example 4
FIG. 9A is a perspective view showing the structure of a biochemical reaction cassette according to the fourth embodiment of the present invention. FIGS. 10A and 10B are a plan view and a cross-sectional view showing the structure of a biochemical reaction cassette according to the fourth embodiment of the present invention.

  The structure of the cassette is such that the first slot portion 2 in the first embodiment is a first partition member 16 and the second slot portion 4 is a second partition member 17. The first partition member 16 and the second partition member 17 have a large number of communication holes through which liquid can pass in the Y direction shown in FIG. Other structures are the same as those in the first embodiment.

  An example of the cassette manufacturing method of the present embodiment is shown in FIG. Groove portions 91 and 92 are provided in the housing 12, and the first partition member 16 and the second partition member 17 are fitted in the groove portions 91 and 92, respectively, and are sandwiched between the housing 12 and the glass substrate 11. However, the manufacturing method is not limited to this, and the first partition member 16 and the second partition member 17 may be bonded and fixed to the housing 112 shown in FIGS.

  With the above-described configuration, the first partition member 16 and the second partition member 17 reduce the flow path cross-sectional area, and exhibit the same effects as the first slot portion 2 and the second slot portion 4. That is, since the nucleic acid sample solution flowing in the reaction chamber 3 and the flow rate of the washing liquid become uniform, the binding ratio between the probe and the target nucleic acid becomes constant regardless of the position, and the detection accuracy can be improved. .

It is a perspective view explaining the structure of the biochemical reaction cassette which concerns on the 1st Example of this invention. It is the top view and sectional drawing explaining the structure of the biochemical reaction cassette which concerns on the 1st Example of this invention. It is a top view explaining the flow of the liquid in the biochemical reaction cassette which concerns on 1st Example of this invention. FIG. 4 is a perspective view for explaining the structure of a biochemical reaction cassette according to the second embodiment of the present invention. FIG. 5 is a plan view and a cross-sectional view for explaining the structure of the biochemical reaction cassette according to the second embodiment of the present invention. FIG. 6 is a plan view for explaining the flow of the liquid in the biochemical reaction cassette according to the second embodiment of the present invention. It is a perspective view explaining the structure of the biochemical reaction cassette which concerns on the 3rd Example of this invention. It is the top view and sectional drawing explaining the structure of the biochemical reaction cassette which concerns on the 3rd Example of this invention. It is a perspective view explaining the structure of the biochemical reaction cassette which concerns on the 4th Example of this invention. It is the top view and sectional drawing explaining the structure of the biochemical reaction cassette which concerns on the 4th Example of this invention. FIG. 11 is a perspective view for explaining the structure of a conventional biochemical reaction cassette. FIG. 12 is a plan view and a sectional view for explaining the structure of a conventional biochemical reaction cassette.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 1st buffer chamber 1a, 2a, 3a, 4a, 5a Ceiling part 2 1st slot part 3, 53, 103 Reaction chamber 4 2nd slot part 5 2nd buffer chamber 6, 56, 106 Inlet 7, 57, 107 Discharge port 10, 60, 110 Cassette 11, 61, 111 Glass substrate 12, 62, 112 Case 13, 63, 113 Probe fixing region 14 First columnar member 15 Second columnar member 16 First partition member 17 Second partition member 51 Buffer chamber 52 Slot portion 54 Tapered portion

Claims (17)

A target nucleic acid detection probe fixing region is provided, a reaction chamber for reacting a sample in the probe fixing region, an inlet for injecting the sample into the reaction chamber, and a sample is discharged from the reaction chamber A biochemical reaction cassette for detecting a target nucleic acid having an outlet for
A flow path configured to include the inlet, the reaction chamber, and the discharge port includes a fluid resistance portion that reduces a cross-sectional area of the flow path, and the fluid resistance portion allows a fluid flow in the reaction chamber to flow. A biochemical reaction cassette characterized by being controlled. The fluid resistance portion is a slot portion formed with a partition portion extending in a vertical direction from a ceiling portion or / and a bottom surface of the flow path,
The biochemical reaction cassette according to claim 1, wherein a buffer chamber separated from the reaction chamber is provided adjacent to the reaction chamber by the partition portion.   The biochemical reaction cassette according to claim 1 or 2, wherein a height of a ceiling portion with respect to the bottom surface of the reaction chamber is constant throughout the reaction chamber.   The biochemical reaction cassette according to claim 2 or 3, wherein the slot portion has a ceiling portion that forms the partition portion, and a height of the ceiling portion with respect to the bottom surface is constant throughout the slot portion.   The biochemical reaction cassette according to claim 2, wherein the partition portion is a protruding member, the protruding member is disposed in a flow path, and the flow path is divided into the reaction chamber and the buffer chamber.   The biochemical reaction cassette according to any one of claims 2 to 5, wherein a width of the buffer chamber and the slot portion with respect to the flow path direction is equal to a width of the reaction chamber.   The biochemical reaction cassette according to any one of claims 2 to 6, comprising the buffer chamber at least upstream of the reaction chamber.   The biochemical reaction cassette according to claim 7, wherein a buffer chamber upstream of the reaction chamber has the inlet.   The buffer chamber is provided on the upstream side of the reaction chamber, and further, the reaction chamber has the discharge port on the downstream side of the probe fixing region, and has a tapered shape with a width narrowing toward the discharge port. A biochemical reaction cassette according to any one of claims 2 to 6.   The biochemical reaction cassette according to claim 9, wherein a buffer chamber on the upstream side of the reaction chamber has the inlet.   The biochemical reaction cassette according to any one of claims 2 to 6, wherein the buffer chamber is provided on each of an upstream side and a downstream side of the reaction chamber.   The biochemical reaction cassette according to claim 11, wherein the upstream buffer chamber has the inlet, and the downstream buffer chamber has the outlet. The fluid resistance portion is a columnar member disposed in the reaction chamber,
The biochemical reaction cassette according to claim 1, wherein the columnar member partitions a part of the reaction chamber as a buffer chamber. The fluid resistance portion is a partition member having a fine hole disposed in the reaction chamber,
The biochemical reaction cassette according to claim 1, wherein the partition member divides a part of the reaction chamber as a buffer chamber. A target nucleic acid detection probe fixing region has a reaction chamber for reacting a sample in the probe fixing region, an inlet for injecting the sample into the reaction chamber, and a sample is discharged from the reaction chamber A biochemical reaction apparatus for detecting a target nucleic acid having an outlet for
The flow path configured to include the injection port, the reaction chamber, and the discharge port includes a fluid resistance portion that reduces a cross-sectional area of the flow channel, and the fluid resistance portion allows a fluid flow in the reaction chamber to flow. A biochemical reactor characterized by being controlled.   A reaction chamber having a reaction site for performing a biochemical reaction; an inlet for injecting a sample into the reaction chamber; and the sample provided between the inlet and the reaction chamber and supplied to the reaction chamber And a buffer chamber for controlling a flow rate of the biochemical reaction cassette.   A reaction chamber having a reaction site for performing a biochemical reaction; an inlet for injecting a sample into the reaction chamber; and the sample provided between the inlet and the reaction chamber and supplied to the reaction chamber And a buffer chamber for controlling the flow rate of the biochemical reaction device.

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