JP5245831B2 - Microchip and microchip inspection system - Google Patents

Description

The present invention relates to a microchip and a microchip inspection system.

  In recent years, a micro total analysis system (μTAS) has been attracting attention, in which a plurality of liquids are mixed and reacted in a microchip in which microchannels are integrated and processed, and the state of the reaction is detected and analyzed. Has been.

  μTAS has advantages such as a small amount of sample, a short reaction time, and a small amount of waste. When used in the medical field, the burden on the patient can be reduced by reducing the amount of specimen (blood, urine, wiping liquid, etc.), and the cost of the test can be reduced by reducing the amount of reagent. In addition, since the amount of sample and reagent is small, the reaction time is greatly shortened, and the efficiency of the test can be improved. Furthermore, since the device is small, it can be installed in a small medical institution, and a test can be performed quickly regardless of location.

In a test using a microchip, heating for promoting the reaction between the specimen and the reagent is often performed. For example, in Patent Document 1, the sample and reagent are driven by a micropump connected to the microchip, the sample and the reagent in the microchip are sent to the reaction unit, and the reaction unit is heated. It is described to promote the reaction.
JP 2006-121934 A

  However, when the microchip is heated, the heated liquid in the flow path becomes vapor, and the vapor reaches the downstream flow path and is then cooled, and condensation may occur in the downstream flow path. In this case, air may be interposed between the liquid leading portion and water droplets generated by condensation on the downstream side. In other words, bubbles are present near the liquid head.

  On the other hand, as described in Patent Document 1, the flow path in the microchip is more than the other flow paths so that the liquid sent through the microchip can be temporarily stopped at a predetermined position. A narrowed channel having a narrow width and a shallow depth may be provided. When the liquid is fed at a pressure lower than a predetermined pressure, the liquid feeding is stopped by the surface tension acting on the surface of the liquid when the liquid is about to exit from the throttle channel to the wide channel. When the liquid is fed at a predetermined pressure or higher, this surface tension is overcome, and the liquid passes through the throttle channel and is sent downstream. Hereinafter, the throttle channel capable of blocking liquid feeding is referred to as a water repellent valve. The water repellent valve is provided, for example, in front of the merging portion of a plurality of flow paths that merge with the merging portion, and is used when liquid feeding is simultaneously started to the merging portion.

  When liquid feeding is started at the predetermined pressure or higher with respect to the liquid blocked by the water repellent valve, if bubbles are present near the liquid leading portion as described above, even if liquid is fed at the predetermined pressure or higher, the liquid The bubbles may contract to absorb the pressure of the liquid feeding, and the liquid may not pass through the water repellent valve. If this happens, stable liquid feeding cannot be performed, and in the example in which a water repellent valve is provided upstream of the above-mentioned merging part, liquid feeding to the merging part may not be performed at the same time, or in some cases, liquid feeding to the merging part may not be possible. become.

  The present invention has been made in view of the above problems, and even when condensation occurs in the flow path and bubbles are present in the liquid, the bubbles are removed and stable liquid feeding is performed. It is an object to provide a microchip and a microchip inspection system that can be performed.

The microchip of the present invention is a microchip having a reaction part in which a reaction of a liquid to be accommodated is performed by heating, and a downstream flow path extending downstream from the reaction part, in the middle of the downstream flow path, The dew trap portion having a flow passage width W2 wider than the flow passage width W1 of the downstream flow passage has a width smaller than the flow passage width W1 of the downstream flow passage. It is characterized by being connected to a narrow channel .

  The microchip inspection system of the present invention includes the microchip of the present invention and a heating unit that can accommodate the microchip and is provided at a position corresponding to the reaction part of the microchip when the microchip is accommodated. And a microchip inspection apparatus.

  According to the present invention, the bubbles confined between the droplet formed by condensation in the downstream flow path of the reaction unit and the liquid in the reaction unit can be released in a wide range of condensation trap units. Can be integrated with the liquid in the reaction section, and stable liquid feeding can be performed.

It is an external view of the test | inspection apparatus using the microchip which concerns on this embodiment. It is a block diagram of the test | inspection apparatus using the microchip which concerns on this embodiment. It is a flow-path block diagram of the microchip which concerns on this embodiment. It is a flow path enlarged view of the reaction part periphery of the microchip which concerns on this embodiment.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Microchip 109 Reaction part 110 Condensation trap part 80 Inspection apparatus 32 Heater

  In the present embodiment, as an example, a case where a sample containing a gene and a reagent used for a gene amplification reaction are heated and reacted on a microchip as in a DNA chip for detecting a specific gene will be described. However, the present invention is not limited to this, and the present invention can be applied to the case where the liquid fed through the microchip is heated.

(Device configuration)
FIG. 1 is an external view of an inspection apparatus using a microchip according to the present embodiment. The inspection device 80 is a device that automatically reacts a sample and a reagent previously injected into the microchip 1 and automatically outputs a reaction result.

  The casing 82 of the inspection device 80 is provided with an insertion port 83 for inserting the microchip 1 into the device, a display unit 84, a memory card slot 85, a print output port 86, an operation panel 87, and an external input / output terminal 88. It has been.

  The person in charge of inspection inserts the microchip 1 in the direction of the arrow in FIG. 1 and operates the operation panel 87 to start the inspection. Inside the inspection device 80, the reaction in the microchip 1 is automatically inspected, and when the inspection is completed, the result is displayed on the display unit 84. The inspection result can be output from the print output port 86 or stored in a memory card inserted into the memory card slot 85 by operating the operation panel 87. Further, data can be stored in the personal computer or the like from the external input / output terminal 88 using, for example, a LAN cable. After completion of the inspection, the inspection person takes out the microchip 1 from the insertion port 83.

  FIG. 2 is a configuration diagram of an inspection apparatus using the microchip according to the present embodiment. FIG. 2 shows a state where the microchip is inserted from the insertion port 83 shown in FIG. 1 and the setting is completed.

  The inspection apparatus 80 includes a driving liquid tank 10 that stores a driving liquid 11 for feeding a specimen and a reagent previously injected into the microchip 1, a micropump 5 for supplying the driving liquid 11 to the microchip 1, a micro pump A packing 6 that connects the pump 5 and the microchip 1 so that the driving liquid 11 does not leak, a temperature control unit 3 that controls the temperature of a necessary part of the microchip 1, a temperature control unit 3 and a packing that do not shift the microchip 1 6, a chip pressing plate 2 for bringing the chip pressing plate 2 into close contact, a pressing plate driving unit 21 for moving the chip pressing plate 2 up and down, a regulating member 22 for positioning the microchip 1 with respect to the micropump 5 with high accuracy, and a sample in the microchip 1. And a photodetection unit 4 for detecting a reaction state between the reagent and the reagent.

  The chip pressing plate 2 is retracted upward from the position shown in FIG. 2 in the initial state. Thereby, the microchip 1 can be inserted / removed in the direction of the arrow X, and the person inspecting inserts the microchip 1 from the insertion port 83 (see FIG. 1) until it comes into contact with the regulating member 22. Thereafter, the chip pressing plate 2 is lowered by the pressing plate driving unit 21 and comes into contact with the microchip 1, and the lower surface of the microchip 1 is in close contact with the temperature adjustment unit 3 and the packing 6.

  The temperature control unit 3 includes a Peltier element 31 and a heater 32 as a heating unit on a surface facing the microchip 1. When the microchip 1 is set in the inspection device 80, the Peltier element 31 and the heater 32 are connected to the microchip. 1 is in close contact. The portion containing the reagent is cooled by the Peltier element 31 so that the reagent is not denatured, or the reaction portion 109 (see FIG. 3) where the specimen and the reagent react is heated by the heater 32 to promote the reaction. To do.

  The light detection unit 4 includes a light emitting unit 4a and a light receiving unit 4b. The light detecting unit 4 irradiates the microchip 1 with light from the light emitting unit 4a, and detects light transmitted through the microchip 1 with the light receiving unit 4b. The light receiving portion 4b is integrally provided inside the chip pressing plate 2. The light emitting unit 4a and the light receiving unit 4b are provided so as to face a detection unit 115 of the microchip 1 described later.

  The micropump 5 is located on the pump chamber 52, the piezoelectric element 51 that changes the volume of the pump chamber 52, the first throttle channel 53 located on the microchip 1 side of the pump chamber 52, and the driving fluid tank 10 side of the pump chamber. The second throttle channel 54 is formed. The first throttle channel 53 and the second throttle channel 54 are narrow and narrow channels, and the first throttle channel 53 is longer than the second throttle channel 54.

  In the case where the driving liquid 11 is fed in the forward direction (direction toward the microchip 1), first, the piezoelectric element 51 is driven so as to rapidly reduce the volume of the pump chamber 52. Then, a turbulent flow is generated in the second throttle channel 54 that is a short throttle channel, and the channel resistance in the second throttle channel 54 is relatively larger than that of the first throttle channel 53 that is a throttle channel. growing. As a result, the driving liquid 11 in the pump chamber 52 is predominantly pushed toward the first throttle channel 53 and fed. Next, the piezoelectric element 51 is driven so that the volume of the pump chamber 52 is gradually increased. Then, the driving liquid 11 flows from the first throttle channel 53 and the second throttle channel 54 as the volume in the pump chamber 52 increases. At this time, since the length of the second throttle channel 54 is shorter than that of the first throttle channel 53, the channel resistance of the second throttle channel 54 is smaller than that of the first throttle channel 53. Thus, the driving liquid 11 flows predominantly from the second throttle channel 54 into the pump chamber 52. When the piezoelectric element 51 repeats the above operation, the driving liquid 11 is fed in the forward direction.

  On the other hand, when the driving liquid 11 is fed in the reverse direction (direction toward the driving liquid tank 10), first, the piezoelectric element 51 is driven so that the volume of the pump chamber 52 is gradually reduced. Then, since the length of the second throttle channel 54 is shorter than that of the first throttle channel 53, the channel resistance of the second throttle channel 54 is smaller than that of the first throttle channel 53. . As a result, the driving liquid 11 in the pump chamber 52 is predominantly pushed out toward the second throttle channel 54 and fed. Next, the piezoelectric element 51 is driven so that the volume of the pump chamber 52 is rapidly increased. Then, the driving liquid 11 flows from the first throttle channel 53 and the second throttle channel 54 as the volume in the pump chamber 52 increases. At this time, turbulent flow is generated in the second throttle channel 54, which is a short throttle channel, and the channel resistance in the second throttle channel 54 is relatively larger than that of the first throttle channel 53, which is a throttle channel. Become bigger. As a result, the driving liquid 11 flows into the pump chamber 52 predominantly from the first throttle channel 53. When the piezoelectric element 51 repeats the above operation, the driving liquid 11 is fed in the reverse direction.

(Microchip channel configuration)
FIG. 3 is a flow path configuration diagram of the microchip 1 according to the present embodiment. It is an example and the present invention is not limited to this.

  In the microchip 1 of this embodiment, as shown in FIG. 3, the two reaction detection flow paths are symmetrically arranged (left half and right half in the figure). For example, the reaction detection channel in the left half is used for detecting the reaction between the sample and the reagent, and the right half is used for detecting the reaction between the internal control and the reagent for monitoring whether the reaction detection is normally performed. . Since the two reaction detection channels are symmetrically arranged and basically have the same channel configuration, only the reaction detection channel for the left half of the sample and the reagent will be described below.

  In the upper part of the microchip 1 in the figure, reagent storage portions 101a, 101b, and 101c that store reagents commonly used in the reaction detection channels in the left half and the right half of the microchip 1 are provided. In the reagent storage units 101a, 101b, and 101c, for example, reagents used for gene amplification reaction are stored in advance. A junction 102 where these reagents merge is provided downstream of the reagent containers 101a, 101b, and 101c.

  A branching unit 103 that branches the mixed reagent from the joining unit 102 into the left half and the right half of the microchip 1 is provided downstream of the joining unit 102. A reagent storage unit 104 that stores the mixed reagent is provided downstream of the branching unit 103.

  A junction 105 is provided downstream of the reagent storage unit 104. In addition to the flow path from the reagent storage section 104, the flow path from the specimen storage section 106 that stores the injected sample and the flow path from the reagent storage section 107 join the merge section 105.

  A mixing flow path 108 is provided downstream of the merging portion 105 for mixing the specimen and the reagent with sufficient molecular diffusion. A reaction unit 109 is provided downstream of the mixing channel 108.

  The reaction unit 109 is a part that heats and reacts the mixed liquid of the sample and the reagent sufficiently mixed in the mixing channel 108. When the microchip 1 is set in the inspection device 80, the reaction unit 109 has a heater for the inspection device 80. 32 are opposed to each other. A gene amplification reaction is performed by heating the reaction unit 109 with the heater 32.

  A dew trap section 110 having a width wider than the width of the flow path from the reaction section 109 that traps dew condensation generated by heating of the reaction section 109 is provided downstream of the reaction section 109.

  A confluence portion 111 is provided downstream of the dew condensation trap portion 110. In addition to the flow path from the dew condensation trap section 110, the flow path from the reagent storage section 112 joins the merge section 111. In the reagent storage unit 112, for example, a reaction stop solution for stopping the gene amplification reaction is stored.

  A junction portion 113 is provided downstream of the junction portion 111. In addition to the flow path from the merge section 111, the flow path from the reagent storage section 114 joins the merge section 113. The reagent container 114 stores, for example, a denaturing solution for denaturing the amplified gene into a single strand.

  A detected portion 115 is provided downstream of the merging portion 113. The detected portion 115 is made of a material such as transparent resin or glass. When the microchip 1 is set in the inspection device 80, the light detection unit 4 of the inspection device 80 faces the detected portion 115. Detection is performed by optically detecting the concentration of the detection target 115 by the light detection unit 4.

  For example, streptavidin or the like that specifically reacts with the gene of the specimen is immobilized on the wall surface of the detection unit 115 in advance. The sample gene is trapped in the detected portion 115 by binding the sample gene to streptavidin.

  In addition to the flow path for supplying the amplification product from the confluence section 113, the flow path from the reagent storage sections 116, 117, and 118 storing the reagents necessary for detection is connected to the detected section 115. Yes. For example, the reagent storage unit 116 stores a probe fluorescently labeled with FITC at the end that binds to the gene trapped in the detection unit 115 by a hybridization reaction. The reagent storage unit 117 stores a colloidal gold solution whose surface is modified with a FITC antibody that specifically reacts with the probe. A cleaning liquid is supplied from the cleaning liquid supply flow path 118 to wash out the detected part 115 and push out the colloidal gold not bonded to the probe from the detected part 115.

  A waste liquid unit 119 that stores the liquid pushed out from the detected unit 115 is provided downstream of the detected unit 115.

  Next, the flow of the specimen and reagent in the microchip 1 will be described. In FIG. 3, each micropump connection part 120 a to 120 k indicated by black circles is a part to which the driving liquid 11 is supplied from the micropump 5 when the microchip 1 is set in the inspection device 80. . The micropump 5 includes a plurality of independent micropumps, and the supply of the driving liquid 11 from the micropump connection portions 120a to 120k is controlled independently.

  First, by supplying the driving liquid 11 from each of the micropump connection parts 120a, 120b, and 120c, the reagents stored in the reagent storage parts 101a, 101b, and 101c are respectively supplied to the joining part 102 and mixed.

  The reagent mixed solution in the merging unit 102 is separated into a left half flow channel and a right half flow channel by the branching unit 103 and then stored in the reagent storage unit 104.

  Next, by supplying the driving liquid 11 from the micropump connection units 120d, 120a to 120c, 120e, respectively, the sample stored in the sample storage unit 106, the reagent stored in the reagent storage unit 104, and the reagent storage Reagents stored in the unit 107 are respectively supplied to the merging unit 105 and mixed. As a result, a sample reagent mixed solution that is a mixed solution of the sample containing the gene and the reagent used for the gene amplification reaction is generated.

  The sample reagent mixed solution in the merging portion 105 is sufficiently diffused and mixed in the mixing channel 108 and then stored in the reaction portion 109.

  The sample reagent mixed solution accommodated in the reaction unit 109 is heated by the heater 32 of the test apparatus 80 to perform a gene amplification reaction, and a gene amplification solution is obtained.

  Next, by supplying the driving liquid 11 from each of the micropump connection parts 120a to 120e and 120f, the gene amplification liquid stored in the reaction part 109 is supplied to the confluence part 111 via the condensation trap part 110. The reagent (reaction stop solution) stored in the reagent storage unit 112 is supplied to the junction unit 111 and mixed with each other. Thereby, the reaction of the gene amplification solution is stopped.

  Next, by supplying the driving liquid 11 from each of the micropump connection parts 120h and 120g, the gene amplification liquid in the merging part 111 and the reagent (denaturing liquid) contained in the reagent containing part 114 are respectively supplied to the merging part 113. Supplied and mixed. As a result, the gene in the gene amplification solution is denatured into a single strand. The denatured gene amplification solution is supplied to the detected portion 115 as an amplification product. At this time, the target gene in the amplification product reacts with streptavidin immobilized on the wall surface of the detected portion 115 and is trapped by the detected portion 115.

  Next, by supplying the driving liquid 11 from the micropump connection unit 120i, the reagent (the probe fluorescently labeled with FITC) stored in the reagent storage unit 116 is supplied to the detection unit 115. As a result, a probe fluorescently labeled with FITC at the end binds to the gene trapped in the detection target 115 by a hybridization reaction.

  Next, by supplying the driving liquid 11 from the micropump connection unit 120j, the reagent (gold colloid liquid whose surface is modified with the FITC antibody) stored in the reagent storage unit 117 is supplied to the detection unit 115. As a result, the gold colloid is bound to the probe bound to the gene trapped in the detected portion 115 by the antigen-antibody reaction.

  Next, the driving liquid 11 as a cleaning liquid is supplied to the detected part 115 by supplying the driving liquid 11 from the micropump connection part 120k. As a result, the detected portion 115 is washed away, and the gold colloid not bonded to the probe is pushed out from the detected portion 115 to the waste liquid portion 119. Thereafter, the concentration of the gold colloid in the detected part 115 is detected by the light detection part 4 of the inspection apparatus 80.

(Flow path configuration around the reaction unit 109)
FIG. 4 is an enlarged view of the flow path around the reaction part of the microchip according to the present embodiment. The flow path from the reaction part 109 to the confluence | merging part 111 is shown.

  As shown in the drawing, the width of the flow path F as the downstream flow path of the present invention between the reaction section 109 and the dew condensation trap section 110 is W1 (for example, 100 μm), and the width of the flow path of the dew condensation trap section 110 is shown. W2 is set to a wider value (for example, 1500 μm) than W1. In the dew trap part 110, the flow path width gradually increases from W1 to W2 at the inlet, and the flow path width gradually decreases from W2 to W1 at the outlet.

  A water repellent valve RB is provided downstream of the dew condensation trap unit 110. As described above, the water-repellent valve RB is a channel whose channel width is narrower than W1 and whose depth is shallower than the channel F. When the liquid is fed at a pressure lower than a predetermined pressure, the liquid feeding is stopped by the surface tension acting on the surface of the liquid when the liquid is about to exit from the throttle channel to the wide channel. When the liquid is fed at a predetermined pressure or higher, this surface tension is overcome, and the liquid passes through the throttle channel and is sent downstream. A merging portion 111 is provided downstream of the water repellent valve RB.

  4A shows a state in which water droplets D due to condensation are generated in the flow path F by heating the liquid L in the reaction unit 109 by the heater 32 of the inspection device 80. FIG. Water droplets D block the flow path F, and bubbles A are interposed between the liquid L and the water droplets D.

  When the liquid L in the reaction unit 109 starts to be fed from the state shown in FIG. 4A, the liquid L and the water droplet D pass through the flow path F with the bubbles A interposed between the liquid L and the water droplet D. It advances and enters into the condensation trap part 110. At this time, since the width W2 of the dew condensation trap part 110 is wider than the width W1 of the flow path F, when the water droplet D blocking the flow path F enters the dew condensation trap part 110, the flow in the dew condensation trap part 110 is increased. It becomes difficult to block the road. If the water droplet D cannot block the flow path in the dew condensation trap part 110, the bubbles A confined between the water droplet D and the liquid L are released, and the air present in front of the water droplet D Merge. FIG. 4B shows this state. The leading part of the liquid L reaches the inlet of the condensation trap part 110, and the water droplet D adheres to the inner wall surface of the condensation trap part 110. The bubble A is united with the air existing in front of the water droplet D and disappears.

  When the liquid feeding is further advanced, the liquid L catches up with the water droplet D adhering to the inner wall surface of the dew condensation trap portion 110, and the water droplet D is absorbed by the liquid L to be integrated. And as shown in FIG.4 (c), the liquid L will be sent from the exit of the dew condensation trap part 110 in the state from which the bubble was removed.

  As a result, the liquid L reaches the water repellent valve RB in a state where the bubbles are removed, so that the bubbles contract and absorb the liquid feeding pressure, and the liquid L cannot pass through the water repellent valve RB. This makes it possible to carry out stable liquid feeding.

  When the flow path width gradually increases from W1 to W2 at the entrance of the condensation trap portion 110 as in the present embodiment, the bubbles A that have entered the condensation trap portion 110 are formed at the corners in the condensation trap portion 110, etc. It is preferable because it is discharged smoothly without staying in the area.

  The flow path width W2 of the dew condensation trap part 110 is preferably 1.2 to 20 times the flow path width W1 of the flow path F. If it is smaller than 1.2 times, there is a high possibility that the flow path is still blocked by the water droplet D even in the condensation trap part 110, and it becomes difficult to release the bubbles A. In order to release the bubbles A, it is better that the flow path width of the dew condensation trap part 110 is larger, and there is basically no upper limit. However, when the flow path width of the condensation trap part 110 becomes too large, the volume of the condensation trap part 110 becomes too large. In this case, the size of the microchip 1 becomes large, and it is necessary to supply a large amount of driving liquid 11 to send the liquid, which takes time for the inspection. For this reason, it is preferable that the flow path width W2 of the dew condensation trap part 110 is 10 times or less with respect to the flow path width W1 of the flow path F. Usually, the flow path width W1 of the flow path F is about 80 to 200 μm, and therefore the flow path width W2 of the dew condensation trap part 110 is preferably 100 to 4000 μm.

  Moreover, it is preferable that the shape of the dew trap part 110 is a vertically long shape (the distance from the inlet part to the outlet part is longer than the flow path width W2) long in the liquid feeding direction. The vertically long shape prevents bubbles A entering from the inlet in the condensation trap portion 110 from reaching the outlet all at once, so that the bubbles A can be reliably removed.

  As described above, according to the present embodiment, the bubble A confined between the droplet D and the liquid L in the flow path F can be released in the wide condensation trap portion 110, so the droplet D Can be integrated with the liquid L, and stable liquid feeding can be performed.

Claims (6)

A reaction part in which a reaction of the liquid to be contained is performed by being heated; and
A downstream flow path extending downstream from the reaction section;
In a microchip having
In the middle of the downstream flow path, there is a condensation trap portion having a flow path width W2 wider than the flow path width W1 of the downstream flow path ,
The microchip according to claim 1, wherein the downstream flow path downstream of the dew condensation trap portion is connected to a flow path narrower than the flow path width W1 of the downstream flow path . 2. The microchip according to claim 1 , wherein the dew condensation trap part gradually widens from a flow path width W <b> 1 of the downstream flow path to a flow path width W <b> 2 of the dew condensation trap part at an inlet portion. 3. The microchip according to claim 1 , wherein a channel width W <b> 2 of the dew trap portion is 1.2 to 20 times larger than a channel width W <b> 1 of the downstream channel. 4. The microchip according to claim 3 , wherein a flow path width W <b> 2 of the dew condensation trap portion is 100 to 4000 μm. The microchip according to claim 1 , wherein the dew condensation trap portion has a vertically long shape that is long in a liquid feeding direction. The microchip according to any one of claims 1 to 5 ,
A microchip inspection apparatus comprising a heating unit capable of accommodating the microchip and provided at a position corresponding to the reaction unit of the microchip when the microchip is accommodated;
A microchip inspection system comprising: