JP2010243230A - Biosample reactor - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To obtain a compact biosample reactor extremely simple in operation. <P>SOLUTION: The biosample reactor includes a reaction substrate 20 and the roller 11 pressing the reaction substrate 20 while moving the same along a definite direction, wherein the reaction substrate 20 includes a plurality of the chambers 203 provided on a first substrate 201, a reaction chamber 205 for performing biosample reaction, and a plurality of the flow channels 204 each keeping one end connected to one chamber 203, and the other end to the reaction chamber 205. The reagent solutions in the chambers 203 are supplied to the reaction chamber 205 through the flow channels 204 by squeezing the chambers 203 on the reaction substrate by the rollers 11. The positions and volumes of the respective chambers 203 arranged on the first substrate 201 correspond to the injection periods and injection amounts of the reagent solutions in a biosample reaction process. <P>COPYRIGHT: (C)2011,JPO&INPIT

Description

  The present invention relates to a biological sample reaction apparatus.

  In recent years, rapid inspections such as quarantine for avian influenza and safety checks of imported foods have been required at airports and harbor facilities. In order to meet such demands, analyzers have been developed that perform the entire process of genetic testing and determination of harmful substances within a biochip.

  Patent Document 1 describes a biochemical reaction microreactor having a plurality of independent reaction chambers formed on the surface of a silicon substrate and capable of simultaneously performing a plurality of biochemical reactions in each reaction chamber. ing.

JP 10-337173 A

  However, in the biochemical reaction microreactor described in Patent Document 1, it is necessary to add several types of reagents and washing solutions to the specimen at appropriate times, and the reagents are quantified by controlling a plurality of microsyringe pumps and complicated switching valves. It was. For this reason, the apparatus is complicated and large. In addition, since several types of reagents and cleaning liquids must be dispensed, it has been desired to simplify the operation.

  Therefore, an object of the present invention is to obtain a small-sized biological sample reaction apparatus that is extremely easy to operate.

The biological sample reaction apparatus according to the present invention includes a reaction substrate and pressing means for pressing the reaction substrate while moving the reaction substrate along a certain direction, and the reaction substrate includes the substrate and a plurality of reagent chambers provided on the substrate. And a reaction chamber for performing a biological sample reaction, and a plurality of flow paths with one end connected to one reagent chamber and the other end connected to the reaction chamber, and the pressing means crushes the reagent chamber on the substrate, The reagent solution in the reagent chamber is supplied to the reaction chamber through the flow path, and the position at which each reagent chamber is arranged on the substrate corresponds to the time when the reagent solution is charged in the biological sample reaction process. The volume corresponds to the input amount of the reagent solution in the biological sample reaction process.
As a result, it is possible to easily control the timing of loading a plurality of reagents into the reaction chamber only by controlling the timing of crushing each reagent chamber by the pressing means. In addition, since the necessary amount of reagent is sealed in each reagent chamber in advance, there is no risk of mistakes in reagent type and determination. As described above, even a complicated process using a plurality of reagents can be performed with an extremely simple operation. Therefore, a highly reliable inspection result can be obtained regardless of the skill level of the operator. In addition, since only the pressing means needs to be controlled, the apparatus can be reduced in size and price.

Further, it is desirable to provide a control means for controlling the timing of starting and stopping the movement of the pressing means and the moving speed.
Thereby, it is possible to deal with various biological sample reaction processes with one apparatus.

The pressing means can be a roller, for example.
By controlling the rotation speed of the roller, the reagent chamber on the substrate can be crushed at an arbitrary timing.

The reagent chamber may include a plurality of micro chambers having the same shape and volume.
Thereby, it is possible to prevent a difference in the discharge amount of the reagent liquid when the reagent chamber is crushed due to the difference in the rigidity of the reagent chamber due to the difference in the size and shape of the reagent chamber.

The schematic diagram which shows the structure of the biological sample reaction apparatus by Embodiment 1 of this invention. 2 is a top view of a reaction substrate according to Embodiment 1. FIG. III-III sectional view taken on the line of FIG. Sectional drawing which shows the structure of the valve | bulb provided in the flow path. The perspective view which showed a mode that the chamber of the reaction substrate was crushed with the roller. Sectional drawing which shows the structure of the snap of a chamber. FIG. 7A is a top view of the reaction substrate according to Embodiment 2 of the present invention, and FIG. 7B is an enlarged view of a part of the chamber. It is a top view of the reaction substrate by the Example of this invention.

Hereinafter, embodiments of the present invention will be described with reference to the drawings.
Embodiment 1 FIG.
FIG. 1 is a schematic diagram showing a configuration of a biological sample reaction apparatus 10 according to Embodiment 1 of the present invention. The biological sample reaction apparatus 10 includes a reaction substrate 20, a roller (pressing means) 11, a synchronization gear 12, a reduction gear 13, a step motor 14, a motor controller 15, a main controller (control means) 16, and a barcode reader 17. Yes.

  The reaction substrate 20 is fixed at a predetermined position of the biological sample reaction apparatus 10 when performing the biological sample reaction. The operator selects the reaction substrate 20 corresponding to the desired biological sample reaction process and sets it on the biological sample reaction apparatus 10. After completion of the reaction process, the used reaction substrate 20 can be removed from the biological sample reaction apparatus 10 and replaced with another reaction substrate 20.

FIG. 2 is a top view of the reaction substrate 20. 3 is a cross-sectional view taken along line III-III in FIG.
The reaction substrate 20 includes a first substrate 201, a second substrate 202, a plurality of chambers (reagent chambers) 203, a plurality of flow channels 204, a reaction chamber 205, a sample inlet 206, a valve 207, a rack 208, and a barcode 209. It has. Each chamber 203 is provided with a snap 2031.

  The first substrate 201 and the second substrate 202 are plastic substrates. The chamber 203 is formed by being affixed to the surface of the first substrate 201 and surrounded by the first substrate 201 and a film such as PET molded into a predetermined shape. In an initial state, each chamber 203 is filled with a reagent used in a biological sample reaction, and the position and volume at which each chamber 203 is arranged correspond to the reagent loading timing and amount in the biological sample reaction process. Decided correspondingly. For example, it is arranged at a position where a chamber containing a reagent that is put into the reaction chamber 205 earlier is crushed earlier.

  The flow path 204 is provided between the first substrate 201 and the second substrate 202. Each flow path 204 has one end connected to the connection port 2032 of one chamber 203 and the other end connected to the reaction chamber 205.

  The reaction chamber 205 is a container for performing a biological sample reaction. The reaction chamber 205 is provided with a sample introduction port 206 for introducing the sample into the reaction chamber 205. In the reaction chamber 205, a biological sample reaction is performed by the reagent supplied from each chamber 203 through the flow path 204 and the sample introduced from the sample introduction port 206.

  The valve 207 controls the flow of the reagent solution in each flow path 204. The valve 207 can prevent the reagent in the chamber 203 from entering the reaction chamber 205 through the flow path 204 at a time other than the reagent charging time in the biological sample reaction process.

  FIG. 4 is a cross-sectional view showing the configuration of the valve 207. As shown in the figure, the second substrate 202 has a recess 2021 for providing the flow path 204. The valve 207 has a bottom plate portion 2071 fitted in the concave portion 2021 of the second substrate 202 and two protruding portions 2072 extending upward. A convex portion 2073 that protrudes upward is formed at a portion of the bottom plate portion 2071 that overlaps the flow path 204. The first substrate 201 is formed with a through hole into which the protrusion 2072 of the valve 207 is inserted. The protrusion 2072 slides in the through hole so that the valve 207 moves up and down with respect to the first substrate 201. Can be moved to. However, the protrusion 2072 is tightly fitted in the through hole, and the valve 207 does not move relative to the first substrate 201 unless a considerably large external force is applied. Therefore, as shown in FIG. 4A, in the initial state, the valve 207 is separated from the bottom surface of the concave portion 2021 of the second substrate 202 against gravity.

  The channel 204 is formed by being surrounded by a film 210 such as PET attached to the back surface of the first substrate 201 and the first substrate 201. As shown in FIG. 4A, in the initial state, the convex portion 2073 of the valve 207 presses the film 210 toward the back surface of the first substrate 201. An adhesive 211 having low adhesive strength is disposed between at least a part of the film 210 and the back surface of the first substrate 201, and the film 210 pressed by the pressing force transmitted from the convex portion 2073 of the valve 207. Is bonded to the back surface of the first substrate 201 with an adhesive 211. In this state, the valve 207 blocks the flow path 204. At this time, the protrusion 2072 penetrates the first substrate 201 and protrudes further upward than the first substrate 201.

  FIG. 4B shows a state in which the protrusion 2072 protruding above the first substrate 201 is pushed down by the roller 11. As shown in FIG. 4B, in this state, the bottom surface of the bottom plate portion 2071 of the valve 207 is in contact with the bottom surface of the concave portion 2021 of the second substrate 202. For this reason, the pressing force that presses the film 210 toward the back surface of the first substrate 201 by the convex portion 2073 of the valve 207 does not work. Further, since the adhesive strength of the adhesive 211 is weak, when the reagent solution enters the flow path 204 in this state, the film 210 adhered to the back surface of the first substrate 201 is peeled off. That is, in this state, the valve 207 opens the flow path 204.

  The rack 208 has holes provided in the first substrate 201 and the second substrate 202 along the movement path of the roller 11 in order to prevent the roller 11 from slipping and slipping. FIG. 5 is a perspective view showing how the chamber 203 of the reaction substrate 20 is crushed by the roller 11. As shown in the figure, by engaging the pinion 111 provided on the roller 11 and the rack 208 provided on the reaction substrate 20, the roller 11 can be driven reliably.

  The barcode 209 is printed on the surface of the first substrate 201. By reading the barcode 209 with the barcode reader 17 of the biological sample reaction apparatus 10, a driving program for the roller 11 corresponding to the reaction substrate 20 is obtained. It is executed by the main controller 16. In addition to the barcode 209, a magnetic label or the like can be used as the identification means of the reaction substrate 20. The reading means of the identification means is not limited to the barcode reader, and a reading means corresponding to the identification means is used. Can do.

  When the barcode 209 of the reaction substrate 20 is read by the barcode reader 17 of the biological sample reaction apparatus 10, a program corresponding to the reaction substrate 20 is read into the main controller 16. In accordance with this program, the main controller 16 supplies commands to the motor controller 15 such as timing for starting and stopping the roller 11 and a feed speed. The motor controller 15 sends out driving pulses for rotating the step motor 14 based on these instructions. The step motor 14 rotates by a certain angle for each step according to the period of the drive pulse sent from the motor controller 15. Further, the rotational speed of the step motor 14 is reduced by the reduction gear 13 in order to obtain a torque necessary for crushing the chamber 203. The rollers 11 are arranged above and below the reaction substrate 20 so as to sandwich the reaction substrate 20. In order to synchronize the rotation of the upper and lower rollers 11, the upper and lower rollers 11 are connected by a synchronization gear 12. When the rotation of the step motor 14 is transmitted to the synchronization gear 12 via the reduction gear 13, the roller 11 moves from the end of the reaction substrate 20 in the direction of the arrow X as shown in FIG. Crush it.

  When the chamber 203 is crushed by the roller 11, the reagent solution in the chamber 203 is pushed out from the connection port 2032 to the flow path 204. The pushed reagent solution is supplied into the reaction chamber 205 through the flow path 204. Here, since the connection port 2032 is provided at the end of each chamber 203 on the side to be crushed at the end, all of the reagent liquid in the chamber 203 can be reliably sent into the flow path 204.

  Further, the valve 207 closing the flow path 204 connected to each chamber 203 is provided in front of the end portion of each chamber 203 on the side to be crushed first along the X direction. In other words, since the projection 2072 of the valve 207 is pushed down immediately before each chamber 203 is crushed, the flow path 204 can be released immediately before the chamber 203 is crushed. Since the flow path 204 is provided between the first substrate 201 and the second substrate 202, there is no fear that the flow path 204 is crushed by the roller 11 and the reagent solution does not flow.

  As shown in FIG. 6, the chamber 203 is provided with a snap 2031. The snap 2031 includes a convex portion 2031 a provided on the film portion constituting the chamber 203 and a concave portion 2031 b provided on the surface of the first substrate 201. The convex portion 2031a and the concave portion 2031b have shapes corresponding to each other, and the width of the convex portion 2031a in the cross section shown in FIG. 6 is the same as or slightly larger than the width of the concave portion 2031b. As shown in FIG. 6A, in a state before the chamber 203 is crushed, the convex portion 2031a and the concave portion 2031b are separated with the reagent solution interposed therebetween. As shown in FIG. 6B, when the chamber 203 is crushed by the roller 11, the convex portion 2031a and the concave portion 2031b are fitted, and the convex portion 2031a and the concave portion 2031b are not separated unless a considerably large external force is applied. Thereby, even after the roller 11 passes, the film portion of the chamber 203 that has been crushed once can be prevented from swelling and the reagent solution from flowing backward.

  The reaction substrate 20 corresponds to a predetermined biological sample reaction process, and the position where each chamber 203 is arranged corresponds to the reagent charging timing in the biological sample reaction process. The volume of each chamber 203 is determined in accordance with the amount of reagent input. For this reason, it is possible to easily control the timing of charging a plurality of reagents into the reaction chamber 205 only by controlling the timing of crushing each chamber 203 by driving control of the roller 11. In addition, since a necessary amount of reagent is enclosed in each chamber 203 in advance, there is no risk of mistakes in reagent type and determination.

  Further, even when a dangerous reagent is used in the reaction process, it is not necessary to directly handle the reagent in a dispensing operation or the like, so that even an unfamiliar worker can perform the operation with peace of mind. The chamber 203 can be filled with various solutions used in the biological sample reaction process such as a cleaning solution as well as a reagent.

  As described above, according to the present embodiment, the biological sample reaction process can be controlled only by the drive control of the roller 11, so that even a complicated process using a plurality of reagents can be performed with a very simple operation. Can do. Therefore, a highly reliable inspection result can be obtained regardless of the skill level of the operator.

Further, since only the roller 11 needs to be controlled, the control device can be simplified, and the device can be reduced in size and price.
In addition, since the biological sample reaction apparatus 10 can automatically select a control program for the roller 11 by providing the barcode 209 on the reaction substrate 20, there is no fear of performing a biological sample reaction with an incorrect control program.

  In addition, since the valve 207 is provided in the flow path 204, the flow path 204 is closed in the initial state, and the flow path 204 is opened immediately before the chamber 203 is crushed. The reagent in the chamber 203 can be prevented from entering the reaction chamber 205 through the flow path 204.

  Further, the snap 2031 is provided in the chamber 203 to prevent the film portion of the chamber 203, which has been crushed once, from swelling, so that the reagent solution delivered from the chamber 203 can be prevented from flowing back.

  The reaction substrate 20 may be provided with an optical or magnetic encoder for confirming the position of the roller 11. Thereby, it is possible to confirm to which stage the reaction process has progressed from the position of the roller 11.

  Further, the main controller 16 may perform not only control of the roller 11 but also temperature control in the reaction chamber 205 and management of information on the sample introduced from the sample introduction port 206.

  In this embodiment, the chamber 203 and the flow path 204 are formed by the film attached to the first substrate 201 and the first substrate 201. However, the chamber 203 and the flow path 204 are integrated with the substrate. It may be molded. In this case, the chamber 203 must be capable of being crushed by the roller 11.

  The present invention can also be used for chemical reaction processes other than biological sample reactions. For example, when used in a synthesis reaction process of a chemical that is difficult to store, such as a chemical having a high decomposition rate, the chemical can be obtained by a simple operation in a necessary amount when necessary.

Embodiment 2. FIG.
FIG. 7A is a top view of reaction substrate 30 according to the second embodiment of the present invention. In the second embodiment, the shape of the chamber 303 is different from the chamber 203 of the first embodiment. As shown in FIG. 7A, the chamber 303 includes a plurality of micro chambers 304 having the same shape and volume. FIG. 7B is an enlarged view of a part of the chamber 303. As shown in FIG. 7B, the individual microchambers 304 are connected by a connection channel 3041. One of the plurality of micro chambers 304 constituting one chamber 303 is connected to the flow path 204 through the connection port 3032. Further, the end of the channel 204 opposite to the end connected to the connection port 3032 is connected to the reaction chamber 205. Therefore, each minute chamber 304 is crushed by the roller 11, whereby the reagent solution is sent from the connection port 3032 to the reaction chamber 205 through the flow path 204. The positions where the respective chambers 303 are arranged and the number of the micro chambers 304 included are determined in accordance with the reagent input timing and the input amount in the biological sample reaction process.

  The rigidity of each chamber may change depending on the size and shape of the chamber. In this case, when the chamber is crushed by the roller 11, there is a possibility that the amount of the reagent solution discharged differs. However, according to the second embodiment, since the chamber 303 is formed of an assembly of micro chambers 304 having the same shape and volume, it is possible to prevent a difference in the amount of reagent solution discharged due to a difference in rigidity. Yes.

FIG. 8 is a top view of the reaction substrate 40 according to an embodiment of the present invention. The reaction substrate 40 is a substrate corresponding to a biological reaction process in which the glucose concentration in serum, plasma, or urine is measured by an enzyme method. The chamber 403a is filled with 260 μL of the enzyme solution A, and the chamber 403b is filled with 130 μL of the enzyme solution B. The glucose kit of Daiichi Chemical Co., Ltd. was used for the experiment. The compositions of enzyme solutions A and B are shown below.
Enzyme solution A: Glucose-6-phosphate dehydrogenase: 3.3 U / mL, disodium adenosine triphosphate, magnesium acetate, Good buffer (pH 8.0): 50 mmol / L
Enzyme solution B: hexonase: 5.7 U / mL, nicotinamide adenine dinucleotide: 11 mmol / L, citrate buffer (pH 6.5): 25 mmol / L

  13 μL of serum (plasma) (4 μL in the case of urine) was introduced into the reaction chamber 205 from the sample inlet 206.

  When the reaction substrate 40 is set in the biological sample reaction apparatus 10, the barcode reader 17 reads the barcode 209 and is automatically identified as a glucose concentration measurement substrate. A program for measuring the glucose concentration is loaded on the main controller 16 and the driving of the roller 11 is started. The roller 11 moves in the arrow X direction.

  First, the protrusion 2072 of the valve 207 of the flow path 204 connected to the chamber 403a is pushed down by the roller 11, and the flow path 204 is opened. When the roller 11 further advances, the chamber 403 a is crushed, and the enzyme solution A (260 μL) in the chamber 403 a is sent to the flow path 204 and introduced into the reaction chamber 205.

  The main controller 16 once stops the rotation of the roller 11 after crushing the chamber 403a. When the rotation of the roller 11 is stopped, the main controller 16 sets the temperature in the reaction chamber 205 to 37 ° C. with a heater (not shown), and holds this state for 5 minutes.

  After that, the main controller 16 resumes the rotation of the roller 11, and the protrusion 2072 of the valve 207 of the flow path 204 connected to the chamber 403 b is pushed down by the roller 11 to open the flow path 204. Further, the chamber 403 b is crushed by the roller 11, and the enzyme solution B (130 μL) in the chamber 403 b is sent to the flow path 204 and introduced into the reaction chamber 205.

  After crushing the chamber 403b, the main controller 16 stops the rotation of the roller 11 again. When the rotation of the roller 11 is stopped, the main controller 16 sets the temperature in the reaction chamber 205 to 37 ° C. and holds the state for 96 to 252 seconds. Thereafter, the glucose concentration of the specimen is calculated by measuring the difference in absorbance between the light with wavelengths of 450 nm and 340 nm of the liquid in the reaction chamber 205.

  10 biological sample reaction device, 11 roller, 111 pinion, 12 synchronization gear, 13 reduction gear, 14 step motor, 15 motor controller, 16 main controller, 17 barcode reader, 20, 30, 40 reaction substrate, 201 first Substrate, 202 Second substrate, 2021 Recess, 203, 303, 403a, 403b Chamber, 2031 Snap, 2031a Protrusion, 2031b Recess, 2032, 3032 Connection port, 204 Channel, 205 Reaction chamber, 206 Sample introduction port, 207 Valve, 2071 Bottom plate part, 2072 Protruding part, 2073 Convex part, 208 rack, 209 Bar code, 210 film, 211 Adhesive, 304 Micro chamber, 3041 Connection flow path

Claims (4)

A reaction substrate;
Pressing means for pressing the reaction substrate while moving along a certain direction,
The reaction substrate is:
A substrate,
A plurality of reagent chambers provided on the substrate;
A reaction chamber for conducting a biological sample reaction;
A plurality of flow paths with one end connected to one of the reagent chambers and the other end connected to the reaction chamber;
When the pressing means crushes the reagent chamber on the substrate, the reagent solution in the reagent chamber is supplied to the reaction chamber through the channel,
The position where each of the reagent chambers is arranged on the substrate corresponds to the timing of introduction of the reagent solution in the biological sample reaction process, and the volume of each of the reagent chambers corresponds to the biological sample reaction process. A biological sample reaction apparatus corresponding to the input amount of the reagent solution.   The biological sample reaction apparatus according to claim 1, further comprising a control unit that controls a timing and a moving speed of movement start and stop of the pressing unit.   The biological sample reaction apparatus according to claim 1 or 2, wherein the pressing means is a roller.   The biological sample reaction apparatus according to any one of claims 1 to 3, wherein the reagent chamber includes a plurality of minute chambers having the same shape and volume.

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