JP2006275713A - Calorimeter, and analytical method using the same - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To solve a problem wherein no reaction heat analyzer for coping with a micro-amount of sample exists in the present now, and a problem wherein a structural size gets large and heat comes in from outside, although capable of detecting reaction heat by using a large amount of the sample, in a technique of measuring the reaction heat as a technique for evaluating a reaction of a biosubstance or a chemical substance, although it is a direct and convenient method for use. <P>SOLUTION: This calorimeter has a sample heat flow sensor and a reference heat flow sensor using a thermo-module, two reaction vessels contacting each other with one-faces thereof, the first heat sink contacting with an opposite face, a temperature sensor for measuring a temperature of the first sink, a Peltier element for precise temperature control contacting with an under face of the first heat sink, the second heat sink contacting with an under face of the Peltier element, and a mixer having a heat insulating shield including the two heat flow sensors thereinto, and for mixing prescribed reagents in the reaction vessel by supplying indirect energy from a heat insulating shield outside. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

    The present invention relates to a calorimeter using a thermo module, and more particularly to a reaction analysis apparatus in which a calorie evaluation sample undergoes a temperature change with a chemical reaction or the like.

  In recent years, with the development of biotechnology, there has been an increasing demand for evaluating various living organisms and chemical substances, such as analysis of biological substances, synthesis of various new drugs, and detection of various environmental substances. Furthermore, there are many very valuable substances that can be synthesized and collected in very small amounts, and analysis methods with limited amounts are required.

  For example, when investigating the function of biological substances, the affinity and inhibition of chemicals, the influence of environmental pollutants on the living body, etc., chemical reactions that can occur are often qualitatively and quantitatively evaluated, including enzyme reactions. And antigen antibodies. Conventionally, when detecting an enzyme reaction or an antigen-antibody reaction, a substance that is secondarily generated by the reaction is decomposed and evaluated, or a third substance is added to newly detect a reaction that is easily detected. Has been done. For example, there are methods such as enzyme electrode and fluorescent substance labeling.

  However, since they contain secondary reaction elements, they are more complicated and not reproducible, or are limited to specific reaction systems. On the other hand, there is a difference in the chemical reaction, but there is always a heat of reaction. If a method for directly capturing the heat of reaction is provided, it will be possible to evaluate and analyze a wide range of chemical substances. However, a highly sensitive reaction heat detection system is required to capture the reaction heat of trace substances.

    A means for capturing heat is generally a heat flow sensor, and one of them is a thermocouple. A thermocouple electrically connects two kinds of metals or thermoelectric semiconductors having different polarities, and generates a voltage by giving a temperature difference between both ends. By detecting this voltage, the temperature of the object in contact with one end or the amount of generated heat can be quantitatively detected. Since a pair of thermocouples generates a low voltage, a large number of thermocouples are often integrated and used. A large number of thermocouples arranged in series and electrically connected in series are generally called a thermo module.

    In addition, the heat measurement reacts sensitively to changes in the environmental temperature and causes large noise, so by making the temperature change that affects the environment very small, it is possible to capture very small heat generation and heat absorption, A high-precision thermal analyzer can be made. For example, Patent Document 1 discloses such a high-precision calorimeter.

    In the reference calorimeter, an extremely high-sensitivity measuring instrument is realized by improving the heat insulation structure, controlling the temperature using a Peltier element, adopting a high-sensitivity heat flow sensor, and an appropriate heat sink. By using this method, it is possible to measure very small heat from a very small amount of sample, and furthermore, since temperature control using a driving part or a fluid is not necessary, a compact and inexpensive apparatus can be realized.

    Unfortunately, this method is very effective as a so-called ultra-sensitive thermal analyzer that captures physical phenomena such as phase transformations associated with temperature changes of materials. Cannot be measured.

On the other hand, some thermal analyzers that capture chemical reactions are also commercially available. For example, an isothermal titration calorimeter VP-ITC (MicroCal) is used. This is a structure in which an evaluation sample is put in a small cell in an insulated thermostat, a reaction solution is dropped from a burette-like conduit, and the heat of reaction is detected using a thermo module in contact with the cell. Yes.

In this case, since the apparatus is large, an evaluation sample is required on the order of mL. In order to cause a stable reaction of the sample, some kind of stirring mechanism is required. In this case, a propeller-like stirring device at the tip of the conduit is used. Since the shaft or conduit of this stirring device is inserted from the outside, it brings in external heat and it is difficult to control the temperature above a certain level. Alternatively, as a countermeasure, in order to stabilize the internal temperature, temperature adjustment is required even in the middle of the shaft, and the apparatus is further enlarged.
JP 2004-20509 A (FIG. 1)

  The conventional problems to be solved by the present invention will be summarized based on the conventional calorimeter. 1) As a method for evaluating the reaction of biological and chemical substances, measurement of reaction heat is a direct and easy-to-use method, but there is no reaction thermal analyzer for a very small amount of sample. 2) Although a method for measuring the calorific value of a small amount of sample has been proposed, there is no means for detecting heat in a chemical reaction. 3) Although it is possible to detect the heat of reaction if a sample is used in a somewhat large amount, it becomes structurally large and heat entry from the outside becomes a problem.

    Accordingly, an object of the present invention is to provide a small and inexpensive calorimeter capable of measuring heat of reaction of a minute evaluation sample with high sensitivity while minimizing heat entry from the outside.

  In order to achieve the above object, the following means are adopted in the structure of the calorimeter of the present invention and the analysis method using the calorimeter.

    The calorimeter of the present invention includes a sample heat flow sensor using a thermo module, a reference heat flow sensor using a thermo module, two reaction vessels in contact with one side of each of the two heat flow sensors, and in contact with opposite surfaces of the two heat flow sensors. A first heat sink, a temperature sensor for measuring a temperature of the first heat sink, a Peltier element for temperature control in contact with the lower surface of the first heat sink, a second heat sink in contact with the lower surface of the Peltier element, and at least two It is a calorimeter having a heat insulation shield containing a heat flow sensor, and has a mixing device for mixing a predetermined reagent inside a reaction container of the sample heat flow sensor.

  And it is preferable to mix a predetermined | prescribed reagent by having a mixing apparatus outside the heat insulation shield, and supplying energy in the heat shield shield indirectly without having mechanical coupling. As the mixing device, at least one of a magnetic stirring device, an ultrasonic vibration stirring device, an oscillating stirring device, and a liquid discharge device may be used.

  Alternatively, the first heat sink preferably has a structure in which materials having relatively different thermal conductivities are laminated, has a plurality of heat shields, and further includes an auxiliary temperature control device that indirectly adjusts the temperature of the first heat sink. Even better.

Further, the analysis method using the calorimeter of the present invention includes a step of maintaining a sample heat flow sensor and a reference heat flow sensor at a predetermined temperature via a first heat sink using a Peltier element, and a reaction of the sample heat flow sensor using a mixing device. A step of mixing two or more kinds of reagents in the container, a step of measuring a signal voltage corresponding to a temperature change caused by a chemical reaction of the reagent from the sample heat flow sensor, and a reference voltage generated in the reference heat flow sensor at the same time And a step of calculating a difference between the signal voltage and the reference voltage to detect a chemical reaction.

  Then, following the step of maintaining the sample heat flow sensor and the reference heat flow sensor at a predetermined temperature via the first heat sink using the Peltier element, it is good to stop the energization to the Peltier element, and two kinds are contained in the reaction container of the sample heat flow sensor. It is more effective if the steps up to mixing the above reagents are performed inside the thermostat.

  The calorimeter of the present invention can mix the sample inside and the reaction solution indirectly without controlling mechanically from the outside after controlling the inside of the heat shield to a constant temperature. That is, in order to mix the sample inside the heat shield, a metal shaft, a glass tube, or the like having relatively good heat conductivity does not enter from the outside of the shield, so that the external temperature disturbance is very small. Therefore, the temperature change inside the shield can be kept very small, and even a very small amount of reaction heat can be detected with high accuracy.

  In addition, since the sensor for measuring the amount of heat can use a high-density and small thermo module, the extra heat capacity is small and a small amount of sample can be analyzed.

    Moreover, since there are few whole components and there is no complicated shape, a very small and cheap calorimeter can be constructed.

    If a liquid discharge device is used among the mixing devices, the reaction solution can be mixed a plurality of times, so that a continuous test such as a titration reaction is possible.

    By using such a calorimeter, it is possible to efficiently analyze very precious and few samples in the bio field and the pharmaceutical field. Furthermore, since this calorimeter can directly capture the heat of reaction, a complicated operation such as introducing a labeling substance as in the conventional analyzer or analyzing a secondary reaction becomes unnecessary.

  From the above, the calorimeter of the present invention is expected to be used as an analyzer in a very wide range of fields such as immunodiagnosis, virus diagnosis, environmental analysis, influence of environmental pollutants on cells, and biocompatibility of new drugs. it can.

  Hereinafter, the optimal embodiment of the calorimeter of the present invention will be described with reference to the drawings. First, FIG. 1 shows a sectional view of a calorimeter of the present invention.

  As shown in FIG. 1, a sample heat flow sensor 11 and a reference heat flow sensor 12 are installed as sensors that are the center of the calorimeter of the present invention. Both heat flow sensors are thermomodules in which many thermocouples are integrated, and the basic structure is the same as that of a Peltier element for temperature control. However, in the present invention, in order to detect the minute amount of heat generated from a very small amount of sample, a micro thermo module in which thermocouples are highly integrated is used. Although it depends on the amount of the sample, for example, the size is 3 mm square and the number of pillars is 18 to 98 pairs.

  The sample heat flow sensor 11 and the reference heat flow sensor 12 detect the heat flow in the vertical direction of FIG. 1, but a reaction vessel 21 is installed on one side of the upper side. The reaction vessel 21 is used to put a sample for evaluation therein, and is made using glass or the like.

  A first heat sink 31 is installed on the lower surface of the sample heat flow sensor 11 and the reference heat flow sensor 12. The first heat sink 31 is installed to suppress the temperature change of the two heat flow sensors as much as possible. For this reason, the first heat sink 31 is made of metal such as copper, which has good heat conduction and a large heat capacity.

  A Peltier element 51 is installed under the first heat sink 31. Since the Peltier element 51 can move the heat in the vertical direction by flowing a direct current, the temperature of the first heat sink 31 in contact with the Peltier element 51 can be precisely controlled by controlling it with an appropriate control device from the outside. Since the Peltier element 51 can only increase and decrease the heat flow, the temperature is controlled based on the temperature information from the temperature sensor 41. Here, the temperature sensor 41 uses a thermistor thermometer or a platinum thermometer.

  A second heat sink 32 is installed below the Peltier element 51. The second heat sink absorbs heat transferred from the Peltier element 51 or serves as a heat reservoir for supplying heat to the Peltier element 51 and is generally larger than the first heat sink 31. Is used. The material is also a metal such as copper. It should be noted that the interface of the reaction vessel 21, the sample heat flow sensor 11 and the reference heat flow sensor 12, the first heat sink 31, the Peltier element 51, and the second heat sink 32 described so far has smooth heat propagation. Since it needs to be performed, it is better to interpose a heat conductive grease or an adhesive mixed with metal or ceramic particles.

  Furthermore, in the calorimeter of the present invention, a first heat shield 61 and a second heat shield 62 are provided in order to stabilize the internal atmospheric temperature. Both are provided to block heat conduction through the outside air. Metals with good thermal conductivity such as copper are also used for both heat shields. Moreover, although the base 70 is also provided in order to install the 2nd heat insulation shield 62 stably, it may not exist depending on the case.

  A mixing device 80 is provided below the base 70. Various types of mixing devices 80 can be used, one of which is a magnetic stirring device 85. A simple cross-sectional structure of the magnetic stirring device 85 is shown in FIG. Inside, a motor 90 and a magnet 91 connected to its rotating shaft are attached. When the electric power is turned on, the motor 90 rotates, and the accompanying rotation is transmitted to the magnet 91 that has been magnetized in advance.

  On the other hand, a minute stirrer 81 is housed in the reaction vessel 21. The stirrer 81 is made of a magnetic material such as iron and has a size of about 0.5 mm in thickness and about 2 mm in length. The stirrer 81 is coated with a resin such as glass or Teflon (registered trademark) so as not to be affected by chemical substances. That is, since the stirrer 81 rotates in the same manner as the magnet 91 by the magnetic force from the magnet 91 inside the magnetic stirrer 85, it becomes possible to stir and mix the reagent inside the reaction vessel 21.

  By using the magnetic stirrer 85 in this way, kinetic energy can be input indirectly from the outside of the two heat shields, and the sample can be stirred and mixed inside. In other words, in order to mix the sample inside the heat shield, a metal shaft, glass tube or other material with relatively good heat conductivity does not enter from the heat shield, so the external temperature disturbance is controlled to be very small. The internal temperature does not change. This effect is the same even when a stirring device 80 described later is used. Although FIG. 1 assumes a magnetic stirrer 85, the stirrer 81 is placed. However, when another stirrer described later is used and the magnetic stirrer 85 is not used, the stirrer 81 is not necessary.

  A second example of the mixing device 80 is shown in FIG. FIG. 3 shows an ultrasonic vibration stirrer 86, which is a simplified cross-sectional view. In the apparatus, an ultrasonic transducer 92 is first provided near the upper surface of the apparatus. The ultrasonic vibrator 92 is made of a piezoelectric element material that is distorted when a voltage is applied. A high frequency power supply 93 that supplies a voltage to the ultrasonic transducer 92 is provided in the apparatus.

  The vibration motion of the ultrasonic vibrator 92 caused by the energy of the high-frequency power source 93 is transmitted to the reaction vessel 21 through the base 70, the heat sink, and the like. That is, the sample in the reaction vessel 21 is stirred and mixed by the vibration. Also in this apparatus, since the stirring energy is transmitted through the apparatus, there is no influence on the internal temperature.

  A third example of the mixing device 80 is shown in FIG. FIG. 4 shows a simplified cross-sectional view of the swing type stirring device 87. A motor 90 is first provided in the apparatus. The rotation of the motor 90 is transmitted to the disk 94 and becomes a large circumferential rotation. The rotation of the disk 94 is converted into a one-dimensional back-and-forth or left-right motion by the mechanical conversion device 95, which rocks the gantry 96. Since the calorimeter is installed on the gantry 96, the entire calorimeter is shaken and the sample inside is agitated. The oscillating stirring device 87 has such a one-dimensional motion, but there are various methods such as a rotational motion, an elliptical motion, and a tilt motion.

  FIG. 5 shows a cross-sectional view of a calorimeter equipped with a liquid discharge type device, which is a fourth example of the mixing device 80. Unlike the above three devices, this is a device that forcibly gives kinetic energy to a reaction solution to be reacted with a sample and mixes them. The liquid discharge type mixing device 80 is installed in contact with the inside of the first heat shield 61 as shown in the figure, and is made uniform with the internal temperature. Details of the mixing device 80 are shown in FIG.

  An upper plate 98 made of a piezoelectric element is bonded to a lower plate 97 in which fine grooves 100 are formed in ceramics. The upper plate 98 is made of a piezoelectric material such as lead zirconate titanate (PZT). Although not shown, the upper plate 98 is provided with a driving thin film electrode. A chemical solution tank 99 is provided at an end of the upper and lower plates, and a reaction solution used for the reaction is stored in the chemical solution tank 99. In order to stabilize the temperature of the liquid inside the tank, the chemical tank 99 is also made of metal or ceramics having good heat conduction. The liquid ejection device 88 basically has such a configuration.

    The liquid in the chemical tank 99 is supplied to the groove 100 of the lower plate 97, and when a voltage is applied to the upper plate 98, the upper plate 98 is deformed to apply pressure to the liquid in the groove 100. Thus, a small amount of liquid can be discharged. This principle is basically a method used in some ink jet printers, and can be assembled very compactly. In the present invention, the liquid ejection device 88 is used with a size of 1 cm or less. An electric wire is necessary to apply a voltage to the liquid ejection device 88, but this time it is not shown. Basically, the electric wire is drawn to the outside of the second heat shield 62, and the control is performed from the outside.

    FIG. 7 shows a cross-sectional view of a calorimeter when a liquid discharge type mixing device 80 having a different shape is used. The mixing device 80 is installed outside the second heat shield 62 of the calorimeter. A pump is mounted in the mixing device 80, and a pipe 101 extends from the fluid outlet of the pump. The pipe 101 reaches the vicinity of the reaction vessel 21 via the second heat sink 32 and the first heat sink. The reaction liquid 102 in the pipe 101 is discharged by the pressure of the pump and can be mixed with the sample in the reaction vessel 21 by the force.

What is characteristic here is that the reaction liquid 102 is only in the vicinity of the tip of the pipe 101, and air is contained in the pipe 101 closer to the pump. Further, although not shown in the figure, the pipe 101 is partially made of a different material, the tip is made of metal, and the side near the pump is made of plastic. The distal end portion of the pipe 101 is in close contact with or in close contact with the first heat sink 31 and is also in contact with the first heat shield 61. For this reason, since the front-end | tip part of the metal piping 101 with favorable heat conductivity becomes substantially the same temperature as a heat flow sensor, the reaction liquid 102 discharged is also controlled to the same temperature.

    The vicinity of the pump of the pipe 101 is made of plastic, and since it is filled with air, there is almost no heat transfer from the outside, so it is possible to mix the sample and the reaction liquid 102 with extremely low thermal noise. It is. When the two liquid ejecting devices 88 are used, the reaction liquid 102 can be mixed several times in a pulsed manner, so that continuous reaction analysis such as titration can be realized. Further, the magnetic stirrer 85, the ultrasonic vibration stirrer 86, the oscillating stirrer 87 and the like described above can be used in combination.

    Now, an analysis method using a calorimeter will be described with reference to FIG. First, an equal amount of the sample for evaluation is dropped into the two reaction vessels 21. Then, a reaction solution that will cause a chemical reaction with the sample is dropped again into the reaction vessel 21 on the sample heat flow sensor 11 side. At this time, the amount and the dropping position are determined in advance so that the sample and the reaction solution are not mixed.

    Subsequently, the first heat shield 61 and the second heat shield 62 are covered to substantially block the outside air from entering and exiting. If it is completely sealed at this time, the internal pressure changes when the internal temperature changes. Then, a current is supplied to the Peltier element 51, and a time is allowed until the first heat sink 31 is controlled to a predetermined temperature and a temperature fluctuation range based on temperature information from the temperature sensor 41.

    Since the first heat shield 31 is on the first heat sink 31, the first heat shield 61 is also controlled to have substantially the same temperature as the first heat sink 31. That is, the inside of the first heat shield 61 is set to a uniform temperature. When the output voltages of the sample heat flow sensor 11 and the reference heat flow sensor 12 are measured at this time, the voltage is very small because there is almost no temperature difference above and below the sensors, that is, at both ends of the thermocouple.

    After the first heat sink 31 is set to a predetermined temperature, the stirring device 80 (here, a magnetic stirring device) is operated. Then, the stirring bar 81 installed in the reaction vessel 21 is rotated, and the sample and the reaction liquid are mixed. As the mixing occurs, if there is reactivity between the two, generation or absorption of reaction heat occurs, so that a temperature difference occurs between the upper and lower sides of the sample heat flow sensor 11 and the voltage output fluctuates. From the presence or absence of this output fluctuation, it is confirmed that there is reactivity between the sample and the reaction solution.

  Further, rotating the stirrer 81 may cause some physical thermal fluctuation. Since there is no reaction solution in the reference heat flow sensor 12, only output fluctuations due to physical thermal fluctuations occur. That is, by calculating the difference between the signal voltages of the sample heat flow sensor 11 and the reference heat flow sensor 12, the amount of heat according to the true reaction heat can be calculated. Further, the generated power can be obtained by integrating the voltage over time until the voltage fluctuation occurs and disappears. That is, if the concentration and amount of the sample and the reaction solution are known, the unit reaction heat quantity of the reaction system can be calculated.

Here, the usage conditions of the calorimeter are not particularly limited, but in order to perform a more precise analysis, it may be used in a temperature-controlled atmosphere such as a thermostatic chamber or a thermostatic chamber. In addition, after the first heat sink 31 reaches a predetermined temperature, the current input to the Peltier element 51 may be stopped to mix the sample and the reaction solution. As long as it is controlled by the Peltier element 51, the temperature rises and falls periodically in the vicinity of the set temperature. However, by stopping the current, the temperature of the heat sink only decreases or rises, so it is possible to remove periodic noise components.

  Further, as shown in FIG. 8, the first heat sink 31 may have a three-layer structure. The first heat sink 31 has a structure in which a glass plate 36 is laminated between two metal plates 35. Once the temperature of the first heat sink 31 is controlled with such a structure, the return of heat from the metal plate 35 in contact with the heat quantity sensor is suppressed by the glass plate 36 having low heat conduction, so that the temperature stability is improved. Is even better.

  Further, there is a method of providing an auxiliary temperature control device 52 in addition to the Peltier element 51. A sectional view of a calorimeter using the auxiliary temperature controller 52 is shown in FIG. In this figure, an auxiliary temperature control device 52 is installed further below the second heat sink 32 below the Peltier element 51. Here, a thermoelectric element is also used for the auxiliary temperature control device 52. The auxiliary temperature control device 52 is also on the third heat sink 33, and a third heat shield 63 is also provided so as to surround them. The auxiliary temperature control device 52 feeds back the temperature of the second temperature sensor 42 attached to the second heat sink 32 and sets the second heat sink 32 to a predetermined temperature. This temperature is set to the temperature of the first heat sink 31 to be set or slightly higher than that.

  Since the temperature inside the second heat sink 32 and the second heat shield 62 that is in contact with the second heat sink 32 has already been kept stable, the temperature adjustment of the first heat sink 31 results in a very small disturbance. The temperature fluctuation range can be controlled small. Here, a thermoelectric element is used for the auxiliary temperature control device 52, but a heater may be used. In addition to the installation under the second heat sink 32, the heater can be brought into contact with the second heat shield 62 or the third heat shield 63.

  By using the calorimeter of the present invention by the method as described above, various chemical reactions can be measured with a very small amount of sample. For example, a case will be described in which a microthermo module including 98 thermocouples of bismuth tellurium materials of 3 mm square is used as a heat flow sensor, and the blood glucose level in blood is measured using a glass reaction vessel 21.

  The standard blood sugar level, that is, the amount of glucose is about 1 mg / mL. Take 1 μL of this sample and put it in the reaction vessel 21, and also put glucose oxidase solution. Since it is an enzyme reaction, the temperature of the first heat sink 31 is set to about 40 ° C. From the unit calorific value of glucose, the calorific value at this concentration is 0.44 mJ, and the temperature of the sample heat flow sensor 11 rises by about 6 mK due to the heat capacity of the reaction vessel 21 of about 60 mJ / K and the heat capacity of the liquid of about 10 mJ / K. To do. As a result, the voltage output from the sensor is about 0.2 mV, which does not require a special device and can be easily measured with a normal voltmeter.

  As described above, when the calorimeter of the present invention is used, a reaction test can be performed with a very small amount on the order of μL of biological materials and the like, and this method can sufficiently deal with a valuable sample.

It is sectional drawing which shows the structure of the calorimeter in embodiment of this invention. It is sectional drawing which shows the structure of the magnetic stirring apparatus in embodiment of this invention. It is sectional drawing which shows the structure of the ultrasonic vibration type stirring apparatus in embodiment of this invention. It is sectional drawing which shows the structure of the rocking | swiveling type stirring apparatus in embodiment of this invention. It is sectional drawing which shows the structure of the calorimeter which mounts the liquid discharge apparatus in embodiment of this invention. It is a perspective view which shows the structure of the liquid discharge apparatus in embodiment of this invention. It is sectional drawing which shows the structure of the calorimeter which mounts the liquid discharge apparatus of the 2nd structure in embodiment of this invention. It is sectional drawing which shows the structure of the heat sink in embodiment of this invention. It is sectional drawing which shows the calorimeter of a different structure in embodiment of this invention.

Explanation of symbols

11 Sample heat flow sensor 12 Reference heat flow sensor 21 Reaction vessel 31 First heat sink 32 Second heat sink 33 Third heat sink 35 Metal plate 36 Glass plate 41 Temperature sensor 42 Second temperature sensor 51 Peltier element 52 Auxiliary temperature control device 61 First heat shield 62 Second heat shield 63 Third heat shield 70 Base 80 Mixing device 81 Stirrer 85 Magnetic stirrer 86 Ultrasonic vibration stirrer 87 Oscillating stirrer 88 Liquid discharge device 90 Motor 91 Magnet 92 Ultrasonic vibrator 93 High frequency power supply 94 Disk 95 Machine conversion device 96 Mounting base 97 Lower plate 98 Upper plate 99 Chemical solution tank 100 Groove 101 Pipe 102 Reaction solution

Claims (8)

    A sample heat flow sensor using a thermo module, a reference heat flow sensor using a thermo module, two reaction vessels in contact with one side of each of the two heat flow sensors which are the sample heat flow sensor and the reference heat flow sensor, and the two heat flow sensors A first heat sink in contact with the opposite surface; a temperature sensor for measuring the temperature of the first heat sink; a temperature control Peltier element in contact with the lower surface of the first heat sink; and a second in contact with the lower surface of the Peltier element A heat meter and a heat meter having a heat insulation shield containing at least two heat flow sensors, and a calorimeter having a mixing device for mixing a predetermined reagent inside a reaction container of the sample heat flow sensor.     The mixing device is provided outside the heat insulation shield, and mixes a predetermined reagent by indirectly supplying energy into the heat insulation shield without mechanical coupling. The calorimeter described in 1.   3. The calorimeter according to claim 1, wherein the mixing device has at least one selected from a magnetic stirring device, an ultrasonic vibration stirring device, an oscillating stirring device, and a liquid discharge device.   The calorimeter according to any one of claims 1 to 3, wherein the first heat sink has a structure in which materials having relatively different thermal conductivities are stacked.   The calorimeter according to any one of claims 1 to 4, further comprising an auxiliary temperature control device that includes a plurality of the heat shields and indirectly adjusts the temperature of the first heat sink.   A step of maintaining the sample heat flow sensor and the reference heat flow sensor at a predetermined temperature via a first heat sink using a Peltier element, and mixing two or more types of reagents in the reaction vessel of the sample heat flow sensor using a mixing device Measuring a signal voltage corresponding to a temperature change caused by a chemical reaction of a reagent from the sample heat flow sensor; measuring a reference voltage generated in the reference heat flow sensor; and An analysis method using a calorimeter having a step of calculating a difference from a reference voltage.   7. The method according to claim 6, further comprising a step of stopping energization of the Peltier element following the step of maintaining the sample heat flow sensor and the reference heat flow sensor at a predetermined temperature via the first heat sink using the Peltier element. An analysis method using the described calorimeter. A step of maintaining the sample heat flow sensor and the reference heat flow sensor at a predetermined temperature via the first heat sink using the Peltier element, and mixing two or more types of reagents in the reaction vessel of the sample heat flow sensor using a mixing device The analysis method using the calorimeter according to claim 6 or 7, wherein the step of performing is performed in a thermostatic chamber.

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