JP4621902B2 - Reaction analyzer - Google Patents

Description

[0001]
[Industrial application fields]
The present invention relates to a technique for causing a predetermined reaction to be performed in a state of weightlessness or near weightlessness and analyzing the state of the reaction.
[0002]
[Prior art]
In order to analyze a phenomenon in a zero gravity state or a microgravity state, it has been generally known to perform a so-called drop test in which an experimental apparatus is dropped and a predetermined experiment or test is performed in the apparatus during the dropping. Yes. When performing this drop test, create a drop test device and use it, or install an experimental device on an aircraft and perform altitude-changing operation (parabolic flight) close to the falling state. It is known to create a near microgravity state. It is extremely important to accurately observe and analyze reaction phenomena in the weightless state in order to predict various industrial activities in the future space.
[0003]
In the drop test apparatus for performing the microgravity experiment, the time during which the experiment can be performed is limited to the time during the drop, and thus the time is short. Therefore, it is necessary to perform the experiment efficiently and to record the course and results of the experiment accurately and efficiently.
[0004]
In addition, sudden acceleration increases at the beginning of the fall, and sudden reduction acceleration occurs at the end of the fall and a drop impact associated therewith is inevitably received. . Furthermore, there is a restriction that the size and weight of the experimental apparatus that can be mounted on the drop test apparatus is limited.
[0005]
As a conventionally known microgravity test, there is a material system drop test related to materials. In addition, drop tests in the fields of life sciences (biochemistry, biomedical, space medicine) are known. However, life science-type drop tests have fewer experiments than materials-type tests. One of the reasons is that the experiment cannot be properly performed due to the structure of the experimental apparatus, and it is difficult to accurately monitor the progress of the experiment and accurately evaluate the result. In particular, in an experimental apparatus mounted on a conventional dropping apparatus, it is difficult to accurately observe the course of the experiment and to analyze the course and results of the reaction.
[0006]
The present invention is configured in view of the above circumstances, and particularly relates to an intracellular biopolymer reaction in a life science system, and can appropriately cause a reaction under microgravity, and the progress and result of the reaction. An object of the present invention is to provide a drop test apparatus capable of accurately evaluating the above.
[0007]
[Means for Solving the Problems]
The above object of the present invention is to provide a reaction analyzer for analyzing a reaction that occurs when at least two kinds of sample liquids are mixed in a microgravity state.
Microgravity recognition means for recognizing the microgravity state;
A sample supply means for introducing the sample liquid into the reactor by a signal from the microgravity detection means;
Sampling means for sampling the state of reaction in the reactor every predetermined time;
This can be achieved by a reaction analysis apparatus provided with a recording means for recording the detection result of the sampling means as electronic information.
[0008]
In a preferred embodiment, the sampling means samples the change in absorbance of the solution in the reactor.
[0009]
In this case, the microgravity recognizing means recognizes the start of the fall of the reaction analyzer by detecting a signal indicating that the change in acceleration due to the drop is sufficiently small. This signal is generated from the drop experiment facility. For example, when this drop experiment facility generates a fall start signal, the signal is received, and the analysis device of the present invention is activated at the timing when the microgravity state is reached after a predetermined time using a timer or the like. Also good. Further, a reaction may be started by providing a μG sensor in the drop experiment facility and receiving a signal from the sensor by the reaction analyzer of the present invention.
[0010]
According to another feature of the present invention, in a reaction analysis method for analyzing a reaction that occurs when at least two types of sample liquids are mixed in a reactor,
Recognizing the microgravity state,
After recognizing the microgravity state, the sample solution is introduced into the reactor,
Sampling the reaction situation in the reactor every predetermined time,
There is provided a reaction analysis method comprising a step of recording the sampling result as electronic information.
[0011]
The present invention is suitable for reaction experiments in the biochemical field where the reaction rate is relatively fast. Since the experiment progress is observed and analyzed by data processing mainly based on electric signals, the experiment progress can be accurately observed. In addition, a computer for processing and analyzing the observation results is installed along with the experimental equipment, so that it is possible to perform continuous processing as needed by remote control and provide a high degree of short time resolution. Thus, the experimental progress can be observed and analyzed substantially continuously.
[0012]
The reaction analyzer of the present invention basically comprises the following components.
That is, the reaction analysis apparatus according to the present invention mounted on the dropping device is used for mixing a liquid feeding part for sending a liquid experimental raw material (biological sample or the like), reacting the sample, reacting, and recovering the reaction product. It has a mixing, reaction, and recovery unit. Some reactions start at about the same time as mixing. In addition, there are those in which the reaction proceeds with sufficient mixing. Therefore, the mixing unit or the reaction unit preferably has a reaction cell having a predetermined volume constituted by a container made of a transparent material such as quartz that can be observed. In the reaction part, a plurality of samples are mixed to cause a reaction, and a reaction product and an unreacted substance remain. The collection unit has a bellows accumulator, collects the sample after completion of the reaction, or stores a reaction result and an unreacted substance. Furthermore, a temperature adjusting unit for adjusting the temperature of the reaction system is provided. Specifically, the temperature adjusting unit adjusts the temperature by circulating constant temperature water, and thus the temperature adjusting unit includes a circulator for the constant temperature water. The dropping device includes a control and a data processing unit for analyzing electronic data relating to a reaction progress and a reaction result. For the observation of the reaction, the outer surface of the reaction part is made of a transparent material, a light source is arranged on one side, a CCD camera is arranged on the other side, and an interface for processing the output of the CCD camera is provided. The reaction can be magnified and observed using a microscope or the like. By adopting such an enlargement means, the reaction can be precisely observed and analyzed. Furthermore, a minute time change of the reaction can be precisely observed and / or analyzed by a high-speed camera. In order to observe the reaction, a change in the reaction can be easily observed by using a dye whose color in the reaction portion changes with the progress of the reaction. For color detection, a reaction change can be detected by a color sensor or a transmitted light amount of tungsten / halogen lamp light. The state of reaction may be observed with a CCD camera.
[0013]
In a preferred embodiment of the present invention, the microgravity test device is mounted on the dropping device. When ready for dropping, the dropping device is dropped. The dropping device generates a falling signal or a signal from the μG sensor. When the fall signal or μG signal is detected, the microgravity device activates a timer, and when the count value of the timer falls below a predetermined value, the valve is opened by a control signal from the controller of the sample supply valve, Supply of the sample to the reaction unit is started. In synchronization with this, the reaction is observed in the reaction part. In this case, the transmittance of light having a specific wavelength is changed. For example, as the reaction proceeds, the sample or the reaction product changes in color, such as coloring or subtractive color, or changes in spectral absorption characteristics at a specific wavelength. That is, when an optical change occurs in the reactant in response to the progress of the reaction, the reaction is observed by observing the optical change. Alternatively, the reaction product is configured to be colored. Data is processed and reaction analysis is performed based on the change in absorbance characteristics over time. Further, depending on the reaction, it may be observed by a change in turbidity. In this case, the transition of the reaction can be observed using a CCD camera. As described above, whether to perform the observation based on the absorbance characteristic or the observation based on the CCD can be appropriately set depending on the type of the sample, the form of the reaction, and the like. Since the reaction rate is greatly affected by the reaction temperature, the temperature of the reaction part is controlled. In this case, constant temperature water is circulated around the reaction part to control the temperature of the reaction part to a predetermined temperature. As described above, in the present invention, a series of operations from the start of the reaction to the analysis of the reaction result can be processed continuously and in real time. It can be played.
[0014]
[Embodiments of the Invention]
Hereinafter, embodiments of the present invention will be described with reference to the drawings.
[0015]
FIG. 1 is a conceptual diagram of a biological sample liquid feeding and mixing apparatus 1 according to an embodiment of the microgravity test apparatus of the present invention. The biological sample liquid feeding and mixing apparatus 1 of this example introduces a liquid feeding part 2 for quantitatively sending a sample liquid as a starting material, a mixing part 3 for uniformly mixing a plurality of sample liquids, and a mixed sample liquid The reaction unit 4 includes a reaction unit 4 in which a predetermined reaction occurs, an observation unit 5 for observing the reaction in the reaction unit 4, and a recovery unit 6 that processes a reaction result and collects a residual raw material.
[0016]
The liquid feeding section 2 includes a liquid feeding cylinder 7 having a delivery capacity (10-20 μl / 50-100 ms). The liquid feeding cylinder 7 contains two syringes 8 and 9 having a capacity of about 2000 μl containing sample liquid. The plungers 10 and 11 are connected to one end. The outlet sides of the two syringes 8 and 9 communicate with one mixing unit 3. The mixing unit 3 communicates with the reaction unit 4. The mixing unit 3 or the side of the reaction unit 4 includes a light source 13 such as a lamp or a laser and an observation device 15 including a spectrophotometer 14 and a CCD camera 17. Another observation device 18 configured is provided. In addition, a fluorescent observation device can be provided on the side of the reaction unit 4.
[0017]
Furthermore, the biological sample liquid feeding and mixing apparatus 1 of the present invention includes a temperature adjusting unit 19 for controlling the temperature of the reaction unit 4. The temperature adjusting unit 19 includes a mechanism for circulating a constant temperature medium around the reaction unit 4. The temperature adjustment unit 19 includes a supply cylinder 20 that sends out a constant temperature medium, generally water. Further, a storage tank 21 for adjusting and storing the temperature of the constant temperature medium and a circulation pump 22 for circulating the constant temperature medium stored in the storage tank 21 are provided. An adjustment valve 23 for adjusting the amount of both of them is provided at the junction of the circulation system and the supply system. The adjustment valve 23 is configured to adjust the amount of the constant temperature medium from the circulation system and the supply system so that the temperature of the constant temperature medium sent to the reaction unit 4 becomes a predetermined temperature.
[0018]
Furthermore, the biological sample liquid feeding and mixing apparatus 1 according to the present invention includes a stop unit 25 for supplying a stop solution for stopping the reaction. The stop unit 25 stops a chemical reaction in the reaction unit. A stop solution is stored. And the stop part 25 is equipped with the liquid feeding cylinder 12 for introduce | transducing this stop liquid into the reaction part via the mixing part 3. FIG. Furthermore, the apparatus of this example includes a liquid feeding cylinder 26 for feeding the third sample. Therefore, in this example, reaction by three types of samples can be caused. The recovery unit 6 is provided with a reaction liquid recovery syringe having a capacity of about 10 ml for separating and collecting a plurality of different components from the reaction unit 4. And a residual component is processed as a waste liquid. In the apparatus of this example, there is provided a controller 28 comprising a computer for recording video from the CCD camera 17 on a recording medium such as a tape or recording sampling data from a spectrophotometer on a recording medium such as a disk. . The controller 28 inputs a signal from the μG sensor 29 (provided outside the microgravity experimental apparatus) to start the operation of the liquid feeding cylinder 7 and the opening / closing of the valve of the experimental apparatus, or to start recording by the CCD camera 17; The recording of sampling data by the spectrophotometer 14 is started, and data analysis is performed.
[0019]
In the case of performing an experiment in which two types of reaction liquids are mixed, the stop unit 25, the constant temperature medium system, the supply cylinder 20, the liquid supply cylinder 12, the stop unit 25, the liquid supply cylinder 26, and the like are unnecessary. That is, what is necessary is just to provide the liquid feeding part 2. FIG.
[0020]
Referring to FIGS. 2 and 3, the biological sample feeding and mixing apparatus 1 of the present invention is shown in the form of a perspective view and an exploded perspective view. As described above, the biological sample liquid feeding / mixing device 1 of the present example introduces the liquid feeding part 2 for sending the sample liquids, the mixing part 3 for mixing the two kinds of sample liquids, and the mixed sample liquids. And a reaction section 4 in which a predetermined reaction occurs. In this example, the liquid feeding cylinder 7 of the liquid feeding part 2 of the biological sample liquid feeding / mixing device 1 is fixed to the substrate 30, and the mixing part 3, the reaction part 4, etc. are fixed to the substrate. The channel-type support base 31 is fixed. The liquid feeding cylinder 7 is accommodated in a space defined by the channel-type support base 31 and the substrate 30.
[0021]
The pair of syringes 8 and 9 constituting the liquid feeding unit 2 of the present example is formed in the mixing cell holder 32 fixed on the support base 31, and each of the syringes 8 and 9 has an internal sample. Plungers 10 and 11 that reciprocate in order to send out are arranged. In order to drive the plungers 10 and 11, the liquid feeding cylinder 7 is provided, and a step motor (not shown) is provided as a drive source of the cylinder 7. The plungers 10 and 11 are fixed to a flange of a reciprocable slider 34 coupled to the liquid feeding cylinder 7 via a coupling member 35. The slider 34 moves along a guide rail 36 extending in the reciprocating direction of the plungers 10 and 11. By reciprocating the plungers 10 and 11, the sample in the syringes 8 and 9 is delivered to the mixing unit 3. The mixing unit 3 and the reaction unit 4 are not strictly distinguished on the apparatus. When the mixed sample starts a reaction immediately and it is important to monitor this progress, the mixing unit 3 is configured to be monitorable. Further, by monitoring the reaction in the reaction unit 4, it is possible to observe the reaction accurately and to analyze the reaction accurately. In the microgravity experiment apparatus of this example, an observation mechanism is provided in each of the mixing unit and the reaction unit. The mixing unit 3 is provided with an imaging mechanism using a CCD camera 17 as an observation device. As shown in FIG. 4, the mixing unit 3 is composed of a transparent material mixing cell so that the internal reaction state can be observed from the outside. Then, the CCD camera 17 is arranged on one side of the mixing cell 33 of the mixing unit 3. In this case, the internal space shape of the volume portion in which the sample in the mixing cell 33 of the transparent material is accommodated has a shape in which the width in the photographing direction of the CCD camera 17 is thin and expands in a direction orthogonal thereto. Due to the unique shape of the internal space, the internal space of the mixing cell 33 can be kept bright throughout. As a result, the progress of the reaction in the internal space can be accurately observed by the CCD camera 17 arranged on the opposite side of the mixing cell 33.
[0022]
The observation part 5 for observing the reaction provided on the side part of the reaction part 4 is constituted by an absorbance measurement mechanism.
[0023]
FIGS. 5 and 6 show mixing cells 37 and 38 having different forms from those in FIG. 4. The mixing cell 38 in FIG. 6 has a single cylindrical passage 39 extending in the sample flow direction. I have. In this structure, sample mixing proceeds relatively slowly. Moreover, in the mixing cell 37 of FIG. 5, the predetermined baffle 40 is provided in the orthogonal direction with respect to the cylindrical channel | path 39 extended in the flow direction as mentioned above. In this example, three places are provided at regular intervals. That is, the most upstream and most downstream baffles 40 are provided so as to be orthogonal to the flow direction in the horizontal direction (X direction), and the middle baffle 40 is orthogonal to the flow direction in the vertical direction (Y direction). It is provided to do. In this structure, as compared with the case of the mixing cell 33 in FIG. 4, the mixing of the sample proceeds relatively rapidly and uniformly.
[0024]
The mixing cell 33 is held in a mixing cell holder 32, and the CCD camera 17 and its light source 16 are connected to the connecting portion 41 of the mixing cell holder 32 on the side thereof so that the inside of the mixing cell 33 can be monitored. Can be attached to each. In this case, the mixing cell 33 is fixed to the mixing cell holder 32 from above and from the front by the fixing member 44 via the mounting plates 42 and 43. Sample replenishment openings 45 and 46 for replenishing the sample in the syringe are formed in the upper part of the mixing cell holder 32. Further, a constant temperature water passage for maintaining the mixing cell 33 and the syringes 8 and 9 at a predetermined temperature is formed inside the mixing cell holder 32, and the constant temperature water passage is provided on both sides of the mixing cell holder 32. Pipes 47 and 48 for connecting water to the constant temperature water passage inside the mixing cell holder 32 are connected. Further, in the drawing, a fixing member 49 and a rectangular intermediate plate 50 are disposed in front of the mixing unit 3, that is, on the downstream side, and a reaction cell holder 52 for holding the reaction cell 51 is disposed on the downstream side. The
[0025]
The reaction cell 51 is made of a transparent material. As shown in FIG. 7, both ends have a small diameter, an intermediate portion extends in the vertical direction, and an internal space having a shape thinned in the optical path direction (lateral direction in this example) is formed. Have. The plate thickness of the container is almost constant. The light source 13 and the spectrophotometer 14 are attached to both sides of the reaction cell holder 52. The reaction cell 51 is fixed to the reaction cell holder 52 by a mounting member 53 from the front side thereof.
[0026]
Next, the operation of the biological sample liquid feeding and mixing apparatus 1 according to the present invention will be described.
[0027]
First, the biological sample liquid feeding and mixing device is mounted and fixed on a dropping device.
In this case, the drop device is vibrated at the start of the drop and subjected to a large gravitational acceleration at the end of the drop, and is thus fixed to the drop device so as to withstand this impact.
[0028]
Next, when the preparation for dropping is completed, the locking member of the dropping device is released. This causes the drop device to begin to fall. Immediately after the start of falling, it is an increased speed motion in which the acceleration increases. After this, a constant speed minute acceleration motion is entered, and the vehicle decelerates at a falling point and stops. The reaction is performed during the constant velocity and minute acceleration motion, and the reaction in the biological sample liquid feeding and mixing apparatus starts and ends. This situation is sampled by the measuring device 58 provided with the spectrophotometer 14 and is observed and recorded by the observation device 15. In the case of this example, the above constant speed and minute acceleration state continues for about 4.5 seconds.
[0029]
The start of dropping is detected by the G sensor 29 attached to the dropping device. When the output of the G sensor 29 becomes sufficiently small, the dropping device generates a predetermined signal, and the biological sample liquid feeding and mixing device according to the present invention starts at an optimal timing by receiving this signal. Is set to That is, the G sensor 29 generates a predetermined signal when detecting that the acceleration is equal to or lower than a predetermined value. When the biological sample feeding and mixing apparatus receives this signal, the controller 28 outputs a signal to the timer 54 and starts the timer 54 as shown in FIG. Then, after a predetermined time has elapsed, the reaction measuring device (spectrophotometer) is operated to start sampling. In this example, the sampling interval is set to 10 milliseconds. Therefore, about 45 frames can be sampled in one drop test.
[0030]
On the other hand, when it is found from the signal from the G sensor that the fall acceleration has become equal to or lower than the predetermined value, the controller 28 operates another timer 56 to operate the liquid feeding cylinder 7 after a predetermined time has elapsed. The motor 57 is driven. The timer value of the timer 54 for operating the measuring device and the timer value of the timer 56 for operating the motor 57 of the liquid feeding cylinder 7 are empirically set separately depending on the reaction type, type, etc. However, it is set so that the measuring device 58 is activated after a predetermined time from the liquid feeding cylinder 7.
[0031]
As described above, the drop time that can be experimented is about 4.5 seconds, which is an extremely short time. Therefore, it is necessary to match the operation start timing of the apparatus with the measurement start timing very accurately. This is because if the measurement is started simultaneously with the start of the reaction, the waste of sampling is eliminated and the reaction state can be monitored efficiently. If the measurement start timing does not coincide with the reaction start timing, the reaction cannot be correctly monitored, and therefore, accurate reaction observation or analysis cannot be performed.
[0032]
Depending on the setting of the controller, the mixing experiment can be repeated several times during a single drop, for example, about 10 times.
[0033]
Referring to FIG. 9 and FIG. 10, there is shown an absorbance characteristic according to a measurement result by a spectrophotometer showing a reaction state in the biological sample liquid feeding and mixing apparatus of the present invention. In the example shown in the figure, the oxidation reaction of blue copper protein azurin by [Fe (CN) 6] 3-complex. FIG. 9 shows an experimental result in a state where the gravitational acceleration is 1 G, that is, on the ground. FIG. 10 shows experimental results in a drop experiment where the gravitational acceleration is substantially 0 G. FIG. 11 is a plot of changes in absorbance characteristics over time, focusing on the absorbance change at a wavelength of 620 nm in FIGS. By analyzing FIG. 11, it becomes clear that the state of reaction differs between the ground gravity state and the microgravity state.
[0034]
Thus, by analyzing the difference in reaction between the microgravity state and the ground gravity state from various angles, the characteristics of the reaction in the microgravity state can be known.
[0035]
FIGS. 12, 13, and 14 are diagrams similar to FIGS. 9, 10, and 11 regarding the 2,6-dichlorophenolindophenol reduction reaction by ascorbic acid (vitamin c).
[0036]
【The invention's effect】
As described above, according to the present invention, the apparatus can be made compact, and in the drop test, the reaction state under microgravity is observed precisely, and the result is already obtained at the end of the drop. Since the recording on the recording medium can be completed, the reaction status can be accurately reproduced by outputting the recording result as it is without further processing the recording. Therefore, a highly accurate reaction analysis can be performed quickly and easily compared to the conventional experimental apparatus.
[Brief description of the drawings]
FIG. 1 is a block diagram of a biological sample liquid feeding and mixing apparatus according to an embodiment of the present invention;
FIG. 2 is a perspective view of a biological sample liquid feeding and mixing apparatus according to an embodiment of the present invention.
FIG. 3 is an exploded perspective view of a biological sample liquid feeding and mixing apparatus according to an embodiment of the present invention.
4 shows an example of a mixing cell according to the invention,
FIG. 5 shows another example of a mixing cell according to the present invention. FIG. 6 shows another example of a mixing cell according to the present invention.
FIG. 7 shows an example of a reaction cell according to the present invention,
FIG. 8 is a schematic block diagram of control of a drop test according to the present invention;
FIG. 9 is a graph showing the absorption spectrum in the ground test of the oxidation reaction of blue copper protein azurin by [Fe (CN) 6] 3-complex;
FIG. 10 is a graph showing an absorption spectrum in a drop test of an oxidation reaction of blue copper protein azurin by [Fe (CN) 6] 3-complex;
FIG. 11 is a graph showing the time course of the absorption spectrum at a wavelength of 550 nm of the oxidation reaction of blue copper protein azurin by Fe (CN) 6] 3-complex.
FIG. 12 is a graph showing an absorption spectrum in a ground test of 2,6-dichlorophenol reduction reaction with ascorbic acid (vitamin c);
FIG. 13 is a graph showing an absorption spectrum in a drop test of 2,6-dichlorophenol reduction reaction with ascorbic acid (vitamin c);
FIG. 14 is a graph showing the change with time of the absorption spectrum at a wavelength of 550 nm in the 2,6-dichlorophenol reduction reaction by ascorbic acid (vitamin c).
[Explanation of symbols]
DESCRIPTION OF SYMBOLS 1 Biological sample liquid feeding and mixing apparatus 2 Liquid feeding part 3 Mixing part 4 Reaction part 5 Observation part 6 Recovery part 7 Liquid feeding cylinder 8, 9 Syringe 10, 11 Plunger 12 Liquid feeding cylinder 13 Light source 14 Spectrophotometer 15, 18 Observation apparatus 17 CCD camera 19 Temperature control unit 20 Supply cylinder 21 Storage tank 22 Circulation pump 23 Control valve 25 Stop unit 26 Liquid feed cylinder 27 Fraction collector 28 Controller 29 G sensor 30 Substrate 31 Support base 32 Mixing cell holder 33 Mixing cell 34 Slider.

Claims (4)

In a reaction analyzer that analyzes the reaction that occurs when at least two types of sample liquids are mixed in a microgravity state,
Microgravity recognition means for recognizing the microgravity state;
A sample supply means for introducing the sample liquid into the reactor by a signal from the microgravity detection means;
Sampling means for sampling the state of reaction in the reactor every predetermined time;
A reaction analysis apparatus comprising: a recording unit that records a detection result of the sampling unit as electronic information or image information. The reaction analysis apparatus according to claim 1, wherein the sampling means samples a change in absorbance of the solution in the reactor. The reaction analysis according to claim 1, wherein the microgravity recognizing means recognizes the microgravity state by detecting a signal indicating that the acceleration change due to the fall is sufficiently small. apparatus. In a reaction analysis method for analyzing a reaction that occurs when at least two kinds of sample liquids are mixed in a reactor in a microgravity state,
Recognizing the microgravity state,
After recognizing the microgravity state, the sample solution is introduced into the reactor,
Sampling the reaction situation in the reactor every predetermined time,
A reaction analysis method comprising a step of recording the sampling result as electronic information.

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