JP5361787B2 - Trace substance detector - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a substance detector for a trace amount of substance that can improve detection accuracy when detecting a trace amount of substance such as DNA by measuring current. <P>SOLUTION: A substance detector for a trace amount of substance comprises a measurement chamber having a pair of compartments partitioned via an ion exchange membrane or an ion permeable membrane to accommodate a specimen and a buffer liquid and a pair of electrodes installed in the pair of compartments of the measurement chamber. While active control is performed to make the voltage generated between the electrode pair to be 0 volts or close to 0 volts, a generated current is measured. The detector stores previously set reference response waveforms and calculates cross-correlations between the previously set and stored reference response waveforms and a measured current waveform obtained by measurement of current to determine presence/absence of a substance to be detected and calculate its concentration in the specimen based on the strength and position of the correlation. <P>COPYRIGHT: (C)2012,JPO&amp;INPIT

Description

  The present invention relates to, for example, a trace substance detection device for detecting trace substances such as DNA (deoxyribonucleic acid), and in particular, configured to detect the presence or amount of trace substances by current measurement, It is configured to detect the amount of generated charges while setting the electrode potential to “0” V or substantially “0” V, and is obtained by measuring the reference reaction waveform and current set and stored in advance. The present invention relates to a device devised so that the presence / absence and concentration of a target substance to be detected in a sample can be detected from the relationship with a measurement current waveform, thereby improving the detection accuracy.

For example, when detecting a trace substance such as DNA, it is common to detect by a voltage measurement.
Moreover, there exist patent document 1, patent document 2, patent document 3, patent document 4, patent document 5, patent document 6, etc. as what is detected not by voltage measurement but by current measurement, for example.
In the case of this type of current measurement, a reference current value is set and stored in advance, and the reference current value is compared with the measured current value obtained by measuring the current to determine whether or not the threshold has been exceeded. The presence or absence of the target substance to be detected in the sample is discriminated, and the concentration of the target substance to be detected in the specimen is calculated from the peak value of the measured current value.

JP 2008-2333050 A JP 2007-108160 A JP 2006-3222 A JP 2005-69836 A JP 2008-134255 A JP 2006-275788 A

The conventional configuration has the following problems. Usually, the measurement current value obtained by measurement includes noise. This type of noise is caused by contamination of the electrodes installed in the measurement chamber, impurities other than the detection target contained in the sample, a difference in concentration between the two buffer solutions, and the like. . Since this noise is superimposed on the detection current body, the presence / absence of the detection target substance in the sample is determined based on the threshold value, and the concentration of the detection target substance in the sample is calculated based on the peak value. There was a problem that an error occurred.
In particular, in the initial stage of detection, an offset DC current that is substantially steady or slowly decaying may be measured for the reasons described above, and thus it is impossible to specify the absolute “0” point position. As a result, there is a problem that high-precision detection is impaired.

  The present invention has been made based on such points, and an object of the present invention is to provide a trace substance detection device capable of improving the detection accuracy when detecting trace substances such as DNA by current measurement. There is.

In order to achieve the above object, a trace substance detection device according to claim 1 of the present invention includes a measurement chamber that includes a pair of chambers partitioned via an ion exchange membrane or an ion permeable membrane and that stores a specimen and a buffer solution, and the above measurement. A pair of electrodes installed in a pair of chambers, and a current generated while canceling a voltage generated due to noise between the pair of electrodes, and a reference reaction waveform set and stored in advance A measurement circuit that calculates a cross-correlation with a measurement current waveform obtained by measuring the current and calculates the presence and concentration of a target substance to be detected in a specimen from the intensity and position of the correlation, To do.
According to a second aspect of the present invention, there is provided a trace substance detection apparatus comprising a pair of chambers partitioned via an ion exchange membrane or an ion permeable membrane, a measurement chamber containing a specimen and a buffer solution, and a pair of chambers of the measurement chamber. The current generated while canceling the voltage generated due to noise between the pair of electrodes and the pair of electrodes is measured, and is obtained by measuring the reference reaction waveform set in advance and stored and the current. And a measurement circuit for calculating the presence / absence and concentration of the detection target substance in the sample from the analysis result .
A trace substance detection device according to claim 3 is characterized in that, in the trace substance detection device according to claim 1 or 2, the reference reaction waveform is changed according to the temperature of the measurement chamber. is there.
Further, the trace substance detection apparatus according to claim 4 is the trace detection apparatus according to any one of claims 1 to 3, which is provided with a plurality of reference reaction waveforms in advance, and the type and assumption of the target substance to be detected. The optimum reference response waveform is selected according to the concentration to be selected.
Further, the trace substance detection device according to claim 5 is the trace substance detection device according to any one of claims 1 to 3, wherein the trace substance detection device includes a plurality of reference reaction waveforms in advance, and currents are present in all of the reference reaction waveforms. The cross-correlation with the measured current waveform obtained by the measurement is calculated, and the presence / absence and concentration of the detection target substance in the sample are determined and calculated from the obtained multiple correlation intensities and positions. It is what.
A trace substance detection device according to claim 6 is the trace substance detection device according to any one of claims 1 to 3, wherein the trace substance detection device includes a plurality of reference reaction waveforms in advance, and currents in all the reference reaction waveforms are present. This is characterized by calculating the wavelet with the measured current waveform obtained by the measurement and determining / calculating the presence / absence and concentration of the target substance in the sample from the obtained multiple analysis results. is there.

As described above, the trace substance detection device according to claim 1 of the present invention includes a measurement chamber that includes a pair of chambers partitioned via an ion exchange membrane or an ion permeable membrane and that stores a specimen and a buffer solution, and the measurement chamber. In a trace substance detection device comprising a pair of electrodes installed in a pair of chambers, a voltage generated between the pair of electrodes is generated while being actively controlled to “0” V or substantially “0” V The reference response waveform is set and stored in advance, the cross-correlation between the reference response waveform set and stored above and the measured current waveform obtained by measuring the current is calculated, and the strength of the correlation is calculated. The presence / absence and concentration of the detection target substance in the sample are calculated from the position and position, so the accuracy of discrimination and detection is greatly improved compared to simply comparing the reference current value with the measured current value. To improve Can do.
According to a second aspect of the present invention, there is provided a trace substance detection apparatus comprising a pair of chambers partitioned via an ion exchange membrane or an ion permeable membrane, a measurement chamber containing a specimen and a buffer solution, and a pair of chambers of the measurement chamber. A trace substance detection device comprising a pair of electrodes, and measuring a current generated while actively controlling a voltage generated between the pair of electrodes to "0" V or substantially "0"V; The reference response waveform is set and stored in advance, and the wavelet between the reference response waveform set and stored in advance and the measured current waveform obtained by measuring the current is calculated. Since it is configured to calculate the presence / absence and concentration of the target substance, the accuracy of discrimination and detection can be greatly improved as compared with the case where the reference current value is simply compared with the measured current value.
According to a third aspect of the present invention, there is provided the trace substance detection device according to the first or second aspect, wherein the reference reaction waveform is changed according to the temperature of the measurement chamber. Highly accurate detection is possible without being affected by the temperature inside.
Further, the trace substance detection device according to claim 4 is the trace substance detection device according to any one of claims 1 to 3, wherein the trace substance detection device is provided with a plurality of reference reaction waveforms in advance, and the type and assumption of the detected object. Since the optimum reference response waveform is selected according to the concentration to be detected, it is possible to detect with high accuracy according to the type of the substance to be detected.
Further, the trace substance detection device according to claim 5 is the trace substance detection device according to any one of claims 1 to 3, wherein the trace substance detection device includes a plurality of reference reaction waveforms in advance, and currents are present in all of the reference reaction waveforms. It is configured to calculate the cross-correlation with the measured current waveform obtained by the measurement, and to determine / calculate the presence / absence and concentration of the detection target substance in the sample from the obtained multiple correlation strengths and positions Therefore, the detection accuracy can be further improved.
A trace substance detection device according to claim 6 is the trace substance detection device according to any one of claims 1 to 3, wherein the trace substance detection device includes a plurality of reference reaction waveforms in advance, and currents in all the reference reaction waveforms are present. It is configured to perform wavelet calculation with the measured current waveform obtained by the measurement and determine / calculate the presence / absence and concentration of the target substance in the sample from the obtained multiple analysis results. The accuracy can be improved.

It is a figure which shows the 1st, 2nd embodiment of this invention, and is a systematic diagram which shows the whole structure of a trace amount substance detection apparatus. It is a figure which shows the 1st, 2nd embodiment of this invention, and is a circuit diagram which shows the structure of the measurement circuit in the control apparatus of a trace amount substance detection apparatus. It is a figure which shows the 1st, 2nd embodiment of this invention, and is a circuit diagram which shows the structure of the current amplification part of the measurement circuit shown in FIG. 2 in detail. It is a figure which shows the 1st, 2nd embodiment of this invention, and is a figure which shows typically DNA of the state by which the magnetic bead was couple | bonded with one end and the oxidoreductase was couple | bonded with the other end. It is a figure which shows the 1st, 2nd embodiment of this invention, and is a schematic diagram which shows the mode of reaction in a measurement chamber. It is a figure which shows the 1st Embodiment of this invention, and is a characteristic view which shows a reference | standard reaction waveform at the time of 25 degreeC. It is a figure which shows the 1st Embodiment of this invention, and is a characteristic view which shows a reference | standard reaction waveform at the time of 40 degreeC. It is a figure which shows the 1st Embodiment of this invention, and is a figure for demonstrating a cross correlation, Comprising: It is a figure which shows a reference | standard reaction waveform. It is a figure which shows the 1st Embodiment of this invention, and is a figure for demonstrating a cross correlation, Comprising: It is a figure which shows a measured current waveform. It is a figure which shows the 1st Embodiment of this invention, and is a figure for demonstrating a cross correlation, Comprising: It is a figure which shows a cross correlation.

Hereinafter, a first embodiment of the present invention will be described with reference to FIGS. FIG. 1 is a system diagram showing a configuration of a trace substance detection apparatus according to this embodiment. First, there is a tube 1. A sample injection unit 3 is provided at one end side (left end in FIG. 1) of the tube 1, and a sample injector 5 is installed in the sample injection unit 3. The sample 6 is injected by the sample injector 5. This specimen 6 contains DNA as a substance to be detected. In addition, a buffer solution injection unit 7 is installed in the tube 1 near the sample injection unit 3, and the buffer solution injection unit 7 is filled with a buffer solution 9. The buffer solution injection part 7 is connected to the tube 1 via a branch tube 11, and a first pushing valve 13 is attached to the branch tube 11.

  The buffer solution 9 is, for example, a liquid obtained by adding an electron mediator such as dichloroindophenol to a mixture of an aqueous solution of potassium dihydrogen phosphate and an aqueous solution of dipotassium hydrogen phosphate. The electron mediator is for mediating the movement of electrons between the oxidoreductase and the electrode.

  A first reagent injection part 15 and a second reagent injection part 17 are connected to the tube 1 via branch tubes 19 and 21, respectively. The first reagent injection part 15 is filled with a first reagent 23. The second reagent injection part 17 is filled with the second reagent 25. The first reagent 23 is provided with a part that specifically binds to DNA as a target substance to be detected in the specimen 6 and has magnetic particles (magnetic beads). The second reagent 25 is provided with a site that specifically binds to DNA as a substance to be detected in the specimen 6 and has an oxidative or reductase enzyme. A second pushing valve 27 is attached to the branch tube 19 and a third pushing valve 29 is also attached to the branch tube 21.

  The tube 1 is provided with a reaction unit 31, and a temperature control heater 33 is installed in the reaction unit 31. The reaction unit 31 is provided with an ultrasonic vibration device 35. The temperature control heater 33 realizes a temperature environment suitable for the reaction. Further, the stirring effect is enhanced by the ultrasonic vibration device 35. The reaction unit 31 is provided with a first magnetic force control unit 37. The first magnetic force control unit 37 adsorbs and holds the substance to be detected (DNA) in the specimen 6 to which magnetic beads are attached. Further, a fourth pushing valve 39 is attached to the tube 1 on the left side of the temperature control heater 33 in FIG.

A measurement chamber 41 is connected to the other end of the tube 1. An ion exchange membrane 43 is installed in the measurement chamber 41, and a pair of chambers 45 and 47 are defined and formed by the ion exchange membrane 43. Electrodes 51 and 53 are installed in the chambers 45 and 47, respectively. The measurement chamber 41 is configured to be vibrated by an ultrasonic vibration device 35 installed in the reaction unit 31 or an ultrasonic vibration device provided separately from the ultrasonic vibration device 35.
In this embodiment, the ion exchange membrane 43 is used, but an ion permeable membrane may be used.

  Branch tubes 53 and 55 are connected so as to sandwich the measurement chamber 41. These branch tubes 53 and 55 are assembled, and a pump 57 is connected thereto. Further, a fifth pushing valve 59, a sixth pushing valve 61, and a seventh pushing valve 63 are attached to the branch tube 53, the tube 1, and the branch tube 55. A second magnetic force control unit 65 is installed in the measurement chamber 41.

  In addition, a control device 71 is installed, and a measurement circuit 73 is provided in the control device 71. Various devices already described, the first pushing valve 13, the second pushing valve 27, the third pushing valve 29, the fourth pushing valve 39, the fifth pushing valve 59, the sixth pushing valve 61, the seventh The pressing valve 63, the temperature control heater 33, the ultrasonic vibration device 35, the first magnetic force control unit 37, the pump 57, and the second magnetic force control unit 65 are all controlled by the control device 71. Further, current measurement and the like via the electrodes 51 and 53 are controlled by the measurement circuit 73 of the control device 71.

According to the above configuration, first, the first pushing valve 13, the fourth pushing valve 39, and the fifth pushing valve 59 are opened, and the pump 57 is driven. As a result, a negative pressure is generated in the tube 1, and the buffer liquid 9 in the buffer liquid injection unit 7 is sucked and filled in the tube 1. In this state, the sample 6 is injected from the sample injector 5 through the sample injection unit 3.
Next, the second pushing valve 27, the third pushing valve 29, the fourth pushing valve 39, and the fifth pushing valve 59 are opened, and the first reagent injection unit 15 A small amount of the first reagent 23 and the second reagent 25 are drawn from the two-reagent injection unit 17. At the same time, the first pushing valve 13 is opened and the buffer solution 9 is drawn out from the buffer solution injection part 7. As a result, the specimen 6, the buffer solution 9, the first reagent 15, and the second reagent 25 are introduced into the reaction unit 31 and mixed.

  Next, the fourth pushing valve 39 and the fifth pushing valve 59 are closed to cause the sample 6 to react with the first reagent 15 and the second reagent 25. At that time, temperature adjustment is performed by the temperature control heater 33 and ultrasonic vibration is applied by the ultrasonic vibration device 35. As a result, a temperature environment suitable for the reaction is provided, and a stirring / mixing action works. By the reaction of the sample 6 with the first reagent 15 and the second reagent 25, a magnetic bead is bonded to one end of DNA as a detection target substance in the sample 6 and an oxidoreductase is bonded to the other end. become. This is shown in FIG. FIG. 4 is a diagram showing a state in which a magnetic bead 83 is bound to one end of DNA 81 and an oxidoreductase 85 is bound to the other end.

  Next, the first magnetic force control unit 37 is turned on to open the first pushing valve 13, the fourth pushing valve 39, and the fifth pushing valve 59, and the temperature is adjusted by the temperature control heater 33. In addition, while the ultrasonic vibration is applied by the ultrasonic vibration device 35, the buffer liquid 9 continues to flow. As a result, the detection target substance in the specimen 6 is adsorbed and held by the first magnetic force control unit 37, and the other substances are washed away.

  Next, the first pushing valve 13, the sixth pushing valve 61, and the seventh pushing valve 63 are opened and suctioned by the negative pressure of the pump 57, thereby being attracted and held by the first magnetic force control unit 37. The target substance to be detected in the sample 6 after the reaction is introduced into the measurement chamber 41. Then, the sixth pushing valve 61 and the seventh pushing valve 63 are closed to perform measurement. After the measurement, the first pressing valve 13 and the seventh pressing valve 63 are opened, and the measured sample 6 and the like are aspirated and washed away by the negative pressure by the pump 57. As a result, the pump 57 itself is also cleaned.

  Next, the configuration of the measurement circuit 73 provided in the control device 71 will be described with reference to FIG. First, there is a current amplifying unit 91, and an A / D converter 93 is connected to the current amplifying unit 91. The signal amplified by the current amplifier 91 is analog / digital converted by the A / D converter 93 and input to the microcomputer 95. The microcomputer 95 is connected with an I / O 97 and a memory 99. The I / O 97 is connected with an input means 101 and a closing sensor 103.

The configuration of the current amplifier 91 will be described in more detail with reference to FIG. The current amplifier 91 includes an operational amplifier 105 and a resistor 107. The resistor 107 functions to convert the generated current into a voltage in cooperation with the operational amplifier 105. The + side terminal Sp is connected to one electrode 51 side in the measurement chamber 41, and the − side terminal Sn is connected to the other electrode 53 side in the measurement chamber 41. The + side terminal Fp and the − side terminal Fn are terminals for current flow.
Incidentally, reference numeral 109 in the figure is a display unit incorporated in the control device 71 shown in FIG. 1, and is for displaying the measurement result.

According to the above configuration, a DC voltage of approximately 10 to 30 mV is generated when the external impedance is sufficiently high due to a current component including noise generated in the measurement chamber 41. In the case of the present embodiment, first, this voltage is detected via the + side terminal Sp and the − side terminal Sn. Next, a current is passed from the operational amplifier 105 through the resistor 107 in the direction from the positive terminal Fp to the negative terminal Fn so as to cancel the generated voltage. In this way, the generated voltage is offset to prevent a decrease in measurement accuracy.
Incidentally, only a very small amount (several picoamperes) of current flows between the + side terminal Sp and the − side terminal Sn, so that a so-called “drop voltage” does not occur.

Here, the reaction in the measurement chamber 41 will be described with reference to FIG. A chemical reaction in the measurement chamber 41 when an oxidase (for example, glucose oxidase) is used as the oxidoreductase 85 will be specifically described. In the chamber 45 above the measurement chamber 41 (upper side in FIG. 5), an oxidase 96 bound to the DNA 81 is present. In the measurement chamber 41, an oxidizable substance 98 and an electron mediator 100 added to the buffer solution are also present. The oxidizable substance 98 is oxidized by the oxidase 96 and releases electrons 102 and cations 104. The electrons 102 are captured by the electron mediator 100, and as a result, the electron mediator 100 is charged “negatively”. The electron mediator 100 charged “negatively” moves toward the electrode 51 having a relatively “positive” potential. On the other hand, the cation 104 passes through the ion exchange membrane 43, is installed on the chamber 47 side, moves to the electrode 53 side having a relatively “negative” potential, and receives the electrons 102. As a result, a potential difference is generated between the chamber 45 and the chamber 47 across the ion exchange membrane 43, and a current flows between the electrode 51 and the electrode 53. By measuring this current, the DNA 81 is detected and quantified.

Further, the present invention is not limited to the above example, and it is conceivable to use a reductase instead of the oxidase 95 as the oxidoreductase 85. In this case, a substance to be reduced is used instead of the substance to be oxidized 98, and electrons are transferred from the electrode 51 to the reductase through the electron mediator to cause a reduction reaction. Therefore, the electrode 51 that receives electrons from the outside is a “positive” electrode, and the electrode 53 that emits electrons to the outside is a “negative” electrode.
The measurement of the substance to be detected can also be performed by measuring the voltage between the electrodes 51 and 53.

Next, determination of presence / absence of a detection target and detection of concentration in the present embodiment will be described. First, the reference response waveform is set and stored in advance in the memory 99 already described. An example of the reference response waveform is shown in FIGS. FIG. 6 is a diagram showing a reference reaction waveform when the temperature of the measurement chamber 41 is 25 ° C., in which time is plotted on the horizontal axis and current value is plotted on the vertical axis, and the change is shown. FIG. 7 is a diagram showing a reference reaction waveform when the temperature of the measurement chamber 41 is 40 ° C., in which time is plotted on the horizontal axis and the current value is plotted on the vertical axis, and the change is shown.
6 and 7 are merely examples, and a plurality of types of reference reaction waveforms are set and stored so as to correspond to various temperatures of the measurement chamber 41.

In the case of the present embodiment, an optimum reference reaction waveform is selected according to the temperature of the measurement chamber 41, and a “cross-correlation” between the reference reaction waveform and an actual measurement current waveform obtained by current measurement is calculated. Then, from the intensity and position of the correlation, the presence or absence of the detection target in the sample 6 is determined, and the concentration of the detection target is calculated.
The term “cross-correlation” used here is used to determine the similarity between two waveforms. If the array of functions of the two waveforms is all “1”, it is “correlated”. If it is all “0”, it means “no correlation”, and if it is all “−1”, it means “negative correlation”. Therefore, the correlation strength is quantified in the range of −1 to 1.

The cross correlation will be described in more detail.
First, the waveform obtained by converting the measured current waveform into digital data is x (i) (i = 0, 1, 2 ---), and the waveform obtained by converting the reference reaction waveform into digital data is y (i) ( If i = 0, 1, 2--), the cross-correlation function R (j) between them is as shown in the following equation (I).

The similarity between the two waveforms is determined by obtaining the cross-correlation based on the above formula (I). For example, a reference response waveform as shown in FIG. 8 is assumed. FIG. 8 is a diagram showing changes in which the horizontal axis represents time and the vertical axis represents current value. On the other hand, assume that the actual measured current waveform is as shown in FIG. FIG. 9 is also a graph showing changes in which the horizontal axis represents time and the vertical axis represents current value. The reference response waveform and the measured current waveform shown in FIGS. 8 and 9 are converted into digital data, and the waveform x (i) (i = 0, 1, 2 ---) and the waveform y (i) (i = 0) are converted. , 1, 2 ---). Then, the two waveforms x (i) (i = 0, 1, 2 ---), y (i) (i = 0, 1, 2 ---) are subjected to processing as shown in the formula (I). Thus, a cross-correlation diagram as shown in FIG. 10 can be obtained. FIG. 10 is a diagram showing changes in which time is plotted on the horizontal axis and cross-correlation values are plotted on the vertical axis.

Then, in the cross-correlation diagram shown in FIG. 10, the maximum correlation value 0 is obtained around time 20 in the cross-correlation diagram even though large noise is superimposed on the actual measured current waveform. .75. This means that a waveform that is very close to the reference response waveform exists from around the time 20 of the measured current waveform, so that it is automatically determined that the waveform exists at the position of the time 20. Will be. Then, the presence / absence and concentration of the detection target substance are obtained from the intensity and position of the cross-correlation.

As described above, according to the present embodiment, the following effects can be obtained.
First, since active control is performed so as to cancel the DC voltage generated in the measurement chamber 41, the current can be detected with high accuracy, and the detection accuracy of the trace substance can be increased.
In addition, chemical noise with relatively weak power is attenuated immediately by the above active control. However, when a substance to be detected exists in the specimen 6, a clear current peak can be seen, and detection with high accuracy is possible. It becomes possible.
Further, since the redox reaction proceeds rapidly, the time required for detection is shortened.
In the case of the present embodiment, since the voltage is actively measured to measure the current, so-called “drop voltage” does not occur, and higher detection accuracy can be realized. it can.
Further, in the case of this embodiment, as in the prior art, by simply comparing the reference current value with the measurement current, it is possible to determine the presence or absence of the detection target substance in the sample 6 and calculate the concentration. Rather than calculating the cross-correlation between the reference response waveform set and stored in advance and the measured current waveform, the presence or absence of the detection target substance in the sample 6 is determined from the correlation strength and position, the concentration Therefore, the accuracy of determination and detection can be improved.

Next, a second embodiment of the present invention will be described with reference to FIGS. In the case of the first embodiment, the cross-correlation between the reference response waveform and the measured current waveform is calculated. In the case of the second embodiment, the reference response waveform and the measurement current waveform are measured. The “wavelet” of the current waveform is calculated, and the presence / absence of the substance to be detected in the sample 6 is determined and the concentration is calculated from the result of the analysis.
The “wavelet” here means a known “wavelet analysis”. Details will be described below.

First, “wavelet” means how a specific waveform (reference response waveform in this embodiment) is mixed with a certain waveform (in this embodiment, a measured current waveform). It is a mathematical method to analyze First, a specific waveform under good conditions and no noise is stored in the memory 99 as a reference response waveform. Then, by calculating the wavelet of the measurement current waveform mixed with noise and the reference response waveform, the “position where the reference response waveform is embedded in the measurement current waveform”, “gain”, and “frequency of the reference response waveform” are obtained. Is.

The above-mentioned “position where the reference reaction waveform is embedded in the measurement current waveform”, but from the point of time when the chemical (buffer solution 9, first reagent 23, second reagent 25) is added (position where the reaction will start). If a component of the reference response waveform is detected within a certain range, it is determined that this measured current waveform includes the reference response waveform. Further, although it is “gain”, it is determined that a “large response” is obtained if a sufficient gain is recognized at the above-mentioned legal position, and a “small response” is obtained if the gain is small. Thereby, the amount of the substance to be detected is calculated. Furthermore, although it is “frequency”, for example, if an output is recognized at a frequency outside a certain range with respect to the reference response waveform, it is determined that the output waveform is not due to a legitimate reaction but due to noise. The

Incidentally, in the case of the cross-correlation described in the first embodiment, a location (time point) where the reference response waveform exists in the measurement current waveform and a gain are obtained from the reference response waveform and the measurement current waveform. However, in the case of the wavelet analysis of the second embodiment, further frequency information can be obtained.

  As described above, even in the case of the second embodiment using the wavelet analysis, the same effect as in the case of the first embodiment can be obtained, and “frequency” information can also be obtained. Therefore, detection with higher accuracy becomes possible.

The present invention is not limited to the one embodiment.
For example, in the case of the above-described embodiment, the explanation has been given by taking the example of active control to be “0” V. However, the present invention is not limited to this, and control to be substantially “0” V. For example, control that is within ± 1 mV is also conceivable.
It is also conceivable to apply a reverse potential.
The circuit configuration is not limited to that shown in FIGS. 2 and 3, and various examples are conceivable.
In the case of the embodiment, the reference reaction waveform is changed according to the temperature of the measurement chamber. At this time, it is arbitrary how many types of reference reaction waveforms having different temperatures are prepared. It is.
It is also conceivable that a plurality of reference response waveforms are provided in advance, and an optimum reference response waveform is selected according to the type of substance to be detected and the assumed concentration.
In addition, a plurality of reference response waveforms are provided in advance, and the cross-correlation with the measured current waveform obtained by measuring the current in all the reference response waveforms is calculated, and the sample is obtained from the obtained plurality of correlation intensities and positions. It may be possible to determine / calculate the presence / absence and concentration of the substance to be detected.
Furthermore, a plurality of reference response waveforms are provided in advance, and wavelets with the measured current waveforms obtained by measuring the current in all the reference response waveforms are calculated, and the detection target in the sample is obtained from the obtained multiple analysis results. It may be possible to determine and calculate the presence and concentration of the target substance.
Further, in the case of the above-described embodiment, the measurement chamber has been described as an example of a type having both a room in which a reaction is performed and a room in which a reaction is not performed. However, the measurement chamber is not limited thereto. is not. For example, a chamber in which a reaction is performed and an electrode is provided, and a chamber having the characteristics of an ion exchange membrane or an ion permeable membrane that is directly or indirectly covered with a resin or a membrane and that has an electrode. It may be a measurement chamber having an arrangement.
The direct means that the resin of the ion exchange membrane or the ion permeable membrane is directly applied to the electrode, and the indirect means that a water-containing resin or the like is provided between the electrode and the resin of the ion exchange membrane or the ion permeable membrane. This means a configuration that intervenes.
In the case of the above embodiment, the case where the electron mediator is placed in the buffer liquid has been described as an example. For example, a third reagent injection part is provided next to the second reagent injection part. Alternatively, an electronic mediator may be placed in the container, and injection may be performed with the pushing valve opened.

  The present invention relates to a trace substance detection apparatus for detecting trace substances, and in particular, in a device configured to detect the presence or amount of trace substances by measuring a current value, the potential of an electrode is set to “0”. V or approximately “0” V is configured to detect the amount of generated charge, and the relationship between the reference response waveform set and stored in advance and the measured current waveform obtained by measuring the current Therefore, it is suitable for detecting DNA (deoxyribonucleic acid), for example, in which the presence / absence or concentration of the target substance in the sample is detected, thereby improving the detection accuracy. .

5 Sample 6 Target substance 41 Measurement chamber 43 Ion exchange membrane 45 Chamber 47 Chamber 51 Electrode 53 Electrode 71 Controller 73 Control circuit 91 Current amplifier 105 Operational amplifier 107 Resistance

Claims (6)

A measurement chamber comprising a pair of chambers partitioned via an ion exchange membrane or an ion permeable membrane and containing a specimen and a buffer solution;
A pair of electrodes installed in a pair of chambers of the measurement chamber;
The current generated while canceling the voltage generated due to noise between the pair of electrodes is measured, and the reference response waveform set and stored in advance and the measured current waveform obtained by measuring the current are mutually A measurement circuit that calculates a correlation and calculates the presence and concentration of a target substance to be detected in a specimen from the intensity and position of the correlation;
Trace substance detection apparatus characterized by comprising a. A measurement chamber comprising a pair of chambers partitioned via an ion exchange membrane or an ion permeable membrane and containing a specimen and a buffer solution;
A pair of electrodes installed in a pair of chambers of the measurement chamber;
The current generated while canceling the voltage generated due to noise between the pair of electrodes is measured, and a wavelet of a reference response waveform preset and stored and a measured current waveform obtained by measuring the current A measurement circuit that calculates the presence and concentration of the substance to be detected in the specimen from the analysis result,
Trace substance detection apparatus characterized by comprising a. In the trace substance detection apparatus according to claim 1 or 2,
The trace substance detection apparatus, wherein the reference reaction waveform is changed according to the temperature of the measurement chamber. In the trace amount detection device according to any one of claims 1 to 3,
A trace substance detection apparatus comprising a plurality of reference reaction waveforms in advance, and selecting an optimum reference reaction waveform according to the type of substance to be detected and the assumed concentration. In the trace amount substance detection device according to any one of claims 1 to 3,
A plurality of reference response waveforms are provided in advance, and the cross-correlation with the measured current waveform obtained by measuring the current in all the reference response waveforms is calculated, and from the plurality of correlation intensities and positions obtained, A trace substance detection apparatus characterized in that the presence / absence and concentration of a target substance to be detected are determined and calculated. In the trace amount substance detection device according to any one of claims 1 to 3,
A plurality of reference response waveforms are provided in advance, and a wavelet with the measured current waveform obtained by measuring the current in all the reference response waveforms is calculated, and the target substance to be detected in the sample is obtained from the obtained multiple analysis results. A trace substance detection device characterized in that the presence / absence and concentration of a substance are determined and calculated.