JP4646813B2 - Biosensor measurement system, viscosity measurement method, and trace mass measurement method - Google Patents

Description

  The present invention measures the viscosity of the solution and the minute adsorption mass of chemicals adsorbed on the electrode surface of the resonator from the change in the electrical characteristics of the sliding wave type piezoelectric resonator during self-excitation in the sample solution. The present invention relates to a biosensor measurement system, a viscosity measurement method, and a trace mass measurement method.

    In recent years, a bio-test chip called Lab-on-a-Chip has been actively developed. This is a configuration in which element structures such as a flow path, a reaction tank, a valve, and a sensor are integrated on a small substrate, and an analysis process is performed on the liquid flowing inside the structure. Since such a bio-test chip is compact and inexpensive, it can be regularly tested with a small amount of sample at home, for example, or applied to environmental measurements outdoors.

  Sensors using various principles have been proposed for the sensor unit which is one of the components of the biotest chip. Among them, a slip-wave type piezoelectric resonator sensor using a slip-wave type piezoelectric resonator, which is small and can be configured in a plate shape and is assumed to be easily mounted on a bio-test chip, has attracted attention.

  The slip wave in the solid vibration is a wave caused by the shear stress of the solid. This slip wave has a property that it is difficult to radiate vibration energy to a liquid such as water as compared with other vibration modes. Therefore, the slip wave type piezoelectric resonator has a great feature that it easily generates electric self-excited vibration even in a liquid. This feature is utilized by a slip wave type piezoelectric resonator sensor. That is, this sliding wave type piezoelectric resonator sensor has a great feature that it can detect the minute mass of the sample adhering to the surface of the piezoelectric resonator and the viscosity of the solution as changes in the electrical characteristics of the resonator in the solution. ing. A representative example of this sliding wave type piezoelectric resonator sensor is an AT-cut quartz crystal resonator sensor using a single crystal crystal, which is a sensor recognized as a QCM (Quartz Crystal Microbalance) type biosensor (for example, non-patent) Reference 1). By the way, this AT-cut quartz resonator has a thickness-slip vibration depending on the thickness of the resonator as its natural frequency, and is a quartz resonator that is widely recognized as the above-described slip-wave type piezoelectric resonator. .

Hereinafter, the case where the AT cut crystal resonator is used as a QCM type biosensor will be described in detail. FIGS. 12A and 12B are schematic views of a QCM type biosensor using an AT cut crystal resonator, and a pair of opposing excitation electrodes 1202 are provided on the surface of the AT cut crystal resonator piece 1201. It is configured. A lead electrode 1203 is integrally formed with the excitation electrode 1202. In the arrangement of the excitation electrode 1202, the AT-cut quartz crystal resonator induces a thickness shear vibration 1204 as a natural frequency. Furthermore, a sensitive film 1205 that captures only the analysis target is immobilized on the surface of one side surface. At this time, the mechanical natural frequency Fr of the AT-cut quartz substrate changes depending on the minute mass that is analyzed by the sensitive film. At this time, it is known that the amount of change ΔF _{1 in} the natural frequency of the AT-cut quartz crystal resonator is proportional to Δm, where Δm is the small mass captured by the sensitive film. This relational expression is derived by Sauerbrey (G. Sauerbrey, Z. Phys. 155, 1959, 206, Non-Patent Document 2), and is given by the following expression.

Here, Fr is the natural frequency of the quartz resonator, A _e is the area of the excitation electrode 1202, C ₆₆ is the shear elastic modulus of the AT-cut quartz resonator piece 1201, and ρ _q is the density of the quartz. Therefore, it has been found that the sensitivity as a mass sensor increases as the natural frequency F _r increases.

  Further, when operating a QCM biosensor in a liquid, if the AT-cut quartz oscillator is vibrated in a solution containing an analysis target or the like, its natural frequency decreases due to the influence of the viscosity of the solution. This relationship is given by

Here, η _s is the viscosity of the solution in which the crystal unit is immersed, and ρ _s is the density of the sample solution. It has been found that the fluctuation ΔF ₂ of the natural frequency is greatly influenced as the natural frequency Fr is higher. Therefore, the change amount ΔF _t of the natural frequency due to the immersion in the solution is obtained from the _equations (1) and (2).

It becomes.
Furthermore, it has been found that the equivalent series resistance (which will be described in detail later), which is one of the electrical equivalent constants of the AT-cut quartz resonator, increases with the viscosity of the immersion solution. At this time, when the increase amount of the electrical equivalent resistance value is ΔR _m , the following equation is given.

Here, A _q is the surface area of the AT-cut crystal resonator piece 1201, and K is the electromechanical coupling coefficient of the AT-cut crystal resonator.

  From the above relational expression, it is found that when a QCM biosensor is used, a minute mass and a viscosity can be detected from the natural frequency and the amount of change in electrical equivalent series resistance. That is, the viscosity can be detected from the amount of change in the equivalent resistance value by using equation (4). The minute mass is determined using the observed amount of change in natural frequency, the viscosity determined from equation (4), and equation (3).

  Actually, in the sensor system using this QCM type biosensor, it is necessary to measure the natural frequency and the electrical equivalent resistance value in a state in which the AT-cut quartz resonator is immersed in the sample solution. There are usually two methods for this purpose.

  FIG. 13 is a schematic diagram showing a first measurement method, and a frequency response having a frequency sweep function represented by an impedance analyzer and an impedance measurement function for a QCM biosensor 1301 immersed in a sample solution 1302. A characteristic measuring device 1303 is connected, and a high frequency signal is applied from the frequency response characteristic measuring device 1303 to the QCM biosensor 1301. As a result, the frequency response characteristic in the sample solution 1302 of the QCM type biosensor 1301 is detected. From the detected frequency response characteristics, the natural frequency and the electrical equivalent resistance value of the QCM biosensor are detected by the natural frequency arithmetic processing unit 1304 and the electrical equivalent constant arithmetic processing unit 1305, respectively.

  The electrical equivalent constant measured by the electrical equivalent constant arithmetic processing unit 1305 is further input to the sample solution viscosity arithmetic processing unit 1306, and the viscosity of the sample solution is detected based on the above equation (4). Is done. Further, the detected viscosity of the sample solution and the natural frequency previously detected by the natural frequency arithmetic processing unit 1304 are input to the minute mass arithmetic processing unit 1307 and are based on the above-described equations (1) to (3). Thus, the minute mass is detected, and finally, the minute mass and the viscosity of the sample solution are mainly used in the output device 1308. This measurement method is a measurement method based on the method of measuring the frequency response characteristics of a piezoelectric vibrator, and is a measurement method called a separate excitation method in the sense that it is excited by a high-frequency signal input from the outside. is there.

  FIG. 14 is a schematic diagram showing a second measurement method, and an inverting amplifier circuit 1401 is connected to a QCM type biosensor 1301 immersed in the sample solution 1302 as in FIG. It is known that when the inverting amplification circuit 1401 is connected to a normal piezoelectric vibrator, the piezoelectric vibrator spontaneously induces an excitation signal at an electrical natural frequency and can be observed as an oscillation frequency. That is, in FIG. 13, the inverting amplifier circuit 1401 and the QCM biosensor 1301 constitute a self-excited circuit 1402. Further, it is known that the oscillation frequency generated from the self-excited circuit using the AT cut crystal resonator is substantially equal to the mechanical natural frequency of the AT cut crystal resonator.

From the oscillation signal output from the self-excited circuit 1402, the self-excited frequency calculation processing device 1403 is input to detect the electrical natural frequency. Similarly, the amplitude component of the oscillation signal is detected by the self-excited signal amplitude calculation processing device 1404. The amplitude component detected by the self-excited signal amplitude calculation processing device 1404 is input by the electrical equivalent resistance calculation processing device 1405, and an electrical equivalent resistance value is detected. This electrical equivalent resistance value is input to the sample solution viscosity calculation processing device 1406, and the viscosity of the sample solution is detected based on the above-described equation (4). Finally, the viscosity and the electric natural frequency detected earlier are input to the minute mass arithmetic processing unit 1407, and the minute mass is detected based on the equations (1) to (3). It is output by the output device 1408. This method is called a self-excitation method because it applies the self-excitation phenomenon of a piezoelectric vibrator. The above is the outline of the conventional sensing system of the QCM type biosensor using the AT cut crystal resonator.
H. Muramatsu, J. et al. M.M. Dicks, E .; Tamiya and I. Karube Anal. Chem. 59 (1987), 2760-2863 G. Sauerbrey, Z .; Phys. 155 (1959), 206

  A QCM type biosensor system using an AT-cut quartz resonator can measure a minute mass and a viscosity of a solution relatively easily. In addition, since the sensor itself is compact and inexpensive, it can be used as a bio-test chip to regularly test the health condition of a small amount of sample at home or to measure the environment outdoors. It is a sensor system that has received much attention.

  For home and outdoor measurement, the other excitation method described above is not suitable for home and outdoor use due to problems such as expensive equipment, large scale, and high power consumption. The self-excited method of the sensor described in FIG. 14 has been studied. However, in this self-excited vibration method, the detection accuracy when detecting the viscosity from the amplitude value of the self-excited vibration is poor, and as a result, there is a problem that the detection accuracy of minute mass is also bad, which is a big problem in the industry. . That is, the problem to be solved by the present invention is to improve the viscosity detection accuracy.

  The biosensor measurement system, the viscosity measurement method, and the micro-mass measurement method according to the present invention are configured to intermittently drive a self-excited circuit configured using a sliding wave type piezoelectric resonator immersed in a solution. By measuring the startup time from the amplitude startup characteristics of the excitation vibration, detecting the electrical equivalent resistance value of the piezoelectric vibrator from this startup time, and measuring the saturation frequency value, the viscosity of the immersion solution and the electrode of the resonator It is characterized by detecting a minute mass adsorbed on the surface.

FIG. 11 is a characteristic diagram showing the effect of the present invention. In the intermittent drive self-excited circuit according to the present invention using an AT-cut quartz resonator as a slip-wave type piezoelectric resonator, the startup time and the viscosity of the immersion solution are shown. FIG. In FIG. 11, the vertical axis represents the self-excitation start time T _{r according to the} present invention, and the horizontal axis represents the viscosity η _s of the immersion solution.

In FIG. 11, the _Tr value is smaller near the origin on the vertical axis, and the _Tr value increases as the distance from the origin increases. As will be described in detail later, a large _Tr value means that the amplitude startup time in the self-excited circuit is long, and the electrical equivalent resistance value of the sliding wave type piezoelectric resonator immersed in the solution is large. ing. On the other hand, a large viscosity η _s of the immersion solution on the horizontal axis means that the viscosity is large. That is, a large viscosity means that the _Tr value is large. When FIG. 11 is viewed in consideration of these relationships, the _Tr value and the viscosity η _s have a high correlation. Can be considered. Therefore, as can be seen from FIG. 11, the present invention can accurately measure the viscosity of the dipping solution, so that a simple, high-precision and small-sized biosensor measurement system can be supplied. It becomes possible to check the health condition regularly and apply it to outdoor environmental measurements.

  FIG. 1 is a block diagram showing the configuration of the biosensor measurement system according to the present invention. The sliding wave type piezoelectric resonator 101 is immersed in a solution tank 103 filled with a sample solution 102. The slip wave type piezoelectric resonator 101 is connected to an inverting amplifier circuit 104, and the inverting amplifier circuit 104 and the thickness slip type resonator 101 constitute a self-excited circuit 105. The power source 106 is supplied to the inverting amplifier circuit 104 through the intermittent drive circuit 107. The self-excited circuit 105 is intermittently driven by the intermittent drive circuit 107. At this time, the intermittent drive circuit 107 has a function of intermittently supplying the power supply voltage at a desired drive interval. Further, the negative resistance variable device 108 is connected to the inverting amplifier circuit 104. The negative resistance variable device 108 selects a negative resistance value corresponding to the viscosity of the immersion solution 102.

  By starting the self-excited circuit 105 intermittently, the starting characteristic of the self-excited signal induced by the self-excited circuit 105 is detected by the starting characteristic measuring device 109. The activation characteristic detected by the activation characteristic measuring device 109 is input to the viscosity calculation processing device 110 according to the present invention. The viscosity calculation processing device 110 includes a saturation amplitude value calculation processing device 111, a startup time calculation processing device 112, an equivalent resistance value calculation processing device 113, and a viscosity coefficient calculation device 114. At this time, the saturation amplitude value calculation processing device 111 determines the amplitude saturation time corresponding to the saturation amplitude value together with the saturation amplitude value of the self-excited vibration.

  The saturation frequency calculation processing device 115 receives the self-excited vibration start characteristics previously measured by the start characteristic measuring device 109 and the amplitude saturation time calculated by the saturation amplitude value calculation processing device 111. A saturation frequency is detected from both. The viscosity of the immersion solution detected by the viscosity calculation processing device 110 is output by the viscosity output device 116 and also input to the adsorption mass calculation processing device 117. A saturation frequency detected by the saturation frequency calculation processing device 115 is also input to the adsorption mass calculation processing procedure 117. The adsorption mass calculation processing device 117 calculates the additional mass adsorbed on the sensitive film formed on the electrode portion of the thickness-sliding piezoelectric resonator from the saturation frequency and the viscosity, and outputs the result by the additional mass output device 118. . The above is the configuration of the biosensor measurement system according to the present invention.

Next, the principle of arithmetic processing performed by the viscosity arithmetic processing unit 110 according to the present invention will be described.
FIG. 2A and FIG. 2B are circuit diagrams showing an electrical equivalent circuit relating to a self-excited circuit using a piezoelectric resonator. FIG. 2A is an equivalent circuit diagram of the self-excited circuit 105 shown in FIG. 1 constituted by the sliding wave type piezoelectric resonator 101 and the inverting amplifier circuit 104. In this figure, the inverting amplifier circuit 104 includes a negative resistance 201, an active capacitor 202, and a fixed capacitor 203. Here, it is known that the negative resistance 201 has an equivalent negative value unlike a normal resistance value, and is greatly dependent on the amplitude value of the induced self-excited current 204. Therefore, the notation −ρ (i ₀ ) is used as the notation in the figure. Here, the amplitude value of the self-excited current 204 is i ₀ . Similarly, the value of the active capacitor 202 also changes depending on the amplitude value i ₀ of the self-excited current. However, since the influence can be ignored for the arithmetic processing according to the present invention, it can be regarded as a fixed value without any problem.

Next, FIG. 2B is a circuit diagram for explaining the electrical equivalent constant of the thickness-sliding resonator 101. The sliding wave type piezoelectric resonator 101 is electrically composed of four components. That is, the equivalent resistance 205, the equivalent capacitance 206, the equivalent inductance 207, and the interelectrode capacitance 208 are represented by the symbols R _m , C _m , L _m, and C ₀ , respectively. Using this electrical equivalent constant, the natural frequency F _r of the sliding wave type piezoelectric resonator can be approximately determined by the following equation.

In addition, the equivalent resistance R _m in the immersion solution is given by the following equation considering the equation (4), where R ₀ is the equivalent resistance value before immersion in the solution.

FIG. 3 is a characteristic diagram showing the relationship between the negative resistance 201 of the inverting amplifier circuit 104 described in FIG. 2A and the amplitude value i ₀ of the self-excited current 204. The vertical axis represents the negative resistance value ρ, The horizontal axis represents the amplitude value i ₀ of the self-excited current 204 induced by the self-excited circuit 105. By the way, the value on the vertical axis is ρ in the sense of the absolute value of the negative resistance value, and actually has a negative value. The value of ρ tends to decrease as the self-excited current increases, and has a maximum value when the self-excited current is zero. This maximum value is ρ _{m shown} in FIG.

Hereinafter, the starting characteristics of the self-excited vibration of the thickness-shear type piezoelectric vibrator will be described with reference to FIGS. In FIG. 2, if the self-excited current 204 generated in the thickness-shear type piezoelectric vibrator 101 is i, the amplitude value is i ₀ as described above, and the time is t,

Can be written. Here, F is a frequency. Both the amplitude i ₀ and the frequency F of this self-excited current are functions of time t. First, it has been found that the current amplitude i ₀ of the self-excited current 204 changes according to the following differential equation with respect to time t, although a detailed theoretical explanation is omitted.

Here, L _m , C ₀ , and R _m are electrical equivalent constants of the thickness-slip type piezoelectric vibrator 101 shown in FIG. 2B, and are equivalent inductance 207, interelectrode capacitance 208, and equivalent resistance 208, respectively. is there. Moreover, ρ (i ₀ ) is the negative resistance 201 shown in FIGS. Further, CL is a load capacity of the self-excited circuit 105, and is a combined capacity of the active capacity 202 and the fixed capacity 203 shown in FIG.

Determined by This equation (8) is a differential equation that determines the starting characteristic of the amplitude of the self-excited current of the thickness-sliding piezoelectric vibrator. The condition that the self-excited current increases with time, that is, the condition that the self-excited vibration is induced is as follows:

It is. When the self-excited vibration reaches a stable state, the amplitude value i ₀ of the self-excited current 204 is saturated and becomes a constant value. If this saturation amplitude value i _m, i _m is determined by the following equation.

Considering FIGS. 3 and (8), when the amplitude value i ₀ is small, the (8) time rate of change of amplitude given by the equation is large, the amplitude rises rapidly, the amplitude saturation value i _m since the time rate of change becomes close when the amplitude value becomes small, the rise of the amplitude is reduced, it is clear tendency to converge to the saturation value i _m and sufficient time has passed.

Next, with respect to the frequency F in the equation (5), the detailed theoretical explanation is omitted as in the case of the amplitude value, but F is saturated to a constant value in conjunction with the saturated amplitude. If this saturation value is F _m

It becomes. This F _m is a saturation frequency in the self-excited circuit, and is generally a physical quantity that is recognized as a self-excited frequency or an oscillation frequency. Changes in the natural frequency F _r due to the minute mass and viscosity determined by the equations (1) to (3) can be detected as changes in the saturation frequency F _m of this self-excited circuit.

The equivalent resistance value R _m of the sliding wave type piezoelectric resonator 101 immersed in the sample solution 102 increases due to the influence of its viscosity as shown by the equation (4). This increase in equivalent resistance gives a change to the starting characteristic determined by equation (8). Therefore, the viscosity can be detected by detecting this starting characteristic.

FIG. 4 is a qualitative characteristic diagram of the starting characteristic according to the present invention determined by the equation (8), and FIG.
The vertical axis in FIG. 4 represents the amplitude i _{0 of the} excitation current, and the horizontal axis represents time T. Time is saturated amplitude becomes sufficiently long, the value is i _m of FIG forth. FIG. 4B is a characteristic diagram that is calculated from the start-up characteristic shown in FIG. 4A, and the vertical axis indicates the time derivative of the amplitude i ₀ of the excitation current given by equation (8). The value di ₀ / dT. In these characteristic diagrams, the start time Tr representing the start characteristic is a time τ _A indicating a value half of the im value in FIG. 4A, and an amplitude amplitude of the excitation current in FIG. 4B. The time τ _B when the time differential value di ₀ / dT of i ₀ becomes the maximum value. As for the starting time for measuring the viscosity according to the present invention, it has been found that the times τ _A and τ _B have a high correlation with the viscosity according to the present invention.

Figure 6 is a characteristic diagram showing the relationship between the startup time T _r the equivalent resistance value R _m, is a calculated value calculated from the equation (8). In the calculation, an AT-cut quartz resonator having a natural frequency of 19.2 MHz having the electrical equivalent constant values described in Table 1 is assumed. FIG. 6 is created based on the startup time τ _B shown in FIG. 4B. However, even if it is created based on the startup time τ _A shown in FIG. It goes without saying that characteristic charts with trends can be obtained.

As the negative resistance of the self-excited circuit, two types of 1 KΩ and 2 KΩ were used with ρ _m as a representative value as shown in FIG. Furthermore C _L value is 100pF value. Further, FIG. 7 is a characteristic diagram showing the relationship of the i _m and an equivalent resistance value R _m, is a calculated value calculated under the same conditions as FIG. Incidentally, FIG. 7 corresponds to a method of detecting the viscosity of the sample solution from the amplitude value of the self-excited vibration described in FIG.

In FIG. 6, the relationship between the start time _Tr and the equivalent resistance R _m is greatly different depending on the value of ρ _m , as shown by the characteristic curve 601 and the characteristic curve 602. Where [rho _m = corresponding curve 1KΩ characteristic curve 601, the curve corresponding to [rho _m = 2K ohms is a characteristic curve 602. Further, the gradient is very large in the region where the R _m value approaches the ρ _m value, that is, in the region I in the characteristic curve 601 and in the region II in the characteristic curve 602. Incidentally, the gradient is 40 μS / Ω in the region I and 15 μS / Ω in the region II, and can be regarded as a sufficiently large value compared to the normal time resolution that can be measured. It turns out that it has detection accuracy. Furthermore, it has also been found that the detection sensitivity can be freely selected by selecting an appropriate negative resistance value for the assumed viscosity of the sample solution.

In contrast, shown in FIG. 7, the relationship between the saturation current amplitude value i _m and equivalent resistance R _m, as indicated by characteristic curve 701 and curve 702, it can be regarded as substantially straight both. Where [rho _m = corresponding curve 1KΩ characteristic curve 701, the curve corresponding to [rho _m = 2K ohms is a characteristic curve 702. Furthermore, the gradient of both characteristic curves is 3 μA / Ω to 8 μA / Ω. Compared with the measurement resolution for ordinary high-frequency signals, it has been found that this value does not provide sufficient detection accuracy. That is, by comparing the FIG. 6 and FIG. 7 to detect the starting characteristic of the self-excited current induced by the self-excited circuit according to the present invention, it is possible to measure the viscosity with higher accuracy compared to the conventional method. Things turned out. The above is the basic explanation of the arithmetic processing performed by the viscosity arithmetic processing unit 110 and the basic features thereof.

  Below, the principal part in the structure of the biosensor measuring system which concerns on this invention shown in FIG. 1 is demonstrated. First, the self-excited circuit 105 and its peripheral devices shown in FIG. 1 will be described. FIG. 8 shows an embodiment of a self-excited circuit composed of an inverting amplifier circuit and a sliding wave type piezoelectric resonator. With reference to this example, the self-excited circuit 105 shown in FIG. This will be described more specifically. A self-excited circuit 801 shown in FIG. 8 (corresponding to the self-excited circuit 105 shown in FIG. 1) is an embodiment of a self-excited circuit constituted by an inverting amplifier circuit using a transistor and a sliding wave type piezoelectric resonator 101. It is. The start-up characteristic measuring device 803 according to the present invention (corresponding to the start-up characteristic measuring device 109 shown in FIG. 1) is provided at both ends of a reference resistor 802 to measure the self-excited current induced in the sliding wave type piezoelectric resonator. It is connected. A bias voltage for driving each transistor is connected to the power source 106 via an intermittent drive circuit 804 (corresponding to the intermittent drive circuit 107 shown in FIG. 1). The voltage is intermittently driven, and as a result, the self-excited circuit 801 is intermittently driven. The negative resistance variable device 805 (corresponding to the negative resistance variable device 108 shown in FIG. 1) changes the resistance, capacitance value, and the like constituting the self-excited circuit shown in FIG. It is possible to vary the negative resistance.

  FIG. 9 shows an embodiment of another self-excited circuit composed of an inverting amplifier circuit and a sliding wave type piezoelectric resonator. The self-excited circuit 901 shown in FIG. 9 (the self-excited circuit 105 shown in FIG. Is an example of a self-excited circuit composed of an inverting amplifier circuit using a CMOS inverter and a sliding wave type piezoelectric resonator 101. A starting characteristic measuring apparatus 903 (corresponding to the starting characteristic measuring apparatus 109 shown in FIG. 1) according to the present invention is provided at both ends of a reference resistor 902 in order to measure a self-excited current induced in a sliding wave type piezoelectric resonator. It is connected. A driving voltage for driving each CMOS is connected to the power source 106 via an intermittent driving circuit 904 (corresponding to the intermittent driving circuit 107 shown in FIG. 1), and the intermittent driving circuit 904 biases the bias voltage. The voltage is intermittently driven, and as a result, the self-excited circuit 901 is intermittently driven. The negative resistance variable device 905 (corresponding to the negative resistance variable device 108 shown in FIG. 1) changes the resistance, capacitance value, and the like constituting the self-excited circuit shown in FIG. It is possible to vary the negative resistance.

It is needless to say that the configuration of the self-excited circuit shown in FIGS. 8 and 9 can be conveniently changed depending on the viscosity of the sample solution to be used, the frequency of the piezoelectric resonator, and the like. Secondly, the startup output waveform output from the startup characteristic measuring apparatus 109 shown in FIG. 1 will be described. FIG. 10 is a diagram for explaining the self-excited waveform measured by the start-up characteristic measuring apparatus 109 according to the present invention and the drive voltage waveform changed by the intermittent drive circuit 107. The vertical axis is output intensity, and the horizontal axis is time. The intermittent operation starts at times T ₁ and T ₂ , and the drive voltage is supplied to the self-excited circuit 105 at both time intervals ΔT. Of the drive voltage waveforms 1001, the first drive state is the drive state 1002, and the second drive state is the drive state 1003. At this time, the self-excitation waveform output corresponding to the driving state 1002 is the self-excitation waveform 1004, and the self-excitation waveform output corresponding to the driving state 1003 is the self-excitation waveform 1008. At this time, the driving state 1002 reflects the reaction state between the immersion solution 102 and the sliding wave type piezoelectric resonator 101 at the time T ₁ , and the driving state 1003 reflects both reaction states at the time T ₂ .

The startup output waveform output from the startup characteristic measuring apparatus 109 shown in FIG. 1 according to the present invention is a drive voltage waveform represented by the drive voltage waveform 1001, the self-excited waveform 1004, and the self-excited waveform 1008 shown in FIG. It is a self-excited waveform. FIG. 10 is a schematic diagram for measuring changes in the viscosity of the immersion solution and the adsorbed minute mass at time T ₁ and time T ₂ .

  Thirdly, each component device constituting the viscosity calculation processing device 110 according to the present invention will be described. First, in the saturation amplitude value calculation processing device 111 as the first component device, the envelope of the self-excited waveform shown in FIG. 10 is determined. That is, in the envelope 1005 and the envelope 1009 illustrated in FIG. 10, the former corresponds to the driving state 1002 and the latter corresponds to the driving state 1003. Furthermore, based on this envelope information, a region where the excitation amplitude is sufficiently saturated is detected. That is, the saturation region 1006 and the saturation region 1010 illustrated in FIG. A saturation amplitude value is determined in the determined saturation region. This saturation amplitude value is the saturation amplitude value 1007 and the saturation amplitude value 1011 shown in FIG.

The start-up time calculation processing device 112, which is the second component device, is a device that performs calculation processing for obtaining the start-up time using the envelope 1005, the envelope 1010, and the drive voltage waveform 1001. The activation time Tr is defined as a time when the time differential value di ₀ / dT of the amplitude i ₀ of the excitation current becomes the maximum value. That is, it is the time when the envelope 1005 and the envelope 1010 shown in FIG. 10 are differentiated with respect to time and the value becomes the maximum value. This time is time T _a and time T _b shown in the figure. Using these two times, the activation time T _r1 in the driving state 1002 is

It becomes. Similarly, the activation time T _r2 in the driving state 1003 is

It becomes.

The above is the arithmetic processing configured based on the activation time τ _B described with reference to FIG. 4B. However, when the activation time τ _A illustrated in FIG. Based on the amplitude value 1007 and the saturation amplitude value 1011, the activation times T _r1 and T _r2 may be obtained from the time corresponding to the half width of both. The criterion for determining the superiority or inferiority of both is the correlation between the startup time and the viscosity according to the present invention, and the higher correlation should be adopted. Since this correlation depends on the apparatus constituting the arithmetic processing unit and the circuit configuration shown in FIGS. 8 and 9, it is merely a design matter.

The equivalent resistance calculation processing device 113, which is a third component device, is a device that performs calculation processing for obtaining an equivalent resistance value in the driving state 1002 and the driving state 1003 using the start times T _r1 and T _r2 . In order to determine the equivalent resistance value R _m , the equation (8) is used as a basis. That is, the negative resistance curve and load capacitance value C _{L of} the self-excited circuit 105 set by the negative resistance variable device 108, and the equivalent inductance 207 and interelectrode capacitance of the slip wave type piezoelectric resonator 101 recorded in advance are recorded. Based on the value of 208 (that is, the values of L _m and C ₀ ), the equivalent resistance R _m is determined by numerically solving equation (8). The determined equivalent resistance value is R _m1 in the driving state 1002 and R _m2 in the driving state 1001.

A viscosity calculation device 114, which is a fourth component device, is a device that performs calculation processing for obtaining the viscosity of the immersion solution using the equivalent resistance values R _m1 and R _m2 . Using the equivalent resistance value R ₀ , the natural frequency F _r , the resonator shape, the electromechanical coupling coefficient, the density of the immersion solution, and the equation (6), which are recorded in advance, of the sliding wave type piezoelectric resonator 101 before the solution immersion, The viscosity of the dipping solution is determined. This determined viscosity is η _S1 in the driving state 1002 and η _S2 in the driving state 1001. At this time, if there is a difference between η _S1 and η _S2 , it means that the viscosity of the immersion solution has changed due to the reaction between the chemical components of the immersion solution and the sensitive film of the sliding wave type piezoelectric resonator 101. The determined viscosity or the amount of change in viscosity is output by the viscosity output device 116.

Fourth, the saturation frequency calculation processing device 115 according to the present invention will be described. This saturation frequency arithmetic processing unit 115 is an arithmetic processing unit for determining the self-excited frequency in the saturation region shown in FIG. 10, that is, the saturation frequency. In order to determine the self-excited frequency in the saturation region, the saturation frequency is measured based on the saturation region 1006 and the saturation region 1010 determined by the saturation amplitude value calculation processing device 111. The determined saturation frequency is F ₁ in the driving state 1002 and F ₂ in the driving state 1003.

Fifth, the adsorption mass calculation processing device 117 will be described. The adsorption mass calculation processing device 117 is a device for determining a minute mass adsorbed on the sensitive film of the sliding wave type piezoelectric resonator 101. To determine the small mass of adsorbing, viscosity eta _S1 of the driving state 1002 is determined by the viscosity ratio calculating unit 114, a viscosity eta _S2 in the driving state 1003, (1) and (2) Is used to determine the adsorption mass. The determined adsorption mass is Δm ₁ in the driving state 1002 and Δm ₂ in the driving state 1003. At this time, if there is a difference between Δm ₁ and Δm ₂ in the same manner as the viscosity, it means that the adsorption amount has changed due to the reaction between the immersion solution and the sensitive film of the sliding wave type piezoelectric resonator 101. The determined amount of change in adsorption amount or viscosity is output by the adsorption mass output device 118.

The above calculation processing has been described mainly with respect to the calculation processing in which intermittent driving is performed at two time points of time T ₁ and time T ₂ , but by performing intermittent driving at more time points instead of two time points, the adsorption mass and viscosity are determined. Needless to say, time-series changes in rates can be measured. Normally, the time during which the amplitude and frequency of the self-excited waveform in FIG. 10 is saturated is several tens of milliseconds (milliseconds) or less. Therefore, the time interval of intermittent driving can be shortened to several hundred mS. Furthermore, the reaction time (adsorption time) between the sensitive film of the sliding wave type piezoelectric resonator and the immersion solution is very slow compared to the time interval of this intermittent drive. Therefore, a substantially continuous change can be measured by narrowing the intermittent drive time interval to some extent. In addition, by adding a function to change the negative resistance curve by the negative resistance variable device 108, the viscosity detection sensitivity can be freely selected, so that the highly sensitive viscosity detection is possible regardless of the type of immersion solution. As a result, highly sensitive detection of the adsorbed minute mass can be realized.

  FIG. 11 showing the effect of the present invention is a result of using a QCM type biosensor using an AT-cut quartz resonator as described above, but is a slip wave type elasticity using a crystal single crystal having a higher self-excited frequency. By using a surface wave resonator (SH wave crystal SAW resonator), a more sensitive sensor system can be realized. When this SH wave quartz SAW resonator is used, the vibration mode becomes a slip wave mode localized on the quartz surface, so that the equations (1) to (4) described above cannot be used as they are. By using the start-up time obtained from the drive type self-excited circuit, it is still possible to measure highly accurate and highly sensitive viscosity.

The block diagram which shows the structure of the biosensor measurement system which concerns on this invention Circuit diagram showing electrical equivalent circuit for self-excited circuit using piezoelectric resonator Characteristic view showing the relationship of the amplitude value i ₀ of the negative resistance and self-excited current according to the present invention Qualitative characteristic diagram of starting characteristics according to the present invention Negative resistance curve diagram for calculating start-up characteristics according to the present invention Characteristic diagram showing the relationship between activation time T _r the equivalent resistance value R _m according to the present invention Characteristic diagram showing the relationship between the amplitude saturation value i _m and equivalent resistance R _m Embodiment of self-excited circuit according to the present invention Another embodiment of the self-excited circuit according to the present invention Characteristics diagram of self-excited waveform and drive voltage waveform measured by the starting characteristic measuring apparatus according to the present invention Characteristics chart showing effects of the present invention Schematic diagram of QCM biosensor using AT-cut quartz crystal Conceptual diagram for explaining a conventional sensing system using another excitation method Conceptual diagram for explaining a conventional self-excited sensing system

Explanation of symbols

DESCRIPTION OF SYMBOLS 101: Sliding wave type piezoelectric resonator 102: Sample solution 103: Solution layer 104: Inversion amplifier circuit 105: Self-excited circuit 106: Power supply 107: Intermittent drive circuit 108: Negative resistance variable device 109: Starting characteristic measuring device 110: Viscosity Rate calculation processing unit 111: Saturation amplitude value calculation processing unit 112: Start-up time calculation processing unit 113: Equivalent resistance value calculation processing unit 114: Viscosity calculation unit 115: Saturation frequency calculation processing unit 116: Viscosity output unit 117: Adsorption mass Arithmetic processor 118: adsorption mass output device

Claims (10)

Using a self-excited circuit comprising a sliding wave type piezoelectric resonator having an excitation electrode on which a sensitive film that adsorbs a specific substance in a sample solution is formed, and an inverting amplifier circuit connected to the sliding wave type piezoelectric resonator In the biosensor measurement system for measuring a minute mass adsorbed on the sensitive film,
A starting characteristic measuring unit that intermittently drives the self-excited circuit to measure a starting characteristic related to an amplitude value and a frequency value of the self-excited vibration;
The startup time from when the self-excited circuit is started up to a predetermined amplitude value is calculated from the measured amplitude characteristic, and the viscosity of the sample solution is calculated using the calculated value. A viscosity calculation processing unit;
The frequency value at the time when the waveform amplitude of the self-excited circuit is saturated from the measured frequency characteristic of the starting characteristic, and using the detected frequency value and the viscosity obtained by calculation An adsorption mass calculation processing unit for calculating the trace mass value adsorbed on the sensitive film;
A biosensor measurement system comprising:   The viscosity calculation processing unit includes a saturation amplitude value calculation processing unit that calculates a saturation amplitude value at a time when the amplitude is saturated from an amplitude characteristic among the measured starting characteristics, and the self-excited circuit is started. An activation time calculation processing unit for calculating an activation time until the amplitude reaches a predetermined amplitude value smaller than the saturation amplitude value, and an equivalent resistance value of the sliding wave type piezoelectric resonator using the calculated activation time value The bio-resistor according to claim 1, further comprising: an equivalent resistance value calculation processing unit that calculates the viscosity, and a viscosity rate calculation unit that calculates the viscosity of the sample solution using the calculated equivalent resistance value. Sensor measurement system.   The biosensor measurement system according to claim 2, wherein the predetermined amplitude value is an amplitude value at which a time differential value of the amplitude value becomes a maximum value.   The biosensor measurement system according to claim 2, wherein the predetermined amplitude value is an amplitude value that is a half value of the saturation amplitude value.   5. The device according to claim 1, further comprising a negative resistance variable device that is connected to the self-excited circuit and changes a negative resistance value so as to correspond to the viscosity of the sample solution. Biosensor measurement system.   The biosensor measurement system according to any one of claims 1 to 5, wherein the sliding wave type piezoelectric resonator is an AT cut type resonator using a crystal single crystal.   6. The biosensor measurement system according to claim 1, wherein the sliding wave type piezoelectric resonator is a sliding wave type surface acoustic wave resonator using a crystal single crystal. A viscosity measurement method for measuring the viscosity of a sample solution,
A first step of measuring a starting characteristic related to an amplitude value and a frequency value of the self-excited vibration obtained by self-excited vibration of a sliding wave type piezoelectric resonator in a sample solution and intermittent driving;
A second step of measuring an activation time from when the self-excited vibration is activated until reaching a predetermined amplitude value from an amplitude characteristic of the measured activation characteristics;
A third step of calculating and measuring the viscosity of the sample solution by calculating using the measured value of the starting time and the equivalent resistance value of the sliding wave type piezoelectric resonator obtained in advance; ,
A method for measuring viscosity, comprising: A trace mass measurement method for measuring a trace mass adsorbed on the sensitive film of a sliding wave type piezoelectric resonator having an excitation electrode formed with a sensitive film that adsorbs a specific substance in a sample solution,
Measuring a starting characteristic related to an amplitude value and a frequency value of the self-excited vibration obtained by self-excited vibration of the sliding wave type piezoelectric resonator in the sample solution and intermittent driving;
The starting time from when the self-excited vibration starts up to a predetermined amplitude value is measured from the measured amplitude characteristics of the starting characteristics, and the viscosity of the sample solution is measured using the measured starting time value. Calculating a rate;
The frequency value at the time when the waveform amplitude of the self-excited vibration is saturated from the measured frequency characteristic of the starting characteristic is used, and the detected frequency value and the viscosity obtained by calculation are used. Calculating and measuring the trace mass value adsorbed on the sensitive membrane;
A method for measuring a minute mass, comprising:   The step of calculating the viscosity of the sample solution is a first step of measuring a start-up time from when the self-excited vibration starts up to a predetermined amplitude value based on the amplitude characteristic among the measured start-up characteristics. And calculating the viscosity coefficient of the sample solution by calculating using the measured value of the starting time and the equivalent resistance value of the sliding wave type piezoelectric resonator obtained in advance. The method according to claim 9, further comprising: a step.

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