JP5102334B2 - Sensing device - Google Patents

Description

  The present invention relates to a sensing device for sensing a sensing object contained in a sample liquid based on an oscillation frequency of a piezoelectric vibrator such as a crystal vibrator.

  As a sensing device for sensing and measuring a trace substance contained in a sample solution, a device using a quartz sensor is known. As this type of sensing device, there is known a device that detects a sensing object while flowing a sample liquid as disclosed in Patent Document 1. On the other hand, as a quartz sensor, a twin sensor type quartz sensor that can reduce the influence of external factors such as temperature on the measurement environment is known (Patent Document 2). In this crystal sensor, a measurement vibration region and a reference vibration region are formed on a crystal piece, and an adsorption layer made of a biological material or the like is formed on the surface of the measurement vibration region. As a result, only the measurement vibration region receives the mass change due to the adsorption of the sensing object, so the difference between the oscillation frequency extracted from the measurement vibration region and the oscillation frequency extracted from the reference vibration region is obtained. Thus, it is possible to obtain a highly accurate measurement result from which external factors have been removed.

  Here, the applicant of the present application, as an example of a technique for measuring a very small amount of substance with higher sensitivity and accuracy, provides the height of the reaction channel (from the surface of the crystal unit to the opposite surface in the case body). To reduce the distance). For example, even if the height of the reaction channel is as small as 1 mm, the ratio of the antigen in the liquid flow on the opposite surface side to react with the antibody is higher than the same ratio of the antigen in the liquid flow on the crystal oscillator side. small. For this reason, among the sensing objects contained in the supplied sample liquid, the ratio of the sensing objects adsorbed on the adsorption layer is small, which cannot be said to be advantageous from the viewpoints of both sensitivity and accuracy.

  Therefore, by setting the distance between the surface of the quartz sensor and the opposing surface, that is, the height of the reaction channel, to be, for example, 0.2 mm or less, the sample liquid is brought into contact with the adsorption layer of the quartz piece or its It is conceivable to increase the proportion of the sample solution flowing in the vicinity. By doing so, the amount of the sensing object contained in the sample liquid adsorbed on the adsorption layer increases, the amount of the sensing object discharged without being adsorbed decreases, and the measurement sensitivity and accuracy of the sensing object. Will improve. In addition, reducing the height of the reaction channel leads to a decrease in the volume of the reaction channel, and the amount of sample solution required for measurement can be kept low, so that dilution of the sample solution is not necessary. This also contributes to improvement in measurement sensitivity and accuracy of the sensing object.

  However, when the case where the height of the reaction channel is made very low is applied to the above-described twin sensor type quartz sensor, two vibration regions (for example, an adsorption layer made of a biological material or the like is formed on the surface). Due to the difference in wettability between the hydrophilic measurement vibration region and the hydrophobic reference vibration region made of gold), as shown in FIG. 13, the more hydrophobic vibration region of the two vibration regions, for example, Bubbles remain without being discharged on the gold reference vibration region side, making it difficult to perform highly reliable measurement.

JP 2008-58086 (paragraph 0014, FIGS. 11 and 13) JP2007-108170

  The present invention has been made under such circumstances, and an object thereof is to provide a sensing device capable of performing a highly reliable measurement in a sensing device that measures a sensing object while flowing a sample liquid. There is.

In the sensing device of the present invention, two pairs of excitation electrodes are arranged in the X direction so as to form a first vibration region and a second vibration region on a common piezoelectric piece, and the excitation electrode on one surface side of the piezoelectric piece is provided. An adsorption layer that adsorbs a sensing object in the sample liquid is formed in the first vibration area, and a sensing object in the sample liquid is formed using a piezoelectric sensor that does not form an adsorption layer in the second vibration area on the one surface side. In a sensing device that senses objects,
The piezoelectric sensor includes a facing surface that is in close contact with the one surface side of the piezoelectric sensor and faces the one surface side including the first vibration region and the second vibration region through a gap, and has a height in a region facing the one surface side. A flow path forming member for forming a reaction flow path of 0.3 mm or less ;
A liquid supply port provided on one end side in the Y direction orthogonal to the X direction in the reaction channel, and for supplying a sample solution to the reaction channel;
A liquid discharge port provided on the other end side in the Y direction in the reaction channel, for discharging the sample liquid from the reaction channel;
In order to control the liquid flow, the first vibration region is hydrophilic and the second vibration region is hydrophobic depending on the presence or absence of the adsorbing layer in order to control the liquid flow. A groove portion formed on the opposite surface so as to extend along the liquid flow direction in the reaction channel ,
The first vibration region and the second vibration region are provided for sensing a sensing object based on the oscillation frequency when the piezoelectric sensor is oscillated by an oscillation circuit. And

Moreover, the following example can be given as a specific example of the sensing device. 1. The groove portion includes a first groove portion extending from the liquid discharge port side toward the first vibration region, and a second groove portion extending from the liquid discharge port side toward the second vibration region, The end portions on the vibration region side in the first groove portion and the second groove portion are positioned so as not to overlap the first vibration region and the second vibration region, respectively.

At this time, in the present invention, the first groove portion and the second groove portion may be formed such that the distance from each other increases from the liquid discharge port toward the vibration region side when viewed from above,
Furthermore, the first groove is close to the line connecting the liquid supply port and the liquid discharge port from the edge of the reaction channel extending from the liquid discharge port to the first vibration region side,
The second groove may be closer to the line side from the edge of the reaction channel extending from the liquid discharge port to the second vibration region side.

  In the present invention, the reaction channel is formed so that the measurement vibration region (first vibration region) and the reference vibration region (second vibration region) face the twin-type piezoelectric sensor arranged in the X direction. In the sensing device in which the inlet and outlet of the sample liquid in the reaction channel are arranged in the Y direction, a groove is formed on the outlet side of the opposing surface of the piezoelectric sensor, and the difference in the wettability between both vibration regions The bias of the liquid flow based on it is suppressed. Therefore, it is possible to prevent bubbles from remaining in the reaction channel and to perform highly reliable measurement.

It is the schematic which shows the whole structure of the sensing apparatus which concerns on this invention. It is a disassembled perspective view which shows the sensor unit provided with the piezoelectric sensor which concerns on this invention. It is a longitudinal side sectional view showing the sensor unit. It is the surface view and back view which show the crystal oscillator which comprises some piezoelectric sensors. It is a block diagram explaining the connection of the crystal oscillator and frequency measurement part which comprise some sensing devices. It is a vertical side sectional view which expands and shows the sensor unit. It is a perspective view which shows the back surface and surface of a flow-path formation member which comprises a part of said sensor unit, respectively. It is explanatory drawing which shows the positional relationship of a groove part and an excitation electrode when the said flow-path formation member and a crystal oscillator contact | abut. It is A-A 'arrow sectional drawing which shows the cross section of the groove part in FIG. It is a top view which shows the inflow state of the liquid for time explaining the effect | action of the said groove part. It is a top view of the flow-path formation member which shows the example of the other implementation shape regarding the said groove part. It is a top view of the flow-path formation member which shows the example of the other implementation shape regarding a recessed part. It is a top view explaining a state when a sample liquid is made to flow in into a sensor unit provided with a channel formation member without a groove part.

  As shown in FIG. 1, an embodiment of the sensing device of the present invention includes a sensor unit 2, a liquid supply system 1 that supplies a liquid (sample liquid or buffer solution) to the sensor unit 2, and a discharge from the sensor unit 2. A liquid discharge system 90 for storing the liquid to be stored, a crystal sensor 7 which is a piezoelectric sensor attached to the sensor unit 2, and an oscillation circuit unit 6 provided adjacent to the sensor unit 2 are provided.

  As shown in FIG. 2, the sensor unit 2 is configured by laminating a support body 21, a wiring board 3, a crystal resonator 4, a flow path forming member 5, and a cover body 24 in this order from the lower side. The support 21 is formed with a support recess 22 for fitting and holding the crystal sensor 7 and the flow path forming member 5. Accordingly, the channel forming member 5 is pressed against the crystal sensor 7 in a state where the crystal sensor 7 is fitted in the support recess 22, so that the lower surface of the channel forming member 5 presses the crystal resonator 4 against the wiring substrate 3. And fixed. Furthermore, the support body 21 is covered with a cover body 24 from above.

  2 and 3, reference numeral 26 denotes a liquid supply pipe, and 27 denotes a liquid discharge pipe as discharge means. The liquid supplied from the liquid supply system 1 to the reaction channel 52 via the liquid supply pipe 26 is a liquid discharge pipe. 27 and the liquid discharge system 90 is discharged. The liquid supply system 1 includes a buffer solution supply unit and a sample solution supply unit that respectively supply a buffer solution and a sample solution as a reference solution to the quartz sensor 7, and the liquid discharge system 90 includes a waste solution storage unit. .

  The crystal sensor 7 is configured by providing a crystal resonator 4 that is a piezoelectric resonator on a wiring board 3. As shown in FIG. 4, the crystal resonator 4 is configured by providing excitation electrodes 42 and 43 on both surfaces of a disk-shaped crystal piece 41 which is a piezoelectric piece. In FIG. 4B, the first excitation electrode 43A and the second excitation electrode 43B are arranged so as to be spaced apart from each other and aligned in the X direction, and the two excitation electrodes 43A are disposed on the surface side (FIG. 4A). , 43B are provided with a common excitation electrode (common electrode) 42. Therefore, the first vibration region 4A is formed by the first excitation electrode 43A and the common electrode 42, and the second vibration region 4B is formed by the second excitation electrode 43B and the common electrode 42. The equivalent thickness of the common electrode 42 and the excitation electrodes 43A and 43B is, for example, 0.2 μm, and the electrode material is, for example, gold or silver. When the crystal sensor 7 is attached to the sensor unit 2, the first excitation electrode 43A and the second excitation electrode 43B have two oscillations via the conductive paths 32 and 34 of the wiring board 3, as shown in FIG. The common electrode 42 is connected to the ground side of the oscillation circuits 6A and 6B through the conductive path 33 of the wiring board 3 while being connected to the circuits 6A and 6B, respectively. Connection terminals 35 to 37 connected to the respective conductive paths 32 to 34 are formed in the end region of the wiring board 3.

  In the region corresponding to the first excitation electrode 43A in the common electrode 42 of the quartz sensor 7, an adsorption layer (reactant) 46 made of an antibody for adsorbing an antigen that is a sensing object is formed. Therefore, for example, when a sensing object in the sample liquid is adsorbed to the adsorption layer 46, the oscillation frequency in the first vibration region 4A is reduced due to the mass load effect. On the other hand, in the second vibration region 4B, the surface of the common electrode is not provided with the adsorption layer 26 and is exposed, so that the sensing object is not adsorbed to the common electrode 42. For this reason, by comparing the oscillation frequencies of the regions 4A and 4B before and after the adsorption of the sensing object, the ambient temperature of the sensor unit 2, the viscosity of the sample liquid, and adhesion of substances other than the sensing object contained in the sample liquid Thus, it is possible to detect the change (decrease) in the oscillation frequency corresponding to the amount of the sensing object adsorbed on the adsorption layer 46 while suppressing the influence of disturbance such as the above. The surface of the common electrode in the region that forms the second vibration region is not exposed, and a blocking layer made of a protein that does not react with the sensing object, for example, may be formed.

  Next, the flow path forming member 5 will be described with reference to FIGS. FIG. 6 is an enlarged vertical cross section of the sensor unit 2. 7A and 7B show the back surface side and the front surface side of the flow path forming member 5, respectively, and the height of the recess 52 is exaggerated. The flow path forming member 5 is formed in a shape corresponding to one end side of the wiring board 3 using an elastic material such as polydimethylsiloxane (PDMS). A recess 52 is formed in the center of the back surface side of the flow path forming member 5, and the flow path forming member 5 and the wiring substrate 3 are overlapped so that the back surface side of the flow path forming member 5 is in contact with the wiring substrate 3. By pressing against the crystal unit 4, the recess 52 becomes a reaction channel. That is, at this time, the ceiling surface of the recess 52 is an opposing surface that opposes the vibration region on the surface side that is one surface side of the crystal resonator 4 via a gap, and the reaction channel is the opposing surface and the crystal resonator 4. Is formed so as to be surrounded by the inner peripheral surface along the contour of the recess 52 in the region between the two. For this reason, reference numeral 52 is used for both the recess and the reaction channel. The recess 52 has a rhombus shape with curved corners so that a dead space does not occur in the liquid flow in the reaction channel 52. The central portion of the recess 52 is set to be slightly larger than the region including the first vibration region 4 </ b> A and the second vibration region 4 </ b> B of the crystal unit 4, and the flow path forming member 5 is brought into contact with the wiring substrate 3. The regions 4 </ b> A and 4 </ b> B are accommodated in the central portion in the recess 52. And the height of the said recessed part 52 is set to 0.3 mm or less, for example, and is set to 0.15 mm in this example. A flow path is formed in a direction (Z direction) perpendicular to the facing surface from the back surface to the front surface of the flow path forming member 5 at locations corresponding to both ends of the diagonal line in the Y direction of the facing surface of the recess 52. ing. One of the channels perpendicular to the facing surface is the liquid supply port 53 and the other is the liquid discharge port 54.

  As shown in FIGS. 8 and 9, two opposing surfaces of the recess 52 extend linearly from the liquid discharge port 54 toward the first vibration region 4 </ b> A and the second vibration region 4 </ b> B. The groove portions 55A and 55B are formed. These groove portions 55A and 55B are formed so as to face the crystal portion of the crystal unit 4, and the end portions on the vibration regions 4A and 4B side of the groove portions 55A and 55B do not face the vibration regions 4A and 4B, respectively. positioned. Further, these groove portions 55A and 55B are formed so that the mutual distance (distance in the X direction) increases from the liquid discharge port 54 toward the vibration regions 4A and 4B, respectively, and the overall planar shape is V-shaped. And from the edge of the reaction channel 52 extending from the liquid discharge port 54 to each vibration region 4A, 4B side, approaching the line side connecting the liquid supply port 53 and the liquid discharge port 54. In this example, the dimensions of the two groove portions 55A and 55B are set to, for example, a length of 3 mm, a width of 0.5 mm, and a depth of 0.5 mm.

  Further, the oscillation circuit unit 6 is electrically connected to the connection terminal portion of the wiring board 3 as shown in FIG. 5 by being inserted into the sensor unit 2. A measurement circuit unit 81 and a data processing unit 82 are provided following the oscillation circuit unit 6. The measurement circuit unit 81 has a function of measuring an oscillation frequency by digitally processing, for example, a frequency signal that is an input signal. The measurement circuit unit 81 may be a frequency counter, and the measurement method can be selected as appropriate. The data processing unit 82 is a part that stores time-series data of measured frequencies and displays the time-series data, and is composed of, for example, a personal computer.

  Next, the operation of the sensing device will be described with reference to FIG. A liquid is supplied from the liquid supply system 1 into the sensor unit 2, and the liquid flows into the reaction channel 52 from the liquid supply port 53. Thereafter, the inflowing liquid radially spreads in both sides toward the liquid outlet 54 in the reaction channel 52 ((a) in FIG. 10), and the first vibration region 4A and the second vibration region 4B. Is reached, the liquid flow is biased toward the hydrophilic first vibration region 4A due to the difference in wettability between the vibration regions 4A and 4B ((b) in FIG. 10). If there is no groove 55A, most of the liquid passes through the hydrophilic vibration region 4A in this way, and a part of the liquid flows from the vicinity of the liquid discharge port 54 toward the hydrophobic vibration region 4B. As a result, bubbles may remain on the hydrophobic vibration region 4B side, which causes a decrease in the reliability of measurement.

  On the other hand, in the present sensing device, when the reaction channel 52 was actually produced with the dimensions of this embodiment, no bubbles were observed. The reason is presumed as follows. Due to the presence of the groove 55A, an action is exerted on the liquid to be spread from the lower side of the groove 55A in the reaction channel 52 to both sides due to the influence of the surface tension. As a result, one-sided outflow of liquid from the hydrophilic region 4A side occurs. It is suppressed. For this reason, the driving force for the penetration of the liquid into the hydrophobic region 4B is increased, and the liquid begins to penetrate into the hydrophobic region 4B ((c) in FIG. 10). Thereafter, the liquid from the hydrophilic region 4A side passes through the extraction electrode 44 made of hydrophobic gold and enters the hydrophobic region 4B side, but again, the liquid is blocked by the groove portion 55B on the hydrophobic region 4B side and enters the groove portion 55B. Along the direction opposite to the liquid discharge port 54.

  Meanwhile, the liquid from the hydrophobic region 4B side also reaches the groove 55B on the hydrophobic region 4B side, and is sandwiched between the groove 55B on the hydrophobic region 4B side and the edge of the reaction channel 52 on the hydrophobic region 4B side. Since the region that has been functioning as a bubble escape route to the end ((d) in FIG. 10), the bubble does not remain, and the uniform extrusion flow in both vibration regions 4A and 4B of the reaction channel 52 Is realized. Thereafter, until the liquid enters the area below the groove portions 55A and 55B, the liquid flow is concentrated near the edge of the reaction channel 52 where bubbles are likely to remain, and the liquid flow velocity is increased. This also contributes to the prevention of residual bubbles. Further, since the grooves 55A and 55B are not applied to both the vibration areas 4A and 4B, respectively, for example, there is no hindrance to the measurement that disturbs the flow of the liquid in the vibration areas 4A and 4B. Then, the liquid that has reached the liquid discharge port 54 is discharged to the liquid discharge system 90.

  On the other hand, the first vibration region 4 </ b> A and the second vibration region 4 </ b> B of the crystal sensor 7 are oscillated by the oscillation circuit unit 6, and their oscillation frequencies are taken into the measurement circuit unit 81. Then, the difference between the oscillation frequency in the first vibration region 4A and the oscillation frequency in the second vibration region 4B is taken out by the data processing unit 82, and the difference is evaluated as the adsorption amount of the sensing object. .

  According to the above-described embodiment, the measurement vibration region (first vibration region) 4A and the reference vibration region (second vibration region) 4B face the twin-type crystal sensor 7 arranged in the X direction. In the sensing device in which the reaction flow channel 52 is formed and the liquid supply port 53 and the liquid discharge port 54 for the sample liquid are arranged in the Y direction, grooves 55A and 55B are provided on the liquid discharge port 54 side on the opposite surface of the crystal sensor 7, respectively. In this way, the unevenness of the liquid flow based on the difference in wettability between the vibration regions 4A and 4B is suppressed. Accordingly, it is possible to prevent bubbles from remaining in the reaction flow path 52 and perform highly reliable measurement.

  In the above example, the groove portion 55 in the flow path forming member 5 is linearly formed from the liquid discharge port 54 toward both the vibration regions 4A and 4B, but is not limited to this configuration. One of the functions of the groove portion 55 is to prevent the liquid from flowing from the hydrophilic region 4A side to the hydrophobic region 4B side. Therefore, as shown in FIG. Only one groove 55 may be formed only on the 4A side. Further, the groove 55 may be curved ((b) in FIG. 11) or may be formed only between the vibration regions 4A and 4B, that is, from the liquid discharge port 54 toward the liquid supply port 53. Good ((c) in FIG. 11).

  In the above-described embodiment, the concave portion 52 has a rhombus shape with curved corners. However, as shown in FIG. 12, the concave portion 52 may have an elliptical shape, or may be a rectangle or a square. .

DESCRIPTION OF SYMBOLS 1 Liquid supply system 90 Liquid discharge system 2 Sensor unit 21 Support body 22 Support body recessed part 24 Cover body 26 Liquid supply pipe 27 Liquid discharge pipe 3 Wiring board 4 Crystal oscillator 41 Crystal piece 4A 1st vibration area 4B 2nd vibration Region 42 Common electrode 42, 43 Excitation electrode 44 Extraction electrode 46 Adsorption layer 7 Quartz sensor 5 Flow path forming member 52 Reaction flow path (recess)
53 Liquid supply port 54 Liquid discharge port 55 Groove part 6 Oscillation circuit units 6A, 6B Oscillation circuit 81 Measurement circuit part 82 Data processing part

Claims (4)

Two pairs of excitation electrodes are arranged side by side in the X direction so as to form a first vibration region and a second vibration region in a common piezoelectric piece, and the first vibration region in the excitation electrode on one side of the piezoelectric piece includes In a sensing device that forms an adsorption layer that adsorbs a sensing object in a sample liquid and senses the sensing object in the sample liquid using a piezoelectric sensor that does not form an adsorption layer in the second vibration region on the one surface side ,
The piezoelectric sensor includes a facing surface that is in close contact with the one surface side of the piezoelectric sensor and faces the one surface side including the first vibration region and the second vibration region through a gap, and has a height in a region facing the one surface side. A flow path forming member for forming a reaction flow path of 0.3 mm or less ;
A liquid supply port provided on one end side in the Y direction orthogonal to the X direction in the reaction channel, and for supplying a sample solution to the reaction channel;
A liquid discharge port provided on the other end side in the Y direction in the reaction channel, for discharging the sample liquid from the reaction channel;
In order to control the liquid flow, the first vibration region is hydrophilic and the second vibration region is hydrophobic depending on the presence or absence of the adsorbing layer in order to control the liquid flow. A groove portion formed on the opposite surface so as to extend along the liquid flow direction in the reaction channel ,
The first vibration region and the second vibration region are provided for sensing a sensing object based on the oscillation frequency when the piezoelectric sensor is oscillated by an oscillation circuit. Sensing device. The groove portion includes a first groove portion extending from the liquid discharge port side toward the first vibration region, and a second groove portion extending from the liquid discharge port side toward the second vibration region, 2. The sensing device according to claim 1 , wherein the end portions on the vibration region side of the first groove portion and the second groove portion are positioned so as not to overlap the first vibration region and the second vibration region, respectively. . The first groove portions and the second groove, when viewed from above, according to claim 2, characterized in that it is formed so that the distance therebetween increases as the fluid outlet toward the vibration area side Sensing device. The first groove portion is close to the line side connecting the liquid supply port and the liquid discharge port from the edge of the reaction channel extending from the liquid discharge port to the first vibration region side,
4. The sensing device according to claim 3 , wherein the second groove portion is closer to the line side from an edge of a reaction flow channel extending from the liquid discharge port to the second vibration region side.

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