JP2004354142A - Biosensor device and measuring method using it - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a biosensor device receiving no effect of evaporation over a long period of time even in a very small amount of a sample, and a measuring method using it. <P>SOLUTION: In the biosensor device equipped with the vibrator constituted of a crystal plate held between electrodes and a cell for holding the sample on the vibrator, the cell is placed under a humidified atmosphere. <P>COPYRIGHT: (C)2005,JPO&NCIPI

Description

[0001]
TECHNICAL FIELD OF THE INVENTION
The present invention relates to a biosensor device used for tracking a chemical reaction and analyzing conditions in the fields of biochemistry, medicine, and food, and a measurement method using the biosensor device.
[0002]
[Prior art]
Conventionally, this type of biosensor device mixes a protein or the like to be measured in a cylindrical cell having an inner diameter of about 10 mm into a buffer solution (biochemical buffer solution) containing NaCl, KCl, etc. The samples before and after are reacted while being vibrated for about 2 hours on a piezoelectric element such as a quartz oscillator or a surface acoustic wave element having electrodes disposed on both sides, and the measurement is performed.
When a large amount of the sample can be used as described above, the amount of evaporation of the sample is about 20 to 30 μl, which is 10% or less of the whole sample, and thus does not significantly affect the measurement.
[0003]
[Problems to be solved by the invention]
However, when the sample is a very small droplet of about several μl, water evaporates before the reaction is completed, which has been a serious problem in measurement.
[0004]
The present invention has been made in view of the above circumstances, and an object of the present invention is to provide a biosensor device and a measurement method using the biosensor device, which are not affected by evaporation for a long time even for a small amount of sample. .
[0005]
[Means for Solving the Problems]
In order to solve the above-mentioned problems, the present inventors have made intensive studies and found that the above-mentioned problems can be solved by setting the periphery of the sample in a humid atmosphere so as to prevent moisture from evaporating from the sample.
That is, as described in claim 1, the biosensor device of the present invention is a biosensor comprising a vibrator formed of a crystal plate sandwiched between electrodes, and a cell for holding a sample on the vibrator. A sensor device, wherein the cell is placed in a humidified atmosphere.
Further, in the biosensor device of the present invention, in the biosensor device according to claim 1, the cell is accommodated in a predetermined space, and a humidification liquid reservoir groove is provided near the cell. It is characterized by having.
The biosensor device according to claim 3 is the biosensor device according to claim 1 or 2, wherein the predetermined space is formed by a cover surrounding the cell from above the cell, and the cover includes And a suction port for introducing the humidified gas.
A biosensor device according to a fourth aspect is the biosensor device according to any one of the first to third aspects, wherein the biosensor device includes a plurality of the cells on an array plate. The array plate is formed by arranging a plurality of the transducers adjacent to each other and sandwiching the transducers between a pair of fixing plate members.
Further, the measuring method by the biosensor device of the present invention comprises, as described in claim 5, a vibrator constituted by a crystal plate sandwiched between electrodes, and a cell for holding a sample on the vibrator. A biosensor device comprising: humidifying a space around the sample to suppress evaporation of the sample.
[0006]
BEST MODE FOR CARRYING OUT THE INVENTION
Next, the present embodiment will be described with reference to the drawings.
FIG. 1A is a cross-sectional view of an array plate 1 of a biosensor device according to an embodiment of the present invention, and FIG. 1B is a plan view of the same. The vibrator indicated by 2 in the figure includes a crystal plate 3 and electrodes 4 and 5 provided on both surfaces thereof. A reaction film 6 is disposed on the surface of the electrode 4 on the side of the electrode 4 on which the sample of the crystal plate 3 is dropped, and reacts with a specific component to be measured, or It is composed of a material that adsorbs specific components.
The vibrator 2 is sandwiched and fixed between two fixing plate members 7 and 8 via a seal member 9. One fixing plate 7 is perforated, and the hole 10 and the quartz oscillator 2 constitute a cell 12 for holding a sample. A liquid storage groove 13 is formed around the hole 10 of the fixing plate 7. A space 5s for accommodating the electrode 5 is formed in the other fixing plate 8.
[0007]
The array plate 1 including the cells 12 is covered with a cover 14 so as to be accommodated in a predetermined space, as shown in FIG. The cover 14 is provided with a suction port 15 for introducing humidified gas and a discharge port 16 for discharging gas in the cover 14.
[0008]
As shown in FIG. 3, the cells 12 of the array plate 1 are respectively connected to analyzers 20 such as a network analyzer and an impedance analyzer. The analyzer 20 is connected to a controller 21 that controls the analyzer 20 and calculates the measurement result.
The analyzer 20 is configured to output an AC signal of a desired frequency to the cell 12 and measure the conductance G of the cell 12 under that frequency. The control device 21 controls the operation of the analysis device 20, changes the frequency of the signal output from the analysis device 20 to the cell 12, associates the frequency with the measurement result, and stores the result together with the calculation result. Have been.
The cell 12, the analyzer 20, and the controller 21 constitute the biosensor device 30 of the present invention.
[0009]
In the above configuration, for example, a sample 11 prepared by mixing a measurement target such as a protein with a biochemical buffer (buffer solution) containing NaCl, KCl, or the like in a pipette is placed on a cell 12. By dropping, vibrating the vibrator 2 and causing it to react, and measuring a frequency variation, an impedance variation, a phase variation, or a combination thereof, the interaction between the biomolecules of the measurement object can be measured.
At this time, in the present embodiment, water is injected into the liquid storage groove 13 provided around the cell 12 to suppress evaporation of water from the sample 11 of the cell 12 during the measurement period. Accurate measurement is possible in the reaction of the period. The depth and position of the liquid storage groove 13 are not particularly limited as long as the evaporation of moisture from the sample 11 in the cell 12 can be suppressed, but as described in the present embodiment. It is more preferable to form the liquid storage groove 13 so as to have an evaporation surface wider than the evaporation surface of the sample 11 of the cell 12 because the amount of evaporation from the liquid storage groove 13 increases.
Further, in the present embodiment, since the upper part of the cell 12 is covered with the cover 14 and the humidified gas is introduced from the inlet 15, the evaporation of moisture from the sample 11 of the cell 12 can be more effectively performed. This makes it possible to perform more accurate measurement. Further, in the present embodiment, by providing the outlet 16 in the cover 14, the humidified gas can be efficiently introduced.
The cell 12 described in the above embodiment is placed in a humidified atmosphere so that the moisture of the sample 11 in the cell 12 does not affect the measurement due to evaporation of the sample 11, and the inside of the atmosphere space of the cell 12 The humidity, temperature, and the like are not particularly specified as long as the amount is less than the saturated water vapor amount.
[0010]
The capacity of the cell 1 described in the above configuration is preferably 5 μl to 10 μl for the purpose of the present invention, and the capacity is adjusted by changing the thickness of the fixing plate 7 or the like. can do.
Further, in the above configuration, the gas introduced from the suction port 15 is, for example, humidified air by atomizing water by vibration of an ultrasonic vibrator, or water is sucked up by a water absorption filter, and the filter is forcibly applied to the filter. Humidified air or the like can be used by passing through.
Further, as the vibrator 2, for example, a piezoelectric element such as a quartz oscillator or a surface acoustic wave element can be used, and the sealing material is, for example, a silicon buffer material (Geltech product name: α-gel) or the like. Can be used. As the fixing plate 5, for example, an acrylic plate or the like can be used.
[0011]
The above materials are merely examples of the materials. Accordingly, materials other than those described above are not particularly limited as long as they meet the purpose of the present apparatus or the measuring method.
[0012]
【Example】
Next, examples of the present invention will be described together with comparative examples.
First, in order to prove that the measurement method of the present invention can measure with higher accuracy than the conventional measurement method, a comparative experiment was performed with the same amount of liquid to be measured.
(Experimental example 1)
In the experimental method, 5 μl of pure water was dropped at the center of the crystal oscillator, and the change in the oscillation frequency of the crystal oscillator was measured. Regarding the experimental conditions, in Example 1, the humidification was performed so that the temperature around the cell was 25 ° C. and the humidity was 90% or more. In Comparative Example 1, humidification was performed under the conditions of Example 1. The conditions were the same except that they were not performed.
The quartz oscillator used was a quartz plate (outside diameter: 8.9 mm) having a frequency of 27 MHz, on which electrodes (outside diameter: 2.5 mm) were formed by vapor deposition at the center on both sides.
[0013]
As a result of the above measurement, as shown in FIG. 4, in Comparative Example 1, the liquid evaporated in 30 minutes, the electrode at the center of the crystal resonator dried, and the viscous load of water disappeared, so that the frequency increased by 10 kHz and measurement became impossible.
On the other hand, in Example 1, the evaporation of the sample was suppressed, and the remaining amount of the liquid was sufficient at 4 μl even after 4 hours.
[0014]
Next, in order to prove that the measurement method according to the present invention is particularly effective in measuring a very small amount of liquid droplets, a comparative experiment was performed with a difference in the liquid amount.
(Experimental example 2)
The measuring method of the second embodiment was the same as that of the first embodiment. The measurement method of Comparative Example 2 was the same as that of Example 2 except that a quartz oscillator was provided on the bottom surface of the cup having a capacity of about 500 μl to form a cell.
Regarding the experimental method, in Example 2, 5 μl of avidin was stirred with 5 μl of the buffer solution in a pipette and then dropped on the quartz oscillator of the cell, and the frequency variation was measured over time. In Comparative Example 2, 5 μl of avidin was dropped into the cell into which 500 μl of the buffer solution had been injected, and the mixture was stirred.
[0015]
The result of the above measurement is shown in FIG.
From FIG. 5, in Example 2, the frequency change after 800 seconds was 390 Hz, whereas in Comparative Example 2, it was about 4 Hz.
From this result, it was found that Example 2 had a sensitivity 100 times higher than that of Comparative Example 2 and that a very small amount of droplets could be measured in a very short time.
[0016]
Next, FIG. 6 shows the results of measuring 2 μl of Block Ace as a measurement target by the same measurement method as in Example 2 and Comparative Example 2 as Example 3 and Comparative Example 3, respectively.
From FIG. 6, it was found that in Example 3, the reaction was completed about 100 times earlier than in Comparative Example 3, so that a small amount of droplets could be measured in a short time.
[0017]
【The invention's effect】
According to the present invention, the reaction can be accurately measured over a long period of time even for a small amount of sample.
Further, according to the present invention, a plurality of cells are generally manufactured on one array plate, compared to a case where a plurality of cells are formed on a large crystal plate at a high manufacturing cost. Since a plurality of crystal plates may be used, manufacturing costs can be reduced. In addition, since it is not necessary to separately provide cells in the device, a plurality of cells can be replaced at a time, and a biosensor device with excellent workability can be provided.
[Brief description of the drawings]
FIG. 1A is a cross-sectional view illustrating an array plate of a biosensor device according to an embodiment of the present invention. FIG. 2B is a plan view. FIG. 2 is a perspective view illustrating a modification of the biosensor device. FIG. 3 is an explanatory diagram for explaining the configuration of the apparatus. FIG. 4 is a graph showing measurement results of Experimental Example 1. FIG. 5 is a graph showing measurement results of Experimental Example 2. FIG. Graph showing the measurement result when the object is changed.
REFERENCE SIGNS LIST 1 Array plate 2 Transducer 3 Crystal plate 4 Electrode 5 Electrode 6 Reaction film 7 Fixing plate 8 Fixing plate 9 Sealing material 10 Hole 11 Sample 12 Cell 13 Reservoir groove 14 Cover 15 Inlet 16 Outlet 20 Analyzer 21 Control Device 30 Biosensor device

Claims (5)

A biosensor device including a vibrator formed of a crystal plate sandwiched between electrodes, and a cell for holding a sample on the vibrator, wherein the cell is placed in a humidified atmosphere. Biosensor device. 2. The biosensor device according to claim 1, wherein the cell is housed in a predetermined space, and a humidification groove is provided near the cell. 3. The said predetermined space is formed by the cover which surrounds the said cell from the said cell upper part, The said cover is provided with the inlet for introducing a humidified gas, The Claim 1 or Claim 2 characterized by the above-mentioned. 3. The biosensor device according to 2. The biosensor device includes a plurality of the cells on an array plate, and the array plate is formed by arranging a plurality of the transducers adjacent to each other and sandwiching the transducers between a pair of fixing plate members. The biosensor device according to any one of claims 1 to 3, wherein the biosensor device is used. A vibrator constituted by a crystal plate sandwiched between electrodes, and a measurement method using a biosensor device including a cell for holding a sample on the vibrator, wherein a space around the sample is A measurement method using a biosensor device, wherein humidification is performed to suppress evaporation of a sample.

Images