JPS61294351A - Biosensor - Google Patents

Abstract

PURPOSE:To quantitatively determine the specific component in a vital sample extremely easily and quickly with high accuracy by coating an electrode system with a porous body carrying oxidation reduction enzyme and electron receptor and uniting the electrode system, substrate and porous body to one body. CONSTITUTION:A conductive carbon paste is printed by screen printing on the insulating substrate 1 consisting of PE terephthalate and is dried by heating to form the electrode system consisting of a counter electrode 2, a measuring electrode 3 and a reference electrode 4. An insulating layer 5 is then formed to partially cover the electrode system so that the electrochemically acting parts 2', 3', 4' of the respective electrodes remain. A holding frame 6 bored with a hole is adhered to the layer 5 and the porous body 7 is held in the hole of the frame 6 so as to cover the electrode systems 2'-4'. A cover 8 having an aperture of the diameter smaller than the diameter of the body 7 is adhered thereto to unite the entire part to one body.

Description

DETAILED DESCRIPTION OF THE INVENTION Field of Industrial Application The present invention relates to a biosensor that can quickly and easily quantify specific components in various minute amounts of biological samples without diluting the sample liquid.

Conventional technology Conventionally, a biosensor as shown in Figure 4 has been proposed as a method for quantifying specific components in biological samples such as blood with high precision without performing operations such as diluting and stirring the sample solution. (For example, Japanese Patent Application Laid-Open No. 59-166852'
). This biosensor has leads 12 and 1 on an insulating substrate 9.
A measurement electrode 10 and a counter electrode 11 each made of platinum or the like having 3 are buried, and the exposed portions of these electrode systems are covered with a porous material carrying an oxidoreductase and an electron acceptor. When the sample liquid is dropped onto the porous body, the oxidoreductase and electron acceptor in the porous body are dissolved in the sample liquid, and an enzymatic reaction proceeds with the substrate in the sample liquid to reduce the electron acceptor. After the enzymatic reaction is completed, the reduced electron acceptor is electrochemically oxidized, and the substrate concentration in the sample solution is determined from the oxidation current value obtained at this time.

Problems to be Solved by the Invention - In such a conventional configuration, the porous body can be easily used for measurement by replacing it after each measurement, but the electrode system requires operations such as cleaning. . On the other hand, if it becomes possible to discard the electrode system after each measurement,
Although the measurement operation is extremely simple, the electrode material such as platinum and the structure make it extremely expensive.

As a result of various studies on these points, the present invention has developed an inexpensive disposable type biotechnology that can extremely easily quantify specific components in biological samples quickly and with high precision by integrating an electrode system and a porous body. It provides a sensor.

Means for Solving the Problems In order to solve the above problems, the present invention provides an electrode system consisting of at least a measurement electrode and a counter electrode on an insulating substrate, reacts an enzyme, an electron acceptor, and a sample solution, and performs the reaction. In a biosensor that electrochemically detects a change in substance concentration with the electrode system and measures the substrate concentration in a sample solution, the electrode system is covered with a porous material carrying an oxidoreductase and an electron acceptor, It is integrated with the electrode system and the substrate.

Function According to the present invention, a disposable type biosensor including an electrode system can be constructed, and the substrate concentration can be extremely easily measured by adding a sample liquid to a porous body.

Example An embodiment of the present invention will be described below.

A glucose sensor will be described as an example of a biosensor. FIG. 1 shows an embodiment of a glucose sensor, and is an exploded view of the constituent parts. A conductive carbon paste is printed on an insulating substrate 1 made of polyethylene terephthalate by screen printing and dried by heating to form a counter electrode 2. Measuring pole 3. An electrode system consisting of a reference electrode 4 is formed. Next, partially cover the electrode system,
2' which becomes the electrochemically active part of each electrode.

An insulating paste is printed in the same manner as described above so that 31 and 47 (1-) are left, and heat treated to form an insulating layer 6.

Next, a resin holding frame 6 with holes is glued to the insulating layer 5,
The porous body 7 is held in the hole so as to cover the electrode systems 2', 3', and 4'. Furthermore, a resin cover having apertures with a diameter smaller than that of the porous body is adhered to integrate the whole body. A cross-sectional view along the measurement electrode 3 of this integrated biosensor is shown in FIG. The porous body used above contained 100 μg of glucose oxidase as an oxidoreductase and 150 μg of potassium ferricyanide as an electron acceptor.
It was prepared by impregnating a nylon nonwoven fabric with the solution of e. dissolved in 1 ml of phosphate buffer and drying it under reduced pressure.

A glucose standard solution is dropped as a sample solution into the porous body of the glucose sensor configured as described above, and after 2 minutes of dropping, the potential of the measurement electrode is changed to 1 V in the direction of the anode with respect to the reference electrode.
/second. In this case, the added glucose reacts with potassium ferricyanide by the action of glucose oxidase supported on the porous material to produce potassium ferrocyanide. Therefore, by the above-mentioned sweep toward the anode, an oxidation current based on the concentration of potassium ferrocyanide produced is obtained, and this current value corresponds to the concentration of glucose, which is the substrate. FIG. 3 shows the relationship between the obtained peak current value and the glucose concentration in the sample. In the figure, B shows the case where an electrode manufactured by the above-mentioned manufacturing method is used, and A shows the case where an electrode in which a conductive carbon paste is printed, heated and dried, and particularly the part that will become the measuring electrode 3' is polished is used. Although the glucose sensor was replaced after each measurement, both cases A and B had good linearity and good reproducibility. On the other hand, polished Al
um had the advantage of being able to obtain a large response current. This difference in response current due to polishing is thought to be due to the resin component contained in the paste as a binder partially covering the carbon surface.

Screen printing as a method for forming electrode systems can produce disposable biosensors with uniform characteristics at low cost, and in particular, it is possible to fabricate electrodes using carbon, which is an inexpensive and stable electrode material. This is a convenient way to form.

The method of integration in the biosensor of the present invention is as follows:
The present invention is not limited to the shapes and combinations of frames, covers, etc. shown in the examples. In addition, the porous body used is
In addition to nylon nonwovens, porous bodies made of cellulose, rayon, ceramic, polycarbonate, etc. can be used alone or in combination. Further, the combination of oxidoreductase and electron acceptor is not limited to the above examples, and any combination can be used as long as it meets the gist of the present invention. On the other hand, in the above embodiment, a three-electrode force type electrode system was described, but measurement can also be performed using a two-electrode force type consisting of a counter electrode and a measurement electrode.

Effects of the Invention The biosensor of the present invention can extremely easily measure the substrate concentration in a biological sample by integrating an insulating substrate, an electrode system, and a porous body carrying an oxidoreductase and an electron acceptor. be able to.

[Brief explanation of the drawing]

Fig. 1 is an exploded perspective view of a biosensor that is an embodiment of the present invention, Fig. 2 is a longitudinal sectional view, Fig. 3 is a response characteristic diagram of the biosensor, and Fig. 4 is a longitudinal sectional view of a conventional biosensor. It is. 1...Substrate, 2...Counter electrode, 3...
... Measuring electrode, 4... Reference electrode, 6... Insulating layer, e... Holding frame, 7... Porous body, 8
······cover. Name of agent: Patent attorney Toshio Nakao and one other person
・ Thread and board with 1 raw board d... Iki 4 pots Iki 4
, 4',...4 wings, pole 1st figure 611. Connection layer/2” 2nd
Figure 3: Glucose Thick 1frN Pi/Aya Figure 4: A

Claims (4)

[Claims] (1) An insulating substrate is provided with an electrode system consisting of at least a measurement electrode and a counter electrode, and the electrode system electrochemically detects a change in substance concentration during the reaction between an enzyme, an electron acceptor, and a sample liquid. A biosensor for measuring the substrate concentration of a liquid, characterized in that the electrode system is covered with a porous body carrying an oxidoreductase and an electron acceptor, and the porous body is integrated with the electrode system and the substrate. (2) The biosensor according to claim 1, wherein the electrode system includes a measurement electrode, a counter electrode, and a reference electrode. (3) The biosensor according to claim 1 or 2, wherein the electrode system is made of a carbon-based material formed by screen printing on an insulating substrate. (4) The biosensor according to claim 3, wherein the electrode system is polished.