JPS58113760A - Injection dilution apparatus for sample - Google Patents

Abstract

PURPOSE:To make injection and dilution of a sample highly accurately with a simple construction, by keeping constant a ratio of each suction quantity of plural pumps. CONSTITUTION:A sample 3 is placed on a sample table 2 of an injection and dilution vessel 1 and the sample 3 is kept in a fixed shape by its surface tension. A fixed flow rate B is kept in an outflow passage 8 in accordance with drive of a pump 10 and a fixed guantity A of a buffer solution 6 is flowed in an inflow passage 5 by a pump 7. Accordingly, the sample 3 on the table 2 is flowed into an inflow passage 4 only the difference (B-A) between the suction quantity B by the pump 10 and that of A by the pump 7. Therefore, diluted sample solution having a dilution ratio expressed by (B-A)/B is obtained in the passage 8.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a sample injection dilution device, and more particularly to a sample injection dilution device that can be easily used for biochemical components.

Many biochemical analyzers are currently commercially available, but they often require sample dilution. For example, in the case of an analyzer that performs blood analysis, an analyzer that uses a cell for one analysis measurement part that will definitely require dilution, has a cell containing a buffer solution that has been quantified in advance. In such a case, where a minute amount of sample is injected and diluted, the amount of sample to be quantified is extremely small, on the order of several tens of micrometers, so it is necessary to prepare an expensive dedicated quantitative meter. Furthermore, in an analyzer that uses a pump to quantify a certain amount of sample and then dilutes it by adding a buffer solution, it is necessary to use a pump with very high precision. If the suction amount and therefore the discharge amount of the pump is on the order of several hundred micrometers, the pump is inexpensive and can be used to accurately quantify the amount. However, pumps that can accurately quantify such minute amounts of samples have been extremely expensive. Therefore, no matter which method is used in the past, when an analysis is performed, the means for injecting and diluting the sample become expensive, making the analyzer expensive as a whole.

Therefore, the present invention provides injection dilution@stomach, which makes it easier to accurately inject and dilute a sample.

Simply put, this invention is based on the first method that communicates with the sample! II
The inlet path and the second inflow path communicating with the 1llir liquid merge, and the outflow path extends from there. B):
A second pump capable of suctioning only (B>A) is provided, and the second
A predetermined amount (B-
A) sample is injected and the outflow lIk: is (B-A>/B
Injection dilution of the sample so as to obtain a diluted sample @stomach.

The above objects and other objects and features of the invention will become more apparent from the following detailed description with reference to the drawings.

FIG. 1 is a diagram showing one embodiment of the present invention. The injection diluter 1 has a sample table 2 with a horizontal part on which a sample can be placed. A sample 3 is placed on the sample table 2, and an eleventh passageway 4 is formed in communication with the sample 3 placed on the sample table 2. This 11th! The 1112 inflow path 5 is joined to the inlet 14. Second
11 The other end of the passage 5 communicates with a buffer solution 6 for dilution.

A first pump 7 is provided in the middle of the second inflow path 5 to suck in the buffer solution 6 by a certain amount 11(A). 181st human route 4
An outflow 11B extends from the confluence of the second inflow path 5 and the second inflow path 5, and an agitation w9 is formed in the middle of this outflow path 8. A second pump 10 is further provided in the outflow path 8 passing through the stirring section 9, and this second pump 10 has a capacity of a certain amount (B). Here, the suction amount B of the second pump 10 is equal to the suction amount B of the first pump 7.
is selected to be larger than the suction amount A, and its M CB
-A) is chosen as the amount of sample to be injected. Outflow channel 8
The other end is brought into drainage 1111.

The outflow path 8 is provided with a **electrode 12 as an example of a measuring means. This enzyme electrode 12 has an immobilized base 13 that can come into contact with the solution passing through the outflow channel 8 . This immobilized enzyme 1113 has immobilized an enzyme that can specifically react with a specific component or substrate contained in one wave passing through the outflow path 8. The immobilized enzyme 1113 is associated with a polarographic electrode capable of amperometrically detecting substances accompanying the reaction of the enzyme. The polarographic electrode includes a reference electrode 14 and a working electrode 15. A constant boularographic potential is applied between the reference electrode 14 and the working electrode 15 by a constant voltage circuit 14a. Working electrode 15
A current-voltage conversion circuit 15a is connected to the working electrode 15, and the signal current flowing through the working electrode 15 is converted into voltage. Note that the functions of these enzyme electrodes 12 will be explained in detail later.

In the above configuration, the sample 3 is placed on the sample table 2 of the injection diluter 1. The sample 3 is held in a fixed shape as shown in the figure by its surface tension. 1st
And the second pumps 7 and 10 are driven. Depending on the driving of the second pump 10, the outflow path 8 has a constant flow rate (B).
will have the following. On the other hand, a constant amount of 1 t (A) of buffer solution 6 flows into the second inflow path 5 by the first pump 7 .

Therefore, the difference (B-A) between the suction amount B of the second pump 10 and the suction 1) A of the first pump 7 is
The sample 3 placed on top will flow into the first inflow path 4. Therefore, a diluted sample solution having a dilution ratio expressed as (B-A)/B is obtained in the outflow path 8. The amount of such a sample (B-A) should be such that the solution can flow continuously through the area for the time period necessary for analysis by the enzyme electrode 12, for example, at least several tens of micrometers. would be necessary, however, even if the sample haze is more than that, resulting in (B-A
)/B is constant, therefore the dilution ratio does not vary and does not affect the analytical measurements.

When the sample 3 on the sample table 2 is injected and disappears from the sample table 2, the next sample is simply placed on the sample table 2, and the injection dilution operation as described above is repeated. . If the sample on the sample table 2 disappears, the outflow channel 8 will contain air bubbles together with the buffer solution, but this may cause the measurement means to drop, for example, the electrode 1.
2 is not affected in any way. Rather, it was found that such bubbles have a cleaning effect on the enzyme electrode. Therefore, the timing of supplying or placing the sample 3 on the sample table 2 is completely free, and analysis and measurement can therefore be performed at any time.

As mentioned above, in this embodiment, the suction by the second pump is
(B) and the suction amount (A) by the first pump 7 are always kept at a constant ratio (B-A)/B. To do so, for example, a pump as shown in FIG. 2 may be used. The first pump 7 and the second pump 10 are rotationally driven by a common interchange motor 16. That is, two sets of two discs are fixed to the output shaft 17 of the common induction motor 16 so that they can rotate together. A plurality of squeezing rollers 18 and 19 are provided at equal intervals between each set of discs. Flexible tubes such as silicone tubes 5a and 8a are wound around these squeezing rollers, respectively. Note that the tubes 5a and 8a were assumed to have the same inner diameter here. Therefore, the ratio of the two pumps 7 and 10 is determined by the radii r1 and 2 (Fig. 2), so by setting r1 and r2 appropriately, , two pumps with different flow rates and a constant flow rate ratio can be easily obtained.Therefore, in the example shown in FIG. 2, in addition to the advantage that only one motor is required for driving, Even if there is a variation in the number, the flow rates of the two pumps 7 and 10 will change, but the ratio of the flow rates will not change, and as a result the dilution ratio will also have the advantage of not changing. For example, 190 for pump 7
If the μ pressure is flowing and, for example, 200 uQ flows into the second pump 10, the outflow path 8 is filled with (B-A)/B, that is, (
A dilute solution of 200-190)/200-20 times is obtained. Even if the rotational speed of the motor 16 increases by 10% and a flow of, for example, 209 μm flows into the second pump 7 and a flow of, for example, 220 μm flows into the second pump 10 at different times, the dilution ratio will be (220-209)/ 220-20, motor 1
It can be seen that the dilution ratio is constant regardless of the variation in the rotation speed. In the experiment, silicone tubes with an inner diameter of 1.0.5 as and an outer diameter of 1.0 - 1 were used as the tubes 5a and 8a, and the tubes were squeezed half 1.0 in. Let r2 be 1°8cgi,
One half of hard work! r i to 2. OC-, and furthermore,
Although it is possible to always obtain a constant suction amount by using separate motors, such as pulse motors, for each pump 7 and 10, according to the embodiment shown in FIG. can be done.

In the experiment, rat blood analysis was performed using the system shown in Figure 1. That is, using rat blood, the blood was diluted approximately 20 times and sent to the enzyme electrode 12 for quantitative determination of glucose, and the intravenous glucose concentration (blood sugar level) contained in the rat blood was measured. The sample, rat blood 3, was supplied onto the sample table 2 for about 20 μm so as to block the 121I inlet 4 located approximately at the center of the sample table 2, and was discharged. The liquid 3 was kept on the sample table 2 for about 1 minute. The foot blood 3 was suctioned in fixed amounts through the first inflow path 4 and diluted, decreasing at a constant rate. Since the time required for the response of the enzyme electrode 12 to detect glucose was approximately 30 seconds, it can be seen that the rat blood sample must be placed on a surface of at least 10 μm.

Note that the dilution buffer supplied through the first pump 7 was approximately 20 cc per hour. If the sample tends to remain, such as blood, 1, the length of the first inlet channel 4 should be set to 1.5 - 1 in order to prevent the sample from blocking the first inlet channel 4 and to clean the sample well. I tried to do this, and the results were good. The diluted rat blood is supplied to the enzyme electrode 12 after passing through the stirring section 9 by the suction force of the second pump 10 . The diluted and agitated rat solution flows while contacting the immobilized enzyme membrane 13 on which glucose oxidase (GOD) is immobilized, and then passes through the pump 10.
The water passes through and is drained into the drainer 11.

Enzyme GOD performs the following enzymatic reaction when the substrate glucose is present in rat blood.

Glucose + σ□・Glucone monoxide The amount of oxygen absorbed or hydrogen peroxide produced is proportional to the concentration of the substrate glucose. Therefore, GOD immobilized 11111113 is deposited on the surface of the polarographic electrode 14.15 to reduce the amount of oxygen that diffuses from the electrode surface or increase the amount of hydrogen peroxide that diffuses to the electrode surface due to the enzyme reaction.゜1
By keeping the value constant at 5, it is possible to know the amount of glucose contained in the rat blood. In the experiment, an increase in hydrogen peroxide was measured as Polarographic Denki 14.15. For this purpose, the reference electrode 14 was made of silver, and the working electrode 15 was made of platinum.

A DC boularography potential of 7 V was applied to the silver reference electrode 14 by the constant voltage circuit 14a. In addition, the enzyme GOD was immobilized on the immobilized enzyme membrane 13 by a strip-horizontal immobilization method using glutaraldehyde on a thin film of acetyl cellulose or polycarbonate. The polarographic reaction on the surfaces of the reference electrode 14 and the working electrode 15 is expressed by the following equation.

Reference electrode: Ot +48" +46--28z O Working electrode: H2O2→2 H"+02+26- A voltage proportional to the signal current flowing through the working electrode 15 is detected by the current-voltage conversion circuit 15a.

Therefore, the voltage from this circuit 15a is obtained as being proportional to the concentration of the substrate, ie, glucose, contained in the solution, ie, rat blood. Once such voltage output is recorded on a recorder (not shown), such glucose concentration can be detected. FIG. 3 shows an example of the experimental results recorded by the recorder, in which the same rat blood was measured three times, and almost the same output voltage was returned each time.

In addition, in the above-mentioned Example, the measuring means was arranged in the outflow path 8 upstream of the second pump 10. However, it goes without saying that this may be placed downstream of the second pump 10.

Furthermore, as the measuring means, various component analysis or measuring instruments can be used in addition to the enzyme electrode used in the examples.

As described above, according to the present invention, a sample can be injected and diluted with high precision using a simple configuration. Since it is not necessary to use a pump with a small flow rate and high precision, such an injection dilution device can be obtained at low cost.

[Brief explanation of drawings]

FIG. 1 is a diagram showing a system according to an embodiment of the present invention. FIG. 2 is an illustrative view showing an example of the pump used in the embodiment shown in FIG. FIG. 3 is a Bolarodalum showing an example of the experimental results using the embodiment shown in FIG. In the figure, 1 is an injection diluter, 2 is a sample table, 3 is a sample, 4 is a 1211th passage, 5 is a second inflow passage, 6 is a liquid solution, 7 is a first pump, 8 is an outflow passage, 10 is the second pump,
12 indicates an enzyme electrode. Patent Applicant Tateishi Electric Co., Ltd. Agent Patent Attorney ± 8i Party Kube (and 2 others) \1 Figure 2. 1 Figure 3

Claims (6)

[Claims] (1) A trial loading section that has a horizontal portion and in which a sample can be placed; a first inlet channel that communicates with the horizontal portion of the pre-allocation material loading section and introduces the sample placed there; , a 21st passage that communicates with the dilution buffer and joins the 111th inlet passage, an outflow passage extending from the confluence of the first and second inflow passages, and a 21st passage provided midway through the 21st passage. A first pump that sucks the liquid III in a corresponding amount (A), and a trowel (B) that is provided in the middle of the outflow path and is capable of sucking only a predetermined amount larger than the amount (A). 2nd pump -
E, the predetermined amount of the sample (B-A) is injected from the 131st passage in response to the suction of the second pump, whereby the sample diluted to (B-A)/B is injected into the outflow passage. Injection dilution of the sample resulting in ms. (2) The first and second pumps are driven by a common motor, and the first and second pumps are characterized by a buffer solution and a diluted sample, respectively, at a constant specific gravity.
Injection dilution of sample as described in section @H. (3) The first pump has a first flexible tube through which the sample passes, and is rotationally driven by the common motor to rotate the first flexible tube to a certain length according to the rotation. and a squeezing roller *i for squeezing each time, and the second pump includes a second squeezing roller through which the diluted sample passes.
and a second squeezing roller that is rotationally driven by the common motor and squeezes the second flexible tube every predetermined length according to the rotation thereof. Injection dilution of the sample described in I1M section 2 @pupil. (4) The first and second pumps are rotated by different motors, so that the eighteenth
and II2 pumps are respectively slow II'I1 at a constant specific gravity.
1 and the second claim featuring a diluted sample.
Sample injection dilution device as described in section. (5) Claims 1 to 4 include a measuring means provided in the outflow path to constant the concentration of a specific component contained in the diluted sample fed into the outflow path. Injection dilution of the sample as described in either @ll. (6) The measuring means is sensitive to an immobilized enzyme on which an enzyme that specifically reacts with a specific component contained in the diluted sample is immobilized, and a reaction between the element and the specific component. 6. A sample injection dilution device according to claim 5, comprising a polar graph electrode.