JPH0471182B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to an automatic immunological analyzer.

In recent years, advances in medical care have made it possible to analyze minute amounts of biological components, which is useful for early diagnosis of various diseases.

Among these, enzyme immunoassay has been attracting particular attention recently because it does not require special equipment or measurement techniques and can be easily performed using a commonly used colorimeter. As enzyme immunoassay methods using a solid phase, competitive methods, Sand-Deutsch methods, and the like are known. The solid phase on which antibodies or antigens that cause an antigen-antibody reaction with the test substance in the sample are immobilized has a size of several μm.
Methods using insoluble carriers such as beads with various particle sizes ranging from 1 to several millimeters, and methods using the inner wall of a reaction vessel directly are known.

To explain this enzyme immunoassay method using an insoluble carrier, in the competitive method, as shown in Figure 1, an antibody or antigen that causes an antigen-antibody reaction with the test substance in the sample is immobilized on the insoluble carrier 1 in advance. This carrier 1 was subjected to an antigen-antibody reaction with the sample, its test substance 2, and a labeling reagent 3 in which the same substance was labeled with an enzyme. After washing, the carrier 1 was competitively bound to the carrier 1 by an antigen-antibody reaction. After separating the test substance 2 and labeling reagent 3 from unbound substances by B and F, for example, a coloring reagent is added as a reagent for measuring enzyme activity that reacts with the labeled enzyme in the labeling reagent 3, and the reaction is carried out. After that, the reaction solution is subjected to colorimetric measurement to determine the enzyme activity of the labeled enzyme, and the amount of the test substance 2 is quantified. In addition, as shown in FIG. 2, in the Sanderutsch method, like the competitive method, an insoluble carrier 5 on which an antibody or antigen that causes an antigen-antibody reaction with the test substance in the sample is immobilized in advance is used.
The test substance 6 in the sample is bound to the carrier 5 by performing an antigen-antibody reaction with the sample, and after washing and separating B and F, the test substance 6 and the antigen-antibody are transferred to the carrier 5. A substance that causes a reaction is reacted with a labeling reagent 7 labeled with an enzyme to cause an antigen-antibody reaction, and then washed again to separate B and F, and then a coloring reagent that reacts with the labeling enzyme in the labeling reagent 7 is added. After addition and reaction, the reaction solution is subjected to colorimetric measurement to determine the enzyme activity of the labeled enzyme, and the amount of the test substance 6 is quantified.

The operation of this conventional enzyme immunoassay automatic analyzer will be explained with reference to FIG. 3 and FIGS. 4A to 4D.

During the first rotation of the reaction tube disk 12, first, at the stop position _S17 , as shown in FIG.
1 into the U-shaped tube 11 one by one from the large opening 11a. A cleaning pump 24 is inserted into the U-shaped tube 11 into which the carrier 21 is placed at the stop position _S22 .
The cleaning liquid is intermittently injected in a shower-like manner from the large opening 11a by the operation of the drain pump 28, and the cleaning liquid is suctioned and discharged through the small opening 11b by the operation of the drainage pump 28 to clean the U-shaped tube 11. position
In _S23 , the cleaning liquid is further discharged almost completely by suctioning through the small opening 11b by the drainage pump 28. By washing the U-shaped tube 11 in this manner, the buffer solution that has preliminarily wetted the carrier 21 is washed away, and the amount of buffer solution in the U-shaped tube 11 after dispensing the buffer solution in the latter stage is kept constant. Next, as shown in FIG. 4B, at the stop position _S24 , the buffer solution dispensing device 25 dispenses a certain amount of the buffer solution 26 from the large mouth portion 11a, and then at the stop position _S1 , the sample dispensing device 13 dispenses a certain amount of the buffer solution 26. A predetermined amount of sample is dispensed from the sample cup 15 located at a predetermined sample suction position of the sampler 14 through the large mouth portion 11a. The U-shaped tube 11 into which the sample has been dispensed at the stop position S ₁ is connected to the stirring air pump 27 at its small opening 11b at the next stopping position S ₂ , and the air pump causes the sample to be ejected through the small opening 11b. The carrier 21, buffer solution 26, and sample housed in the U-shaped tube 11 are stirred to start the first antigen-antibody reaction. This stirring is also performed sequentially at stop positions S ₃ , S ₄ and S ₅ . Note that the carrier input device 20, the buffer solution dispensing device 25, the sample dispensing device 13, and the sampler 14 are operated once for each U-shaped tube and then left inactive.

In the second rotation after the U-shaped tube 11 receives the carrier 21 at the stop position S ₁₇ and then moves once again to the stop position S ₁₇ , it first rotates at the stop position S ₂₂ as shown in FIG. 4B. The reaction liquid in the U-shaped tube 11 is suctioned and discharged by the drainage pump 28 through the small opening 11b, and the washing liquid is dispensed intermittently in a shower-like manner from the large opening 11a by the washing pump 24. The cleaning solution is applied to the stop position S ₂₂
And in the same way at the next stop position S ₂₃
The U-shaped tube 11 and the carrier 21 are washed by suctioning and discharging the liquid through the drain pump 28 through the liquid drain 1b, and the first B/F separation is performed. After that, at the stop position _S3, as shown in FIG. 4C, the large mouth part 11a
A fixed amount of the enzyme-labeled reagent 17 is dispensed by the reagent dispensing device 16, and at the same time, air is ejected from the small opening 11b by the stirring air pump 27 at the stop position _S3 and the next sequential stop positions _S4 and _S5 . Then, the carrier 21 and the enzyme labeling reagent 17 are stirred, and the second antigen-antibody reaction is started.

In this way, the U-shaped tube 11, which received the dispensing of the enzyme labeling reagent 17 at the stop position _S3 and started the second antigen-antibody reaction, moves to the stop position _S17 and starts the second antigen-antibody reaction.
When it reaches the rotation point, the cleaning liquid is dispensed by the cleaning pump 24 and the U-shaped pipe 11 is discharged by the drain pump 28 in the same manner as described above at the stop positions S ₂₂ and S ₂₃ .
The reaction liquid and dispensed washing liquid inside are suctioned and discharged to wash the U-shaped tube 11 and carrier 21, and a second B/F separation is performed. Then stop position S ₄
As shown in FIG. 4D, a fixed amount of the coloring reagent 19 is dispensed from the large opening 11a by the reagent dispensing device 18, and the stop position _S4 and the next stop position are
In S ₅ , air is ejected by the stirring air pump 27 to stir the carrier 21 and the coloring reagent 19 to initiate a reaction between the labeled enzyme in the enzyme labeling reagent 17 bound to the carrier 21 and the coloring reagent 19. .

When the U-shaped tube 11 that has received the dispensing of the coloring reagent 19 moves to the stop position _S17 and enters the fourth rotation, first, the reaction liquid in the U-shaped tube 11 is passed through the colorimeter 22 at the stop position _S19 . aspirate and measure colorimetrically. Colorimeter 22
For example, as shown in FIG. 4D, a light source 22b and a detector 22c are arranged through a flow cell 22a through which the reaction solution passes, and the light from the light source 22b is projected onto the flow cell 22a through an interference filter 22d.
It can be configured such that the transmitted light from the flow cell 22a is received by the detector 22c via the light guide 22e. Next, at the stop position S ₂₀ , U
The carrier 21 remaining in the tube 11 is taken out from the large opening 11a by the carrier extractor 23. Thereafter, at the stop position S ₂₂ , the cleaning pump 24 intermittently dispenses the cleaning liquid in a shower-like manner, and the dispensed cleaning liquid is transferred to the stop position S ₂₂ and the next stop position.
At S ₂₃ , the U-shaped tube 11 is cleaned by suction and discharge by the drain pump 28, in preparation for loading the carrier in the next sample analysis.

In the enzyme immunoassay method as mentioned above,
During the analysis of one test substance, quantitative dispensing of buffer solution,
Requires quantitative dispensing of enzyme labeling reagents and quantitative dispensing of enzyme activity measurement reagents. For this reason, conventional devices in which each meter is provided with a fixed-quantity dispenser have the disadvantage that the devices are large, complicated, and expensive.

An object of the present invention is to solve the above-mentioned problems and provide an automatic immunological analyzer with good measurement accuracy and a simple configuration.

The present invention provides an apparatus for immunologically analyzing a test substance in a sample by causing an antigen-antibody reaction in a reaction container, in which the reaction container is used to conduct an antigen-antibody reaction in a sample containing one of the test substances contained in the reaction container. a reaction container transport means for repeatedly transporting and stopping at a predetermined position at least twice during analysis of two measurement items; One for selecting one of at least two types of reagents necessary for analysis from a plurality of reagent containers containing enzyme activity measurement reagents for measuring the enzyme activity of a labeled enzyme and dispensing it into a reaction container. The present invention is characterized in that it has a reagent dispenser, and at least the enzyme labeling reagent and the enzyme activity measuring reagent are dispensed with one reagent dispenser.

The present invention will be described below with reference to the drawings. Fifth
The figure shows an example of an enzyme immunoassay automatic analyzer using an insoluble carrier as a solid phase, which implements the present invention.
This method employs the Sanderch method shown in FIG.

The reaction vessel uses 24 U-shaped tubes 11 having a large opening 11a and a small opening 11b, and these are held on the same circumference of the reaction tube disk 12 at equal intervals. The reaction tube disk 12 is intermittently rotated at a predetermined pitch (for example, every 15 seconds) in the direction indicated by the arrow in a horizontal plane while the U-shaped tube 11 is immersed in the constant temperature bath 10 (FIG. 4). The stopping positions of the U-shaped tube 11 due to the intermittent rotation of the reaction tube disk 12 are indicated by symbols S ₁ to S ₂₄ . In this example, the sample is selectively dispensed from the sample cup 15 at a predetermined sample suction position of the sampler 14 by the sample dispensing device 13 into the U-shaped tube 11 at the stop position _S1 . The sampler 14 has 24 tubes, which is the same number as the number of U-shaped tubes held in the reaction tube disk 12.
Sample cups are held at equal intervals on the same circumference and rotated intermittently in the direction of the arrow in synchronization with the rotation of the reaction tube disk 12. A carrier feeder 20 is inserted into the U-shaped tube 11 at the stop position _S17 from its large opening 11a.
One of the insoluble carriers 21, such as synthetic resins such as plastics and glass beads, which are housed in large numbers in the container, is selectively introduced. The carrier 21 has a size that allows it to be easily taken in and taken out from the large opening 11a of the U-shaped tube 11, but does not enter the small opening 11b, and has an antibody on its surface that causes an antigen-antibody reaction with the test substance in the sample as described above. Alternatively, the antigen is immobilized in advance and the carrier input device 20 is moistened with a buffer solution. In addition, the U-shaped tube 11 at the stop position S ₁₉
Then, the reaction solution contained in this is measured using a colorimeter 2.
2, and from the U-shaped tube 11 at the stop position _S20 , the carrier 21 accommodated therein is selectively taken out and discharged by the carrier extractor 23.
Furthermore, a washing pump 24 selectively injects a washing liquid such as ion exchange water, immunoassay buffer, physiological saline, etc. into the U-shaped tube 11 located at the stop position _S22 .

Further, the U-shaped tube 11 at the stop position S2 has its small end 11b removably connected to the stirring air pump 27, and similarly, each U-shaped tube 11 located at the stopped positions _S22 and _S23 has its small end connected to the stirring air pump 27. The portions 11b are removably connected to a common drain pump 28, respectively.

Stop position S ₂₄ has U-shaped tube 1 that stops at this position.
1 is provided with a reagent dispenser 29 for dispensing any one of a buffer solution 30, an enzyme labeling reagent 31, or a coloring reagent 32. That is, the reagent dispenser 29
In this embodiment, a syringe type dispenser is used to enable precise dispensing.

One end of the reagent tube is connected to this reagent dispenser 29, and the other end, the nozzle 35, is transferred to the stop position _S24 and the cleaning tank 33 by a moving means (not shown). Switching valve 34 in the middle of the reagent tube
is connected to each tube whose other end is connected to the buffer solution storage container 30, the enzyme labeled reagent storage container 31, and the coloring reagent storage container 32.
This switching valve 34 allows the reagent dispenser 29
A buffer solution storage container 3 is placed in a flow path consisting of a nozzle 35 and a nozzle 35.
0, the oxygen labeling reagent storage container 31 and the coloring reagent storage container 32 are selectively communicated with each other.

Next, the operation of the apparatus shown in FIG. 5 will be explained.

At the first rotation of the disk 12, the stop position is first reached.
At _S17 , the carrier 21 is placed in the U by the carrier input device 20.
It is thrown into the tube 11. U where the carrier 21 is introduced
The cleaning pump 2 is connected to the pipe 11 at the stop position S ₂₂ .
4 and the drain pump 28..., the U-shaped tube 11 and the carrier 21 are cleaned. At the next stop position _S24 , the reagent dispenser 29 causes the switching pulp 34 to select the buffer container 30, and then a fixed amount of buffer is dispensed into the U-tube. At stop position S ₁ sample dispenser 1
3. Sample cup 1 due to the function of sampler 14
A certain amount of sample 11 is dispensed from 5 into the U-shaped tube 11, and the first reaction begins. At the stop position _S2 , the test liquid in the U-shaped tube is stirred by the action of the stirring air pump 27. In the second rotation after performing the above operations for all U-shaped tubes, the carrier injector 20, sample dispenser 13, and sampler 14 are kept inactive. In the second rotation, at the stop position _S22 , due to the action of the cleaning pump 24 and the drain pump 28...U-shaped pipe 1
1 and carrier 21 are washed, and the first B/F separation is performed. Thereafter, at the stop position _S24 , the reagent dispenser 29 causes the switching valve 34 to select the enzyme-labeled antibody container 31, and then a fixed amount of the enzyme-labeled antibody is dispensed into the U-shaped tube, and the second reaction begins. At _S2 , the test liquid in the U-shaped tube is stirred by the stirring air pump 27.

At the third rotation, at the stop position _S22 , the U-shaped pipe 11 is
Then, the carrier 21 is washed and a second B/F separation is performed. Thereafter, at the stop position _S24 , the switching valve 34 selects the coloring reagent container 32 by the reagent dispenser, and then a fixed amount of the coloring reagent 32 is dispensed into the U-shaped tube 11, and the third reaction begins. The enzyme reaction is promoted by stirring at the stop position _S2 . Thereafter, at the fourth rotation stop position _S19 , the test liquid in the U-shaped tube 11 is sucked into the colorimeter 22 for colorimetric measurement.

Next, at the stop position _S20 , the carrier 21 remaining in the U-shaped tube 11 is taken out by the carrier extractor 23.
Thereafter, the U-shaped pipe 11 is cleaned by the cleaning pump 24 and the drain pump 28 at the stop position _S22 .

Further, at the stop position _S23 , the remaining liquid is discharged by the drain pump 28 in preparation for the next analysis. During the above operation, the nozzle 35 is transferred to the cleaning tank 33 after dispensing each reagent, and the inner and outer walls of the nozzle are cleaned by suction and discharge operations of the syringe 29.

As explained above, according to the present invention, two or more different enzyme labeling reagents and enzyme activity measurement reagents can be quantified using a single reagent dispenser installed in the reaction line during the analysis of each sample. By combining dispensing, the number of dispensers can be reduced, and the reaction line can be shortened, so the entire automatic analyzer can be made compact, inexpensive, and simple in configuration.

Furthermore, in the conventional apparatus, it was not possible to unify the first, second, and third reaction times because the positions of the reagent dispensers were different, but in the present invention, the reagents are dispensed from the same position. Therefore, the reaction time can be uniformly extended, which has the great effect of improving measurement accuracy and simplifying the device configuration.

[Brief explanation of drawings]

Figure 1 is a diagram for explaining the enzyme immunoassay method using the competitive method, Figure 2 is a diagram for explaining the enzyme immunoassay method using the Sand-Deutsch method, and Figure 3 is a diagram for explaining the enzyme immunoassay method using the Sand-Deutsch method. A diagram showing an example configuration,
4A to 4D are diagrams for explaining its operation, and FIG. 5 is a diagram showing the configuration of an example of an enzyme immunoassay automatic analyzer implementing the present invention. 10... Constant temperature bath, 11... U-shaped tube, 12... Reaction tube disk, 13... Sample dispensing device, 14
...Sampler, 15...Sample cup, 16,
18... Reagent dispensing device, 17... Enzyme labeling reagent,
19...Coloring reagent, 20...Carrier injector, 21...
... carrier, 22 ... colorimeter, 23 ... carrier extractor,
24...Washing pump, 25...Buffer solution dispensing device,
26... Buffer solution, 27... Stirring air pump,
28...Drainage pump, 29...Reagent dispenser, 30
... Buffer solution storage container, 31 ... Enzyme labeling reagent storage container, 32 ... Coloring reagent storage container, 33 ... Washing tank, 34 ... Switching valve, 35 ... Nozzle.

Claims (1)

[Scope of Claims] 1. In an apparatus for immunologically analyzing a test substance in a sample by causing an antigen-antibody reaction in a reaction container, a reaction container transport means for repeatedly transporting and stopping at a predetermined position at least twice during analysis of one measurement item of a substance; and an enzyme labeled with a predetermined antibody or antigen each time the reaction container stops at the predetermined position. Select one of at least two types of reagents necessary for analysis from a plurality of reagent containers containing a labeled reagent and an enzyme activity measurement reagent for measuring the enzyme activity of the labeled enzyme, and dispense it into a reaction container. 1. An automatic immunological analyzer comprising one reagent dispenser for dispensing at least the enzyme labeling reagent and the reagent for enzyme activity measurement using the one reagent dispenser. 2. The automatic immunological analyzer according to claim 1, wherein the reagent for measuring enzyme activity is a coloring reagent.