JPH0756490B2 - Immunological automatic analysis method - Google Patents

Description

TECHNICAL FIELD The present invention relates to an immunological automatic analysis method.

(Prior Art) In recent years, with the progress of medical treatment, it has become possible to analyze a very small amount of biological components, which is useful for early diagnosis of various diseases. For example, malignant tumors represented by α-fetoprotein, carcinoembryonic antigen, etc., abnormal secretion of hormones represented by insulin, thyroxine, etc., immune disorders represented by immunoglobulin, etc. Not only early diagnosis of various diseases, but also monitoring after treatment of those diseases,
Alternatively, recently, low molecular haptens (incomplete antigens) such as drugs can be measured, which is also useful for drug administration planning.

Many of these biological components are analyzed by an immunochemical method utilizing an antigen-antibody reaction, and as an analysis method utilizing such an immunochemical reaction, for example, synthesis of a predetermined antibody or an antigen-immobilized plastic etc. A labeling reagent involved in an antigen-antibody reaction using an insoluble carrier such as resin or glass beads and a labeling reagent in which a predetermined antibody or antigen is labeled with a marker having high detection sensitivity such as a radioisotope, a fluorescent substance, an enzyme, etc. Heterogeneous to separate B and F by washing operation and those that are not involved and quantify the test substance in the sample based on the labeling reagent after this B and F separation
There is an immunoassay method. This immunoassay method is being generalized because it can be properly analyzed regardless of whether the test substance is a low molecule or a high molecule, and the analysis target is extremely wide.

As such a heterogeneous immunoassay method, a competitive method, a Sangerci method, etc. are known. In the competitive method, as shown in FIG. 1, an antibody or an antigen that causes an antigen-antibody reaction with a test substance in a sample is pre-immobilized on an insoluble carrier 1, and the carrier (carrier reagent) 1 and the sample and the test For example, the same substance as the substance 2 is allowed to undergo an antigen-antibody reaction with, for example, an enzyme-labeled labeling reagent 3 and then washed to compete with the carrier 1 by the antigen-antibody reaction and bind to the test substance 2 and the labeling reagent 3, After separating B and F from those that are not bound, a coloring reagent that reacts with the labeling enzyme in the labeling reagent 3 is added and reacted, and then the reaction solution is subjected to colorimetric measurement to obtain the enzyme activity of the labeling enzyme. The test substance 2 is quantified. As shown in FIG. 2, the Sangertian method uses an insoluble carrier 5 on which an antibody or an antigen that causes an antigen-antibody reaction with a test substance in a sample is immobilized in advance, as shown in FIG. Carrier reagent) 55 and the sample are allowed to undergo an antigen-antibody reaction to bind the test substance 6 in the sample to the carrier 5, and then washing is performed to separate B and F, and then the carrier 5
A substance which causes an antigen-antibody reaction with the test substance 6 is reacted with a labeling reagent 7 labeled with an enzyme to cause an antigen-antibody reaction, and then washing is performed again to separate B and F, and then the labeling reagent 7 is added. After a color-forming reagent that reacts with the labeling enzyme is added and reacted, the reaction solution is subjected to colorimetric measurement to obtain the enzyme activity of the labeling enzyme to quantify the test substance 6. Incidentally, in the Sangeertichi method, the carrier 5, the sample and the labeling reagent 7 may be reacted at the same time, in which case the BF separation is performed once.

The present applicant has developed an apparatus for carrying out such a heterogeneous immunoassay method, but as is clear from the above description, the carrier, labeling reagent, etc., depend on the analyte to be analyzed in the sample. Is different. For this reason, in developing such an automatic analyzer, it is better to make it possible to exchange the carrier, labeling reagent, etc., and analyze for each analysis item, thereby simplifying, downsizing, and controlling the device. Is advantageous in terms of easiness of

(Object of the invention) In view of the above points, an object of the present invention is to enable efficient exchange of a carrier, a labeling reagent, etc. for switching analysis items, and therefore, to efficiently analyze a plurality of items, It is intended to provide an immunological automatic analysis method that can be continuously performed.

(Summary of the Invention) The present invention provides a multi-step reaction for each analysis item, including a carrier reagent on which a predetermined antibody or antigen is immobilized and a labeling reagent on which the predetermined antibody or antigen is labeled with a predetermined substance. Using multiple types of reagents related to, the multi-step reaction results including the antigen-antibody reaction are measured in multiple reaction vessels that are circulated, and the test substances of specified items in the sample are immunologically automatically In the analysis, a plurality of types of reagents related to the multi-step reaction necessary for analyzing a predetermined analysis item, a step of setting in a dispensing part in a state in which they can be exchanged and set, and Distributing the sample to be analyzed and the plurality of kinds of reagents into the plurality of reaction containers in an order according to the reaction step while moving the plurality of reaction containers a plurality of times, and a portion of the plurality of kinds of reagents. Count the number of injections for each reagent For the reagents corresponding to the analysis step and the analysis items that need to be replaced, the dispensing operation is performed when the number of dispensing of each reagent according to the reaction step reaches the number of samples to be analyzed. And a step of displaying the completion for each type of reagent, and based on the display, a reagent corresponding to an analysis item different from the reagent set in the aliquoting unit is transferred from the reagent for which the aliquoting is completed. It is characterized in that a plurality of items can be analyzed continuously and efficiently by sequentially exchanging and setting in the dispensing section.

(Example) FIG. 3 is a diagram showing the construction of an example of an enzyme immunoassay automatic analyzer for carrying out the method of the present invention, which employs the Sangertisch method shown in FIG.

In this example, 24 U-shaped tubes 11 each having a large opening 11a and a small opening 11b are used as reaction vessels, and these reaction tubes 12 are used.
Hold at equal intervals on the same circumference. The reaction tube disk 12 is rotated intermittently in a horizontal plane in a direction indicated by an arrow with a predetermined pitch (for example, 15 seconds) while immersing the U-shaped tube 11 in a constant temperature bath. U-tube 11 by intermittent rotation of this reaction tube disk 12
The stop positions of are indicated by symbols S _{1 to} S ₂₄ . In this example, stop position S ₁
A sample dispenser 13 is used to sample the U-shaped tube 11 in
A sample is selectively dispensed from the sample cup 15 at a predetermined sample suction position of 14. The sampler 14 holds 24 sample cups of the same number as the number of U-shaped tubes held on the reaction tube disk 12 at equal intervals on the same circumference, and in the direction of the arrow in synchronization with the rotation of the reaction tube disk 12. Rotate intermittently.

The U-shaped tube 11 at the stop position S ₁₇ is provided with a large number of synthetic resins such as plastics and insoluble carriers 17 such as glass beads, which are accommodated in the carrier feeder 16 from the large opening 11a, under the control of the carrier loading controller 18. Selectively throw in one. The carrier 17
Is a size that can be easily put in and taken out from the large mouth portion 11a of the U-shaped tube 11 and does not fit into the small mouth portion 11b. On the surface thereof, as described above, an antibody or an antigen that causes an antigen-antibody reaction with the test substance in the sample The carrier is fixed in advance and moistened with the buffer solution in the carrier feeder 16. In addition, the U-tube 11 at the stop position S ₁₉ selectively sucks the reaction liquid contained therein into the colorimeter 19, and the U-tube 11 at the stop position S ₂₀ collects the reaction liquid. The carrier 17 accommodated therein is selectively taken out and discharged by the carrier take-out device 20. Furthermore, the U-tube 11 at the stop position S ₂₂ by the washing pump 21, ion exchange water, immunoassay buffer, selectively injecting washing liquid such as physiological saline.

Further, the U-shaped pipe 11 at the stop position S ₂ has a small opening 11b.
A stirring air pump 22 is detachably connected to the U-tube 11 and a common drainage pump 23 is detachably connected to each of the U-shaped pipes 11 at the stop positions S ₂₂ and S ₂₃ at the small port 11b. .

Furthermore, the U-shaped tube 11 at the stop position S ₂₄ is provided with a syringe-type reagent dispenser 24 capable of precise dispensing, by which the first reagent container
First reagent (buffer solution) 26 in 25, second reagent in second reagent container 27
Reagent (enzyme labeling reagent) 28 and third reagent container 29 in the third
Any one of the reagents (color-forming reagents) 30 is dispensed through the nozzle 31. The nozzle 31 is configured so that it can be transferred to the stop position S ₂₄ , the suction position of each reagent and the washing tank 34 via the nozzle transfer device 33 by the nozzle transfer control device 32, and at least the suction position of each reagent and the cleaning tank 34. It is configured to be able to move up and down.

In this example, in the carrier charging control device 18, the carrier charging device 16
When the number of loaded carriers 1 contained in the sample is counted, and when the count value is equal to the number of samples set in the sampler 14, it is indicated that the carrier loading has been completed for all the samples for the analysis item based on the information. Is displayed on the display device 35. Similarly, in the nozzle transfer control device 32,
When the number of times of dispensing each of the first reagent 26, the second reagent 28, and the third reagent 30 is counted and the count value of each reagent is equal to the number of samples set in the sampler 14, the analysis is performed based on the information. The display device 35 displays that the dispensing of the reagent has been completed for all the samples corresponding to the items. As the display device 35, a known device such as CRT or LED is used.

Next, the operation of the enzyme immunoassay analyzer shown in FIG. 3 will be described. First, the carrier 17 and the first to third reagent containers 25, 27, 29 corresponding to a certain analysis item are set. Further, the number of suppli cups set in the sampler 14, that is, the number of samples to be analyzed is previously input to the carrier charging control device 18 and the nozzle transfer control device 32. Reaction Tube Disk 1 of 12
At the turning point, first, at the stop position S ₁₇ , the carrier loading device 16
Thus, the carrier 17 corresponding to the analysis item is put into the U-shaped tube 11. The U-shaped tube 11 into which the carrier 17 has been charged is washed by the operation of the washing pump 21 and the drainage pump 23 at the stop position S ₂₂ , and the first reagent 26 by the reagent dispensing 24 at the stop position S ₂₄ .
Is dispensed in a fixed amount. After that, by the operation of the sample dispensing device 13 at the stop position S ₁ , a fixed amount of sample is dispensed from the sample cup 15 at the predetermined sample suction position, and by the operation of the stirring air pump 22 at the next stop position S ₂ . The first reaction begins with stirring. After the carrier loading operation, the first reagent dispensing operation, and the sample dispensing operation have been performed the number of times equal to the number of samples set in the sampler 14, these operations are not performed. This reaction tube disk 12
In the first rotation of the above, when the carrier inserter 16 finishes the operation of inserting the carrier 17 the number of times equal to the number of samples set in the sampler 14, the fact is displayed on the display device 35 at that time, and the reagent dispensing is performed similarly. When the container 24 finishes the dispensing operation of the first reagent 26 the number of times equal to the number of set samples, the fact is displayed on the display device 35 at that time.

At the second rotation of the reaction tube disk 12, the U-shaped tube 11 is cleaned by the action of the cleaning pump 21 and the drainage pump 23 at the stop position S ₂ , the first BB / F separation is performed, and then the stop position S 2. _{At 24} , the reagent dispenser 24 dispenses a fixed amount of the second reagent 28 to start the second reaction. The test liquid in the U-shaped tube 11 into which the second reagent 28 has been dispensed is stirred by the stirring air pump 22 at the stop position S ₂ . In the second rotation of the reaction tube disk 12, when the dispensing operation of the second reagent 28 of the number equal to the number of the set samples by the reagent dispensing device 24 is completed, that effect is displayed on the display device 35 at that time. It

At the third rotation of the reaction tube disk 12, at the stop position S ₂₂ , the action of the cleaning pump 21 and the drainage pump 23 causes a U-shaped tube.
11 second time B · F separated and washed is carried out, then the third reagent 30 of the nozzle transferring device 34 by the reagent dispensing unit 24 selects the coloring reagent container 29 at the stop position S ₂₄ is constant amount The third reaction begins when poured. The test liquid in the U-shaped tube 11 into which the third reagent 30 has been dispensed is stirred at the stop position S ₂ to promote the enzymatic reaction. At the third rotation of the reaction tube disk 12, when the dispensing operation of the third reagent 30 by the number equal to the number of the set samples by the reagent dispensing device 24 is completed, the display device 35 at that time is displayed.

After that, at the fourth rotation of the reaction tube disk 12, the stop position S
_{At 19} , the test liquid in the U-shaped tube 11 is sucked into the colorimeter 19 for colorimetric measurement, and then at the stop position S ₂₀ , the carrier 17 remaining in the U-shaped tube 11 is taken out by the carrier take-out device 20. . Then, at the stop position S ₂₂ , the cleaning pump 21 and the drainage pump 23
After U-shaped tube 11 is cleaned by the remaining liquid is discharged by the discharge pump 23 at the next stop position S ₂₃ is prepared for the next analysis. During the above operation, the nozzle 31 is transferred to the cleaning tank 34 after dispensing each reagent, and the inner and outer walls of the nozzle 31 are cleaned by the suction and discharge operation of the reagent dispenser 24.

As described above, in the present embodiment, the carrier 17, the first reagent 26, and the second reagent which need to be exchanged according to the analysis item.
For each of the 28 and the third reagent 30, the fact that the loading operation and the dispensing operation have been completed for the set sample is displayed at the end time of each operation. The carrier and reagent used in the analysis item can be efficiently exchanged and set. Therefore, it is possible to efficiently analyze a plurality of items for a plurality of samples.

It should be noted that the present invention is not limited to the above-mentioned examples, and many modifications and variations are possible. For example, in the above-mentioned embodiment, the number of operations is counted, but a final identification of the sample cup 15 set in the sampler 14 or an appropriate identification member is provided in the vicinity of the set position, which is used for optical measurement. Alternatively, the number of times each reagent is dispensed may be indirectly measured based on the detection signal, and the end of each operation for the final sample may be displayed. In addition, although the enzyme immunoassay according to the Sangertisch method is carried out in the above-mentioned examples, it can be similarly applied to the analysis by the competitive method, and the radioimmunoassay using a radioisotope as a marker and the fluorescence as a marker. It can be similarly applied to a fluorescent immunoassay using a substance. Further, the reaction container does not necessarily have to be held on the disk, and for example, a snake chain or gondola type transfer device can be used, and the sampler can be changed in various ways such as using a rack. Further, in the above-mentioned example, the colorimetric measurement was carried out by guiding the finally obtained test solution to the colorimetric cell.However, the colorimetric measurement was carried out in a state where the test solution was present in the reaction vessel using a transparent reaction container. It is also possible to use the direct metering method. In this case, if the carrier remaining in the reaction vessel interferes with the photometry, the carrier can be removed before the photometry. Further, when such a direct photometric method is adopted, the carrier can be discharged together with the test solution after the photometry, so that the carrier discharging device becomes simple.
Further, in the above-mentioned embodiment, the reaction container is used repeatedly, but this is not always necessary, and the reaction container used for the analysis can be disposable. Further, various dispensing positions, carrier loading / unloading positions, colorimetric measurement positions, etc. are not limited to those in the above-described embodiment, and various changes can be made.

(Effects of the Invention) As described above, in the immunological automatic analysis method of the present invention, a reagent corresponding to a plurality of types of reaction steps including a carrier reagent and a labeling reagent corresponding to the analysis item to each reaction container is used. The fact that the number of samples dispensed for each analysis is completed for each type of reagent is displayed, which reduces efficiency even during analysis when multi-step reactions are being performed during multiple rounds. It is possible to replace the reagents of the kind to be exchanged in the dispensing section in order without needing to do so, and thus it is possible to continuously and efficiently perform the analysis of a plurality of items for a plurality of suples.

[Brief description of drawings]

FIG. 1 is a diagram showing the process of enzyme immunoassay by the competitive method, FIG. 2 is a diagram showing the process of enzyme immunoassay by the Sangerci method, and FIG. 3 is an automatic analyzer for carrying out the assay method according to the present invention. It is a diagram showing an example of a configuration. 11 …… U-shaped tube, 12 …… Reaction tube disk 13 …… Sample dispenser, 14 …… Sampler 15 …… Sample cup, 16 …… Carrier feeder 17 …… Carrier, 18 …… Carrier charging controller 19 …… Colorimeter, 20 …… Carrier extractor 21 …… Washing pump 22 …… Stirring air pump 23 …… Drainage pump, 24 …… Reagent dispenser 25 …… First reagent container, 26 …… 1 reagent 27 ...... second reagent container, 28 ... second reagent 29 ... third reagent container, 30 ... third reagent 31 ... nozzle 32 ... nozzle transfer control device 33 ... nozzle transfer device, 34 ... … Cleaning tank 35 …… Display device

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Claims (1)

[Claims] 1. A plurality of multi-step reactions relating to each analysis item, including a carrier reagent on which a predetermined antibody or antigen is immobilized and a labeling reagent on which the predetermined antibody or antigen is labeled with a predetermined substance. When immunologically and automatically analyzing a test substance of a specified item in a sample by measuring multi-step reaction results including antigen-antibody reaction in multiple reaction vessels that are orbited using different types of reagents A step of setting a plurality of types of reagents related to the multi-step reaction required for analyzing a predetermined analysis item in a dispensing part in a state in which they can be exchanged and set, and the plurality of reaction vessels During a plurality of circular movements, a step of dispensing the sample to be analyzed and the plurality of types of reagents into a plurality of reaction vessels in an order corresponding to the reaction step, and the number of dispensing times of the plurality of types of reagents The process of measuring each Regarding the reagents corresponding to the analysis items that need to be exchanged and set, the dispensing operation should be completed when the number of dispensing of each reagent according to the reaction stage reaches the number of samples corresponding to the number of samples to be analyzed. And a step of displaying for each type of reagent, and based on the display, the reagents corresponding to the analysis items different from the reagent set in the dispensing unit are dispensed in order from the reagent which has been dispensed. An immunological automatic analysis method, characterized in that a plurality of items can be continuously and efficiently analyzed by exchanging and setting them in a section.