JPS6411910B2 - - Google Patents

Description

Detailed Description of the Invention The present invention provides a method for dispensing particle samples, blood cell components such as red blood cell types, white blood cell types, platelets, lymphocyte shapes and types, various antibodies, antigens, specific proteins, viruses, etc. The present invention relates to a method for separating components and foreign substances in blood serum by an agglutination reaction using blood cell particles, latex particles, carbon particles, etc., and a particle sample dispensing method particularly suitable for application to an apparatus.

Conventionally, it has been widely practiced to detect and analyze various components in blood (blood types, various antibodies, various proteins, etc.) and foreign substances such as viruses by determining the aggregation patterns of blood cell particles, latex particles, and carbon particles. ing. However, in conventional immunological analysis methods using agglutination reactions, the reaction vessels and analysis procedures used vary depending on the properties of each component. For example, the most common conventional method for determining ABO blood type is a method that uses a test tube as a reaction vessel. In this method, serum and blood cells are separated by centrifugation, the blood cells are collected to create a 2-5% blood cell suspension, which is aliquoted into test tubes, and then anti-A serum or anti-B serum is tested. After dispensing a fixed amount into a tube and centrifuging it, it is visually determined whether aggregates are formed after shaking the test tube. In this case, the sample blood that agglutinated with anti-A serum but not with anti-B serum is type A, and the blood sample that did not agglutinate with anti-A serum but agglutinated with anti-B serum is type B. Those that agglutinate with both anti-A and anti-B sera are determined to be type AB, and those that do not agglutinate with both anti-A and anti-B sera are determined to be type O. In addition, as a method for detecting HBs antigen, there is a method using a microplate having many holes with a V or U bottom. This method is
For example, a 10 x 12 well microplate is used to detect and measure HBs antigen using the method described below.

1 Add one drop (0.025 ml) of R-PHA buffer to each well of the microplate.

2. Take the sample into a diluter and dilute it twice in two series up to 10 tubes.

3. Add one drop (0.025 ml) of R-PHA buffer solution to the first column of the sample dilution series, and one drop (0.025 ml) of R-PHA inhibition solution to the second column.

4 After shaking thoroughly for 10 seconds with a micro mixer,
Incubate at ℃ for 1 hour.

5 1 drop of R-PHA cell (1% suspension 0.025ml)
Add to each hole.

6 Shake thoroughly for 10 seconds with a micro mixer, and
- Uniformly suspend PHA cells.

7. Detect the aggregation pattern after leaving at room temperature for 1 hour avoiding vibration.

Furthermore, in testing for syphilis using the TPHA method,
Make a dilution series of the test serum in a microplate,
A reagent containing syphilis pathogen body components bound to sheep red blood cells is mixed with this, and after natural sedimentation, it is visually determined whether aggregates have been formed.

As described above, in immunoagglutination-based analysis methods, the reaction containers and analysis procedures used vary depending on the analysis item.

On the other hand, a microtiter method is known, which is a partially mechanized analysis method based on an agglutination reaction that uses a microplate as a reaction vessel. In this system, for example, quantitative dispensing of a specimen, quantitative dispensing of a reagent, and detection of the presence or absence of agglutination are performed mechanically, and other parts are performed manually. That is, when using a microplate, the instruments and operations used are complicated, making it extremely difficult to automate the entire process. However, this microtiter method also has various drawbacks. For example, quantitative dispensing of sample serum utilizes the capillary phenomenon caused by the diluter, so a volume of diluent is dispensed into each hole of the microplate in advance, and the tip of the diluter from which the sample was collected is inserted into the diluent. It is necessary to rotate and stir. Since such an operation is considerably more complicated than the dispensing operation in a typical biochemical analyzer, there is a drawback that the configuration and control device thereof are extremely complicated. In addition, since capillary action is used, the amount dispensed is always constant, making it impossible to obtain an arbitrary amount of dispensing, and the diluted sample is partially removed due to capillary action after dispensing. Therefore, samples are wasted. Such a problem becomes a major drawback in multi-item processing of a large number of samples. Furthermore, unless the blood cell reagent is dispensed after the sample serum is dispensed, an unspecified amount of the blood cell sample itself will be removed by the direuter due to capillary action. Therefore, in the microtiter method, it is necessary to sequentially perform dilution liquid dispensing, serum dispensing, and blood cell dispensing, which has the disadvantage that mechanical arrangement is limited. Furthermore, when determining blood type by this microtiter method, a blood cell suspension of around 1% must be prepared, which makes the operation much more complicated than in the above-mentioned test tube method.

The applicant is Japanese Patent Application No. 54-53370
No. 146044), we proposed a blood type determination method that can sufficiently accurately determine blood types using natural antibodies with strong aggregation and binding strength, as well as blood types using irregular antibodies with extremely weak aggregation and binding strength, using an extremely small amount of blood.
Such a blood type determination method uses, for example, a reaction container with a conical bottom, dispenses blood corpuscles and a standard antiserum reagent into which the blood type is to be determined, and stirs the resulting mixture for a relatively short period of time. The blood type is determined by detecting the agglutination pattern after the device is allowed to stand (for about 30 minutes). In this method, when the blood cells to be tested react with the antiserum reagent, the aggregated blood cells settle and are deposited thinly like snow on the bottom of the conical shape, but when the blood cells and the antiserum reagent do not react, The blood cell particles do not aggregate, but settle as discrete particles, and when they reach the conical bottom, they roll down the slope and collect in the center of the conical bottom. Therefore, the blood type can be determined by photoelectrically detecting the difference in the pattern of precipitated blood cells depending on the presence or absence of reaction with the antiserum reagent formed on the bottom of the cone. In addition to determining blood type, this method can also be used to
Various antigens and antibodies such as HBs antigen and syphilis antibody can also be effectively detected.

The present inventors have also found that by performing such a method automatically, it is possible to improve analysis efficiency and accuracy, and in particular, to perform various immunological analyzes on blood, ie, red blood cells, using the same device. Blood cell components such as type, white blood cell type, platelet, lymphocyte type and type, various antibody antigens, specific proteins, components in serum such as viruses, and foreign substances such as blood cell particles, latex particles, carbon particles, etc. Patent application No. 55-155021 for an analyzer based on immunological agglutination reaction, which is appropriately configured to enable selective analysis depending on the agglutination reaction used.
(Japanese Patent Application Laid-Open No. 57-79450).

This analyzer uses a microplate with a large number of reaction vessels arranged in a matrix on a substrate, each of which has a slightly inclined bottom. , in an analyzer that performs immunological analysis of blood type, various antigens, antibodies, etc., based on an agglutination pattern formed on the bottom surface of a reaction container as a result of an antigen-antibody binding reaction of particles that settle due to natural sedimentation, the microplate. means for sequentially supplying from the inlet side of the reaction line, and means for dispensing a predetermined amount of the blood sample to be analyzed, that is, serum or blood cells or a suspension thereof, into at least one reaction container of the microplate on the reaction line. , a means for dispensing a predetermined amount of a reagent, that is, a particle reagent or a serum reagent, according to an analysis item into the reaction vessel, and a microplate with the blood reagent dispensed into the reaction vessel in this way, approximately along the reaction line. means for transporting it in a stationary state;
After the sample and reagent are dispensed into the reaction container, the antigen-antibody binding reaction is carried out while the sample and reagent are transferred on the reaction line in an almost stationary state, and the agglutination pattern formed on the bottom of the reaction container is photoelectrically detected. A detection means, a circuit that performs immunological analysis by determining the presence or absence of particle aggregation due to natural sedimentation based on the output signal obtained from the photometric detection means, and a microcomputer that conducts an immunological analysis for all reaction vessels. and means for sequentially discharging the plates from the outlet side of the reaction line.

In such a blood analyzer, whole blood collected from a subject is centrifuged to separate a blood sample and a blood cell sample, and a predetermined amount of each sample is diluted and dispensed to prepare a test solution. Dispensing of this internal blood cell sample can be carried out in various ways, including aspirating a predetermined amount with a dispensing nozzle, discharging it into a reaction vessel, and subsequently dispensing the diluted liquid through the same nozzle, or dispensing it into a reaction vessel. Generally, a predetermined amount of diluent is dispensed in advance and the blood cell sample is discharged into it. However, in the former method, when a blood cell sample is discharged into an empty reaction container, scattered blood cells adhere to the inner wall of the container. Even if the amount of this adhesion is small, the total amount of blood cell sample is small, resulting in a relatively large error, which has the drawback of lowering the accuracy of analysis. In addition, in the latter dispensing method, the diluent is stored in the reaction container in advance, which reduces the number of blood cells adhering to the inner wall, but the blood cells are not well dispersed in the diluent. Therefore, after dispensing the blood cells, it is necessary to stir the reaction container by vibration or other operations to uniformly disperse the blood cells in the diluent.

The present invention aims to eliminate these drawbacks and provide a method for dispensing particle samples such as blood cells, which prevents particle samples from adhering to the inner wall of a reaction vessel and allows them to be uniformly dispersed in a diluent. It is something.

When diluting and dispensing particle samples such as blood cell particles, latex particles, carbon particles, glass particles, etc., the present invention provides a first dispenser that aspirates and dispenses a predetermined amount of a particle sample, and a first dispenser that dispenses a predetermined amount of a diluent. After the second dispenser starts dispensing the diluent, the first dispenser dispenses the particle sample, and the diluent continues to flow until after dispensing the particle sample. It is characterized by continuous dispensing.

The present invention will be described in detail below with reference to the drawings.

FIG. 1 shows the sequential steps of an analytical method for blood type determination suitable for applying the particle sample dispensing method according to the present invention. Whole blood BL collected from the subject 1 is stored in a test tube 2. This test tube 2 is centrifuged using a centrifuge in the usual manner, and is separated into serum S, which is a supernatant fluid, and blood cell (erythrocyte) particles R, which are deposits. Normal blood has a blood cell content of approximately 40%. Next, from the test tube 2 using the dispenser 3-1 shown as a microsyringe,
Collect 30μ blood cell particles R and transfer to another test tube 4-1
Dispense. This test tube 4-1 has another pipettor 5.
-1 the diluent DL stored in the diluent container 6;
For example, dispense 1970μ of physiological saline. Therefore, test tube 4-1 contains 1.5% blood cell suspension RD.
2000μ obtained. Also, from the test tube 2 to the dispenser 3-
Dispense 200 μ of serum S into test tube 4-2 according to step 2, and add 600 μ of diluted solution into it using dispenser 5-2.
Dispense the DL to obtain 800 µl of 25% serum dilution SD.

Next, 25 micrometers of the thus obtained blood cell suspension RD in the test tube 4-1 is collected by the dispenser 7-1 and dispensed into a reaction container 8-1 having a conical bottom. 25 μ of the 12.5% anti-A serum reagent A-S contained in the container 10-1 is dispensed into the reaction container 8 using another dispenser 9-1. A further 25μ is sampled from the blood cell suspension RD in the test tube 4-1 using the dispenser 7-1 and dispensed into another reaction container 11-1. This reaction vessel 11-1 has another dispenser 12-
1, dispense 25μ of the 12.5% anti-B serum reagent B-S contained in the container 13-1. In this way, the test solution A-T-1 containing the test blood cell particles and the anti-A serum reagent is placed in the reaction vessels 8-1 and 11-1.
and test solution B-T-1, which is a mixture of test blood cell particles and anti-B serum reagent, are obtained.

On the other hand, 25μ of the diluted sample serum SD obtained in the test tube 4-2 is sampled using the dispenser 7-2 and dispensed into a reaction container 8-2.
2, dispense 25μ of the 1.2% A blood cell reagent A-R contained in the container 10-2. This serum dilution SD
Further, the dispenser 12-2 dispenses 25μ into the reaction container 11-2, and here the B blood cell reagent B-R contained in the container 13-2 is dispensed using the dispenser 12-2.
Dispense 25μ. In this way, reaction vessel 8-2
and 11-2, test serum dilution solution and A
Test solution A-, which is a mixture of blood cell reagent and B blood cell reagent.
T-2 and B-T-2 are obtained, respectively. As will be described later, it is suitable for liquid processing that these reaction vessels are formed by forming a microplate in which a large number of reaction vessels are arranged in a matrix. Next, these test solutions A-T-1, B-T
-1, reaction vessels 8-1, 11-1, 8-2 and 11-2 containing A-T-2 and B-T-2
Stir by vibrating each. However, reagent A-
When dispensing R, B-R and antiserum reagents A-S, B-S into the reaction vessels 8 and 11, if both liquids are sufficiently stirred, there is no need to stir them again. do not have. After stirring, the microplate is gently transferred along the reaction line so as not to apply strong vibrations to the reaction vessels. During this time, natural sedimentation of blood cell particles takes place. This standing time can be, for example, 30 minutes. In this example, it is assumed that the sample blood is type A. Therefore, in the test solution A-T-1 to which the anti-A serum reagent A-S has been added, blood cells are aggregated and deposited in a thin layer on the bottom of the reaction container 8-1. On the other hand, test solution B to which anti-B serum reagent B-S was added
- Blood cells do not aggregate in T-1, and reaction vessel 11-1
It settles to the bottom of the mountain, rolls down its slope, and collects in the center. If the blood cells aggregate in this way, the pattern will be uniform with blood cells existing on the entire bottom surface, as shown in the drawing, and if they do not aggregate, there will be almost no blood cells in the periphery, and a large number in the center. The result is an accumulation pattern in which blood cells gather together. Blood type can be determined by photoelectrically detecting such a pattern. Generally, this determination is called table determination. In order for unagglutinated blood cells to roll down to the center, it is preferable that the angle of the bottom surface be approximately 30° with respect to the horizontal.

On the other hand, in reaction vessel 8-2, aggregation does not occur and an accumulation pattern is formed on the bottom surface, and in reaction vessel 11-2, aggregation occurs and a uniform deposition pattern is formed on the bottom surface. Blood type can be determined by photoelectrically detecting these patterns. This determination is generally called a tail determination. Since it is possible to perform both front and back determination in this manner, the determination accuracy is significantly improved. The microplates for which pattern determination has been completed for all reaction vessels are sequentially discharged from the reaction line.

In Figure 1, it is shown that the reagent is dispensed after the sample blood cell suspension and diluted serum are dispensed into the reaction container, but these dispensing orders may be reversed, or they may be dispensed at the same time. Of course it's a good thing.

According to this blood type determination method, since the reaction container is left still and an agglutination pattern is formed by natural sedimentation, it is possible to accurately determine blood type using irregular antibodies with weak aggregation binding strength. can. The reason for this is that once bound blood cells are not separated by shaking the reaction container as in the conventional method. Furthermore, the blood cell suspension only needs to be extremely dilute at 0.5 to 2%, and the amount thereof is as small as 25 microns, so the amount of blood required is also very small. In contrast, conventional methods require a considerable amount of 2-5% blood cell suspension. Furthermore, according to this determination method, other types of blood types and immunological analyzes other than blood types can also be effectively determined using the same method.

FIG. 2 is a plan view diagrammatically showing the configuration of an example of an analyzer that implements the dispensing method according to the present invention and automatically performs the above-described analysis method. Reference numeral 15 designates the entire sampler, in which a rack cassette 16 is loaded with a number of racks 17, and each rack is loaded with a number of sample containers 18. These sample containers 18
are three sample containers 18-1, 18- arranged in a row.
2,18-3 make one set, and one rack 17 has
Attach 12 rows of sample containers. Sample containers 18-1-1, 18-2-1, 18-3 at the bottom of each row
-1... contains centrifuged sample blood (whole blood), and the remaining two sample containers 18-1...
-2,18-1-3;18-2-2,18-2-
3; Leave blank. The rack 17 with the rows of sample containers is transported at a predetermined pitch in the direction indicated by arrow A and enters the extension of the sample guide line 19.
When one rack 17 has entered the sample guide line 19, the rack 17 moves one pitch in the direction of arrow A. A pair of endless belts 20-1 and 20-2 are provided on the sample guide line 19, and are constantly rotated at a relatively high speed in the direction of arrow B, so that the rack 17 entering the line 19 is sent to the left. . This rack 17 is indexed to the sample dilution position C by the stopper 21-1, and the blood cells and serum samples contained in the sample container 18-1-1 are transferred to the dispenser 21.
and discharges a predetermined amount into empty sample containers 18-1-2 and 18-1-3. This dispensing device 22 has two dispensing nozzles 22-1 and 22-
2, each of which is connected to an arm 22-
3 and 22-4, and their tips are inserted to different depths into the sample container 18-1-1 containing centrifuged blood, and then suction is performed to aspirate blood cells and serum, respectively. Next, after pulling up the nozzle, move it horizontally and position it above the empty sample containers 18-1-2 and 18-1-3, respectively, and transfer the previously aspirated blood cells and serum into the respective sample containers. Exhale. At the same time, add a predetermined amount of physiological saline contained in the diluent container 6 to the dispenser 3.
00 and discharged from nozzles 301 and 302 into sample containers 18-1-2 and 18-1-3 to prepare predetermined amounts of a 1.5% blood cell suspension and a 25% serum dilution. Finally, nozzle 22-1,
22-2 is immersed in the cleaning tank 22-5 to be cleaned. When the above-described dispensing and diluting operation for the samples in the first row in the rack 17 is completed, the stopper 21-1 is driven, and the sample containers in the next row are indexed to the dilution position C, and the same dispensing and diluting is performed. . In this example, the pitch of sequential movement of the sample container rows is 15 seconds. Therefore
The dispensing and dilution of all the samples stored in one rack 17 is completed in 15×12=180 seconds=3 minutes. This rack 17 is attached to belts 20-1 and 20-2.
The sample is further sent in the direction B and reaches the sample dispensing position D. A stopper 21-2 is also provided at this position so that successive rows of sample containers can be indexed at intervals of 15 seconds. A sample dispensing device 23 having four dispensing nozzles is provided at this sample dispensing position D. The detailed structure and operation of this sample dispensing device will be described later.
Here we will explain its functions.

As shown in FIG. 2, a reaction line 24 is provided in parallel with the sample guide line 19, in which three reaction line passages 24-1 to 24-3 are arranged parallel to each other. Therefore, the reaction vessels are transferred in a zigzag pattern along these three reaction line passages. The left end of the first reaction line passage 24-1 is the entrance of the reaction line, from which a microplate 26 with a large number of reaction vessels 25 arranged in a matrix is inserted into a frame-shaped cassette 27, as shown in FIG. The combined products will be supplied sequentially. Hereinafter, in FIG. 2, for convenience, the combination of a microplate and a cassette will be simply referred to as a microplate. Various types of autoloading mechanisms can be used for this microplate autoloading mechanism, but in this example, as will be described later, an elevator-type autoloader is used to load a large number of microplates 26.
are stacked vertically and stored and supplied to the reaction line 24 in order from the upper microplate. Therefore, in FIG.
The microplate at the left end of 1 has not yet been placed on the reaction line. A pair of endless belts 28-1, 28-2; 29 are provided in each passage 24-1, 24-2 and 24-3 of the reaction line.
-1, 29-2; 30-1, 30-2 are provided, and everything except the belt 29 always rotates at a relatively high speed in the direction indicated by the arrow.

The microplate 26 supplied to the reaction line 24 is conveyed by belts 28-1 and 28-2, and is moved to the first reagent dispensing position E by the stopper 21-3 before reaching the sample dispensing position D mentioned above. It will be indexed. Microplate 26 in this example has 8×12
8 reaction vessels 25 are formed so that eight analyzes can be performed on each sample. There are various types of these analyses, but in this example, we will use the front and back ABOs explained in Figure 1.
Blood type determination, Rn blood type, and antibody screening will be performed. That is, the first and second reaction vessels 25-1-1 and 25-1
Table determination is performed using -2, Rh formula determination is performed using the third and fourth reaction vessels 25-1-3 and 25-1-4, and Rh formula determination is performed using the fifth and sixth reaction vessels 25-1- 5 and 25-1-6, and the seventh and eighth reaction vessels 25-1-7 and 25-1-8
ABO-style tails determination shall be performed. For this purpose, a first reagent dispensing device 31 is provided at the first reagent dispensing position E, and its eight dispensing nozzles 31-
1 to 31-8 are selectively operated to open the reagent container 32.
A predetermined amount of the required reagents from -1 to 32-8 is dispensed. That is, in this example, 12.5% anti-A serum (diluted with physiological saline) and 12.5% anti-B serum are placed in the first to fifth reagent containers 32-1 to 32-5, respectively.
Contains serum (diluted with saline), 12.5% anti-D serum (diluted with phosphate buffer), phosphate buffer and 0.44% bromelin. First, the arm supporting the dispensing nozzle is retracted and the nozzles 31-1 to 31-8 are moved to the reagent container 32-1.
After positioning it on ~32-8, lower the nozzle,
Predetermined amounts of 25 μ of the above-mentioned anti-A serum and anti-B serum, 12.5 μ of anti-D serum and phosphate buffer, and 12.5 μ of bromelin are sucked into the nozzles 31-1 to 31-5. Next, after raising the nozzles, the arm is extended and the nozzles 31-1 to 31-8 are
Eight reaction vessels 25-1 located on reagent dispensing position E
-1 to 25-1-8, and discharge the reagent to reaction containers 25-1-1 to 25-1-5. When the reagent dispensing to one row of reaction containers is completed, the stopper 21-3 operates and the microplate 26 is
Move it by a pitch and operate in the same way. When the reagent dispensing operation for all the reaction container rows is completed, the microplate 26 is moved to the belt 28-1, 28-2.
is sent to the right by the next stopper 21-4 and indexed to the above-mentioned sample dispensing position D. A sample dispensing device 23 is provided at this position, and its four nozzles 23-1 to 23-4 collect the blood cell suspension and diluted serum from the sample containers 18-1-2 and 18-1-3. Suction and reaction vessel 25-1-
1 to 25-1-8. In this example, nozzle 2
3-1, 23-2 into the sample container 18-1-2 to aspirate the blood cell suspension, and the nozzles 23-3, 2
3-4 into the sample container 18-1-3 to aspirate the diluted serum. The arms 23-5 to 23-8 supporting these nozzles are appropriately driven to move the nozzles 23-1 to 23-4 into the reaction vessels 25-1-1, 25-1-3, 25-1-5 and 25-1, respectively. Position it above -1-7. Here, the syringe of the dispensing device 23 is operated, and the reaction container 25 is
Dispense 25μ of 1.5% blood cell suspension into -1-1 and 25-1-3, and dispense 25μ of 25% serum dilution into reaction vessels 25-1-5 and 25-1-7. . Next, move the arms 23-5 to 23-8 to connect the nozzles 23-1 to 23-4 to the reaction vessel 2.
5-1-2, 25-1-4, 25-1-6, and 25-1-8 respectively, and operate the syringe of the dispensing device 23 again to fill the reaction vessel 25.
Dispense 25μ of blood cell suspension into reaction vessels 25-1-6 and 25-1-2 and 25-1-4.
Dispense 25% serum dilution into Steps 1-8. After dispensing,
Move the arms 23-5 to 23-8 to remove the nozzle 2.
3-1 to 23-4 are immersed in a cleaning tank 23-9 to clean the nozzles.

A row of sample containers 18-1-2 and 18-1-
The blood cell suspension and diluted serum contained in the microplate 26 are transferred to a row of reaction vessels 25-1-
After dispensing to 1 to 25-1-8, stopper 21
-2 and 21-4 are driven to move the rack 17 containing the sample containers one pitch to the left and move the microplate 26 containing the reaction containers one pitch to the right. Both of these movement periods are 15 seconds, but the movement amounts are different. In this way, the dispensing operation for the samples is performed sequentially, and when the dispensing of all the samples to one rack 17 is completed, the rack 17 is moved to the left by the belts 20-1 and 20-2 and stored in the rack. It is stored in a cassette 33. The cassette 33 containing the rack 17 is transported in the direction indicated by arrow F at a predetermined pitch. On the other hand, the microplate 26, which has received the samples dispensed into all the reaction vessels, is then indexed to the second reagent dispensing position G by the stopper 21-5. This second reagent dispensing device is provided with a reagent dispensing device 34 having exactly the same configuration as the reagent dispensing device 31. There are eight reagent containers 35-1 to 35-8 corresponding to eight nozzles 34-1 to 34-8, and the third to eighth reagent containers 35-3 to 35-8 contain 0.44%. Bromelin (phosphate buffer dilution), 0.44% Bromelain (phosphate buffer dilution), 2.4% O blood cell reagent, 2.4%
It contains 1.2% O blood cell reagent, 1.2% A blood cell reagent (diluted with physiological saline), and 1.2% B blood cell reagent (diluted with physiological saline). Nozzle 34-3
Add bromelin to a 12.5μ reaction vessel 25-1.
-3, bromelin was dispensed into the 12.5μ reaction container 25-1-4 through nozzle 34-4, and O blood cell reagent was dispensed through nozzle 34-5 and 34-6.
12.5μ reaction vessels 25-1-5 and 25-1-
6, and add A blood cell reagent through nozzle 34-7.
Dispense into 25μ reaction vessel 25-1-7 and use nozzle 3.
4-8, add the B blood cell reagent to a 25 μ reaction vessel 25-
Dispense into portions 1-8.

This second reagent dispensing position G is the reaction start position,
From this point on, the binding and aggregation reaction is carried out by leaving the mixture almost stationary for 30 minutes. A fixed stopper is provided at the right end of the first passage 24-1 of the reaction line 24, and at a predetermined timing after the microplate 26 reaches this position, it is moved to a second stopper by a microplate transfer device 35, which will be described later. It is moved to the right end of passage 24-2. This conveying device 35 has belts 28-1, 2
It has a plate-shaped arm 35-1 that freely passes between the arms 8-2 and can reciprocate in the vertical direction in FIG. That is, arm 3
5-1 is located below the first passage 24-1, and when the microplate 26 comes, it rises, supporting the microplate 26 in this process, and supporting the belt 28-1.
1, lift it above 28-2. Next, as shown by the imaginary line, the microplate 26 is moved downward in FIG. 2 and transported above the second passage 24-2. Next, attach the arm 35-1 to the belts 29-1, 29
Descend below -2 and place the microplate on the belt. In this way, the first passage 24-
The microplate can be moved from the end point of the first passage 24-2 to the starting point of the second passage 24-2 in a substantially stationary state. In this example, it is assumed that this movement also requires 3 minutes.

Microplate 2 in second passage 24-2
6 is moved to the left by belts 29-1 and 29-2, but since no dispensing is performed in this path, the speed is set very slowly, for example about 15 mm/
Move in 15 seconds. A fixed stopper is also provided near the end point of the second passage 24-2, and when the microplate 26 comes to this position, a microplate conveyance device 36 having the same configuration as the above-mentioned microplate conveyance device 35 moves the microplate 26 to the third passage 24-2. −3 is moved to the starting point. The microplate 26 transferred to the third passage 24-3 is transferred to the right by belts 30-1 and 30-2 moving at a relatively high speed, and is positioned at the pattern detection position H by the stopper 21-6. . At this detection position H, a pattern formed on the bottom surface of the reaction vessel is photoelectrically detected by a pattern detection device 37, which will be described later. The microplate 26 moves from the reaction start position G to the detection position H.
It takes 30 minutes to reach this point, so the reaction time is 30 minutes. The pattern-detected signal is supplied to a determination circuit 38, which will be described later, where various determinations are made.
The results are displayed on the display device 39. At the pattern detection position H, the microplate 26
The reaction vessels 25 inside are processed one row at a time.

The microplate 26, on which the patterns of all the reaction containers 25 have been determined, is further transferred to the right and is stopped at the stopper 21-7 at the photographing position I.
will be stopped by At this photographing position I, an illumination lamp is placed above the microplate 26 and a camera is placed below so that the patterns of all reaction vessels 25 on the microplate 26 can be photographed at once from the back side of the plate. . The photographed microplate 26 is then indexed to a visual observation position J by a stopper 21-8. At this position, the operator can visually observe the pattern formed on the bottom surface of the reaction vessel.
In this viewing section, an illumination lamp is installed below the microplate. Reaction line passage 24-3
The microplates that have reached the right end are ejected from there and stored in the microplate storage device 40 in a stacked manner. For this reason, the storage device 40 is provided with a plate 40-1 that moves up and down.

FIG. 3 is a flowchart showing the sequential operations of the above-mentioned analyzer, and since the details are clear from the above explanation, they will be omitted.

FIG. 4 shows a dispensing device 22 and a diluent dispenser 300 for dispensing blood cells and serum samples contained in sample containers into two empty sample containers according to the dispensing method of the present invention. 1 shows the configuration of an example of a sample diluter that has a sample diluter. The dispensing device 22 is 2
having book dispensing nozzles 22-1 and 22-2,
These are arms 22-3 and 22-4 respectively.
Connect to. Dispensing nozzles 22-1 and 22-2
are made to have different lengths so as to penetrate into the centrifuged blood cells and serum sample contained in the sample container 18-1-1, respectively. Arm 2
2-3, 22-4, arm 22-3 is arm 22
The dispensing nozzles 22-1 and 22-2 are coaxially arranged so as to be able to slide inside the dispensing nozzles 22-1 and 22-2. A protrusion 308 is provided at the rear end of arm 22-3, and this protrusion 308 projects from an opening 310 formed at the rear end of arm 22-4. A spring 312 is provided between the rear end and the arm 22-3 to urge the arm 22-3 to the left in the figure, so that the protrusion 308 moves into the opening 31.
0 is brought into contact with the tip end side 310-1. Further, the arm 22-4 is held movably in the horizontal direction via a slide bearing 316 provided on a vertical transfer shaft 314, and a rack 318 is formed on the outer wall of the arm 22-4.
A pinion 322 fixed to the output shaft of the arm 22-4 is engaged with the output shaft of the arm 22-4 by driving the motor 320.
to move horizontally. Furthermore, a light shielding plate 324 is provided protruding from the rear end of the arm 22-4, and the dispensing nozzles 22-1 and 22-2 are arranged in the horizontal movement path of the light shielding plate 324 at the cleaning position, the discharge position, and the suction position. arranging photoelectric switches 326, 328 and 330 for positioning;
These photoelectric switches detect the intrusion of the light shielding plate 324 and control the drive of the motor 320. Also,
A stopper 332 is provided in the movement path of the protrusion 308 provided at the rear end of the arm 22-3 so as to come into contact with the protrusion 308, and this stopper 332 positions the suction position of the dispensing nozzle 22-1.

The vertical transfer shaft 314 is a slide bearing 334
It is held movable vertically through the This vertical transfer shaft 314 has a rack 3 extending in the vertical direction.
36, and a motor 338 is connected to this rack 336.
A pinion 340 fixed to the output shaft of is engaged,
The vertical transfer shaft 314 is driven by the motor 338, and therefore the above-mentioned dispensing nozzles 22-1, 22-2 and arm 22- held on the vertical transfer shaft 314
3, 22-4, etc. are configured to move together in the vertical direction. This vertical transfer shaft 314 has a protrusion 3
42 is provided, and the micro-switches 344 and 346 are actuated by the protrusion 342 to move the vertical transfer shaft 314 and hence the dispensing nozzles 22-1, 22-2.
The upper limit position and lower limit position are respectively regulated.
Further, a micro switch 347 is further provided between the micro switches 344 and 346, and the sample container 18-1-
When aspirating only the serum sample from the dispensing nozzle 22-1, this micro switch 347
Position the tip of the tube into the serum sample.

Dispensing nozzle 22-1 is connected to cleaning liquid container 358-1 containing cleaning liquid 356-1 via syringe 348-1, valve 350-1, syringe 352-1, and valve 354-1. Syringe 34
The pistons 8-1 and 352-1 are configured to be driven by pulse motors 360-1 and 362-1, respectively, and the top dead center and bottom dead center of these pistons are set by micro switches 364-1 and 364-1, respectively.
66-1; regulated by 368-1, 370-1.
In addition, tube pinching 372 is provided at any position in the flow path from the dispensing nozzle 22-1 to the valve 350-1 to change the flow path volume and form an air layer at the tip of the dispensing nozzle 22-1. -1 is provided.

The dispensing nozzle 22-2 is also the dispensing nozzle 22 mentioned above.
-1 as well as syringe 348-2 and valve 350
-2, syringe 352-2 and valve 354-
The pistons of each syringe 348-2 and 352-2 are driven by pulse motors 360-2 and 362-2, respectively. point and bottom dead center using micro switches 364-2, 366-2, 368-, respectively.
2,370-2. Also,
In the flow path from the dispensing nozzle 22-2 to the valve 350-2, the dispensing nozzle 22 is connected by changing the flow path volume.
A tube pinching 372-2 is provided at the tip of the tube 372-2 to form an air layer. In addition, valve 3
The flow path after 54-2 and the flow path after valve 354-1 of dispensing nozzle 22-1 can also be made common.

On the other hand, the diluent dispenser 300 has two sample containers 18-1-2 and 18-1 located at the sample dilution dispensing position.
-2 fixed dispensing nozzles 30 facing each side
1 and 302. These dispensing nozzles 30
1,302 are dilution rate selectors 374-
1,374-2, valve 376-1,376-
2. Diluent 382- via syringe 378-1, 378-2 and valve 380-1, 380-2
Diluent container 384- containing 1,382-2
Connect to 1,384-2. Valves 376-1 and 380-1, as well as valves 376-2 and 380-2, are configured so that when one is open, the other is closed. Syringes 378-1 and 378
-2 pistons are driven by motors 386-1 and 386-2, respectively, and microswitch 388
-1 and 388-2, a constant amount of diluent 382-1 and 382-2 is always supplied to the syringe 3.
78-1 and 378-2. In addition, in the above description, two diluent containers 384-1 and 384-2 are used, but one diluent container may be used in common.

Further, a cleaning liquid container 394 containing a cleaning liquid 392 is connected via a pump 390 to the cleaning tank 22-5 for cleaning the dispensing nozzles 22-1 and 22-2, so that the cleaning liquid 392 is supplied into the cleaning tank. At the same time, the old cleaning liquid in the tank is configured to be discharged through a valve 396. Further, a nozzle (not shown) is provided in the upper part of the cleaning tank 22-5, and this nozzle is connected to a compressor 398 via a valve 397 to inject air.
1 and 22-2 from the cleaning tank 22-5, the cleaning liquid adhering to their outer walls is blown away.

The operation of the sample diluter shown in FIG. 4 will be explained below.

FIG. 4 shows a state in which the dispensing nozzles 22-1 and 22-2 are in a cleaning position located above the cleaning tank 22-5, and in this state, the light shielding plate 324 is at the position of the photoelectric switch 326. Also, the dispensing nozzle 22-
1 and 22-2 are filled with cleaning liquids 356-1 and 356-2,
An air layer is formed at the tips of the dispensing nozzles 22-1 and 22-2. When the motor 320 is driven to horizontally move the arms 22-3 and 22-4 to the left in the figure, the protrusion 308 provided on the arm 22-3 first comes into contact with the stopper 332, and the dispensing nozzle 22-1 releases the sample. After being positioned at the suction position, only the arm 22-4 is moved leftward while compressing the spring 312. When the light shielding plate 324 blocks the optical path of the photoelectric switch 330, the motor 320 is deenergized by a signal from the photoelectric switch 330, and the dispensing nozzle 22-2 is also positioned at the sample suction position. Next, drive the motor 338,
The vertical transfer shaft 314 is lowered until the micro switch 346 is activated, and the dispensing nozzles 22-1 and 22-1
2-2 into the sample container 18-1-1, and their tips are located in the blood cell and serum samples, respectively. In this state, valve 350-1,350-
2, close the pulse motor 360-1, 360-
2, close the pulse motor 360-1, 360-
2 to aspirate predetermined amounts of blood cells and serum samples, respectively.

After aspirating the sample, the motor 338 is driven in the opposite direction to the above direction until the micro switch 344 is activated.
1 and 22-2, and then the motor 320 is similarly driven in the opposite direction to the above. motor 320
When the arm 22-3 is driven, the arm 22-3 is attached to the tip side 310-1 of the opening 310 formed in the arm 22-4.
until the protrusion 308 provided on the arm 22 comes into contact.
Only -4 is transferred to the right, and after contact, arm 2
2-3 and 22-4 are moved together to the right.
When the light shielding plate 324 activates the photoelectric switch 328, the motor 320 is deenergized and the dispensing nozzle 22
-1 and 22-2 are respectively sample containers 18-1-
2, positioned at the sample discharge position on 18-1-3. Thereafter, the motor 338 is driven again to transfer the dispensing nozzles 22-1, 22- through the vertical transfer shaft 314.
2, and insert the dispensing nozzle 22-1 into the sample container 1.
8-1-2, insert the dispensing nozzle 22-2 into the sample container 1.
After entering into 8-1-3, pulse motor 3
60-1 and 360-2 are driven to start discharging the aspirated blood cells and serum sample.

On the other hand, while the dispensing nozzles 22-1 and 22-2 go from the washing position to the sample suction position to the sample discharge position as described above, the diluent dispenser 300 closes the valves 376-1 and 376-2. , valve 380-
1,380-2 is open, motor 386-1,3
86-2 for a predetermined period of time to discharge the syringe 378-
A predetermined amount of diluent 382-1 in 1,378-2,
Absorbs 382-2. Dispensing nozzle 22-1, 2
2-2, the valves 376-1 and 376-2 are opened, the valves 380-1 and 380-2 are closed, and the motor 38
6-1 and 386-2 are driven again to start discharging prescribed amounts of diluted liquid from the amounts of diluted liquid sucked through dilution rate selectors 374-1 and 374-2 and fixed dispensing nozzles 301 and 302, respectively. . In the present invention, among these dilution liquid discharge periods, the dilution liquid discharge period by the co-dispensing nozzle 301 is as follows:
It is set to be longer than the subsequent period for discharging the blood cell sample by the dispensing nozzle 22-1, and to continue even after the period for discharging the blood cell sample ends. In this way, if the blood cell sample discharge period is set within the dilution liquid discharge period, the blood cell sample will be delivered to the sample container 18-1-2.
Since the diluent and the blood cell sample can be stirred at the same time without adhering to the inner wall of the blood cell, a highly accurate blood cell suspension can be prepared in a short time. Note that this also applies to serum samples.

After discharging blood cells and serum samples, motor 3
38 to raise the dispensing nozzles 22-1, 22-2 via the vertical transfer shaft 314, and then drive the motor 320 to move the arms 22-3, 22-
4 to the right, dispensing nozzles 22-1, 22
-2 is positioned above the cleaning tank 22-5. After that, the motor 338 is driven again to move the vertical transfer shaft 31
The arms 22-3 and 22-4 are lowered through the
-Infiltrate into 5. In this state, the cleaning tank 22-5
At the same time, the valve 396 connected to the valve 350-1, 350-2 is closed and the pump 390 is operated to supply the cleaning liquid 392 into the cleaning tank.
is opened, valves 354-1 and 354-2 are closed,
The cleaning liquid 356-1, 356-2 is dispensed into the nozzle 22 by driving the pulse motors 362-1, 362-2.
-1, 22-2 and is discharged into the cleaning tank. Note that the cleaning liquids 356-1, 356-2 are supplied to the pulse motors 362-1, 362-2 by closing the valves 350-1, 350-2 and opening the valves 354-1, 354-2 during the series of steps described above. is driven to aspirate into the syringes 352-1 and 352-2. In this way, the dispensing nozzles 22-1, 22-2
By discharging the cleaning liquid from the nozzles 22-1 and 22-2, the inner and outer walls of the dispensing nozzles 22-1 and 22-2 can be reliably cleaned, and contamination between successive samples can be effectively prevented. Dispensing nozzle 22-
After cleaning 1 and 22-2, the cleaning liquid in the cleaning tank is discharged by opening the valve 396, and the motor 33
8 to raise the dispensing nozzles 22-1 and 22-2 via the vertical transfer shaft 314. During this rising period, valve 397 is opened and compressor 398
The airflow from the
The cleaning liquid adhering to the outer walls of the dispensing nozzles 22-1, 22-2 is blown away, and the tube pinching 372-1, 372-2 is operated for a short time to remove the dispensing nozzles 22-1, 22-2. Form an air space at the tip and complete the preparation for the next diluted aliquot of blood cells and serum samples.

In the sample diluter described above, the syringe 348-1 for dispensing blood cells and serum samples;
348-2 and pulse motor 360-1, respectively.
360-2, the amount of blood cells and serum samples to be dispensed can be easily changed by simply changing the number of driving pulses.

The diluent dispenser 300 also includes a dilution rate selector 3.
74-1 and 374-2, the discharge amount of the diluent can be arbitrarily changed, so that a blood cell suspension and diluted serum with a desired concentration can be created.

[Brief explanation of drawings]

FIG. 1 is a diagram showing the sequential steps of an analytical method based on immunological agglutination reaction suitable for applying the dispensing method according to the present invention, and FIG. FIG. 3 is a plan view schematically showing the configuration of an embodiment of the analyzer, FIG. 3 is a flowchart showing the operation thereof, and FIG. 4 is a diagram showing the configuration of an example of the sample diluting device. 24-1 to 24-3...Reaction line passage, 22
... Sample dispensing device, 25 ... Reaction container, 26 ...
Microplate, 27...cassette, 33...
Rack storage cassette, 35, 36...Microplate transport device, 37...Pattern detection device, 3
8... Judgment circuit, 39... Display device, 40... Microplate storage device, 300... Diluted liquid dispenser.

Claims (1)

[Claims] 1. A first dispenser that aspirates and dispenses a predetermined amount of a particle sample when diluting and dispensing a particle sample such as blood cell particles, latex particles, carbon particles, glass particles, etc.;
using a second dispenser that dispenses a predetermined amount of the diluent, and after the second dispenser starts dispensing the diluent, the first dispenser discharges the particle sample;
A method for dispensing a particle sample, characterized in that dispensing of a diluent is continued until after dispensing of the particle sample is completed.