JPH09274040A - Method and apparatus for automatic immunity analysis - Google Patents

Abstract

PROBLEM TO BE SOLVED: To enhance the reliability of a measured result by a method wherein, according to the compared result of areas in ranges which are designated arbitrarily on the basis of a reaction process absorbence, the existence of a reaction suppression phenomenon due to the excess of an antigen is judged. SOLUTION: A sampling disk 2 is turned, a sample container 1 into which a sample is put is moved to a sampling position, and a sample in a constant is put is sucked by a sampling mechanism 3 so as to be discharged into a reaction container 7. The reaction container 7 is moved to a reagent dispensing position in a next cycle so as to be stopped, and a reagent is added by a reagent disk 4a and a reagent pipetting mechanism 5a. Then, whenever a reaction disk 8 is turned, the reaction container 7 crosses an optical axis, and an absorbance is measured by a photometer 10, a wavelength selector 11 and the like. A CPU 15 compares areas in two ranges which are disignated arbitrarily on the basis of a reaction process absorbance (i.e., computes an area ratio), and the existence of a reaction suppression phenomenon due to the excess of an antigen is judged accordingly. The measured absorbance is contorted into a concentration so as to be output.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to an automatic immunoassay method and apparatus.

[0002]

2. Description of the Related Art In an immunoserologic test based on a binding reaction between an antigen and an antibody, when a component (for example, antigen) in a sample and a reagent (for example, antibody) are reacted with each other, an optimal amount ratio of the antigen and the antibody is required. The strongest aggregation occurs. However, if the concentration of the component in the sample is extremely high, the antigen becomes excessive, the reaction is suppressed, and the measured value is output at an apparently low value. For example, when the concentration is in the normal range, the absorbance increases with the reaction time as shown in FIG. 2A, but when a sample with an abnormally high concentration is measured, the reaction is suppressed as shown in FIG. 2B. After the absorbance increases only immediately after the addition of the reagent, the change in the absorbance with the reaction time hardly occurs. Since the concentration is obtained by multiplying the absorbance at the end of the reaction by a constant calibration coefficient, the measured value is output at a lower value than the sample in the normal value range, despite the abnormally high concentration.
To solve this problem, there has been attempted a method of observing the absorbance in the process of reacting a sample with a reagent over time to determine the reaction suppression phenomenon due to excess antigen. That is, as described above, the reaction rate when the reaction suppression phenomenon occurs tends to be fast immediately after adding the reagent to the sample and then slow. On the contrary, there is also a tendency that the speed becomes slower as the antigen becomes excessive. In either case, the tendency of the reaction rate to change is determined by the rate ratio of two ranges of the initial rate of the absorbance in the reaction process and the overall rate or the rate after a fixed time. This method has an advantage that the conventional measurement method can be used as it is without any special operation for the analyst. However, it is necessary to capture the initial velocity of the reaction when there is an excess of antigen, and it is necessary to shorten the measurement time of the absorbance as soon as possible after adding the reagent, and to shorten the reaction time immediately after adding the reagent or immediately after stirring. It is not possible to accurately grasp the presence or absence of the reaction suppression phenomenon due to variations in the photometric system such as fluctuations in light source due to stability and shortening of measurement time. Also, after reacting the components in the sample with the reagents for a certain period of time, the same components as the components in the sample are added as reagents,
There are antigen re-addition method that determines excess antigen based on difference in absorbance before and after addition, and method of measuring one sample with two types of reagents with different sensitivities. There are drawbacks such as delays.

[0003]

The present invention accurately captures the reaction suppression phenomenon while using the conventional measurement method as it is in order to solve the problems of the above-mentioned conventional examples, and is arbitrarily designated from the absorbance of the reaction process. By calculating the area ratio of the two ranges and making a determination, and by outputting the presence or absence of the reaction suppression phenomenon,
The purpose is to improve the reliability of the measurement result so that an apparently low value is not mistakenly reported. Another object of the present invention is to provide a method and apparatus capable of promptly determining the dilution ratio of a sample when a reaction inhibition phenomenon occurs and a dilution retest is performed.

[0004]

The above-mentioned object uses the following technique in an automatic analysis in which the state of a reaction solution when a sample and a reagent are mixed is optically measured and the absorbance in the reaction process is measured at multiple points. Can be achieved by That is, (1) Two photometric points that determine the range for determining the area and the allowable value for determining the reaction suppression phenomenon can be input.

(2) It has an area for storing the measured absorbance at several points, and as shown in FIG. 2, it inputs two points (l, m).
A line is used to connect between them, and a calculation function for calculating the ratio of the area S1 on the side of the reaction absorbance and the area S2 of the triangle, that is, the judgment value X of Equation 1 is provided.

[0006]

[Equation 1] X = S1 / S2 × 100 (%) (Equation 1) (3) A function for comparing the judgment value with the allowable value is provided, and when the allowable value is exceeded, the measured value, the judgment value and the alarm are given. It has a function to print the mark.

In the automatic analysis for measuring the absorbance of the reaction process at multiple points, from the measured absorbances of several points of the reaction process, the previously input photometric points l and m are connected by a line to calculate the next area ratio. It has an arithmetic function, and an example of this calculation will be described with reference to FIG. First, the absorbance at each photometric point in the Y-axis direction (reaction time) is A ₁ , A ₂ ... _Am, and the distances in the X-axis direction between the two points _Am and _Am-1 are equidistant. This is D
Then, the approximate area S of the whole is calculated by Equation 2.

[0008]

## EQU00002 ## S = .SIGMA. (Ai.times.D) (Equation 2) Here, the area of the triangular portion protruding from the reaction absorbance is subtracted from the area S by Equation 3 to obtain S '.

[0009]

Equation 3] S '= S-Σ {( A m -A m-1) × D / 2} ... ( Equation 3) Next, any two points of the reaction absorbance (l, m) during connected by a line The area S2 of the triangle formed by the three points, 1, l, m and n is calculated by the equation 4.

[0010]

[Equation 4] S2 = T _w × A _m / 2 (Equation 4) Therefore, the area S1 on the side of the reaction absorbance is

[0011]

## EQU00005 ## Since S1 = S'-S2 (Equation 5) is obtained, the ratio X of the area S1 on the reaction absorption side and the area S2 of the triangle can be obtained by Equation 6.

[0012]

[Equation 6] X = S1 / S2 × 100 (%) (Equation 6) This area ratio becomes larger than the area S1 in the reaction process of the normal reaction as shown in FIG. In the reaction in which the reaction suppression phenomenon occurs as shown in FIG. 5, the area S1 approaches the width of the area S2, and the area ratio increases. Therefore, by comparing the determined area ratio value with the previously entered allowable value and printing the area ratio value and the alarm mark together with the measured value when the allowable value is exceeded, the analyst can confirm that the measured value is normal. It is possible to know whether the measured value is a value in or the value is an apparently low value due to the reaction suppression phenomenon caused by excess antigen, and thus the reliability of the measurement result can be improved.

[0013]

BEST MODE FOR CARRYING OUT THE INVENTION An embodiment of an automatic immunoassay device using the present invention will be described. As shown in FIG. 6, the device is
A sample disk 2 on which a sample container 1 is installed, a sampling mechanism 3 for collecting a fixed amount of a sample, reagent disks 4a and 4b on which a reagent is installed, reagent pipetting mechanisms 5a and 5b for dispensing a reagent, and a sample and a reagent are mixed. Stirring mechanism 6a, 6b, reaction disk 8 holding a plurality of reaction vessels 7 also functioning as optical cells, and light intensity for measuring the absorbance of the reaction fluid by passing light emitted from a light source lamp 9 through the reaction fluid in the reaction vessel. Total 10, wavelength selector 11, logarithmic conversion amplifier 12, A
A / D converter 13, a reaction container cleaning mechanism 14 for cleaning the used reaction container, and a central processing unit (microcomputer) 15 for controlling the entire mechanical system and performing data processing such as concentration calculation. . There are another printer 16, a CRT 17, an operation panel 18, a sample dispensing mechanism drive circuit 19, a reagent dispensing mechanism drive circuit 20, etc., all of which are connected to the central processing unit 15 via an interface 21.

The operation principle of these configurations will be described below. When the start switch on the operation panel 18 is pressed, the reaction container cleaning mechanism 14 starts cleaning the reaction container, and the operation of the reaction disk 8 rotates counterclockwise by half a rotation + one container and temporarily stops. ,
The reaction container advances to the sample discharge position. Here, the sample disc 2 rotates and the sample container 1 containing the sample to be measured
Moves to the sampling position, and a certain amount of the sample is sucked and discharged into the reaction container by the operation of the sampling mechanism 3. The reaction container from which the sample has been discharged moves to the reagent dispensing position and stops in the next cycle, and the reagent disk 4a and the reagent pipetting mechanism 5a operate so that a certain amount of the first reagent solution in the reaction container containing the sample. Is added. When the reaction disk rotates one rotation + two containers after the first reagent solution is added, the stirring mechanism 6a operates to stir the sample and the reagent. Thereafter, the reaction disks rotate one after another, and each time the reaction disk crosses the optical axis, the absorbance is measured each time. Also, the reaction container is rotated 15 times from the sample dispensing position +15
The second reagent is added to the reaction container containing the sample and the first reagent by the operation of the reagent disc 4b and the reagent pipetting mechanism 5b when the container is rotated to the position where it is rotated, that is, the second reagent dispensing position. Then, stirring is performed by the stirring mechanism 6b. For example, when the reaction time is 10 minutes and the cycle is 6 seconds, a total of 50 times of photometry are performed from the discharge of the sample to the end of the photometry, and the reaction vessel after the photometry is the reaction vessel cleaning mechanism 1
The sample is washed according to No. 4 and is ready for the next measurement. The absorbance measured by 50 times of photometry is converted into the concentration by the central processing unit 15 by the analysis parameter input in advance, and the measured value is output from the printer 16.

In the present invention, the two photometric points (l, m) input by the analysis parameter together with the output of this measurement value are used.
Then, as shown in FIG. 1, 1 and m are connected by a line, and the central processing unit 15 calculates the ratio of the area S1 on the reaction absorption side and the area S2 of the triangle. When the obtained area ratio exceeds the allowable value set in advance by the analysis parameter, the alarm mark and the area ratio are output together with the measured value.

Next, FIG. 7 shows the relationship between the change in absorbance and the judgment value (area ratio) of the present invention when a sample with a high concentration of immunoglobulin IgA is measured from a low concentration to a high concentration. This figure shows the system in which the reaction rate slowly progresses during the reaction suppression phenomenon due to excess antigen. Here, since the measured value is output by multiplying the absorbance by a constant coefficient, the change in the absorbance is directly reflected in the measured value. In the normal reaction range (1000 to 9000 mg / dl) where the absorbance increases with increasing concentration of IgA, the judgment value shows a constant value, but the absorbance decreases with increasing concentration. In the concentration range where a low value is output because the reaction suppression phenomenon occurs even though the concentration should be output, the judgment value also tends to decrease proportionally.
Therefore, according to this example, by setting the numerical value of 60 as the allowable value in advance in the analysis parameter and printing the alarm mark when the judgment value is less than that, the analyst causes the reaction suppression phenomenon due to excess antigen. You can easily understand what is going on and prevent false reports. further,
Since the concentration and the judgment value are in a proportional relationship in the excess antigen region, the judgment value can be printed together with the alarm mark to estimate the concentration of the sample and promptly determine the dilution ratio of the sample at the time of retesting.

[0017]

EFFECTS OF THE INVENTION According to the present invention, the reaction suppression phenomenon due to excess of antigen (or antibody), which has been the biggest problem in immunoserologic measurement, can be accurately grasped. Moreover, since the concentration of the sample in the antigen (or antibody) excess region can be estimated, the dilution retest can be performed only once without performing multiple times at a plurality of dilution ratios, leading to a reduction in measurement time and a reduction in reagent cost.

[Brief description of drawings]

FIG. 1 is a characteristic diagram showing an area ratio (S1 / S2) which is a method of determining a reaction suppression phenomenon in the present invention.

FIG. 2 is a characteristic diagram showing a difference in absorbance in a reaction process of samples having different concentrations.

FIG. 3 is a characteristic diagram showing a method of calculating an area ratio.

FIG. 4 is an area of absorbance (S1,
The characteristic view which shows the ratio of S2).

FIG. 5 is a characteristic diagram showing the ratio of the areas (S1, S2) of the absorbance in the reaction process in which the reaction suppression phenomenon occurs.

FIG. 6 is an explanatory diagram showing an embodiment of the automatic immunoassay device according to the present invention.

FIG. 7 is a characteristic diagram showing the effect of the area ratio-based determination method of the present invention.

[Explanation of symbols]

DESCRIPTION OF SYMBOLS 1 ... Sample container, 2 ... Sample disk, 3 ... Sampling mechanism, 4a, 4b ... Reagent disk, 5a, 5b ... Reagent pipetting mechanism, 6a, 6b ... Stirring mechanism, 7 ... Reaction container, 8 ... Reaction disk, 9 ... Light source lamp, 10 ... Photometer, 11 ... Wavelength selector, 12 ... Logarithmic conversion amplifier, 13 ...
A / D converter, 14 ... Reaction container cleaning mechanism, 15 ... Central processing unit, 16 ... Printer, 17 ... CRT, 18 ... Operation panel, 19 ... Sample dispensing mechanism drive circuit, 20 ... Reagent dispensing mechanism drive circuit, 21 ... Interface.

 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Inventor Masami Hayashi 832 Nagakubo, Horiguchi, Hitachinaka City, Ibaraki Prefecture 2 Hitachi Measurement Engineering Co., Ltd. (72) Inventor, Seven characters, 832 Nagakubo, Horiguchi, Hitachinaka, Ibaraki 2 Hitachi Measurement Engineering Co., Ltd.

Claims (3)

[Claims] 1. An immunoturbidimetric reaction utilizing an antigen-antibody reaction or a latex agglutination method to react a target component in a sample with a reagent, the reaction solution is optically measured, and the absorbance in the reaction process is measured at multiple points. In the automatic immunoassay method, the presence or absence of a reaction suppression phenomenon due to excess antigen is determined according to the result of comparing the areas of two arbitrarily designated areas from the absorbance of the reaction process. 2. A measurement device for measuring the reaction process absorbance of a reaction solution formed by a sample and a reagent, and a measurement result when the area ratio of two ranges designated in terms of time exceeds a preset allowable range. An automatic immunoassay device characterized in that it is also provided with a control device for outputting an alarm mark. 3. The automatic immunoassay device according to claim 2, wherein when the area ratio exceeds a preset allowable range, the area ratio is output together with the measurement result.