JPH063359A - Method and apparatus for measuring antigen or antibody in sample - Google Patents

Abstract

(57) [Summary] (Modified) [Configuration] The light source 10, the sample mounting portion 12, and the light receiving portion 13 are provided, and the substantially central portion of the sample housed in the liquid reservoir portion of the inspection plate mounted on the sample mounting portion 12 A light source 10 irradiates light from the light source 10, and a light receiving unit 13 senses the transmitted light to measure the light transmittance of the sample, a driving unit 21 provided with a driving source 15, a measuring unit 20, and a driving unit. Drive source 1 by connecting with the part 21
5 and a driving force transmission means 22 for rotating the measuring unit 20 in the horizontal direction. [Effect] There is no need for visual judgment, and there is no variation or error in judgment. Using this measuring device, a quick and accurate measurement is possible.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a method for measuring an antigen or an antibody in a sample by an immunological agglutination reaction, and a measuring device used in the measuring method.

[0002]

2. Description of the Related Art Since immunological measuring methods have high specificity and sensitivity, they have become extremely useful clinically in the medical field in recent years. In particular, the method utilizing an immunological agglutination reaction is used for various diagnostic purposes such as fecal occult blood and rheumatoid factor detection, pregnancy diagnosis and the like, because the test can be carried out easily.

Conventionally, in order to carry out these inspections,
A suspension (reagent) of latex particles sensitized with an antibody or an antigen corresponding to an antigen or an antibody that is a test substance, and a test liquid are in the liquid reservoir of the test plate (a recess provided in the test plate. Alternatively, a method of observing the presence or absence of agglutination with the naked eye by mixing in a test plate within a portion surrounded by a hydrophobic ring) and rocking the reaction solution by hand or a rocking device for several minutes. ing. Usually, the one in which aggregation is observed is defined as positive, and the one in which aggregation is not observed is defined as negative.

Since these methods make a final judgment by observing the presence or absence of agglutination with the naked eye, individual differences among the judges are likely to occur. Especially in the vicinity of the concentration (so-called judgment pending region) where delicate positive / negative determination is required, it is necessary to judge whether or not weak agglomeration occurs, so the light amount (brightness) during observation, There is a possibility that the judgment may be mistaken depending on the reaction time, the degree of rocking, etc.

As a method for solving the problems of variations and erroneous determinations caused by the naked eye determination, various methods have been proposed in which measurement is performed using an apparatus including optical means. For example,
Japanese Patent Publication No. 58-11575 proposes a method in which an agglutination reaction is carried out in a cell for measuring absorbance, and an increase in absorbance of a reaction solution due to the agglutination reaction is measured.

However, since this method measures the absorbance at the same time as the reaction progresses, it is necessary to make the reaction liquid uniform by stirring or shaking, and it is easily affected by the insoluble matter derived from the sample. Further, in the measurement of a sample containing a high concentration of an antigen or an antibody, the absorbance may not correctly correspond to the amount of the antibody or the antigen, and the measurable concentration range is limited. Therefore, when measuring a high-concentration sample, it is necessary to pre-dilute the sample, which has a drawback that the operation becomes complicated and an error due to dilution is likely to occur.

[0007]

An object of the present invention is to provide a new measuring method capable of quantitatively measuring an antigen or an antibody in a sample without making a visual judgment. A further object of the present invention is to provide a measuring device that is preferably used in the above measuring method.

[0008]

Means for Solving the Problems The inventors of the present invention have realized that when the agglutination reaction is carried out in the liquid reservoir of the inspection plate and the reaction liquid in the liquid reservoir is horizontally swirled, Focusing on the fact that the light transmittance of the reaction solution in the center once decreases and then increases as shown in FIG. 6, this phenomenon can be used to measure the amount of the antigen or antibody in the sample. The inventors have found that it is possible and completed the present invention.

The method for measuring an antigen or an antibody in a sample of the present invention is a method in which a sample containing an antigen or an antibody is allowed to undergo an agglutination reaction with an antibody corresponding to the antigen or the antibody or carrier particles sensitized with the antigen. A method for measuring an antigen or an antibody, in which the agglutination reaction is performed in the liquid reservoir of the test plate, and the reaction liquid in the liquid reservoir is horizontally swirled while the reaction liquid in the liquid reservoir is substantially transmitted. It is characterized in that the amount of the antigen or antibody is measured by continuously measuring the rate after a lapse of a predetermined time T _{1 and} obtaining the characteristic value of the time change of the light transmittance.

Further, an apparatus for measuring an antigen or an antibody in a sample according to the present invention, which is preferably used in the above method, comprises a light source, a sample mounting portion, and a light receiving portion, and comprises an inspection plate mounted on the sample mounting portion. A measuring unit for irradiating light from the light source to the substantially central portion of the sample contained in the liquid reservoir, and measuring the light transmittance of the sample by the light receiving unit sensing the transmitted light, and a driving unit provided with a driving source. And a driving force transmitting means for connecting the measuring unit and the driving unit and transmitting the driving force of the driving source to rotate the measuring unit in the horizontal direction, preferably the measuring unit is A lid provided with a light source, an inspection plate mounting base arranged so that the light emitted from the light source passes through substantially the center of the liquid reservoir, and the center of the liquid reservoir provided under the base. A slit that allows the light transmitted through the section to pass downward,
The measuring unit body includes a light receiving unit that is disposed immediately below the slit and that senses light that has passed through the slit. More preferably, the light receiving unit is connected to an arithmetic device, and the light measured by the measuring unit is used. It is configured to calculate the amount of the antigen or antibody in the sample from the transmittance.

The present invention will be described in more detail below. FIG. 6 shows the change in light transmittance associated with the agglutination reaction. A suspension (reagent) of carrier particles obtained by sensitizing a sample containing an antigen or an antibody with the antibody corresponding to the antigen or the antibody or the antigen.
When the solution is dropped, the light transmittance is small because the reagent is initially near the center of the liquid reservoir, but by swirling horizontally, the light transmittance is increased until time T ₁ because it is dispersed in the entire liquid reservoir. After the lapse of T ₁ , the particles aggregate with the progress of the aggregation reaction,
Since the apparent particle size increases and the scattered light increases, the light transmittance decreases. On the other hand, the agglomerated particles are horizontally swirled to gradually move to the periphery of the liquid reservoir due to the centrifugal force, so that the central portion becomes transparent and the light transmittance increases. Since the latter phenomenon occurs later than the former phenomenon, the light transmittance decreases and then increases.

As shown in FIG. 7, when the amount of the antigen or antibody in the reaction solution is large, the light transmittance curve approximated to the quadratic curve becomes sharp after the passage of T ₁ , and conversely the amount of the antigen or antibody is increased. When is small, the curve of light transmittance becomes gentle. Therefore, the measurement method of the present invention is as follows: (1) The light transmittance of the reaction solution is continuously measured after the elapse of a predetermined time T ₁ , and the light transmittance is quadratic y = a ₀ + a ₁ x + a
₂ x ² [wherein x represents time and y represents light transmittance], and the amount of the antigen or antibody is measured by the value of the quadratic coefficient a ₂ .

Further, as shown in FIG. 7, when the amount of the antigen or antibody in the reaction solution is large, the time until the light transmittance becomes the minimum in the curve of light transmittance after the passage of T ₁ is short, and When the amount of antibody is small, the time required to reach the minimum is long. Therefore, another measuring method of the present invention is as follows: (2) The light transmittance of the reaction solution is continuously measured after a lapse of a predetermined time T ₁ , and the amount of the antigen or antibody is measured by the time when the light transmittance is minimized. To do. The time at which the light transmittance becomes the minimum may be determined based on an actual measurement value, or the light transmittance measured continuously as in (1) above may be calculated by a quadratic equation y =
a ₀ + a ₁ x + a ₂ x ² [where x is time and y is light transmittance] and is obtained by differentiating this quadratic expression.
= −a ₁ / 2a ₂ may be set as the time when y becomes the minimum.

Furthermore, the measuring method of the present invention is based on the above-mentioned light transmission.
Paying attention to the characteristic of the curve of the reflectance, the light transmittance in FIG.
From the time when it becomes small to the predetermined time T₂Degree of increase in light transmittance of
(L ₂-L₁) And FIG. 6 show the minimum light transmittance.
Predetermined time T after₃Value of light transmittance at
(L₃) To determine the amount of antigen or antibody. Sanawa
Then, another measuring method of the present invention is: (3) The light transmittance of the reaction solution is set to a predetermined time T₁Continuous after passage
For a predetermined time from the time when the light transmittance becomes the minimum.
T₂Antigen by measuring the increase in light transmittance of
The amount of antibody is measured. Furthermore, another measuring method of the present invention is: (4) The light transmittance of the reaction solution is measured for a predetermined time T₁After the passage
Continuously measured, and a predetermined value after the time when the light transmittance becomes minimum.
Time T₃Depending on the value of light transmittance in
It is a measure of quantity. Light transmission also in these measurement methods (3) and (4)
The time when the rate becomes the minimum may be determined based on the actual measurement value.
X = −a obtained by differentiating the above-mentioned quadratic equation₁/
2a₂May be the time when y is minimized.

Here, the light transmittance is the ratio of the intensity of the transmitted light to the incident light, and the light transmittance T (%) is expressed by the following equation. T = (I / I ₀ ) × 100 (where I is the intensity of transmitted light and I ₀ is the intensity of incident light). In this specification, “continuously measuring” means continuous. It is meant to include not only the measurement while actively monitoring, but also the measurement at fixed time intervals.

In all of these methods, the light transmittance is measured in a standard solution dilution series of known concentration to prepare a calibration curve, and the light transmittance of an unknown sample is measured under the same conditions as the standard dilution series measurement. Can be measured and compared with a calibration curve to quantify the amount (or concentration) of the antigen or antibody in the unknown sample.

The measuring methods of the present invention are all based on the light transmittance, but the transmittance (T) and the absorbance (A) have a relationship of A = -log (1 / T). ,
Also by measuring the absorbance, the amount of the antigen or antibody in the sample can be similarly measured by using the methods (1) to (4) of the present invention. Therefore, the case where the absorbance is used instead of the light transmittance is also included in the scope of the claims of the present invention.

According to the measuring method of the present invention, the antigen or the antibody in the sample can be quantitatively measured without the naked eye judgment. In addition, since the change in light transmittance at the approximate center of the liquid reservoir is measured, even when insoluble matter is mixed in the sample, the insoluble matter moves to the peripheral portion of the liquid reservoir due to centrifugal force during swirling and The measurement of the transmittance has the advantage that it is not disturbed.

The antibody or antigen to which the method of the present invention can be applied is not particularly limited, and the present invention includes, for example, measurement of serum proteins such as fecal occult blood reaction and tumor markers, measurement of hormones such as pregnancy diagnosis, and measurement of viruses and autoantibodies. Used for measurement etc.

The carrier particles used in the present invention are not particularly limited, and any carrier particles can be used as long as they are biologically inactive, insoluble in water and function as a carrier for an antigen or an antibody. However, the average particle size is 0.05 to 10 μm, preferably 0.1 to 1 because the light scattering of the carrier particles themselves in the light transmittance measurement is small.
Latex particles of 0 μm are preferably used. The type of latex is not particularly limited, but a latex of styrene type, olefin type, vinyl type, acrylic acid ester type, methacrylic acid ester type, diene type, etc. and a modifying monomer such as acrylic acid or acrylonitrile are reacted with these latexes. Examples of the latex thus obtained include polystyrene latex and the like.

As the aqueous solvent in which the carrier particles are dispersed, a pH buffer solution (eg, phosphate buffer solution, borate buffer solution, etc.) is preferably used, and sodium chloride, a preservative, bovine serum albumin are appropriately used. You may add proteins, such as.

The inspection plate used in the present invention is made of a material that does not affect the measurement of light transmittance. For example, a transparent test plate provided with a liquid reservoir having a hydrophobic ring on the peripheral portion, a glass or plastic plate provided with a shallow U-shaped liquid reservoir, etc. A plastic transparent inspection plate provided with a mold liquid reservoir is preferably used because it has little influence on the measurement of light transmittance.

The shape of the liquid reservoir of the inspection plate is
The horizontal cross-sectional shape is preferably a circle or an ellipse. Further, it is preferable that the side wall of the liquid reservoir has a smooth curved surface. This is to make the swirling smoothly when swirling the reaction liquid in the liquid reservoir in the method of the present invention. In addition, since the reaction liquid swirling swirls along the side wall of the liquid reservoir, the shape of the transparent or translucent portion near the center of the liquid reservoir due to this swirling is the horizontal cross-sectional shape of the liquid reservoir. Shows a shape similar to. In order to measure the light transmittance at the approximate center of the liquid reservoir, it is desirable that the transparent or semitransparent portion has a perfect circle or a shape close to a perfect circle. Therefore, in the present invention, the inspection plate in which the horizontal cross-sectional shape of the liquid reservoir is a perfect circle or a circle close thereto is most preferably used.

In the present invention, the light transmittance is measured by using an aqueous solvent used for the agglutination reaction and a light beam having a wavelength which is less absorbed by carrier particles, and is usually 400 to 2400.
nm, preferably using light having a wavelength near 600 to 1300 nm. The region for measuring the light transmittance is preferably about 1 to 25%, preferably about 1 to 10%, with the liquid surface area of the reaction liquid being 100% near the center of the liquid reservoir. When a plate having a circular cross section with a diameter of 20 mm is used, it is preferable to measure the light transmittance in a concentric area having a diameter of about 2 to 7 mm.

The rotation speed and the radius of gyration of the reaction solution are
The optimum number of rotations and the radius of rotation vary depending on the shape of the liquid reservoir, the amount of the reaction liquid, the specific gravity of the latex particles, etc., and therefore the number of rotations is appropriately set, but normally the number of rotations is 20 to 200r.
pm, preferably 80-140 rpm, with a turning radius of 5
It may be selected from the range of 50 mm, preferably 10 to 30 mm.

As a light source for irradiating the above-mentioned light rays, a tungsten lamp, a halogen lamp, a silicon carbide rod, a nichrome wire or the like combined with a filter for setting the above wavelength, or a light emission having a specific wavelength characteristic A diode (LED) or the like is used, and an LED is particularly preferable because it can emit sharp linear light without scattered light.

In the present invention, a sample containing an antigen or an antibody is mixed with the antibody corresponding to the antigen or the antibody or carrier particles sensitized with the antigen in the liquid reservoir of the test plate to cause an agglutination reaction. At the same time, since the reaction liquid in the liquid reservoir is swung in the horizontal plane, it is necessary to swivel the inspection plate in the horizontal direction at a constant cycle. For this purpose, an oscillating device that gives a rotational motion to the reaction liquid in the liquid reservoir is required. However, if the tilting motion or the translational motion is added to the rotary motion, the formation of a transparent or translucent portion of the reaction liquid generated in the approximate center of the liquid reservoir is hindered, which is not preferable. As such an oscillating device, for example, a shaft is fixed perpendicularly to the axial direction of a rotating power take-off shaft, and the shaft is provided at an arbitrary position different from the power take-off shaft center in parallel with the axial direction of the power take-off shaft. It is possible to use a device in which the shaft is fixed, and the shaft is rotatably supported on the bearing portion of the rotary table for mounting the inspection plate so that the table is held horizontally and swung in the horizontal direction.

FIG. 1 is a plan view showing the movement of the inspection plate mounted on a rotary table (not shown) of the swinging device and being horizontally swung. In the figure, 1 is an inspection plate having 10 liquid reservoirs 2. A sample liquid for measurement (not shown) is contained in the liquid reservoir 2. For example, with the position indicated by I in the figure as the start position, the inspection plate is horizontally rotated in the order of I → II → III → IV → I, so that the sample liquid becomes horizontal in the liquid reservoir 2. It can be turned.

FIG. 2 shows a part of the AA 'cross section at I of the inspection plate 1 which is rotating as described above. The sample liquid 3 which is swung horizontally in the liquid reservoir 2.
Is in the state shown in FIG. 2 at the I position. As the sample liquid continues to swirl in this manner, the agglomerated particles generated by the agglutination reaction move to the peripheral portion of the liquid reservoir 2 due to the centrifugal force, and as a result, the agglomeration of the agglomerated particles occurs in the approximate center of the liquid reservoir 2. It is considered that transparent or semi-transparent parts are formed depending on the degree.

As an apparatus that can be suitably used in the above method, the inspection plate can be horizontally swung in a constant cycle, and a rocking portion for imparting a rotational motion to the sample liquid in the liquid reservoir, and a liquid The sample solution in the reservoir is irradiated with light to measure the light transmittance thereof. The present invention provides an apparatus for measuring an antigen or an antibody in a sample, which has the above two functions. Hereinafter, this measuring device will be described in detail with reference to the drawings showing one embodiment.

In FIG. 3, (a) is a perspective view of a measuring apparatus showing an embodiment of the present invention, and (b) is a partially cutaway front view thereof. In FIG. 3 (a), S is a light transmittance measuring device, which includes a lid 11 having a light source 10, an inspection plate mounting portion 12 and a light receiving portion 13, and measures the light transmittance of the sample liquid. The measuring unit 20 including the main body 14, the driving unit 21 in which the driving source 15 is disposed, the measuring unit 20 and the driving unit 21 are connected to each other, and the driving force of the driving source 15 is transmitted to the measuring unit. Driving force transmission means 2 for rotating 20 in the horizontal direction
2 and.

As shown in FIG. 4, the measuring unit 20 has a lid 11 for blocking light from the outside, and the lid 11.
A light source 10 attached to the inspection plate 1, an inspection plate attachment base 16 arranged so that the irradiation light from the light source 10 passes through substantially the center of the liquid reservoir 2 of the inspection plate 1, and a lower portion of the base 16. A slit 16a which is provided at the center of the liquid reservoir 2 and guides the light transmitted through substantially the center of the liquid reservoir 2 downward,
The measuring unit main body 14 includes a light receiving unit 13 which is disposed immediately below a and senses the transmitted light that has passed through the slit 16a.

The above-mentioned light source is used as the light source 1, and an LED is particularly preferably used. As shown by phantom lines in FIG. 4, slits 17 are provided above the pedestal 16.
If the shielding plate 17 having a formed thereon is installed and the irradiation light from the light source 1 is guided, the liquid reservoir 2 of the inspection plate 1
It is preferable because the light can be accurately radiated to the approximate center.

As the drive source 15, the power take-off shaft 18 is used.
Any means can be used as long as it can give a rotational motion to the body and a force for rotating the measuring unit 20, and an electric motor or the like is generally used.

As the driving force transmission means 22, a shaft 19 is fixed to a power takeoff shaft 18 directly connected to the drive source 15 perpendicularly to the axial direction thereof, and the shaft center of the power takeoff shaft on the shaft 19 is fixed. One end of the connecting shaft 19a is fixed in an eccentric arbitrary position in parallel with the axial direction of the power take-off shaft 18, and the other end is a bearing portion 19 provided on the bottom of the measuring unit main body.
The measuring unit 20 is supported by being supported by b.

Next, the operation of this apparatus will be described with reference to FIG.
In the above, the power take-off shaft 18 is rotated by the operation of the drive source 15 such as a motor, and the shaft 19 is rotated with this rotation.
Rotates. This shaft 19 has an eccentric connecting shaft 1
Since 9a is fixed, the connecting shaft 19a makes a circular motion about the shaft center of the power take-off shaft 18. This connecting shaft 19
Since a is pivotally supported on the bottom of the measuring unit body 14, this circular movement causes the measuring unit body 14 to rotate in the horizontal direction.
Therefore, the sample liquid contained in the liquid reservoir 2 of the inspection plate 1 attached to the pedestal 16 of the measuring unit body 14 shown in FIG. 4 swirls along the side wall of the liquid reservoir 2, Aggregates in the center move to the periphery.

On the other hand, light is emitted downward from the light source 10 at the same time as the turning operation of the measuring unit 20 or after a certain time. The irradiation light is transmitted through a substantially central portion of the sample liquid 3 which is attached to the pedestal 16 and swirls along the side wall of the liquid reservoir 2, and the transmitted light is detected by a light receiving portion 13 arranged below the transmitted light. The light transmittance of the sample liquid 3 is measured.

In this apparatus, the light transmittance is calculated as described above from the irradiation amount emitted from the light source 10 and the transmitted light amount sensed by the light receiving unit 13, and from this result, the time of the light transmittance is calculated. The antigen or antibody can be measured by determining the characteristic value of the change.

According to the above-mentioned measuring device, the inspection plate can be horizontally swung in a constant cycle, and the swinging device for giving a rotational motion to the reaction liquid in the liquid reservoir and the reaction liquid in the liquid reservoir are exposed to light. It becomes possible to precisely measure the light transmittance of the reaction solution by irradiating with.

As shown in the system diagram of FIG. 5, when the light receiving unit 13 is connected to the arithmetic unit 32, the amount of the antigen or antibody in the sample is automatically determined based on the value of the light transmittance. It is particularly preferable because the antibody or the antigen can be measured more accurately because the calculation is performed in an accurate manner. At this time, it is preferable to use the amplifier 30 and the A / D converter 31 together in the connection circuit.

[0041]

EXAMPLES Examples of the present invention will be shown below and will be described more specifically. Example 1 Measurement of human hemoglobin (Material) To 10 ml of 5% carboxylated polystyrene latex (average particle size 0.30 μm), 1 mg / ml of 1-ethyl-3- (3-diaminopropyl) carbodiimide 1
After adding 0 ml and reacting for 20 minutes, 0.01 mol /
It was washed twice with a liter borate buffer (pH 8.0) by centrifugation. To this latex, 7 ml of anti-human hemoglobin antibody (rabbit IgG, 5 mg / ml) was added, and the mixture was reacted for 5 hours.
After centrifugation and washing with a 1 / liter borate buffer (pH 8.0) three times, an anti-human hemoglobin antibody-sensitized latex reagent having a latex concentration of 1% was obtained. Diameter 2 for inspection plate
A transparent polystyrene plate provided with a U-shaped liquid reservoir having a 0 mm round opening was used.

(Apparatus) As the measuring apparatus, the light transmittance measuring apparatus shown in FIG. 3 is used, and L having a wavelength of 940 nm is used.
The ED was used as a light source and the silicon photodiode was used as a light receiving portion. The optical axis of the light source is installed vertically on the inspection plate, and it is concentric with the center of the liquid reservoir (4.8
(mmφ) area was set to irradiate with light.

(Measurement) Human hemoglobin was treated with 0.1% bovine serum albumin, 0.9% sodium chloride, 0.1%
It was dissolved in a 0.1 mol / liter borate buffer solution (pH 8.0) containing sodium azide to obtain a standard solution. The standard solution was appropriately diluted with the buffer solution to prepare a standard solution dilution series. Standard solution diluted into the test plate reservoir 1
After adding 25 μl of the above-prepared latex reagent to 00 μl, attach it to the pedestal of the measuring device, start the measuring device, and swivel the measuring part horizontally at a radius of gyration of 25 mm and a rotation speed of 110 rpm, and light from the LED. The light transmitted through the sample irradiated and injected into the liquid reservoir was detected by the silicon photodiode, and the light transmittance was measured at intervals of 5 to 6 seconds for 3 minutes. FIG. 7 shows the time change of light transmittance at each antigen concentration.

(1) Minimum light transmittance after 33 seconds
Quadratic formula y = a by the square method₀+ A₁x + a ₂x²[In the formula, x
Is the time, and y is the light transmittance.]₂
Find the value of. The results are shown in Table 1.

[0045]

[Table 1]

A calibration curve obtained by plotting the above data on the graph with the antigen concentration of the standard solution as the horizontal axis and the value of the secondary coefficient a ₂ as the vertical axis is shown in FIG. The antigen concentration and the value of the secondary coefficient a ₂ show a good correspondence.

(2) Minimum light transmittance after 33 seconds has elapsed
Quadratic formula y = a by the square method₀+ A₁x + a ₂x²[In the formula, x
Is the time, and y is the light transmittance.]
The time x at which the light transmittance y is minimized is calculated (that is,
Chi x = -a₁/ 2a₂Ask). The results are shown in Table 2.

[0048]

[Table 2]

A calibration curve obtained by plotting the above data on the graph with the antigen concentration of the standard solution as the horizontal axis and the light transmittance minimum time x as the vertical axis is shown in FIG. There is a good correspondence between antigen concentration and time x.

(3) The time for which the light transmittance is minimum for each density is determined based on the measured value, and the increase in the light transmittance for 22 seconds is obtained from that time. The results are shown in Table 3.

[0051]

[Table 3]

FIG. 10 shows a calibration curve obtained by plotting the above data on the graph with the antigen concentration of the standard solution as the horizontal axis and the increase (%) in light transmittance as the vertical axis. There is a good correspondence between the antigen concentration and the increase in light transmittance.

(4) For each density, the time when the light transmittance becomes the minimum is determined based on the measured value, and it is after that time.
Table 4 shows the values of the light transmittance at 25 seconds.

[0054]

[Table 4]

FIG. 11 shows a calibration curve obtained by plotting the above data on the graph with the antigen concentration of the standard solution as the horizontal axis and the light transmittance (%) at 125 seconds as the vertical axis. There is a good correspondence between the antigen concentration and the light transmittance value.

[0056]

Industrial Applicability According to the present invention, it is possible to quantitatively measure an antigen or an antibody in a sample without performing visual determination, and variations in determination results by an inspector as in the conventional visual determination method. The problem of erroneous determination in the determination pending region is solved, and highly reproducible and accurate measurement is possible. Unlike the conventional method, the method of the present invention measures the change in the light transmittance of the liquid reservoir substantially in the center while rotating the reaction solution. Since the insoluble matter derived from the sample moves to the peripheral portion of the liquid reservoir due to the centrifugal force during the swirling, there is an advantage that the measurement is not disturbed by these insoluble matter. It is preferably used for measurement of a sample such as a fecal occult blood reaction in which insoluble matter derived from a sample may be mixed.

There is also an advantage that the test plate and the reagent used in the conventional naked eye determination method can be used as they are. In the conventional method of measuring the increase in absorbance while stirring, a sample containing a high concentration of antigen or antibody may be measured at a concentration lower than the actual concentration. According to the measuring method of the present invention, accuracy is slightly lowered even in a high-concentration sample, but it is easy to determine whether or not an antigen or antibody at a certain concentration or more is contained (negative or positive determination). The sample can be measured without pre-dilution and the operation is simple.

Further, according to the measuring apparatus of the present invention, the sample can be irradiated with light while the inspection plate is horizontally swung at a constant cycle, so that the light transmittance of the sample solution can be continuously and precisely measured. it can. Further, when the light transmittance measuring device is connected to the arithmetic device, the arithmetic processing of the measurement result is automatically performed, so that the quick measurement can be performed. Therefore, it is not necessary to perform a conventional human operation or a visual determination, and the time can be shortened and a precise and accurate determination can be performed.

[Brief description of drawings]

FIG. 1 is a plan view showing the movement of a horizontally rotating inspection plate.

FIG. 2 A- in I of the inspection plate 1 during turning
It shows a part of the A ′ cross section.

FIG. 3A is a perspective view of a measuring apparatus showing an embodiment of the present invention, and FIG. 3B is a partially cutaway front view thereof.

FIG. 4 is a partial cross-sectional view showing a measuring unit of the above apparatus.

FIG. 5 is a system diagram of a measuring apparatus showing another embodiment of the present invention.

FIG. 6 is a view of horizontally swirling the reaction liquid in the liquid reservoir,
It is a graph which shows the time change of the light transmittance of the liquid storage part substantially center.

FIG. 7 is a graph showing changes in light transmittance with time of each antigen concentration in Example 1.

FIG. 8 is a graph showing a calibration curve obtained in Example 1 (1).

FIG. 9 is a graph showing a calibration curve obtained in Example 1 (2).

FIG. 10 is a graph showing a calibration curve obtained in Example 1 (3).

FIG. 11 is a graph showing a calibration curve obtained in Example 1 (4).

[Explanation of symbols]

 DESCRIPTION OF SYMBOLS 10 light source 12 inspection plate mounting part 13 light receiving part 15 driving source 20 measuring part 21 driving part 22 driving force transmitting device S light transmittance measuring device

 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Inventor Kino Miyamoto 4-chome, Imamiya 11-34, Minoh City, Osaka Prefecture

Claims (7)

[Claims] 1. A method for measuring an antigen or antibody in a sample by causing an agglutination reaction of a sample containing the antigen or antibody with an antibody corresponding to the antigen or antibody or carrier particles sensitized with the antigen, The agglutination reaction is performed in the liquid reservoir of the inspection plate, and while the reaction liquid in the liquid reservoir is horizontally swirled, the light transmittance of the reaction liquid in the approximate center of the liquid reservoir is continuously passed after a predetermined time T _{1 has} elapsed. The light transmittance is measured and the light transmittance is quadratic.
= A ₀ + a ₁ x + a ₂ x ² [wherein x represents time and y represents light transmittance], and the amount of the antigen or antibody is measured by the value of the quadratic coefficient a _2. A method for measuring an antigen or antibody in a sample. 2. A method for measuring an antigen or an antibody in a sample by causing an agglutination reaction of a sample containing the antigen or the antibody with an antibody corresponding to the antigen or the antibody or carrier particles sensitized with the antigen, The agglutination reaction is performed in the liquid reservoir of the inspection plate, and while the reaction liquid in the liquid reservoir is horizontally swirled, the light transmittance of the reaction liquid in the approximate center of the liquid reservoir is continuously passed after a predetermined time T _{1 has} elapsed. A method for measuring an antigen or an antibody in a sample, which comprises measuring and measuring the amount of the antigen or the antibody according to the time when the light transmittance becomes minimum. 3. A method of measuring an antigen or an antibody in a sample by causing an agglutination reaction of a sample containing the antigen or the antibody with an antibody corresponding to the antigen or the antibody or carrier particles sensitized with the antigen, The agglutination reaction is performed in the liquid reservoir of the inspection plate, and while the reaction liquid in the liquid reservoir is horizontally swirled, the light transmittance of the reaction liquid in the approximate center of the liquid reservoir is continuously passed after a predetermined time T _{1 has} elapsed. A method for measuring an antigen or an antibody in a sample, which comprises measuring the amount of the antigen or the antibody by measuring the increase in the light transmittance for a predetermined time T ₂ from the time when the light transmittance is minimum. . 4. A method for measuring an antigen or an antibody in a sample by causing an agglutination reaction of a sample containing the antigen or the antibody with an antibody corresponding to the antigen or the antibody or carrier particles sensitized with the antigen, the method comprising: The agglutination reaction is performed in the liquid reservoir of the inspection plate, and while the reaction liquid in the liquid reservoir is horizontally swirled, the light transmittance of the reaction liquid in the approximate center of the liquid reservoir is continuously passed after a predetermined time T _{1 has} elapsed. A method for measuring an antigen or antibody in a sample, which comprises measuring and measuring the amount of the antigen or antibody by the value of the light transmittance at a predetermined time T ₃ after the time when the light transmittance becomes the minimum. 5. A light source, a sample mounting section, and a light receiving section are provided, and light is emitted from the light source to approximately the center of the sample contained in the liquid reservoir section of the inspection plate mounted on the sample mounting section, and transmitted. The light receiving section senses the light to measure the light transmittance of the sample, the driving section in which the driving source is provided, and the measuring section and the driving section are connected to transmit the driving force from the driving source. Device for measuring an antigen or an antibody in a sample, which comprises a driving force transmitting means for horizontally rotating the measuring section. 6. A lid body having a light source, a pedestal for mounting an inspection plate arranged so that light emitted from the light source passes through substantially the center of the liquid reservoir, and a lower portion of the pedestal. And a measuring unit main body provided with a slit for passing the light passing through the substantially central portion of the liquid reservoir downward and a light receiving unit arranged immediately below the slit for sensing the light passing through the slit. The measuring device according to claim 5. 7. The measuring device according to claim 5, wherein the light receiving part is connected to a computing device and the amount of the antigen or antibody in the sample is calculated from the light transmittance measured by the measuring part.