Biological sample analyzing apparatus

In a biological sample analyzing apparatus and method, an assay sample is prepared by mixing a reagent with a biological sample which may contain assay material. Then, a first information relating to the assay material is collected from the assay sample, and when the first information satisfies predetermined condition, the assay material is analyzed based on the first information. However, when the first information does not satisfy the predetermined condition, a second information related to the assay material is collected from the assay sample, and the assay material is analyzed based on the second information.

This application claims priority under 35 U.S.C. §119 to Japanese Patent Application No. 2004-308701 filed Oct. 22, 2004, the entire content of which is hereby incorporated by reference.

FIELD OF THE INVENTION

The present invention relates to an apparatus and method for analyzing assay material that contained in a biological sample.

BACKGROUND

Methods for quantifying assay material contained in a biological sample include methods in which material in an assay sample is agglutinated using a substance to induce an antigen-antibody reaction with the assay material, and the concentration of the assay material is calculated based on the degree of agglutination. Examples of such methods include immunoturbidity, immunonephelometry, counting immunoassay (CIA) and the like.

A calibration curve, which represents the relationship between the concentration of the assay material and information reflecting the degree of agglutination of the material in the assay sample, is used when calculating the concentration of the assay material by these methods.FIG. 14shows an example of a calibration curve when the information reflecting the degree of agglutination of carrier particles is the absorbance in the immunoturbidity method using carrier particles. InFIG. 14, when the concentration of the assay material is in the low concentration range, the absorbance gradually increases in conjunction with the increase in the concentration of the assay material. However, above a certain concentration of the assay material (high concentration range), there is a reduction in the absorbency (referred to as a ‘zone phenomenon’). As shown inFIG. 14, when the zone phenomenon occurs, two concentrations (C1and C2) are obtained in the low concentration range and high concentration range of a single absorbency level A, such that the concentration of the assay material cannot be ultimately determined. Therefore, the concentration range in which the assay material can be measured is limited.

Japanese Laid-Open Patent Publication No. 61-280568 discloses a method for measuring an assay material even when the specimen contains a high concentration of assay material. This method utilizes CIA using carrier particles as the principle of measurement. CIA first mixes a specimen including an assay material and carrier particles on which antibody or antigen against the assay material is immobilized, and agglutinates the carrier particles by an antigen-antibody reaction. After a predetermined reaction time, the degree of agglutination is detected and the carrier particle distribution is obtained, then the degree of agglutination of the carrier particles is analyzed from the particle distribution, and the concentration of the assay material is calculated based on the degree of agglutination. The predetermined reaction time in this case is the time interval from the initiation of the antigen-antibody reaction until agglutination is detected.

In the method disclosed in Japanese Laid-Open Patent Publication 61-280568, a calibration curve is prepared across the entire region including the low concentration range and high concentration range, and the agglutination of the carrier particles is determined for the reaction time T1and reaction time T2of the antigen-antibody reaction. Then, a concentration is calculated from the degree of agglutination and calibration curve at reaction time T1, and another concentration is calculated from the degree of agglutination and calibration curve at reaction time T2. In this way the concentrations are compared for the reaction time T1and reaction time T2, and a concentration common to both reaction time T1and reaction time T2is designated as the final assay material concentration.

The method of Japanese Laid-Open Patent Publication No. 61-280568 is described below usingFIG. 15.FIG. 15shows the calibration curves at reaction time T1and reaction time T2(where T1<T2). InFIG. 15, calibration curve T1is the calibration curve at reaction time T1, and calibration curve T2is the calibration curve at reaction time T2. For example, when the agglutination at reaction time T2is designated B, the concentration of the assay material is either C1or C2. Likewise, when the agglutination at reaction time T1is A1, the concentration C1, which is common to each reaction time on the calibration curves, becomes the concentration of the assay material in the specimen. Similarly, when the agglutination at reaction time T1is A2, the concentration C2, which is common to each reaction time on the calibration curves, becomes the concentration of the assay material in the specimen.

However, the value of the degree of agglutination at reaction time T2is indispensable for determining the concentration of the assay material in this method. That is, the degree of agglutination at both reaction times T1and T2are invariably required in this method.

BRIEF SUMMARY

The scope of the present invention is defined solely by the appended claims, and is not affected to any degree by the statements within this summary. The present invention provides a biological sample analyzing apparatus and biological sample analyzing method providing improved efficiency in biological sample analysis over the conventional art.

A first aspect of the present invention relates to a biological sample analyzing apparatus comprising: an assay sample preparing mechanism for preparing assay sample by mixing testing reagent and a biological sample; a measuring unit for collecting information relating to an assay material contained in the biological sample from the assay sample; a first control means for controlling the measuring unit such that the measuring unit collects a first information relating to the assay material; a first analyzing means for analyzing the assay material based on the first information when the first information satisfies a predetermined condition; a second control means for controlling the measuring unit such that the measuring unit collects a second information relating to the assay material when the first information does not satisfy the predetermined condition; and a second analyzing means for analyzing the assay material based on the second information.

A second aspect of the present invention relates to a biological sample analyzing apparatus comprising: an assay sample preparing mechanism for preparing assay sample by mixing testing reagent and a biological sample; a measuring unit for detecting transmitted light from an assay sample, and collecting information of change in absorbance based on the detected transmitted light; a first control means for controlling the measuring unit so as to collect a first information of change in absorbance based on the transmitted light detected at a first time and a second time, wherein the first time and the second time are times after the assay sample preparing mechanism has prepared the assay sample; a first analyzing means for analyzing an assay material contained in the biological sample based on the first information when the first information satisfies a predetermined condition; a second control means for controlling the measuring unit so as to collect a second information of change in absorbance based on the transmitted light detected at a third time and a fourth time when the first information does not satisfy the predetermined condition, wherein the third time and the fourth time are times after the assay sample preparing mechanism has prepared the assay sample; and a second analyzing means for analyzing the assay material based on the second information.

A third aspect of the present invention relates to a biological sample analyzing method comprising the steps of: (a) preparing an assay sample by mixing testing reagents and a biological sample; (b) collecting a first information relating to an assay material contained in the biological sample from the assay sample; (c) analyzing the assay material based on the first information when the first information satisfies a predetermined condition; (d) collecting a second information relating to the assay material from the assay sample when the first information does not satisfy the predetermined condition; and (e) analyzing the assay material based on the second information.

In a biological sample analyzing apparatus and method of an embodiment, first an assay sample is prepared by mixing a reagent with a biological sample which may contain assay material. Then, a first information relating to the assay material is collected from the assay sample, and when the first information satisfies predetermined condition, the assay material is analyzed based on the first information. However, when the first information does not satisfy the predetermined condition, a second information related to the assay material is collected from the assay sample, and the assay material is analyzed based on the second information. Therefore, the first information and the second information need not be collected for all biological samples, and the second information is collected only when the first information does not satisfy predetermined condition. In this way, it is possible to improve efficiency when analyzing biological samples.

The method of analyzing biological samples comprises a method in which an assay material contained in a biological sample, such as blood and the like, is measured using an antigen-antibody reaction. Biological sample analyzing methods in which an assay material is measured using an antigen-antibody reaction include, for example, immunoturbidity methods, immunonephelometry, counting immunoassay (CIA) and the like.

The biological samples used as the biological samples in the aforesaid analyzing method are not specifically limited, and may be, for example, urine, and blood samples such as whole blood, plasma, and serum and the like.

The reagents used in the aforesaid analysis method are not specifically limited, and may be, for example, reagents containing substances that induce an antigen-antibody reaction with the assay material. Substances that induce an antigen-antibody reaction with the assay material may be an antigen that produces a specific antigen-antibody reaction with the antibody when the assay material is an antibody, and may be an antibody that produces a specific antigen-antibody reaction with the antigen when the assay material is an antigen. For example, an anti CRO antibody may be used in the case of an assay material that is a CRP antigen marker for infection and myocardial infarction.

Furthermore, Immunoturbidity method that use carrier particles, immunonephelometry method that use carrier particles, and counting immunoassay method use carrier particles on which the substance that produces an antibody-antigen reaction with the assay material are immobilized. For example, when the assay material is a CRP antigen, carrier particles on which anti CRP antibody is immobilized may be used. The carrier particles typically used in the aforesaid analyzing method includes, for example, latex particles, magnetic particles, metal particles, dendrimer and the like.

The information relating to the assay material comprises information which generally used in methods that measure an assay material contained in a biological sample using an antigen-antibody reaction. For example, transmitted light and absorbance in the immunoturbidity methods and scattered light in the immunonephelometry methods may be used as the information relating to the assay material. Furthermore, the change in transmitted light, absorbance or scattered light per a predetermined time may be used. The rate of agglutination of carrier particles in the CIA method may also be used. The rate of agglutination in the CIA method can be determined based on information that reflects the size of particles (hereinafter referred to as ‘size information’). When unagglutinated carrier particles (hereinafter referred to as ‘independent particles’) and clusters formed by a plurality of agglutinated carrier particles (hereinafter referred to as ‘aggregates’) are compared, the apparent size of the aggregate is larger. Therefore, independent particles and aggregates can be differentiated and counted separately and the rate of agglutination of carrier particles can be determined by detecting the size information. For example, the value of P/T can be used as the agglutination rate. The value of P/T is calculated based on the total number of particles (T) obtained by adding the number of independent particles (M) and the number of aggregates (P). The optical information of the scattered light can be used as the size information. Furthermore, electrical information representing the direct current resistance obtained when the particles cross between electrodes through which a direct current flow may be used as an alternative to optical information.

In the method for analyzing biological samples, a first information related to an assay material is collected from an assay sample, and the assay material is analyzed based on the first information when the first information satisfies predetermined condition. However, when the first information does not satisfy the predetermined condition, a second information related to the assay material is collected from the assay sample, and the assay material is analyzed based on the second information. The predetermined condition is used to determine whether or not the assay material can be measured using the first information. For example, when a threshold value X is provided for the first information and a condition is set such that “the assay material is measured based on the first information when the first information is equal to or greater than X.” Then the assay material is measured based on the first information when the first information obtained by a measurement is equal to or greater than X, and the assay material is measured based on a second information when the first information is less than X. This condition may be suitably set in accordance with the type of measurement method and type of assay material while considering the reliability of the result obtained by the measurement based on the first information.

The biological sample analyzing apparatus1of one embodiment of the present invention is described hereinafter. The biological sample analyzing apparatus1uses the measurement principles of the CIA method.

The biological sample analyzing apparatus1prepares an assay sample by mixing a carrier particle suspension and reaction buffer solution with a specimen, such as blood, urine or the like. Carrier particles suspended in a suitable fluid, such as water, buffer solution or the like, may be used as the particle suspension. If an assay material is present in the specimen, the carrier particles agglutinate by an antigen-antibody reaction when the carrier particle suspension is added to the specimen. The reaction buffer solution may be added to the carrier particle suspension and specimen to adjust the environment in which the antigen-antibody reaction is produced. The biological sample analyzing apparatus1illuminates the prepared assay sample with laser light, detects optical information emitted from the assay sample, and calculates the carrier particle agglutination rate based on the detected optical information. The optical information is detected at a predetermined reaction time. The predetermined reaction time in this case is the time interval from the initiation of the antigen-antibody reaction until agglutination is detected. The biological sample analyzing apparatus1calculates the carrier particle agglutination rate based on the optical information detected at reaction time T1. Then, the value of the agglutination rate at reaction time T1is compared to a predetermined threshold value, and a determination is made as to whether or not measurement at reaction time T2(T1<T2) is required. When the value of the agglutination rate at reaction time T1is equal to or greater than the predetermined threshold value, the assay material is measured based on a calibration curve and the agglutination rate at reaction time T1without performing a measurement at reaction time T2. However, when the value of the agglutination rate at reaction time T1is less than the predetermined threshold value, a measurement is performed at reaction time T2to obtain a measurement result of higher reliability since the measurement result at reaction time T1has a low reliability. In this case, the optical information at reaction time T2is detected, and the agglutination rate at reaction time T2is calculated based on the detected optical information. Then, the assay material contained in the specimen is measured based on a calibration curve and the agglutination rate at reaction time T2. The calibration curves represent the relationship between the agglutination rate and the assay material, and are prepared by measuring a standard solution consisting of a fluid containing a known concentration of the assay material. When preparing a calibration curve, a plurality of standard solutions are used which have graduatedly different concentrations of included assay material. In the biological sample analyzing apparatus1, forward scattered light is used as the optical information.

(General Structure of Biological Sample Analyzing Apparatus1)

FIG. 1is an external view of the biological sample analyzing apparatus1. A liquid crystal touch panel2for inputting various settings and displaying measurement results, assay sample preparation unit cover3, and start switch4are arranged on the front of the apparatus1.FIG. 2shows the internal structure of the biological sample analyzing apparatus1. A controller5for controlling the operation and analyzing process of the apparatus is arranged in a space at the right side of the apparatus1. A measuring unit6for detecting signals from the assay sample is arranged in a space on the lower left side of the apparatus1. An assay sample preparation unit7for preparing the assay sample is arranged in the remaining space.

(Structure of the Assay Sample Preparation Unit)

FIG. 3shows the assay sample preparation unit7. The assay sample preparation unit7includes a specimen placement unit8, standard solution placement unit9, reagent placement unit10, reaction unit11, dispensing unit12, and fluid delivery device13. An operator places a container containing a specimen in the specimen placement unit8by opening the assay sample preparation unit cover3. The operator places a container containing standard solution in the standard solution placement unit9. The operator respectively places a container14containing reaction buffer solution and a container15containing carrier particle suspension in the reagent placement unit10. A container is placed in the reaction unit11, an assay sample is prepared by mixing the reaction buffer solution and carrier particle suspension with the specimen or standard solution in the container. Although not shown in the drawing, the reaction unit11is provided with a mixing device for mixing the fluids in the container, and a temperature regulating device for maintaining the fluid in the container at a constant temperature. The tip of the dispensing unit12suctions and discharges a predetermined amount of fluids, and is also movable in vertical and front-to-back directions by means of a drive device not shown in the drawing. The fluid delivery device13includes a suction tube16for suctioning the assay sample, delivery tube17for delivering assay sample suctioned from the suction tube16to the measuring unit6shown inFIG. 4, and a pump18for suctioning and delivering the assay sample to the measuring unit6. Furthermore, the fluid delivery device13is movable in vertical and front-to-back directions by a drive device not shown in the drawing, so as t insert the suction tube16in the container placed in the reaction unit11and suction a predetermined amount of assay sample. The suctioned assay sample is delivered through the fluid delivery system17to the measuring unit6.

(Structure of the Measuring Unit)

FIG. 4shows the measuring unit6. The measuring unit6is provided with a sheath flow cell19, laser light source20, condenser lens21, collective lens22, pinhole23, and photodiode24. The sheath flow cell19is provided for the flow of the assay sample prepared by the previously described assay sample preparation unit7shown inFIG. 3. As shown inFIG. 5, the sheath flow cell19is provided with a sample nozzle25for spraying assay sample upward toward a fine bore part28, a sheath fluid inlet26, and waste fluid outlet27. The collective lens22collects the forward scattered light obtained from each individual particle in the sample illuminated by laser light. The photodiode24receives the forward scattered light, subjects the light to photoelectric conversion, and outputs the resulting electrical signal. Each output signal is transmitted to the controller5.

(Structure of the Controller)

FIG. 6is a block diagram showing the structure of the controller5, and the relationship between the controller5and each unit. The controller5has a microcomputer provided with a central processing unit (CPU), and memory devices such as ROM, RAM and the like, and circuits for processing the signals received from the measuring unit6. The controller5functions as the analyzing unit30, and operation controller31. The memory unit29stores an analysis program for analyzing the signals obtained from the particles in the sample, and a control program for controlling the operation of each part of the apparatus. The memory unit29also stores the data of the signals detected by the measuring unit6, and the processing results of the analysis program. The analyzing unit30analyzes the signals detected by the measuring unit6based on the analysis program, and generates data related to each particle contained in the assay sample. The data generated by the analyzing unit30are output to the liquid crystal touch panel2. The operation controller31controls the operation of each part of the apparatus based on the control program stored in the memory unit29.

The operation of the biological sample analyzing apparatus1is described in detail below. An operator first places the standard solution, specimen, and reagents at the predetermined positions in the assay sample preparation unit7. A standard solution is used at reaction time T1and a standard solution is used at reaction time T2; each standard solution consists of a plurality of standard solutions having graduatedly different concentrations of included assay material. The respective standard solutions can be placed in the standard solution placement unit9of the assay sample preparation unit7shown inFIG. 3by opening the assay sample preparation unit cover3shown inFIG. 1. The specimen can be placed in the specimen placement unit8of the assay sample preparation unit7. Furthermore, the container14containing reaction buffer and the container15containing carrier particle suspension can be respectively placed in the reagent placement unit10of the assay sample preparation unit7.

FIG. 7is a flow chart showing the general control flow of the control program. In step S1(mode setting process), a condition setting screen is displayed on the liquid crystal touch screen2. In the biological sample analyzing apparatus1, there are two measurement modes, which include the a calibration curve mode for measuring the standard solutions and preparing calibration curves, and a specimen mode for measuring a specimen and quantifying the assay material contained in the specimen; each mode is selectable by the operator in accordance with the measurement to be performed. The operator enters various settings in the displayed condition setting screen, for example, the measurement mode, reaction time which is time from the start of the antigen-antibody reaction by the addition of the carrier particle suspension until agglutination is detected, concentrations of the assay material contained in each standard solution and the like. When setting input of step S1ends, step S2(discrimination of the calibration curve mode selection), step S3(standard solution measuring process), step S4(calibration curve preparation process), step S5(α(T1) setting process), step S6(discrimination of the specimen mode selection), step S7(specimen measuring process), step S8(all specimens completion), and step S9(output process) are sequentially executed.

For example, when only calibration curve preparation is performed, only the calibration mode is selected in the mode setting process of step S1. In this case, in the subsequent step S2, ‘calibration mode is set’ is discriminated, and then in step S3the measurement of each standard solution is sequentially performed. When the measurement of all standard solutions is completed in step S3, the process continues to step S4and calibration curves are respectively prepared at reaction times T1and T2. (Hereinafter, the calibration curve at reaction time T1is referred to as calibration curve T1, and the calibration curve at reaction time T2is referred to as calibration curve T2.) When the calibration curves have been prepared in step S4, the process continues to step S5. In step S5, the threshold value of the agglutination rate at reaction time T1is set at α(T1). In the biological sample analyzing apparatus1, the value of the agglutination rate at reaction time T1is compared with the predetermined threshold value, and a determination is made as to whether or not to perform a measurement at reaction time T2(T1<T2). The value α(T1) set in step S5is equivalent to the predetermined threshold value. When α(T1) is set in step S5, the process advances to step S6. In step S6, “specimen mode not set” is determined, and the process advances to step S9and the data, such as the standard solution agglutination rate and calibration curves and the like, are output.

When the calibration curves have already been prepared and only specimen measurements are to be performed, only the specimen mode is selected in the mode setting process of step S1. In this case, in the subsequent step S2, ‘calibration mode is not set’ is discriminated, and then α(T1) is set in step S5. When α(T1) is set in step S5, the process advances to step S6. In step S6, “specimen mode is set” is determined, and the process advances to step S7and the specimen is measured. When measuring a plurality of specimens, “measurement of all specimens not completed” is determined in step S8, and step S7is repeated. When the measurement of all specimens has been completed, the process advances to step S9and the data, such as the specimen measurement results and the like, are output.

When both calibration curve preparation and specimen measurement are performed, both the calibration curve mode and specimen mode are selected in the mode setting process of step S1. In this case, the calibration curves are prepared through steps S1, S2, S3, and S4, and thereafter α(T1) is set in step S5. When α(T1) is set in step S5, the process advances to step S6. In step S6, “specimen mode is set” is determined, and the process advances to step S7and the specimens are measured through step S8. When the measurement of all specimens has been completed, the process advances to step S9and the data, such as the calibration curves and specimen measurement results and the like, are output. The assay sample preparation unit7, measuring unit6, and analyzing unit29are controlled by the control program, and their sequential operations are automatically performed from steps S1through S9. Each step is described below.

The mode setting process is described below referring toFIGS. 8 and 9.FIG. 8shows the screens displayed on the liquid crystal touch panel2during the mode setting process. Various types keys are displayed on the screens, and a key is selected when the operator uses a finger or the like to touch a position at which a key is displayed on the liquid crystal touch panel2.FIG. 9is a flow chart showing the flow of the mode setting process. Each step of the flow chart is described below.

S101: The screen A shown inFIG. 8Ais displayed on the liquid crystal touch panel2. Then, the process advances to step S102.

S102: the measurement mode and reaction time settings are entered on the screen A. A key for operator selection of the measurement mode is provided at the upper left side of the screen A. Calibration curves are prepared when the [calibration mode] key is selected, and a specimen is measured when the [specimen mode] key is selected. Furthermore, when both the [calibration mode] key and the [specimen mode] key are selected, the specimen is measured after the calibration curves have been prepared. Boxes for the operator to enter the reaction times T1and T2are provided in the lower left of the screen A. A ten-key pad is provided on the right side of the screen A for entering numerical values in each box. When setting input on screen A has been completed, the process moves to step S103.

S103: An [Accept] key is displayed on the screen A, and the selection of the [Accept] key by the operator is received in step S103. When the [Accept] key has been selected, the process continues to step S104.

S104: When [calibration mode] is selected in step S102, the process continues to step S105. However, when [calibration mode] is not selected in step S102, the process advances to step S108.

S105: The screen B shown inFIG. 8Bis displayed on the liquid crystal touch panel2. Then, the process continues to step S106.

S106: The setting inputs related to the calibration curve, such as the number and concentrations of each standard solution used to prepare the calibration curves, are received on the screen B. Boxes for the operator to enter the number and concentrations of each standard solution are provided on the left side of the screen B. A ten-key pad is provided on the right side of the screen B for entering numerical values in each box. When setting input on screen B has been completed, the process moves to step S107.

S107: The [Accept] key is displayed on the screen B, and the operator selection of the [Accept] key is received in step S107. When the [Accept] key has been selected, the process continues to step S108.

S108: When the [specimen mode] is selected in step S102, the process continues to step S109. When the [specimen mode] has not been selected in step S102, however, the mode setting process ends.

S109: The screen C shown inFIG. 8Cis displayed on the liquid crystal touch panel2. Then, the process advances to step S110.

S110: Setting inputs related to the specimen mode, such as specimen number and specimen name and the like, are received on screen C. Boxes for the operator to enter the number and name of each specimen are provided on the left side of the screen C. A ten-key pad is provided on the right side of the screen C for entering numerical values and text in each box. When setting input on screen C has been completed, the process moves to step S111.

S111: The [Accept] key is displayed on the screen C, and the operator selection of the [Accept] key is received in step S111. When the [Accept] key has been selected, the mode setting process ends.

In step S2, a determination is made as to whether or not the calibration mode has been selected based on the measurement conditions input in the mode setting process of step S1. When [calibration mode] has been selected in step S1, the process continues to step S3(standard solution measuring process). When [calibration mode] is not selected in step S1, however, the process advances to step S5(α(T1) setting process).

In step S3, the standard solutions containing known concentrations of assay material are measured.FIG. 10is a flow chart showing the flow of the standard solution measuring process. In the standard solution measuring process, step S301(assay sample preparation process), step S302(measuring process), and step S303(analysis process) are sequentially performed. Steps S301, S302, and S303are described below.

The operation of the assay sample preparation unit7in step S301is described referring toFIG. 3. First, the dispensing unit12suctions standard solution from the container placed in the standard solution placement unit9, and dispenses 10 μL to the container placed in the reaction unit11. Then, the dispensing unit12suctions reaction buffer from the container14placed in the reagent placement unit10, and dispenses 80 μL to the container placed in the reaction unit11. Then, the dispensing unit12suctions carrier suspension from the container15placed in the reagent placement unit10, and dispenses 10 μL to the container placed in the reaction unit11. The antigen-antibody reaction is started by the addition of the carrier particle suspension. The assay sample in the container in the reaction unit11is agitated while maintained at a temperature of 45° C. Then, the fluid delivery device13suctions 14.5 μL of the assay sample in the container in the reaction unit11, and delivers the sample to the sheath flow cell19of the measuring unit6.

When preparing a calibration curve, a plurality of standard solutions are used which have graduatedly different concentrations of included assay material. Therefore, a plurality of T1standard solutions are placed in the standard solution placement unit9. Then, assay samples are sequentially prepared from the standard solutions of each concentration placed in the standard solution placement unit9. Thus, when assay samples are prepared from the standard solutions, the subsequent steps S302and S303described later are sequentially executed. Then the agglutination rate at reaction time T1is calculated for the assay samples prepared from the T1standard solutions, and the agglutination rate at reaction time T2is calculated for the assay samples prepared from the T2standard solutions.

The operation of the measuring unit6in the measuring process is described below referring toFIGS. 4 and 5. In the measuring process, after the carrier particle suspension has been added and the antigen-antibody reaction has started, 14.5 μL of the assay sample is suctioned from the container in the reaction unit11by the fluid delivery device13and delivered to the sheath flow cell19of the measuring unit6. The assay sample delivered to the sheath flow cell19is discharged from the sample nozzle25into the sheath flow cell. Sheath fluid is also discharged from the sheath fluid inlet26into the sheath flow cell at the same time as the aforesaid operation. In this way the sample fluid is encapsulated in sheath fluid within the sheath flow cell, and then the flow is narrowly constricted by the fine bore part28. The sample fluid flows in the fine bore part28and the particles contained in the sample fluid can be adjusted to form a line by the constricting the flow to the same degree as the particle diameter.

Laser light emitted from the laser light source20is constricted by the condenser lens21, and irradiates the sample flowing through the fine bore part28. The forward scattered light from each individual particle in the sample illuminated by the laser light is collected by the collective lens22and passes through the pinhole23. The forward scatter light that passes through the pinhole23is received by the photodiode24, subjected to photoelectric conversion, and output as a forward scattered light signal. Each output signal is transmitted to the controller5, and stored in the memory unit29as data for each particle. Thus, the forward scattered light from the assay sample is detected at predetermined reaction times in step S302. These predetermined reaction times are the reaction time T1and reaction time T2set in step S1.

When the forward scattered light is detected in step S302, analysis is executed by the analyzing unit30based on the analysis program. The operation of the analysis program in step S303is described below using the flow chart ofFIG. 11. Each step of the flow chart is described below.

Step S303-1: The data of the forward scattered light signals are read from the memory unit29. Then, the process moves to step S303-2.

S303-2: The forward scatter light intensity (Fsc) is calculated for each particle in the sample fluid based on the data of the forward scattered light signals. Then, the process continues to step S303-3.

S303-3: A carrier particle histogram is prepared.FIG. 16shows an example of a histogram prepared based on the carrier particle Fsc; the number of particles are plotted on the vertical axis, and the Fsc is plotted on the horizontal axis. Then, the process continues to step S303-4.

S303-4: the agglutination rate is calculated based on the histogram prepared in step S303-3. First, the independent particles and aggregates are differentiated based on the histogram prepared in step S303-3. The detected particles are distributed to positions corresponding to the size of the carrier particles, that is, independent particles, two aggregate particles, and three aggregate particles. Actual particles are not distributed at locations smaller than independent particles, locations between the independent particles and two aggregate particles, locations between two aggregate particles and three aggregate particles, and locations larger than three aggregate particles, as indicated by v, w, x, and y in theFIG. 16. In this histogram, the threshold value is set between the forward scattered light intensity corresponding to the size of independent particles and the forward scatter light intensity corresponding to the size of two aggregate particles; and the number of independent particles (M) and the number of aggregate particles (P) can be calculated by identifying the particles distributed in the range below the threshold value as independent particles, and identifying the particles distributed within the range greater than the threshold value as aggregates. Furthermore, the total number of particles (T) can be determined by adding M and P, such that P/T can be calculated as the agglutination rate. Then, the process continues to step S303-5.

S303-5: The histogram prepared in step S303-3and the data of the agglutination rate calculated in step S303-4are stored in memory.

The aforesaid is shown in the flow chart of step S3(standard solution measuring process). In this way the T1standard solutions and T2standard solutions are measured, and the agglutination rate is calculated for each standard solution.

In step S4, the calibration curves are prepared based on the data of the agglutination rate for each standard solution obtained in step S3. The calibration curves prepared in step S4include a calibration curve T1prepared based on the data obtained by measurements performed at reaction time T1, and calibration curve T2prepared based on data obtained by measurement performed at reaction time T2. The calibration curve T1is prepared based on the concentration of the assay material in the T1standard solutions input in step S1, and the agglutination rate obtained by measuring the T1standard solutions in step S3. The calibration curve T2is prepared based on the concentration of the assay material in the T2standard solutions input in step S1, and the agglutination rate obtained by measuring the T2standard solutions in step S3. The operation of the analysis program in the calibration curve preparation process is described below using the flow chart ofFIG. 12. Each step of the flow chart is described below.

Step S401: Data of the concentration of the assay material in each T1standard solution and agglutination rate of each T1standard solution calculated in step S3of the analysis are read from the memory unit29. Then, the process continues to step S402.

S402: The calibration curve T1is prepared based on the concentrations and agglutination rates. Then, the process continues to step S403.

S403: The calibration curve T1prepared in step S402is stored in memory. Then, the process continues to step S404.

S404: Data of the concentration of the assay material in each T2standard solution and the agglutination rate of each T2standard solution calculated in step S3of the analysis are read from the memory unit29. Then, the process continues to step S405.

S405: The calibration curve T2is prepared based on the concentrations and agglutination rates. Then, the process continues to step S406.

S406: The calibration curve T2prepared in step S405is stored in memory.

The aforesaid steps are shown in the flow chart of the calibration curve preparation process. Thus, the calibration curve T1and calibration curve T2are prepared in this manner.

In step S5, a threshold value α(T1) is set for the agglutination rate at reaction time T1. In general, when calculating concentrations using a calibration curve, the concentration range in which the calculation of a reliable value is possible is a range in which the calibration curve is linear. For this reason, value have been determined based on the lower limit value of the range ensuring the linearity of the calibration curve T1and the upper limit value of the range ensuring the linearity of the calibration curve T2, and have been stored as threshold value data in the memory unit29. In the present step, the threshold value data are automatically read from the memory unit29, and set as α(T1). When α(T1) is set in this way, the routine advances to step S6(specimen mode selection determination).

In step S6, a determination is made as to whether or not the specimen mode has been selected based on the measurement conditions input in the mode setting process of step S1. When [specimen mode] has been selected in step S1, the process continues to step S7(specimen measuring process). When [specimen mode] is not selected in step S1, however, the process advances to step S9(output process).

In the specimen measuring process, a specimen is measured, and the concentration of the assay material contained in the specimen is calculated based on the agglutination rate and the calibration curves prepared in step S4.FIG. 13is a flow chart showing the flow of the specimen measuring process. In the specimen measuring process, step S701(assay sample preparation), step S702(T1measurement), step S703(T1analysis), step S704(determination whether to perform T2measurement), step S705(T2measurement), step S706(T2analysis), and step S707(quantification) are sequentially performed under the conditions input in the mode setting process of step S1.

The operation of the assay sample preparation unit7in the assay sample preparation process is described below referring toFIG. 3. The dispensing unit12first suctions specimen from the container placed in the specimen placement unit8, and dispenses 10 μL of specimen into the container placed in the reaction unit11. Then, the dispensing unit12suctions reaction buffer from the container placed in the reagent placement unit10, and dispenses 80 μL to the container placed in the reaction unit11. Then, the dispensing unit12suctions carrier suspension from the container15placed in the reagent placement unit10, and dispenses 10 μL to the container placed in the reaction unit11. The antigen-antibody reaction is started by the addition of the carrier particle suspension. The assay sample in the container in the reaction unit11is agitated while maintained at a temperature of 45° C. Then, the fluid delivery device13suctions 14.5 μL of the assay sample in the container in the reaction unit11, and delivers the assay sample to the sheath flow cell19of the measuring unit6. When the assay sample is suctioned from the container in step S702, the reaction unit11thereafter continues to agitate the assay sample in the container while maintaining it at a temperature of 45° C.

In step S702, the forward scattered light signal is detected at reaction time T1input in step S1. When the reaction time T1input in step S1is, for example, 20 seconds, then the operation of the assay sample preparation unit7and the measuring unit6is controlled by the control program so as to detect the forward scattered light 20 seconds after the start of the antigen-antibody reaction. The operation of the measuring unit6in step S702is identical to the operation in step S302(measuring process), that is, the forward scattered light signals are detected and the detected signals are stored in the memory unit29.

When the forward scattered light is detected in step S702, T1analysis is executed by the analyzing unit30based on the analysis program. The operation of the analysis program in step S703is identical to the operation in step S303(analysis process), that is, the agglutination rate A is calculated based on the detected forward scattered light signals. Then, the data of the calculated agglutination rate A and the prepared histogram are stored in the memory unit29.

S704(Determination of Whether to Perform T2Measurement)

The agglutination rate A calculated in step S703is compared to the threshold value α(T1) set in step S5. When the value of the calculated agglutination rate A is greater than α(T1), a measurement at reaction time T2is not required because it is possible to determine the assay material concentration using the agglutination rate A at reaction time T1. In this case, therefore, the temperature maintenance and agitation in the container of the reaction unit11ends in the assay sample preparation unit7, and the process advances to step S707. When the value of the calculated agglutination rate A is less than α(T1), a measurement at reaction time T2is required because it is impossible to determine the assay material concentration using the agglutination rate A at reaction time T1. At this time, therefore, the process advances to step S705. Also at this time, the temperature maintenance and agitation in the reaction unit11is continued thereafter as before until reaction time T2.

When the value of the agglutination rate A is less than α(T1) in step S704, the next step S705is executed. In step S705, the forward scattered light signal is detected at reaction time T2input in step S1. When the reaction time T2input in step S1is, for example, 15 minutes, then the operation of the assay sample preparation unit7and the measuring unit6is controlled by the control program so as to detect the forward scattered light 15 minutes after the start of the antigen-antibody reaction. The operation of the measuring unit6in the T2measurement is identical to the operation in step S302(measuring process), that is, the forward scattered light signals are detected and the detected signals are stored in the memory unit29.

When the forward scattered light is detected in step S705, T2analysis is executed by the analyzing unit30based on the analysis program. The operation of the analysis program in step S706is identical to the operation in step S303(analysis process), that is, the agglutination rate B is calculated based on the detected forward scattered light signals, and the data of the calculated agglutination rate B and the prepared histogram are stored in the memory unit29.

In step S707, the concentration of assay material contained in the specimen is calculated based on the calibration curves preciously prepared in step S4and the agglutination rate obtained in step S703or S706. When the T2measurement and T2analysis are not performed, the concentration of the assay material is calculated based on the calibration curve T1and the agglutination rate A at reaction time T1. However, when the T1measurement and T2analysis are performed, the concentration of the assay material is calculated based on the calibration curve T2and the agglutination rate B at reaction time T2. Then, the concentration data calculated in step S707is stored in the memory unit29.

When a plurality of specimens are assayed, step S7is repeated until [measurement of all specimens has been completed] has been determined in step S8. When the measurement of all specimens has been completed, the process advances to step S9.

The standard solution agglutination rate data stored in step S3, the calibration curve data stored in step S4, and specimen agglutination rate and concentration data stored in step S7are output and displayed on the liquid crystal touch panel2.

The aforesaid steps are shown in the flow chart of the general control in the present embodiment. As described above, the biological sample analyzing apparatus1is an automatic analyzer that performs automatically from the preparation of the assay sample to the quantification of the assay material.

The example describes the preparation of calibration curves using the previously described biological sample analyzing apparatus1. The reaction time were set at T1=20 seconds, and T2=15 minutes.

A latex reagent containing latex particle on which anti-CRP antibody is immobilized was used as the carrier particle suspension. This reagent was prepared by the following method. First, 50 μl of 10% (w/v) polystyrene latex (commercial product) was added to 450 μl of GTA buffer solution (0.53 mg/ml 3,3-dimethylglutaric acid, 0.4 mg/ml trishydroxymethylaminomethane, 0.35 mg/ml 2-amino-2-methyl-1,3-propane diol; pH 7.0) containing 100 μg of anti-CRP antibody (mouse monoclonal antibody, a commercial product), and allowed to rest for 2 hours. The solution was centrifuged for 10 minutes at 10,000×g, and 1 ml GTA buffer solution containing 1% (w/v) bovine serum albumin (commercial product) was added to the centrifuge precipitate, then the solution was subjected to an ultrasound process for dispersion. The process from centrifuging to dispersion was repeated a plurality of times, and the supernatant was removed after the final centrifugation, and 1 ml GTA buffer solution (pH 6.2) containing 220 mg/ml glycerin and 0.3% (w/v) bovine serum albumin was added and subjected to ultrasound processing. The solution was used as the latex reagent. A suitable size of the carrier particles in the present embodiment is a particle diameter ranging from 0.1˜1.0 μm. The particle diameter of the polystyrene latex used in the present example was 0.7 μm.

The standard solutions were prepared by the following method. A PBS solution (2.4 mg/ml trishydroxymethylamino methane, 2.4 mg/ml sodium chloride; pH 7.5) containing 5% (w/v) bovine serum albumin was prepared, and purified CRP (commercial product) was added to produce concentrations of 4.05×102, 1.215×103, 5×103, 2×104, 6×104, 1.8×105, and 3×105ng/mL as the T1standard solutions, and purified CRP (commercial product) was added to produce concentrations of 5, 15×10, 4.5×10, 1.35×102, 4.05×102, 1.215×103ng/mL as the T2standard solutions.

FIG. 17shows the calibration curves obtained by measuring the T1and T2standard solutions.FIG. 17Ais a calibration curve obtained by measuring the agglutination rate of the T1standard solutions at a reaction time of 20 seconds.FIG. 17Bis a calibration curve obtained by measuring the agglutination rate of the T2standard solutions at a reaction time of 15 minutes. In both cases, the agglutination rate is plotted on the vertical axis, and the CRP concentration is plotted on the horizontal axis.

When the calibration curve T1is used, it can be understood fromFIG. 17Athat CRP can be measured in a measurement range of 4×102ng/mL to 3×105ng/mL. When the calibration curve T2is used, it can also be understood from T2inFIG. 17that CRP concentration can be measured in a measurement range from 5 ng/mL to 1×103ng/mL. From the above information it can be understood that, in the biological sample analyzing apparatus1, CRP concentration can be measured in an extremely broad measurement range from 5 ng/mL to 3×105ng/mL by using both calibration curve T1and calibration curve T2.

Next, the calibration curve T2was prepared at various reaction times T2using the previously described biological sample analyzing apparatus1.

Each type of reagent and the T2standard solutions used in the present example were identical to the reagents and T2standard solutions used in measurement example 1.

InFIG. 18, the change in the agglutination rate per unit concentration increases in conjunction with the lengthening of reaction time T2. In this way the agglutination rate sensitivity becomes greater as the time set as the reaction time T2increases. That is, effective measurement is possible by setting the reaction time T2in accordance with the measurement range required by the operator without increasing the reaction time longer than is necessary.

Moreover, it is understood that the calibration curve e inFIG. 18describes an increasing agglutination rate in conjunction with an increase in CRP concentration in the concentration range from 10 ng/mL to 1×103ng/mL. In this way even when the reaction time T2was set at a short 95 seconds, the measurement could be used in the calibration curve, and in this case CRP was measurable even in specimens with a relatively low concentration of 10 ng/mL.

The measurement results shown were taken when the reaction time T1was set at 20 sec, and reaction time T2was set at 95 sec, using the previously described biological sample analyzing apparatus1. In the present example, the agglutination rate threshold value α(T1) was set at 7%. In this way the concentration of CRP contained in a specimen could be calculated based on the agglutination rate A and calculation curve T1at reaction time T1when the agglutination rate A was 7% or greater. Moreover, the concentration of CRP contained in a specimen could be calculated based on the agglutination rate B at reaction time T2when the agglutination rate A was less than 7%.

Each type of reagent and standard solutions used in the present example were identical to the reagents and standard solutions used in measurement example 1. In the present example, blood plasma 1˜5 and blood sera 1˜3 containing various concentrations of CRP are used as specimens. The same specimens were assayed using a Dimension R×L automatic bioanalyzer, manufactured by Dade Behring, Inc., to determine the concentration of CRP contained in each specimen.

Table 1 shows the results when each specimen was measured using the biological sample analyzing apparatus1. Among the categories of Table 1, [A (%)] are values representing the agglutination rate A (%) at reaction time T1, and [B (%)] are values representing the agglutination rate B (%) at reaction time T2. The [calibration curve] represents the calibration curve used to calculate the concentration, T1indicating that the calibration curve T1was used, and T2indicating that the calibration curve T2was used. Furthermore, [concentration I] represents the CRP concentration (ng/mL) obtained using the biological sample analyzing apparatus1, and [concentration II) represents the CRP concentration obtained using Dimensional R×L.

It can be understood from Table 1 regarding the specimens having relatively high concentrations of CRP (plasma 3, plasma 4, plasma, 5, serum 2, serum 3) that the CRP concentrations calculated from assays using the biological sample analyzing apparatus1approximate the CRP concentrations determined beforehand in assays using the Dimension R×L. In regard to the specimens with relatively low concentrations of CRP (plasma 1, plasma 2, and serum 1), however, the assays using the biological sample analyzing apparatus1calculated CRP concentrations of higher sensitivity than did the assays using the Dimension R×L.

The measurement results in this case sows the results when whole blood was assayed using the previously described biological sample analyzing apparatus1. The whole blood further contained hemocytes. Therefore, when whole blood is used as the specimen and the same quantity is used as when assaying serum and plasma, the assay values are lower because they reflect the hemocyte component (hemocyte volume). Therefore, the hemocyte volume must be compensated when assaying whole blood. In the present example, this compensation is made using the method disclosed in U.S. Application Patent Publication No. 2003-0082662. Specifically, whole blood specimens were assayed using a completely automated hemocyte analyzer model XE-2100 manufactured by Sysmex Corporation to determine the number of hemocytes beforehand. Compensation was accomplished using the following equation.
C=C0/{1−(B/A)}

(Where C represents the concentration of assay material after compensation, C0represents the concentration of the assay material when whole blood is measured, B represents the number of hemocytes contained in the whole blood, and A represents a constant.) The constant A can be determined experimentally from the correlation between the number of hemocytes and the hematocrit value. The hematocrit value is equivalent to the total number of hemocytes when the hematocrit is assumed to be 100% (that is, when the entire whole blood sample is the hemocyte component); since red blood cells comprise nearly all the hemocytes in whole blood, the hematocrit value can be used as the percentage volume of hemocytes present in a constant amount of whole blood. The constant A is equivalent to the total number of hemocytes when the hematocrit value is assumed to be 100% (that is, the hemocyte component is the entire whole blood sample). The percentage hemocyte component in the collected whole blood sample can be determined by calculating B/A.

In the present example, reaction time T1=20 seconds, T2=95 seconds, and threshold value α(T1) was set at 7%. The reagents and standard solutions used were identical to the reagents and standard solutions used in measurement example 1. The specimens included whole blood 1˜4 containing various concentrations of CRP, and plasma 1˜4 obtained by centrifuging the whole blood 1˜4 (8,000 rpm for 5 minutes).

Table 2 shows the results obtained when each specimen was measured suing the biological sample analyzing apparatus1. In Table 2, the [calibration curve] represents the calibration curve used to calculate the concentration, T1indicating that the calibration curve T1was used, and T2indicating that the calibration curve T2was used. Furthermore, [concentration] represents the CRP concentration (ng/mL) obtained using the biological sample analyzing apparatus1.

When the CRP concentrations of whole blood 1 and plasma 1, whole blood 2 and plasma 2, whole blood 3 and plasma 3, whole blood 4 and plasma 4 were compared, in each case the whole blood CRP concentration approximated the plasma CRP concentration. This indicated that whole blood could be used for measurements.

In the present embodiment, measurements were performed at different reaction times (reaction time T1and reaction time T2). When the relationship of reaction time T1and reaction time T2was T1<T2, the reaction was relatively more stable at reaction time T2than reaction time T1. Therefore, the measurement of the relatively stable reaction at reaction time T2has better sensitivity and reproducibility than the measurement of the relatively unstable reaction at reaction time T1. The reason for this difference is that the calibration curve T2at reaction time T2was used to calculate the particularly low concentration of the assay material. However, the measurement at reaction time T1was less affected by the zone phenomenon than the measurement at reaction time T2. The reason for this difference is that the calibration curve T1at reaction time T1was used to calculate the particularly high concentration of the assay material.

In the present embodiment, predetermined conditions are provided relating to the measurement results at reaction time T1, such that the calibration curve T1is used when calculating the concentration of assay material that has a high concentration, and the calibration curve T2is used when calculating the concentration of assay material that has a low concentration. For this reason, measurements at both the first reaction time (reaction time T1) and the second reaction time (reaction time T2) are not always necessary, and measurement at reaction time T2is performed only when the measurement results for reaction time T1do not meet predetermined conditions. In this way the efficiency of the measurement is improved.

In the present embodiment, the reaction time T1and reaction time T2can be set in accordance with the measurement range deemed necessary by the operator. In this way measurement can be performed efficiently without performing measurements longer than necessary.

Although the reaction time T1is set at 10 seconds, and the reaction time T2is set at either 95 seconds or 15 minutes in the present embodiment, the reaction time T1and reaction time T2are not limited to these times. The time of the reaction time T1and the reaction time T2differ depending on the assay material. For this reason, the times of the reaction time T1and the reaction time T2are set at times suitable for the material being measured.

The biological sample analyzing apparatus41of another embodiment of the present invention is described below. The biological sample analyzing apparatus41employs the immunoturbidity method using latex particles as the measurement principle.

The biological sample analyzing apparatus41first prepares an assay sample by mixing a carrier particle suspension and reaction buffer solution with a specimen, such as blood, urine or the like. Then, the prepared assay sample is irradiated with light, the light transmitted through the assay sample is detected, and the absorbance is determined based on the detected transmitted light.FIG. 19shows an example of the change in absorbance during the reaction time. The biological sample analyzing apparatus41detects the absorbance at reaction times T1aand T1a′, and calculates the amount of change A in absorbance (A=a′−a) during the reaction time span T1based on each absorbance (a, a′) measurement. Then, the value of the amount of change A is compared to a predetermined threshold value, and a determination is made as to whether or not to perform measurements during reaction time span T2. When the amount of change A exceeds the predetermined threshold value, the assay material contained in the specimen is quantified based on the amount of change A without performing measurements at reaction time span T2. When the amount of change A is less than the predetermined threshold value, measurements are performed during the reaction time span T2to obtained measurement results of higher reliability since the measurement results in reaction time span T1have low reliability. In this case, the absorbance is detects at reaction times T2band T2b′, and the amount of change B in the absorbance during the reaction time span T2is calculated (B=b′−b) based on the detected absorbances (b, b′). Then, the assay material is quantified based on the amount of change B.

(General Structure of Biological Sample Analyzing apparatus41)

An external view of the biological sample analyzing apparatus41provides a liquid crystal touch panel, assay sample preparation unit cover, and start switch similar to the biological sample analyzing apparatus1shown inFIG. 1. The internal structure of the biological sample analyzing apparatus41has a controller, measurement unit, and assay sample preparation unit similar to the biological sample analyzing apparatus1shown inFIG. 2.

(Structure of the Assay Sample Preparation Unit)

FIG. 20shows the assay sample preparation unit of the biological sample analyzing apparatus41. The assay sample preparation unit includes a specimen placement unit42, standard solution placement unit43, reagent placement unit44, reaction unit45, and dispensing unit46. The operator places a container containing the specimen in the specimen placement unit42by opening the cover of the assay sample preparation unit. The operator respectively places a containers of standard solution in the standard solution placement unit43. The operator respectively places a container47containing reaction buffer and container48containing carrier particle suspension in the reagent placement unit44. A light-transmitting container is placed in the reaction unit45, and the assay sample is prepared by mixing the reaction buffer and carrier particle suspension with the specimen or standard solution. Although not shown in the drawing, the reaction unit45is provided with a temperature controlling mechanism for maintaining the solution in the container at a constant temperature, and a mixing mechanism for agitating the solution in the container. The dispensing unit46suctions and dispenses a predetermined amount of fluid from the tip, and the unit is movable vertically, laterally, and back-and-forth by means of a drive device not shown in the drawing.

(Structure of the Measuring Unit)

FIG. 21shows the measuring unit. The measuring unit is provided with a container49, light source50, filter51, and photodiode52. The light from the light source50is diffracted into a spectrum at 800 nm by the filter51. The diffracted light passes through the assay sample in the container49, and the transmitted light reaches the photodiode52. The photodiode52subjects the received transmitted light to photoelectric conversion, which is output as electrical signals. The output signals are transmitted to the controller.

(Structure of the Controller)

The structure of the controller of the biological sample analyzing apparatus41includes a central processing unit (CPU), microcomputer provided with storage devices such as ROM, RAM and the like, and circuits for processing the signals sent from the measuring unit. The Controller functions as an analysis unit and operation controller. The memory unit stores an analysis program for analyzing the signals obtained from the particles in the sample, and a control program for controlling the operation of each unit. The memory unit further stores the data of the signals detected by the measuring unit, and processing results of the analysis program. The analysis unit analyzes the signals detected by the measuring unit based on the analysis program, and generates data. The data generated by the analysis unit are output to the liquid crystal touch panel. The operation controller controls the operation of each unit based on the control program stored in the memory unit.

The operation of the biological sample analyzing apparatus41is described in detail below. First, the operator places standard solutions, specimens, and reagents at the predetermined positions in the assay sample preparation unit. The standard solutions include standard solutions used for reaction time T1and standard solutions used for reaction time T2, and the plurality of standard solutions have graduatedly different concentrations of included assay material. Then, the standard solutions can be placed in the standard solution placement unit43of the assay sample preparation unit shown inFIG. 20by opening the cover of the assay sample preparation unit of the apparatus41. The specimen can be placed in the specimen placement unit42of the assay sample preparation unit. Furthermore, the container47containing reaction buffer and the container48containing carrier particle suspension can be respectively placed in the reagent placement unit44of the assay sample preparation unit.

FIG. 22is a flow chart showing the general control flow of the control program. In step SS1(mode setting process), a condition setting screen is displayed on the liquid crystal touch panel. In the biological sample analyzing apparatus41, there are two measurement modes, which include the a calibration curve mode for measuring the standard solutions and preparing calibration curves, and a specimen mode for measuring a specimen and quantifying the assay material contained in the specimen; each mode is selectable by the operator in accordance with the measurement to be performed. The operator enters various settings in the displayed condition setting screen, for example, the measurement mode, reaction time which is time from the start of the antigen-antibody reaction by the addition of the carrier particle suspension until agglutination is detected, concentrations of the assay material contained in each standard solution and the like. When setting input of step SS1ends, step SS2(discrimination of the calibration curve mode selection), step SS3(standard solution measuring process), step SS4(calibration curve preparation process), step SS5(γ(T1) setting process), step SS6(discrimination of the specimen mode selection), step SS7(specimen measuring process), step SS8(all specimens completion), and step SS9(output process) are sequentially executed.

The assay sample preparation unit, measuring unit, and analysis unit are controlled by the control program, and the sequential operations of steps SS1to SS9are performed automatically. Each step is described below.

The mode setting process in the biological sample analyzing apparatus41is described below with reference toFIGS. 23 and 24.FIG. 23shows the screen displayed on the liquid crystal touch panel during the mode setting process. Various types keys are displayed on the screens, and a key is selected when the operator uses a finger or the like to touch a position at which a key is displayed on the liquid crystal touch panel2.FIG. 24is a flow chart showing the flow of the mode setting process. Each step of the flow chart is described below.

SS101: The screen A shown inFIG. 23Ais displayed on the liquid crystal touch panel. The process then advances to step SS102.

SS102: The measurement mode and reaction time setting inputs are received on the screen A. A key for operator selection of the measurement mode is provided at the upper left side of the screen A. Calibration curves are prepared when the [calibration curve mode] key is selected, and a specimen is measured when the [specimen mode] key is selected. Furthermore, when both the [calibration curve mode] key and the [specimen mode] key are selected, the specimen is measured after the calibration curves have been prepared. Boxes for the operator to enter the reaction times T1and T2are provided in the lower left of the screen A. The biological sample analyzing apparatus41determines the change in absorbance during the reaction time span T1from reaction time T1ato reaction time T1a′, and determines the change in absorbance during reaction time span T2from reaction time T2ato reaction time T2a′. Therefore, boxes for setting the reaction times are provided at four locations T1a, T1a′, T2a, and T2a′. A ten-key pad is provided on the right side of the screen A for entering numerical values in each box. When setting input on screen A has been completed, the process moves to step SS103.

SS103: An [Accept] key is displayed on the screen A, and the selection of the [Accept] key by the operator is received in step SS103. When the [Accept] key has been selected, the process continues to step SS104.

SS104: When [calibration mode] is selected in step SS102, the process continues to step SS105. However, when [calibration mode] is not selected in step SS102, the process advances to step SS108.

SS105: The screen B shown inFIG. 23Bis displayed on the liquid crystal touch panel. Then, the routine advances to step SS106.

SS106: The setting inputs related to the calibration curve mode, such as the number and concentrations of each standard solution used to prepare the calibration curves, are received on the screen B. Boxes for the operator to enter the number and concentrations of each standard solution are provided on the left side of the screen B. A ten-key pad is provided on the right side of the screen B for entering numerical values in each box. When setting input on screen B has been completed, the process moves to step SS107.

SS107: The [Accept] key is displayed on the screen B, and the operator selection of the [Accept] key is received in step SS107. When the [Accept] key has been selected and pressed on screen B, the process continues to step SS108.

SS108: When the [specimen mode] is selected in step SS102, the process continues to step SS109. When the [specimen mode] has not been selected in step SS102, however, the mode setting process ends.

SS109: The screen C shown inFIG. 23Cis displayed on the liquid crystal touch panel. Then, the routine advances to step SS110.

S110: Setting inputs related to the specimen mode, such as specimen number and specimen name and the like, are received on screen C. Boxes for the operator to enter the number and name of each specimen are provided on the left side of the screen C. A ten-key pad is provided on the right side of the screen C for entering numerical values and text in each box. When setting input on screen C has been completed, the process moves to step SS111.

SS111: The [Accept] key is displayed on the screen C, and the operator selection of the [Accept] key is received in step SS111. When the [Accept] key has been selected, the mode setting process ends.

In step SS2, a determination is made as to whether or not the calibration mode has been selected based on the measurement conditions input in the mode setting process of step SS1. When [calibration mode] has been selected in step SS1, the process continues to step SS3(standard solution measuring process). When [calibration mode] is not selected in step SS1, however, the process advances to step SS5(γT1) setting process).

In step SS3, the standard solutions containing known concentrations of assay material are measured.FIG. 25is a flow chart showing the flow of the standard solution measuring process. In the standard solution measuring process, step SS301(assay sample preparation process), step SS302(measuring process), and step SS303(analysis process) are sequentially performed. Steps SS301, SS302, and SS303are described below.

The operation of the assay sample preparation unit in step SS301is described referring toFIG. 20. First, the dispensing unit46suctions standard solution from the container placed in the standard solution placement unit43, and dispenses 15 μL to the container placed in the reaction unit45. Then, the dispensing unit46suctions reaction buffer from the container placed in the reagent placement unit44, and dispenses 140 μL to the container placed in the reaction unit45. Then, the dispensing unit46suctions carrier suspension from the container48placed in the reagent placement unit44, and dispenses 50 μL to the container placed in the reaction unit45. The antigen-antibody reaction is started by the addition of the carrier particle suspension. The assay sample in the container in the reaction unit45is agitated while maintained at a temperature of 37° C.

When preparing a calibration curve, a plurality of standard solutions are used which have graduatedly different concentrations of included assay material. Therefore, a plurality of T1standard solutions and T2standard solutions are placed in the standard solution placement unit43. Then, assay samples are sequentially prepared from the standard solutions of each concentration placed in the standard solution placement unit43. Thus, when assay samples are prepared from the standard solutions, the subsequent steps SS302and SS303described later are sequentially executed. Then, using the assay samples prepared from the T1standard solutions, the absorbance a at reaction time T1aand the absorbance a′ at reaction time T1a′ are calculated, and the amount of change A in absorbance (A=a′−a) during the reaction time span T1is calculated based on the absorbances a and a′.

Then, using the assay samples prepared from the T2standard solutions, the absorbance b at reaction time T2band the absorbance b′ at reaction time T2b′ are calculated, and the amount of change B in absorbance (B=b′−b) during the reaction time span T2is calculated based on the absorbances b and b′.

The operation of the measuring unit in step SS302is described below usingFIG. 21. In step SS302, the absorbance is determined during a predetermined reaction time after the antigen-antibody reaction has been started by the addition of the carrier particle suspension. The light from the light source50is diffracted into a spectrum at 800 nm by the filter51, and irradiates the container49. The diffracted light passes through the assay sample in the container49, and the transmitted light reaches the photodiode52. The photodiode52receives the transmitted light, subjects the light to photoelectric conversion, and outputs the resulting electrical signal. Each output signal is transmitted to the controller. The output signals are sent to the controller and stored in the memory unit as data. Thus, the transmitted light from the assay sample is detected at predetermined reaction times in step SS302. The predetermined reaction times are the reaction times T1a, T1a′, T2b, and T2b′ set in step SS1.

When the transmitted light is detected in step SS302, analysis is executed by the analyzing unit based on the analysis program. The operation of the analysis program in step SS303is described below using the flow chart ofFIG. 26. Each step of the flow chart is described below.

SS303-1: The data of the transmitted light signals are read from the memory unit. Then, the process advances to step SS303-2.

SS303-2: The absorbance is determined based on the transmitted light signal data. Then, the process continues to step S303-3.

SS303-3: The amount of change in absorbance in the reaction time span is calculated based on the absorbances calculated in step SS303-2. The amount of change A in absorbance at reaction time span T1is calculated based on the absorbance a at reaction time T1aand absorbance a′ at reaction time T1a′, and the amount of change B in absorbance at reaction time span T2is calculated based on the absorbance b at reaction time T2band absorbance b′ at reaction time T2b′. Then, the process continues to step SS303-4.

SS303-4: the absorbance data calculated in step SS303-2and the amount of change data calculated in step SS303-3are stored in the memory unit.

The aforesaid is shown in the flow chart of step SS3(standard solution measuring process). In this way the T1standard solutions and T2standard solutions are measured, and the amount of change in absorbance is calculated for each standard solution.

In step SS4, calibration curves are prepared based on the data of the amount of change in absorbance for each standard solution obtained in step SS3. The calibration curves prepared in step SS4are calibration curve T1prepared based on the data of the amount of change in absorbance during reaction time span T1, and calibration curve T2based on the data of the amount of change in absorbance during the reaction time span T2. The calibration curve T1is prepared based on the concentration of the assay material in the T1standard solutions input in step SS1, and the amount of change in absorbance at reaction time T1obtained by measuring the T1standard solutions in step SS3. The calibration curve T2is prepared based on the concentration of the assay material in the T2standard solutions input in step SS1, and the amount of change in absorbance at reaction time T2obtained by measuring the T2standard solutions in step SS3. The operation of the analysis program in the calibration curve preparation process is described below using the flow chart ofFIG. 27. Each step of the flow chart is described below.

SS401: The concentration of the assay material in each T1standard solution, and the data on the amount of change in absorbance during reaction time span T1calculated in step SS3are read from the memory unit. Then, the process advances to step SS402.

SS402: The calibration curve T1is prepared based on the concentrations and amount of change in absorbance. Then, the process advances to step SS403.

SS403: The calibration curve T1prepared in step SS402is stored in memory. Then, the process advances to step SS404.

SS404: The concentration of the assay material in each T2standard solution, and the data on the amount of change in absorbance during reaction time span T2calculated in step SS3are read from the memory unit. Then, the process advances to step SS405.

SS405: The calibration curve T2is prepared based on the concentrations and amount of change in absorbance. Then, the routine advances to step SS406.

SS406: The calibration curve T2prepared in step SS405is stored in memory.

The aforesaid is shown in the flow chart in step SS4(calibration curve preparation process). Thus, the calibration curve T1and calibration curve T2are prepared in this manner.

In step SS5, a threshold value γ(T1) is set for the amount of change in absorbance during the reaction time span T1. In general, when calculating concentrations using a calibration curve, the concentration range capable of calculating a reliable value is a range in which the calibration curve is linear. For this reason, value have been determined based on the lower limit value of the range ensuring the linearity of the calibration curve T1and the upper limit value of the range ensuring the linearity of the calibration curve T2, and have been stored as threshold value data in the memory unit of the apparatus beforehand. In the present step, the threshold value data are automatically read from the memory unit, and set as γ(T1). When γ(T1) is set in this way, the routine advances to step SS6(specimen mode selection determination).

In step SS6, a determination is made as to whether or not the specimen mode has been selected based on the measurement conditions input in the mode setting process of step SS1. When [specimen mode] has been selected in step SS1, the process continues to step SS7(specimen measuring process). When [specimen mode] is not selected in step SS1, however, the process advances to step SS9(output process).

In step SS7, a specimen is measured, and the concentration of the assay material contained in the specimen is calculated based on the calibration curve prepared in step SS4, and the amount of change in the absorbance.FIG. 28is a flow chart showing the flow of the specimen measuring process. In the specimen measuring process, step SS701(assay sample preparation), step SS702(T1measurement), step SS703(T1analysis), step SS704(determination whether to perform T2measurement), step SS705(T2measurement), step SS706(T2analysis), and step SS707(quantification) are sequentially performed under the conditions input in the mode setting process of step SS1.

The operation of the assay sample preparation unit is described referring toFIG. 20. The dispensing device46first suctions specimen from the container placed in the specimen placement unit42, and dispenses 15 μL of specimen into the container placed in the reaction unit45. Then, the dispensing unit46suctions reaction buffer from the container placed in the reagent placement unit44, and dispenses 10 μL to the container placed in the reaction unit45. Then, the dispensing unit46suctions carrier suspension from the container48placed in the reagent placement unit44, and dispenses 50 μL to the container placed in the reaction unit45. The antigen-antibody reaction is started by the addition of the carrier particle suspension. The assay sample in the container in the reaction unit45is agitated while maintained at a temperature of 37° C.

In step SS702, the transmitted light signals at reaction times T1aand T1a′ input in step SS1are detected. When assuming the reaction time T1ais 10 seconds and reaction time T1a′ is 60 seconds input in step SS1, the operation of the assay sample preparation unit and measuring unit are controlled by the control program such that the transmitted light is detected at reaction time T1a10 seconds after the start of the antigen-antibody reaction, and transmitted light is detected at reaction time T1a′60 seconds after the start of the antigen-antibody reaction. The operation of the measuring unit in step SS702is identical to the operation in step SS302(measuring process), that is, the transmitted light signals are detected and the detected signals are stored in the memory unit.

When the transmitted light is detected in step SS702, T1analysis is executed by the analyzing unit based on the analysis program. The operation of the analysis program in step SS703is identical to the operation in step SS303(analysis process), that is, the absorbance a detected at reaction time T1aand the absorbance a′ detected at reaction time T1a′ are calculated. Then, the amount of change A in absorbance during reaction time span T1is calculated based on absorbances a and a′. The data of the absorbances and the amount of change A in absorbance are stored in the memory unit.

SS704(Determination of Whether to Perform T2Measurement)

The amount of change A calculated in step SS703is compared to the threshold value γ(T1) set in step SS5. When the amount of change A exceeds the threshold value γ(T1), measurements are not required at reaction times T2band T2b′ since the concentration of the assay material can be determined using the amount of change A during the reaction time span T1. In this case, therefore, the temperature maintenance and agitation in the container of the reaction unit45ends in the assay sample preparation unit, and the process advances to step SS707. When the amount of change A is less than the threshold value γ(T1), measurements are required at reaction times T2band T2b′ since the concentration of the assay material cannot be determined using the amount of change A during the reaction time span T1. At this time, therefore, the process advances to step SS705. Also at this time, the temperature maintenance and agitation in the reaction unit45is continued thereafter as before until reaction time T2b′.

SS705is executed when the amount of change A is less than the threshold value γ(T1) in step SS704. In step SS705, the transmitted light signal is detected reaction time T2band reaction time T2b′ input in step SS1. When assuming the reaction time T2bis 60 seconds and reaction time T2b′ is 180 seconds input in step SS1, the operation of the assay sample preparation unit and measuring unit are controlled by the control program such that the transmitted light is detected at reaction time T2b60 seconds after the start of the antigen-antibody reaction, and transmitted light is detected at reaction time T2b′180 seconds after the start of the antigen-antibody reaction. The operation of the measuring unit at the T2measurements is identical to the operation in step SS302(measuring process), that is, the transmitted light signals are detected and the detected signals are stored in the memory unit.

When the transmitted light signals are detected in step SS705, T2analysis is executed by the analyzing unit based on the analysis program. The operation of the analysis program in step SS706is identical to the operation in step SS303(analysis process), that is, the absorbance b detected at reaction time T2b, and the absorbance b′ detected at reaction time T2b′ are calculated. Then, the amount of change B in the absorbance during reaction time span T2is calculated based on absorbances b and b′. The data of the absorbances and the amount of change B in absorbance are stored in the memory unit.

In step SS707, the concentration of assay material contained in the specimen is calculated based on the amount of change in absorbance obtained in step SS703or step SS706. When T2measurement and T2analysis are not performed, the concentration of the assay material contained in the specimen is calculated based on the amount of change A during the reaction time span T1, and the calibration curve T1. When the T2measurement and T2analysis are performed, however, the concentration of the assay material contained in the specimen is calculated based on the amount of change B during the reaction time T2, and the calibration curve T2. Then, the concentration data calculated in step SS707are stored in the memory unit.

When a plurality of specimens are assayed, step SS8is repeated until [measurement of all specimens has been completed] has been determined in step SS8. When the measurement of all specimens has been completed, the process advances to step SS9.

The data of the amount of change in absorbance of the standard solutions stored in step SS3, and the data of the concentration and amount of change in absorbance of specimens stored in step SS7are output and displayed on the liquid crystal touch panel.

The aforesaid steps are shown in the flow chart of the general control in the present embodiment. As described above, the biological sample analyzing apparatus41is an automatic analyzer that performs automatically from the preparation of the assay sample to the quantification of the assay material.

The measurement results shown were taken when the reaction time T1awas set at 10 sec, T1a′ was set at 60 seconds, and reaction time T2bwas set at 60 sec, and T2b′ was set at 180 seconds using the previously described biological sample analyzing apparatus41. In the present example, the threshold γ(T1) was set at 0.01 for the amount of change in absorbance. In this way if the amount of change A exceeds 0.01, the CRP concentration contained in the specimen is calculated based on the amount of change A at reaction time T1and the calibration curve T1. In this way if the amount of change A is less than 0.01, the CRP concentration contained in the specimen is calculated based on the amount of change B at reaction time T2.

COBAS reagent CRPLX (manufactured by Roche Diagnostics) was used in the measurement of this example. This reagent is provided in a reagent kit, which includes latex reagent containing latex particle on which anti-human CRP mouse monoclonal antibody is immobilized and TRIS buffer solution. The buffer solution was used as the reaction buffer, and the latex reagent was sued as the carrier particle suspension.

The standard solution was LZ test for CRP standard serum Eiken' manufactured by Eiken Chemical Co., Ltd. The standard solutions contained CRP concentrations of 0, 0.5, 4, 12, 22, and 30 mg/dL. CRP concentrations of 0, 0.5, 4, and 12 mg/dL were used in T1standard solutions, and CRP concentrations of 0, 0.5, 4, 12, 22, and 30 mg/dL were used in T2standard solutions.

In the present example, plasmas 1˜16 containing various concentrations of CRP were used as specimens.

These same specimens were measured to determine concentration of CRP contained in the specimens using a model TBA-80FR NEO2 Clinical Autochemical Analyzer manufactured by Toshiba Corporation. CRP latex (II) ‘Seiken’ X2 manufactured by Denka Seiken Co., Ltd. was used as the reagent. (Hereinafter measurements using the TBA-80FR NEO2 are referred to as ‘comparative measurements’.) In the comparative measurements, absorbance was measured at a reaction times of 50 seconds and 158 seconds, and the amount of change in absorbance was calculated during the time 50 seconds after to the start of the reaction to 158 seconds after the start of the reaction based on these absorbances.

Table 3 shows the results obtained by measuring each specimen using the biological sample analyzing apparatus41. In Table 3, column [A] shows the values of amount of change A during reaction time span T1, and column [B] shows the values of amount of change B during reaction time span T2. [Calibration curve] indicates the calibration curve used when calculating concentration, T1indicating Calibration curve T1was used, and T2indicating calibration curve T2was used. [Concentration I] indicates the CRP concentration (mg/dL) obtained by measurements using the biological sample analyzing apparatus41, and [concentration II] indicating CRP concentration (mg/dL) obtained by measurements of the comparative examples.

According to Table 3, the CRP concentrations calculated using the measurements of the biological sample analyzing apparatus41approximate the concentrations determined by the comparative measurements. In the biological sample analyzing apparatus41, the CRP concentration calculation for specimens with relatively high CRP concentrations (sera 8˜16) is based on the amount of change A during reaction time span T1. It can also be understood from Table 3 with regards to specimens having relatively high CRP concentrations (sera 8˜16), that the CRP concentrations calculated using the measurements of the biological sample analyzing apparatus41approximate the concentrations determined using the comparative measurements. Accordingly, when measuring specimens with high concentrations, the biological sample analyzing apparatus41can calculate concentrations faster than calculations by the comparative measurements.

The measurement results indicated here are from whole blood measurements using the biological sample analyzing apparatus41described above.

When whole blood was used directly as a specimen, there was concern that the measurement would be affected by the hemocyte component (mainly red blood cells) contained in the whole blood, such that an accurate measurement result could not be obtained. In the present example, a hemolytic agent, STROMATOLYSER-WH, manufactured by Sysmex Corporation, was diluted three fold using a dilution fluid also manufactured by Sysmex corporation, and this was mixed with whole blood in a hemolytic process to obtain a specimen used for measurement.

When whole blood is used as a specimen in the same quantity as when plasma and serum is used for measurement, the hemocyte component (hemocyte volume) is reflected in the low measurement values. Therefore, the hemocyte volume must be compensated when measurements are performed using whole blood. In the present example, this compensation is accomplished by the following method. Specifically, whole blood specimens were measured using a model XE-2100 fully automatic hemocyte analyzer manufactured by Sysmex corporation to determine the hemocyte count, and the hematocrit value was determined based on this hemocyte count. (The hematocrit value represents the percentage volume of red blood cells in a constant volume of whole blood.) The equation below was then used for the compensation.
C=C0/{1−(H/100)}

(Where C represents the compensated concentration of the assay material, Co represents the concentration of the assay material obtained by measuring whole blood, and H represents the hematocrit value (%).)

The reaction time in the measurements of the present example were T1a=10 seconds, T1a′=60 seconds, T2b=60 seconds, and T2b′=180 seconds. Each type of reagent and standard solution used in the present example were identical to the reagents and standard solutions used in measurement example 5. Furthermore, in the present example, whole blood 1˜11 containing various concentrations of CRP were prepared, and 100 μL of three-fold diluted STROMATOLYSER-WH was added to 20 μL whole blood to obtain usable specimens. For comparison purposes in the present example, plasma 1˜11 were prepared by centrifuging (8,000 rpm for 5 minutes) the whole blood 1˜11, and 100 μL three-fold diluted STROMATOLYSER-WH was added to 20 μL of the plasma to obtain usable specimens.

Table 4 shows the results obtained by measuring each specimen using the biological sample analyzing apparatus41. [Calibration curve] indicates the calibration curve used when calculating concentration, T1indicating calibration curve T1was used, and T2indicating calibration curve T2was used. [Concentration] indicates the CRP concentration (mg/dL) obtained by measurements using the biological sample analyzing apparatus41.

When the CRP concentrations of the whole blood specimens and plasma specimens of the same reference numbers (Whole blood 1 and plasma 1, whole blood 2 and plasma 2, whole blood 3 and plasma 3) in Table 4 were compared, it was observed that the whole blood CRP concentrations approximated the plasma CRP concentrations in all cases. This indicated that whole blood could also be measured.

In the present embodiment, measurements were performed during different reaction time bands (reaction time span T1and reaction time span T2). When the reaction time T1a, T1a′, T2b, and T2b′ are set such that T1a<T1a′ and T2b<T2b′, and T1a<T2band T1a′<T2b′, the reaction during the reaction time span T2is generally comparatively stable compared to the reaction during reaction time span T1. Therefore, the measurements during the reaction time span T2, which has a relatively stable reaction, have between reproducibility and sensitivity than measurements during the reaction time span T1, which has a relatively unstable reaction. For this reason, the calibration curve T2during the reaction time span T2is particularly useful when calculating the concentration of assay material of low concentration. Measurement during the reaction time span T1, however, are less easily affected by the zone phenomenon than measurements during the reaction time span T2. For this reason, the calibration curve T1during the reaction time span T1is particularly useful when calculating the concentration of assay material of high concentration.

In the present embodiment, predetermined conditions are provided related to measurement results during the reaction time span T1, such that the calibration curve T1is used when calculating the concentration of assay material that has high concentration, and the calibration curve T2is used when calculating the concentration of assay material that has low concentration. For this reason, measurements at both a first reaction time span (reaction time span T1) and a second reaction time span (reaction time span T2) are not always necessary, and measurements during the reaction time span T2are performed only when the measurement result during the reaction time span T1do not satisfy the predetermined conditions. In this way measurements can be accomplished more efficiently on a time basis.

In the present embodiment, the reaction time span T1and reaction time span T2can be set by the operator in accordance with the measurement range required. In this way measurement can be accomplished more efficiently without measuring over a time period longer than necessary.

Although reaction time T1a=10 seconds, T1a′=60 seconds, T2b=60 seconds, and T2b′=180 seconds in the above embodiments, the reaction times are not limited to these settings. The times of suitable reaction times T1a, T1a′, T2b, T2b′ may differ depending on the assay material. For this reason, suitable reaction times T1a, T1a′, T2b, T2b′ may be set in accordance with the assay material to be measured.

Although the transmitted light is detected by illuminating an assay sample with light having a wavelength of 800 nm in the above embodiments, light of an optimum wavelength may be selected and used for measurement in accordance with the measurement conditions and reagents.

Although serum, plasma, and whole blood collected from humans are used as specimen in the two embodiments, the present invention is not limited to these. In the present invention, other biological sample, such as urine and the like, may be used as specimen.

Latex on which anti-CRP antibody is immobilized is used as the carrier particle in the above two embodiments, the present invention is not limited to this arrangement. Other carrier particles may be used insofar as such carrier particles on which antibody or antigen against the assay material is immobilized. The antibody or antigen immobilized on carrier particles are not specifically limited insofar as they are detectable using an antigen-antibody reaction. The method for immobilization of antigen or antibody on carrier particle may be well known methods in the art. For example, physical absorption methods, chemical bonding methods and the like may be used.

Although CRP is detected as the assay material in the above two embodiments, the present invention is not limited to this arrangement. Other materials may be detected as assay materials insofar as such assay materials are detectable in immunoassays using carrier particles. Examples of assay materials include immunoglobulin (IgG, IgA, IgM, IgD, IgE) complements (C3, C4, C5, C1q), α-fetoprotein (AFP), β2-microglobulin, carcinoembryonic antigen (CEA), prostate specific antigen (PSA), anti HCV antibody, insulin, ferritin and the like.

Although the amount of change in absorbance obtained by immunoturbidity and the agglutination rate in CIA are used as information related to the assay material in the above two embodiments, it is to be noted that the information related to the assay material is not limited to such information. For example, since transmitted light, absorbance, scattered light obtained by illuminating an assay sample with light, or the amount of change in these parameters per predetermined unit time varies depending on the degree of agglutination in the assay sample, information related to the assay material in the assay sample may be used.

Although the immunoturbidity method employing carrier particles and CIA is used as the method for quantifying assay material in the above two embodiments, the present invention is not limited to these methods. For example, immunonephelometry employing carrier particles also may be used. Furthermore, immunoturbidity methods that do not use carrier particles and immunonephelometry methods that do not use carrier particles also may be used.

Although data of the threshold values α(T1) and γ(T1) stored beforehand in the memory unit of the apparatus, and data stored when setting the threshold values are automatically read and set as the threshold values in the above two embodiments, the present invention is not limited to this arrangement. For example, a program capable of automatically setting threshold values based on calibration curve data may be installed in the apparatus, the program may be automatically started when setting a threshold, the threshold value may then be automatically calculated based on the data such as a calibration curve, and the calculated value can be set as a threshold value.