WO2012176596A1 - 核酸増幅装置及び核酸分析装置 - Google Patents
核酸増幅装置及び核酸分析装置 Download PDFInfo
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- WO2012176596A1 WO2012176596A1 PCT/JP2012/064037 JP2012064037W WO2012176596A1 WO 2012176596 A1 WO2012176596 A1 WO 2012176596A1 JP 2012064037 W JP2012064037 W JP 2012064037W WO 2012176596 A1 WO2012176596 A1 WO 2012176596A1
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L7/00—Heating or cooling apparatus; Heat insulating devices
- B01L7/52—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
- B01L2200/14—Process control and prevention of errors
- B01L2200/148—Specific details about calibrations
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L2300/00—Additional constructional details
- B01L2300/02—Identification, exchange or storage of information
- B01L2300/024—Storing results with means integrated into the container
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L2300/00—Additional constructional details
- B01L2300/08—Geometry, shape and general structure
- B01L2300/0809—Geometry, shape and general structure rectangular shaped
- B01L2300/0829—Multi-well plates; Microtitration plates
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L2300/00—Additional constructional details
- B01L2300/18—Means for temperature control
- B01L2300/1805—Conductive heating, heat from thermostatted solids is conducted to receptacles, e.g. heating plates, blocks
- B01L2300/1822—Conductive heating, heat from thermostatted solids is conducted to receptacles, e.g. heating plates, blocks using Peltier elements
Definitions
- the present invention relates to a nucleic acid amplification device equipped with a plurality of temperature control blocks that can be individually temperature controlled, and a nucleic acid analyzer that uses the nucleic acid amplification device as a part of the device.
- the temperature control block needs to accurately control the absolute temperature value to the target temperature.
- a calibrated temperature measurement probe is used to measure the temperature absolute value of the temperature control block, and if the measured temperature is different from the target temperature, a method of correcting the measured temperature to match the target temperature has been adopted. Has been.
- the limit accuracy of temperature correction using a calibrated temperature measurement probe is ⁇ 0.25 ° C.
- a plurality of temperature control blocks that can be individually controlled in temperature may be mounted on one apparatus. Also in this case, if the temperature of each temperature control block is individually corrected using a calibrated temperature measurement probe, the temperature control accuracy of each temperature control block after the correction is ⁇ 0.25 ° C. Therefore, the temperature difference between the temperature control blocks is theoretically 0.5 ° C. at the maximum.
- thermochromic liquid crystal a method using a test piece using a thermochromic liquid crystal has been proposed for temperature correction of the temperature control block (see, for example, Patent Document 1).
- the method is based on the principle that the temperature of the test piece is calibrated by detecting the color change of the liquid crystal when the test piece is controlled to the test temperature by mixing the thermochromic liquid crystal into the test piece in contact with the fluid sample. To do.
- the PCR method includes (1) a heat denaturation temperature of 95 ° C., (2) an annealing temperature of about 55 ° C. to 65 ° C., and (3) an extension reaction temperature alternately n times (hereinafter referred to as “temperature cycle”). ) To amplify the target nucleic acid sequence 2n times.
- the annealing temperature and extension reaction temperature vary depending on the target sequence, and the required accuracy of the temperature is generally within ⁇ 0.5 ° C. However, the higher the accuracy and reproducibility of temperature, the better the amplification efficiency and amplification reproducibility of DNA.
- the real-time PCR method refers to a method of measuring amplification by PCR in real time by using a fluorescent dye and quantifying the amount of nucleic acid in the reaction solution from the amplification rate (number of cycles).
- a real-time PCR apparatus in which a plurality of reaction vessels (wells) are arranged in a space where temperature is controlled by one temperature control block is used.
- the temperature difference between the plurality of reaction vessels (wells) can be suppressed to ⁇ 0.2 ° C. or less.
- This real-time PCR device can individually control the temperature of a plurality of temperature control blocks, and can simultaneously execute a plurality of temperature cycles.
- this type of apparatus also requires a temperature control accuracy equivalent to that in the case where only one temperature control block is used between a plurality of reaction vessels (wells).
- HRM analysis refers to the temperature of the amplified product amplified by the real-time PCR method in the range of about 60 ° C to 95 ° C with a resolution of 0.1 ° C or less, and the melting temperature of the amplified product (the two amplified products) This is an analysis method for determining the melting temperature of the double-stranded bond of the strand nucleic acid.
- the melting temperature is different for each amplified sequence, and theoretically, it is known that even a difference of 1 base differs.
- nucleic acids can be separated and detected for each sequence from an amplification reaction solution in which a plurality of different sequences are mixed.
- the conventional temperature correction method using a calibrated temperature measurement probe has a temperature difference of 0.5 ° C. at the maximum, and it is difficult to satisfy the above-described requirements.
- nucleic acid fragment amplified by the PCR method was collected and mixed well in the same container, and then the mixed solution was dispensed into 96 reaction containers (wells) as shown in FIG. 1A.
- the nucleic acid fragment was subjected to HRM analysis using a real-time PCR apparatus having a temperature uniformity between reaction vessels (wells) of 0.05 ° C. or less to determine the melting temperature.
- FIG. 1B the inventors discovered that the variation in melting temperature of nucleic acid fragments amplified by PCR was very small, and the variation was within ⁇ 0.05 ° C.
- the inventors utilize this discovery, a plurality of temperature control blocks that can be individually temperature controlled, a real-time fluorescence measurement unit that performs fluorescence measurement in real time on a sample in a reaction vessel temperature-controlled by each temperature control block, A storage unit for storing a reference melting temperature of a temperature calibration sample dispensed in one or a plurality of reaction vessels whose temperature is controlled in each temperature control block, and a temperature stored in each reaction vessel corresponding to each temperature control block.
- the melting temperature measurement unit that measures the melting temperature of the calibration sample as the measured melting temperature, the measured melting temperature corresponding to each temperature control block and the reference melting temperature are compared, and the temperature of each temperature control block is based on each difference value.
- a temperature correction unit that corrects the absolute value is mounted on the nucleic acid amplification device.
- the nucleic acid amplification device having the above configuration is mounted on the nucleic acid analyzer.
- temperature uniformity among a plurality of temperature control blocks that can be individually temperature controlled can be realized with the same accuracy as the temperature control resolution.
- FIG. 2 shows a functional block configuration of the nucleic acid amplification device according to the embodiment.
- the nucleic acid amplification apparatus shown in FIG. 2 includes a plurality of temperature control blocks 1 that can be individually temperature controlled, a real-time fluorescence measurement unit 3, and a control unit 5 that controls them.
- a mechanism part including a plurality of temperature control blocks 1 and a real-time fluorescence measurement unit 3 that can individually control the temperature is referred to as a real-time fluorescence measurement mechanism 15.
- the base body of the temperature control block 1 is formed of a material having excellent thermal conductivity, and the reaction vessel is accommodated in a holding mechanism formed on the base body.
- a temperature sensor and a heat source are also arranged on the base.
- the substrate for example, copper, aluminum, and various alloys are used.
- the temperature sensor uses a thermistor, a thermocouple, a resistance temperature detector, or the like. The temperature sensor is disposed in the vicinity of the holding mechanism of the reaction container in order to measure the temperature of the sample in the reaction container.
- a Peltier element is used as the heat source.
- the Peltier element is a thermoelectric element and is used for heating or cooling the substrate.
- a radiation fin is disposed on the base.
- temperature control it is not necessary to mount a heat source in the temperature control block 1.
- the plurality of temperature control blocks 1 are arranged on a pedestal made of a material having excellent heat insulation properties such as plastic. Accordingly, the propagation of temperature from one temperature control block 1 to another temperature control block 1 can be ignored. That is, the mutual interference of temperatures between the plurality of temperature control blocks can be ignored.
- the real-time fluorescence measurement unit 3 measures the sample in the reaction container whose temperature is controlled by each temperature control block 1 in real time.
- the sample is fluorescently labeled.
- the real-time fluorescence measurement unit 3 includes an excitation light source that generates excitation light that irradiates the reaction container, and a fluorescence detector that measures fluorescence generated from the sample irradiated with the excitation light.
- a light emitting diode (LED), a semiconductor laser, a xenon lamp, a halogen lamp, or the like is used as the excitation light source.
- a photodiode, a photomultiplier, a CCD, or the like is used as the fluorescence detector.
- the control unit 5 executes temperature control of each temperature control block 1, processing of measurement data of the real-time fluorescence measurement unit 3, and the like.
- the control unit 5 stores the melting temperature of the temperature calibration sample used when correcting the temperature of each temperature control block 1 in the storage unit 11.
- the melting temperature here (hereinafter also referred to as “reference melting temperature”) is taken into the control unit 5 through various routes, as will be described later.
- the control unit 5 has a melting temperature measurement unit 7 and a temperature correction unit 9 as functions used when correcting the temperature of each temperature control block 1.
- the melting temperature measurement unit 7 measures the melting temperature of the temperature calibration sample accommodated in each reaction container corresponding to each temperature control block 1.
- the measurement of the melting temperature is determined as the measurement temperature of the temperature sensor (hereinafter also referred to as “measured melting temperature”) when the real-time fluorescence measurement unit 3 detects the melting of the temperature calibration sample.
- the measured melting temperature is given from the melting temperature measuring unit 7 to the temperature correcting unit 9.
- the temperature correction unit 9 compares the measured melting temperature with the reference melting temperature stored in the storage unit 11, and corrects the temperature absolute value of each temperature control block 1 so that the difference is eliminated. The difference between the measured melting temperature and the reference melting temperature detected for each temperature control block 1 gives the temperature control accuracy of the temperature control block 1.
- FIG. 3 shows a specific example of a nucleic acid amplifier that corrects the absolute temperature values of a plurality of temperature control blocks 1 using one type of temperature calibration sample.
- the device configuration shown in FIG. 3 represents the device configuration shown in FIG. 2 in more detail.
- the turntable 22 here corresponds to the base of the first embodiment.
- the turntable 22 is made of a material having excellent heat insulation. Accordingly, the mutual interference of temperatures between the plurality of temperature control blocks can be ignored.
- the turntable 22 is fixed with respect to a rotating shaft (not shown), and can freely rotate both in the clockwise direction and in the counterclockwise direction as indicated by an arrow.
- the rotating shaft is rotationally driven by a stepping motor (not shown).
- reaction vessel 21 is made of a member that is transparent to light in the fluorescence wavelength band, and is attached so that its bottom is exposed from the back side of the turntable 22.
- the real-time fluorescence measurement unit 3 is disposed on the back side of the turntable 22 and irradiates the bottom of the reaction vessel 21 with excitation light generated from an excitation light source.
- the real-time fluorescence measurement unit 3 detects fluorescence generated in the sample in the reaction vessel 21 irradiated with the excitation light with a fluorescence detector, and outputs the fluorescence intensity to the data processing unit 23 as fluorescence measurement data.
- the data processing unit 23 processes the fluorescence measurement data sequentially input from the fluorescence detector and the temperature data measured by the temperature sensor, and outputs the data to the storage / calculation unit 24.
- the storage / calculation unit 24 is composed of, for example, a general-purpose computer, and executes analysis processing for analyzing the melting temperature of each temperature block 1 and calculation processing for calculating a correction value.
- the storage / calculation unit 24 sets the measurement temperature at the time when the melting of the temperature calibration sample is detected from the fluorescence measurement data as the measurement melting temperature.
- the storage / calculation unit 24 calculates a correction value for the absolute temperature value based on the difference value between the measured melting temperature and the reference melting temperature.
- the measured temperature of the temperature control block 1 is also given to the device control unit 25.
- the reference melting temperature is stored in advance in the storage / calculation unit 24.
- the device control unit 25 controls each temperature control block 1 to a target temperature so that a temperature change necessary for real-time fluorescence detection can be obtained. Specifically, the heat generation amount of the heat source mounted on the temperature control block 1 is controlled. At this time, the apparatus control unit 25 acquires the measured temperature from the temperature sensor mounted on the temperature control block 1 and performs feedback control so that the measured temperature matches the target temperature. The measured temperature here is also given to the storage / calculation unit 24 as described above.
- the apparatus control part 25 changes the temperature of the reaction container 21 in the range of at least 50 degreeC to 95 degreeC in the case of real-time fluorescence detection.
- the data processing unit 23, the storage / calculation unit 24, and the device control unit 25 in FIG. 3 correspond to the control unit 5 in FIG. In FIG. 3, the data processing unit 23, the storage / calculation unit 24, and the device control unit 25 are shown as independent devices, but may be configured as one device.
- the temperature control block 1 is mounted on the outer edge of the turntable 22, and fluorescence is detected when the turntable 22 rotates and the temperature control block 1 passes in front of the real-time fluorescence measurement unit 3.
- fluorescence is detected when the turntable 22 rotates and the temperature control block 1 passes in front of the real-time fluorescence measurement unit 3.
- the pedestal side on which the temperature control block 1 is mounted may be fixed and the real-time fluorescence measurement unit 3 side may be rotated or moved. In this case, fluorescence detection is performed when the real-time fluorescence measurement unit 3 passes a position facing the temperature control block 1.
- FIG. 4 shows another device configuration example of the nucleic acid amplification device that corrects the temperature of the temperature control block 1 using one type of temperature calibration sample.
- the apparatus configuration shown in FIG. 4 also represents the apparatus configuration shown in FIG. 2 in more detail.
- the number of the real-time fluorescence measurement units 3 is smaller than the number of the temperature control blocks 1. For this reason, the structure which fixed either one of the base which mounts the temperature control block 1, and the real-time fluorescence measurement part 3, and rotationally controlled the other was employ
- the temperature control block 1 and the real-time fluorescence measurement unit 3 correspond one-to-one and a plurality of these are provided, the apparatus configuration shown in FIG. 4 is possible.
- the temperature control block 1 represents an example in which a reaction plate 26 in which a plurality of reaction vessels 21 are arranged in a matrix is placed.
- reaction vessel 21 and the reaction plate 26 may be of any material and shape as long as they can transmit the fluorescence wavelength and can conduct the heat of the temperature control block 1. More preferably, it is desirable to use a PCR tube (Gleiner, Germany) or a 96-well PCR plate that does not contain DNase or RNase.
- the temperature calibration sample in the present embodiment only needs to contain a nucleic acid fragment capable of HRM analysis and a detection dye.
- a nucleic acid fragment capable of HRM analysis and a detection dye.
- DNA, RNA, or PNA can be used as the nucleic acid fragment. More preferably, a sample obtained by amplifying any one kind of nucleic acid fragment by the PCR method is used. More preferably, a nucleic acid fragment in which the coincidence between the absolute temperature of the aqueous solution and the melting temperature is confirmed is used.
- nucleic acid fragments capable of HRM analysis and a detection dye it is only necessary to include two types of nucleic acid fragments capable of HRM analysis and a detection dye. More preferably, amplification products obtained by individually amplifying two types of nucleic acid fragments by PCR are used.
- the temperature calibration sample is preferably contained in a container having a barcode attached to the outer wall.
- the barcode information includes at least melting temperature information of the temperature calibration sample.
- a barcode may be attached to the reaction plate 26 itself.
- the bar code information includes at least the melting temperature information of the temperature calibration sample.
- the nucleic acid amplification device corrects variations in temperature absolute values existing between a plurality of temperature control blocks through the following three processing steps.
- a sample containing a nucleic acid fragment having a predetermined melting temperature amplified by a nucleic acid amplification method such as a PCR method is dispensed into a plurality of reaction containers as a temperature calibration sample. Then, this reaction container is installed or installed in the holding mechanism of the several temperature control block 1 in the nucleic acid amplifier which carries out temperature calibration. Alternatively, the temperature calibration sample is dispensed into a reaction vessel previously installed or installed on the holding mechanism of the plurality of temperature control blocks 1.
- the melting temperature of the temperature calibration sample is actually measured for each temperature control block 1.
- a known method is applied to the measurement of the melting temperature here.
- the fluorescence intensity is measured in real time while changing the temperature of the temperature control block 1 from a low temperature (eg, 60 ° C.) to a high temperature (eg, 95 ° C.).
- the fluorescence intensity is measured by changing the temperature with a temperature resolution equal to or higher than the temperature accuracy required for the nucleic acid amplification device. For example, if the temperature accuracy required for the nucleic acid amplifier is ⁇ 0.1 ° C or less, the target temperature is varied in increments of 0.1 ° C or less, and the melting temperature is measured by fluorescence.
- the control unit 5 corrects the absolute temperature value managed for each temperature control block 1 so that the measured value of the melting temperature matches the melting temperature stored in the storage unit 11. . Note that measurement error may occur in the measurement of the melting temperature. Therefore, in a more preferred embodiment, it is desirable to repeat step 2 and step 3 twice or more to improve temperature uniformity between the temperature control blocks.
- FIG. 5 shows a melting temperature information confirmation processing procedure executed prior to temperature correction.
- the description will be made assuming that the storage / calculation unit 24 executes the processing. But you may perform using the other control part which comprises a nucleic acid amplifier, and the external control part connected to a nucleic acid amplifier.
- the storage / calculation unit 24 tries to obtain melting temperature information of the temperature calibration sample via the network (step S1). If available, the melting temperature information is read out via the network and stored in a predetermined storage area (step S2).
- the network here includes the Internet as well as the LAN.
- the storage / calculation unit 24 requests the user to input melting temperature information (step S3).
- the user inputs the melting temperature using bar code input or the like in addition to keyboard input.
- the storage / calculation unit 24 stores the input melting temperature information in a predetermined storage area (step S4).
- FIG. 6 shows the entire temperature correction operation including the melting temperature information confirmation process (FIG. 5).
- the storage / calculation unit 24 is described as executing a series of processes. However, a series of other control units constituting the nucleic acid amplification device or an external control unit connected to the nucleic acid amplification device is a series. Processing may be executed.
- the storage / calculation unit 24 confirms melting temperature information of the temperature calibration sample (step S11).
- the processing operation shown in FIG. 5 is executed.
- the storage / calculation unit 24 measures the melting temperature of the installed temperature calibration sample (step S12). Specifically, the temperature of the temperature control block 1 is changed from a low temperature (for example, 60 ° C.) to a high temperature (for example, 95 ° C.) in predetermined increments, and the fluorescence intensity emitted from the temperature calibration sample at that time is measured in real time. To do. When detecting the melting of the sample from the fluorescence intensity, the storage / calculation unit 24 stores the measured temperature at that time in the predetermined storage area as the measured melting temperature.
- a low temperature for example, 60 ° C.
- a high temperature for example, 95 ° C.
- the storage / calculation unit 24 compares the reference melting temperature confirmed in advance with the measured melting temperature of each temperature control block 1 (step S13). At this time, the storage / calculation unit 24 calculates a difference value between the reference melting temperature confirmed in advance and the measured melting temperature of each temperature control block 1.
- the storage / calculation unit 24 uses the difference value calculated for each temperature control block 1 so that the measured melting temperature of each temperature control block 1 becomes the reference melting temperature of the temperature calibration sample. The value is corrected (step S14).
- the following is obtained by measuring the melting temperature with a resolution of 0.1 ° C or lower when using the temperature control block 1 that can set the temperature with a resolution of 0.1 ° C or lower and the temperature calibration sample with a melting temperature of 87.3 ° C.
- the melting temperature curve will be explained.
- the melting temperature of the sample accommodated in each reaction container is determined as the temperature at the fluorescence intensity value (0.2) having the largest decay rate (the amount of decrease in fluorescence intensity per unit time).
- the method for determining the melting temperature is not limited to this method, and for example, the analysis method shown in Non-Patent Document 1 can also be used.
- FIG. 7A shows a measurement example of melting temperature curves measured for a plurality of temperature control blocks 1 when no temperature correction is performed.
- the maximum temperature difference between the plurality of temperature control blocks was 1.7 ° C.
- the measured values are plotted with the horizontal axis as temperature and the vertical axis as fluorescence intensity. The same applies to FIGS. 7B and 7C.
- FIG. 7B shows a measurement example of melting temperature curves measured for a plurality of temperature control blocks 1 when the temperature absolute value of each temperature control block 1 is corrected based on the melting temperature of 87.3 ° C.
- the maximum temperature difference between the plurality of temperature control blocks converges to 0.1 ° C. or less.
- FIG. 7C shows a measurement example of melting temperature curves measured for a plurality of temperature control blocks 1 when a conventional temperature setting method using a calibrated temperature probe is used.
- the maximum temperature difference between the plurality of temperature control blocks is 0.53 ° C.
- the nucleic acid amplification apparatus is equipped with a function for correcting the temperature absolute value of the temperature control block 1 using the temperature calibration sample.
- the maximum temperature difference between the plurality of temperature control blocks can be made uniform with the same accuracy as the temperature control resolution of each temperature control block 1.
- the maximum temperature difference between the plurality of temperature control blocks can be uniformized to ⁇ 0.05 ° C. or less. That is, the temperature difference between the temperature control blocks can be corrected to the same extent as the temperature control resolution in each temperature control block. For this reason, even when nucleic acid amplification is performed using a plurality of temperature control blocks, the influence of the difference in temperature control blocks on the analysis accuracy can be ignored.
- a part having a large change rate at which the fluorescence intensity changes abruptly (a part having the largest fluorescence intensity decay rate (in the case of FIG. 7B, the fluorescence intensity value is 0.2)) is determined as the melting temperature. .
- the melting temperature may be determined using other methods.
- the melting temperature may be determined using a measurement curve in which temperature is plotted on the horizontal axis and the change rate of fluorescence intensity is plotted on the vertical axis. Specifically, the temperature at which the change in fluorescence intensity is the largest may be determined as the melting temperature. In this case, the melting temperature can be specified more clearly.
- 8A corresponds to FIG. 7A
- FIG. 8B corresponds to FIG. 7B.
- the detection of the melting temperature is executed by the storage / calculation unit 24.
- Example 3 the temperature at which the measured decay rate of the fluorescence intensity is the highest or the temperature at which the change rate of the fluorescence intensity is the largest is used as the “melting temperature”.
- any determination method may be used. . That is, the method is not limited to the method for determining the melting temperature from the measurement curve of the temperature calibration sample.
- Example 4 In the above-described embodiment, the temperature calibration method using the temperature calibration sample having one melting temperature has been described. If the temperature characteristics of each temperature control block 1 are the same so that the temperature characteristics of each temperature control block 1 can be ignored even with a single temperature calibration sample, the maximum temperature difference of the temperature control block 1 is uniform to ⁇ 0.05 ° C or less for temperatures other than the melting temperature. Can be
- each temperature control block 1 generally has a unique temperature characteristic. Therefore, in the present embodiment, by adopting the following method, the temperature absolute values between the plurality of temperature control blocks are made uniform even when controlling the plurality of temperature control blocks 1 that can be individually controlled to an arbitrary temperature. .
- each temperature control block 1 when the temperature characteristics of each temperature control block 1 are measured in advance and stored in the storage / calculation unit 24 and the temperature is controlled to a temperature other than the melting temperature, the temperature characteristics and the melting temperature are used.
- the temperature of each temperature control block 1 is controlled based on the control error.
- the temperature characteristics may be measured in the temperature range used for nucleic acid amplification.
- the target temperature may be set from about 50 ° C. to about 100 ° C.
- FIG. 9 shows an example of actually measured temperature characteristics.
- FIG. 9 shows the relationship between the measured temperatures of the temperature control block 1 measured by the temperature sensor when the target temperature of each temperature control block 1 is varied in increments of 1 ° C.
- the vertical axis in FIG. 9 is the measured temperature, and the horizontal axis is the target temperature.
- the temperature characteristic unique to each temperature control block 1 can be defined by the slope and intercept of a straight line.
- the temperature characteristics specific to each temperature control block 1 are measured and stored in the storage area, and the error between the target temperature and the measured temperature with respect to the melting temperature is corrected. It can be accurately controlled at any temperature. That is, for any temperature other than the melting temperature, the temperature absolute value can be made uniform among the plurality of temperature control blocks.
- each temperature control block 1 can be controlled to an arbitrary absolute value by using the measured temperature characteristic as it is.
- Example 5 a temperature correction function having a function of evaluating the accuracy of the corrected temperature will be described.
- the measured melting temperature of the temperature control block 1 should match the reference melting temperature (strictly, the difference should be below the same level as the temperature control resolution).
- the temperature correction function described below is proposed.
- FIG. 10 shows an example of a processing procedure corresponding to the temperature correction function.
- the storage / calculation unit 24 performs the above-described reference melting temperature confirmation process (step S21). This processing operation is the same as steps S1 to S4 shown in FIG.
- step S22 the storage / calculation unit 24 performs temperature correction of each temperature control block 1 (step S22).
- This processing operation is the same as steps S12 to S14 shown in FIG. Specifically, the target temperature of the temperature control block 1 is varied from around 50 ° C. to around 100 ° C., and the melting temperature is measured and the absolute temperature value is corrected.
- the storage / calculation unit 24 remeasures the melting temperature of the temperature calibration sample for each temperature control block 1. This operation is performed automatically.
- the storage / calculation unit 24 determines whether or not the temperature difference between the measured melting temperature and the reference melting temperature is within the target accuracy (whether or not it is equal to or less than a determination threshold) (step S23).
- the threshold value for giving the target accuracy may be set in advance by the user or may be assigned as an initial value.
- the storage / calculation unit 24 displays the accuracy that each temperature control block 1 satisfies, and ends the correction operation (step S24).
- the accuracy the temperature absolute value measured for each temperature control block 1 after correction and the accuracy between wells are also displayed.
- step S23 if it is determined in step S23 that the temperature difference exceeds the target accuracy, the storage / calculation unit 24 compares the temperature correction count (repetition count) that has already been executed with a threshold (step S25).
- the threshold value here gives the upper limit value of the number of corrections of the melting temperature measurement and the temperature absolute value of each temperature control block 1.
- the threshold value may be set in advance by the user or may be assigned as an initial value.
- step S25 When there is a temperature control block 1 for which a negative result is obtained in the determination process of step S25 (that is, when the number of temperature corrections for the corresponding temperature control block has not reached a predetermined threshold), the storage / calculation unit 24 The process returns to step S22.
- the storage / calculation unit 24 executes a temperature absolute value correction process based on the temperature difference between the measured melting temperature value and the original melting temperature for each temperature control block 1. When there is a temperature control block that does not fall within the target accuracy, a series of operations are repeatedly executed until the specified number of repetitions is reached.
- the storage / calculation unit 24 determines that the corresponding temperature control block 1 is identified and an alarm indicating temperature control abnormality is displayed (step S26). Also in this case, the storage / calculation unit 24 displays the temperature absolute value measured for each corrected temperature control block 1 and the accuracy between wells.
- normal operation means all operations that can be performed by the nucleic acid amplification apparatus, in addition to the apparatus operation for temperature calibration.
- nucleic acid amplification apparatus equipped with a function for correcting temperature absolute values of a plurality of temperature control blocks 1 that can individually control the temperature.
- the basic configuration of the nucleic acid amplification device according to this embodiment is the same as that of the nucleic acid amplification device described in Embodiment 1.
- the two types of temperature calibration samples in this embodiment need only be separated from each other by at least 5 ° C. More preferably, a nucleic acid fragment having a melting temperature of about 60 ° C. (eg, 50 ° C. to 70 ° C.) is used as the first type of temperature calibration sample, and a melting temperature of about 90 ° C. (eg, 80 ° C.) is used as the second type of temperature calibration sample. ( ⁇ 100 ° C) nucleic acid fragment.
- Each temperature calibration sample may measure the melting temperature separately, or two melting temperatures may be measured simultaneously by measuring the melting temperature once using a mixture of each temperature calibration sample.
- FIG. 11 shows a processing procedure example corresponding to a temperature correction function using N (N ⁇ 2) temperature calibration samples.
- N N ⁇ 2
- FIG. 11 shows a processing procedure example corresponding to a temperature correction function using N (N ⁇ 2) temperature calibration samples.
- the storage / calculation unit 24 performs confirmation processing of all N melting temperature information (step S31). This processing operation is the same as steps S1 to S4 shown in FIG. 5 except that the number of melting temperatures to be confirmed is N.
- the temperature of the control block 1 is actually measured (step S32).
- the storage / calculation unit 24 compares the actually measured melting temperature with the original melting temperature of the i-th temperature calibration sample (step S33). Further, the storage / calculation unit 24 displays information on the original melting temperature acquired in advance for the i-th temperature calibration sample (step S35).
- the storage / calculation unit 24 corrects the absolute temperature value of each temperature control block 1 so that the actually measured melting temperature for each temperature control block 1 matches the original melting temperature (step S34).
- the storage / calculation unit 24 measures the melting temperature of the i-th temperature calibration sample for each temperature control block 1 and determines whether or not the difference value between the measured melting temperature and the reference melting temperature is within the target accuracy. (Step S36).
- the storage / calculation unit 24 displays the accuracy that each temperature control block satisfies, and proceeds to processing for the next temperature calibration sample (step S37). Specifically, the temperature absolute value measured for each temperature control block 1 after correction and the accuracy between wells are displayed.
- step S36 when it is determined in step S36 that the temperature difference exceeds the target accuracy, the storage / calculation unit 24 compares the temperature correction count (repetition count) that has already been executed with a threshold value (step S38).
- step S38 When there is a temperature control block 1 for which a negative result is obtained in the determination process of step S38 (that is, when the number of temperature corrections for the corresponding temperature control block has not reached a predetermined threshold), the storage / calculation unit 24 The process returns to step S32.
- the storage / calculation unit 24 executes again the temperature absolute value correction processing based on the temperature difference between the measured melting temperature value and the original melting temperature for each temperature control block 1. If the temperature difference between the temperature control blocks does not fall within the target accuracy, a series of operations are repeatedly executed until the specified number of repetitions is reached.
- step S38 If the accuracy between the temperature control blocks does not fall within the target accuracy even if the series of operations is repeated a specified number of times (if a positive result is obtained in step S38), the storage / calculation unit 24 selects the corresponding temperature control block 1 And an alarm indicating a temperature control abnormality is displayed (step S39).
- step S40 the storage / calculation unit 24 determines whether or not the correction operation has been completed for all the temperature calibration samples. If a negative result is obtained, the storage / calculation unit 24 returns to step S32 and executes a correction operation for the next i + 1th temperature calibration sample. When a positive result is obtained in step S40, the series of processes is terminated.
- Example 7 (Function block configuration of nucleic acid analyzer)
- a nucleic acid analyzer mounting the nucleic acid amplification device according to each of the above-described embodiments will be described.
- the nucleic acid analyzer include a genetic testing device.
- FIG. 12 shows a specific example of the nucleic acid analyzer according to this embodiment.
- the nucleic acid analyzer includes a preprocessing unit, a real-time fluorescence measurement mechanism 15, and a control unit (not shown).
- the pretreatment unit includes at least a dispensing mechanism 31, a reaction container transport mechanism 32, a sample erection position 33, a nucleic acid extraction reagent erection position 34, a nucleic acid amplification reagent erection position 35, a consumables erection position 36, and a consumables disposal hole 37.
- the reaction vessel disposal hole 38 is provided.
- the dispensing mechanism 31 is provided with a dispensing tip for dispensing reagents and samples.
- the apparatus configuration shown in FIG. 12 corresponds to the case where the real-time fluorescence measurement mechanism 15 having the configuration shown in FIG. 3 is incorporated. That is, it corresponds to the case of using the real-time fluorescence measurement mechanism 15 having a rotation drive system.
- FIG. 13 shows another specific example of the nucleic acid analyzer according to this embodiment.
- the nucleic acid analyzer shown in FIG. 13 corresponds to the case where the real-time fluorescence measuring mechanism 15 having the configuration shown in FIG. 4 is incorporated. That is, this corresponds to the case of using the real-time fluorescence measurement mechanism 15 that does not use a rotational drive system.
- FIG. 14 shows a processing operation procedure executed by the nucleic acid analyzer shown in FIGS.
- the same reference numerals are given to the portions corresponding to FIG. 10.
- the user installs a temperature calibration reagent and consumables necessary for the operation of the nucleic acid analyzer at predetermined positions. Thereafter, the user inputs an instruction for temperature correction of the plurality of temperature control blocks 1 that can be individually controlled or temperature confirmation of the temperature control block 1.
- the nucleic acid analyzer that detected the previous instruction input confirms the melting temperature of the temperature calibration reagent and stores it in the storage area (step S51).
- the melting temperature here is taken into the nucleic acid analyzer via the network, input from a barcode attached to the container of the temperature calibration reagent, or both, or manual input by the user.
- the dispensing mechanism 31 dispenses a predetermined amount of the temperature calibration sample installed at the reagent installation position 33 into the reaction container (step S52).
- the predetermined amount here may be any capacity as long as it is a capacity that can be measured by the real-time fluorescence measurement mechanism 15. However, if the real-time fluorescence measurement mechanism 15 does not have a function of preventing evaporation of the reaction solution, it is preferable to add mineral oil to the upper layer of the temperature calibration sample at this stage.
- reaction vessel is closed and conveyed to the real time fluorescence measurement mechanism 15 through the reaction vessel conveyance mechanism 32. Thereafter, the temperature correction operation according to each embodiment described above is executed (steps S22 to S26).
- FIG. 15A shows a measurement example of melting temperature curves measured for a plurality of temperature control blocks 1 when no temperature correction is performed.
- FIG. 15A shows a melting temperature curve when the melting temperature is measured for a mixture of two types of temperature calibration samples (for example, a sample having a low-temperature side melting temperature (60 ° C.) and a high-temperature side melting temperature (95 ° C.)). A measurement example is shown.
- the horizontal axis represents temperature and the vertical axis represents temperature change rate.
- a maximum temperature difference of 1.5 ° C. is observed for the low temperature side melting temperature
- a maximum temperature difference of 1.7 ° C. is recognized for the high temperature side melting temperature.
- FIGS. 15B and 15C when the temperature is corrected based on the melting temperature, as shown in FIGS. 15B and 15C, the temperature difference between the plurality of temperature control blocks can be controlled to 0.1 ° C. or less.
- FIG. 15B is a melting temperature curve measured when the temperature of the plurality of temperature control blocks 1 is corrected for the low-temperature side melting temperature
- FIG. 15C is the correction of the temperature of the plurality of temperature control blocks 1 for the high-temperature side melting temperature. It is a melting temperature curve measured in the case of.
- each temperature control block 1 can be made equal to the melting temperature, and extremely high uniformity can be realized as compared with the conventional method.
- a target accuracy evaluation function it is possible to automatically determine the temperature control block 1 in which the temperature control of the liquid temperature has become abnormal. If the information of the temperature control block 1 in which the temperature control abnormality is determined is stored in the system side, it is excluded from the inspection area so that the temperature control block 1 in which abnormality is recognized in the temperature control is not used in the normal inspection. It is possible to realize control that does or does not use the inspection result.
- FIG. 16 shows a system configuration according to this embodiment.
- a temperature calibration sample is used for temperature correction.
- the system shown in FIG. 16 includes a network information database 100, an information management apparatus 101, a nucleic acid amplification apparatus 102, and a service information management apparatus 103.
- temperature calibration sample information (specifically, the melting temperature of the temperature calibration sample) and temperature calibration result information of each nucleic acid amplification device 102 are stored.
- the temperature calibration sample information is stored in the network information database 100 from the information management apparatus 101 via the network, and read out to the N nucleic acid amplification apparatuses 102 via the network.
- the temperature calibration result information is stored in the network information database 100 from the N nucleic acid amplification devices 102 via the network, and is further read out by the information management device 101.
- the temperature correction operation using the temperature calibration sample having the same melting temperature is executed in each nucleic acid amplification device 102, whereby the temperature absolute value among the N devices can be made uniform. Further, the information management apparatus 101 can collectively manage the temperature calibration result information of each nucleic acid amplification apparatus 102. For this reason, when an abnormality in temperature control is recognized in a certain nucleic acid amplification device 102, information regarding the nucleic acid amplification device 102 in which the abnormality is found is provided to the service information management device 103 that manages service information. Customer support is possible. Of course, the arrangement positions of the service information management apparatus 103 and the nucleic acid amplification apparatus 102 that is the service providing destination may be the same or different.
- each of the above-described configurations, functions, processing units, processing means, and the like may be partly or entirely realized as, for example, an integrated circuit or other hardware.
- Each of the above-described configurations, functions, and the like may be realized by the processor interpreting and executing a program that realizes each function. That is, it may be realized as software.
- Information such as programs, tables, and files for realizing each function can be stored in a memory, a hard disk, a storage device such as an SSD (Solid State Drive), or a storage medium such as an IC card, an SD card, or a DVD.
- control lines and information lines indicate what is considered necessary for explanation, and do not represent all control lines and information lines necessary for the product. In practice, it can be considered that almost all components are connected to each other.
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Abstract
Description
(核酸増幅装置の機能ブロック構成)
図2に、形態例に係る核酸増幅装置の機能ブロック構成を示す。図2に示す核酸増幅装置は、個別に温度制御可能な複数の温度制御ブロック1と、リアルタイム蛍光測定部3と、それらを制御する制御部5で構成される。本明細書では、個別に温度制御可能な複数の温度制御ブロック1とリアルタイム蛍光測定部3を含む機構部分をリアルタイム蛍光測定機構15と呼ぶ。
図3に、1種類の温度校正試料を使用して複数の温度制御ブロック1の温度絶対値を補正する核酸増幅装置の具体例を示す。図3に示す装置構成は、図2に示した装置構成をより詳細に表したものである。
図4に、1種類の温度校正試料を使用して温度制御ブロック1の温度を補正する核酸増幅装置の他の装置構成例を示す。図4に示す装置構成も、図2に示した装置構成をより詳細に表したものである。
本形態例の場合、反応容器21や反応プレート26は蛍光波長を透過でき、かつ、温度制御ブロック1の熱を伝導可能な材質であれば、いかなる材質や形状でも良い。より好ましくは、DNase、RNaseが混入していないPCRチューブ(グライナー社、ドイツ)又は96穴のPCRプレートを用いることが望ましい。
本形態例における温度校正試料は、HRM解析が可能な核酸断片と検出色素が含まれていれば良い。核酸断片には、DNA、RNA、PNAを利用することができる。より好ましくは、任意の1種類の核酸断片をPCR法で増幅した試料を用いる。さらに好ましくは、水溶液の絶対温度と融解温度の一致を確認した核酸断片を用いる。
本形態例に係る核酸増幅装置は、以下に示す3つの処理工程を通じ、複数の温度制御ブロック間に存在する温度絶対値のバラツキを補正する。
PCR法等の核酸増幅法により増幅した所定の融解温度を有する核酸断片を含む試料を、温度校正試料として複数の反応容器に分注する。その後、この反応容器を、温度校正する核酸増幅装置内の複数の温度制御ブロック1の保持機構に設置又は架設する。または、複数の温度制御ブロック1の保持機構に予め設置又は架設された反応容器内に、温度校正試料を分注する。
次に、温度校正試料の融解温度を、温度制御ブロック1毎に実際に測定する。ここでの融解温度の測定には、公知の方法を適用する。例えば低温(例えば60℃)から高温(例えば95℃)まで、温度制御ブロック1の温度を変化させながら、リアルタイムで蛍光強度を測定する。このとき、核酸増幅装置に要求される温度精度と同等以上の温度分解能で温度を変化させ、蛍光強度を測定する。例えば核酸増幅装置に要求される温度精度が ±0.1℃以下の場合、 0.1℃刻み以下で目標温度を可変し、融解温度を蛍光測定する。
融解温度の測定が終了すると、融解温度の測定値が、記憶部11に記憶されている融解温度に一致するように、制御部5は、各温度制御ブロック1について管理する温度絶対値を補正する。なお、融解温度の測定には、測定誤差が生じる可能性がある。従って、より好ましい実施の形態では、工程2と工程3を2回以上繰り返し、温度制御ブロック間の温度均一性を高めることが望ましい。
図5に、温度補正に先立って実行される融解温度情報の確認処理手順を示す。ここでは、記憶・演算部24が当該処理を実行するものとして説明する。もっとも、核酸増幅装置を構成する他の制御部や核酸増幅装置に接続される外部の制御部を用いて実行しても良い。
以上のように、本形態例に係る核酸増幅装置には、温度校正試料を用いて温度制御ブロック1の温度絶対値を補正する機能を搭載する。このため、本形態例に係る核酸増幅装置によれば、複数の温度制御ブロック間の最大温度差を個々の温度制御ブロック1の温度制御分解能と同等精度に均一化することができる。例えば、図1Bに示したように、複数の温度制御ブロック間の最大温度差を±0.05℃以下に均一化することができる。すなわち、温度制御ブロック間の温度差を各温度制御ブロックにおける温度制御分解能と同等程度に補正できる。このため、複数の温度制御ブロックを用いて核酸増幅を行う場合にも、温度制御ブロックの違いが分析精度に与える影響を無視することができる。
前述した形態例の場合には、蛍光強度が急激に変化する変化率の大きい部分(蛍光強度の減衰率が最も大きい部分(図7Bの場合、蛍光強度値が0.2))を融解温度に決定した。
前述の形態例においては、測定された蛍光強度の減衰率が最も大きい温度又は蛍光強度の変化率の最も大きい温度を「融解温度」として使用した。
前述の形態例においては、融解温度が1つの温度校正試料を用いる温度校正方法について説明した。融解温度が1つの温度校正試料でも、各温度制御ブロック1の温度特性が無視できる程度に同じであれば、融解温度以外の温度についても温度制御ブロック1の最大温度差を±0.05℃以下に均一化することができる。
ここでは、補正後の温度の精度を評価する機能を有する温度補正機能について説明する。理想的には、前述した温度補正が終了すると、温度制御ブロック1の測定融解温度は基準融解温度に一致するはずである(厳密には、差分が温度制御分解能と同レベル以下となるはずである)。ただし、デバイスの故障等のため、温度補正後も誤差が残る可能性がある。そこで、以下に説明する温度補正機能を提案する。
前述の形態例の場合には、1種類の温度校正試料の使用を前提とする温度補正機能を搭載する核酸増幅装置について説明した。
(核酸分析装置の機能ブロック構成)
ここでは、前述した各形態例に係る核酸増幅装置を実装する核酸分析装置について説明する。核酸分析装置には、例えば遺伝子検査装置がある。
図12に、本形態例に係る核酸分析装置の具体例を示す。核酸分析装置は、前処理部と、リアルタイム蛍光測定機構15と、不図示の制御部とを有している。ここでの前処理部は、少なくとも分注機構31、反応容器搬送機構32、試料架設ポジション33、核酸抽出試薬架設ポジション34、核酸増幅試薬架設ポジション35、消耗品架設ポジション36、消耗品廃棄穴37、反応容器廃棄穴38を有している。なお、分注機構31には、試薬や試料を分注する分注チップが取り付けられている。図12に示す装置構成は、図3に示す構成のリアルタイム蛍光測定機構15を組み込む場合に対応する。すなわち、回転駆動系を有するリアルタイム蛍光測定機構15を使用する場合に対応する。
図13に、本形態例に係る核酸分析装置の他の具体例を示す。図13に示す核酸分析装置は、図4に示す構成のリアルタイム蛍光測定機構15を組み込む場合に対応する。すなわち、回転駆動系を用いないリアルタイム蛍光測定機構15を使用する場合に対応する。
図14に、図12及び図13に示す核酸分析装置で実行される処理動作手順を示す。なお、図14には、図10との対応部分に同一符号を付して示している。
前述した形態例の説明では、単一の核酸増幅装置又は核酸分析装置内で複数の温度制御ブロック間の温度絶対値を均一化する場合について説明した。
なお、本発明は上述した形態例に限定されるものでなく、様々な変形例が含まれる。例えば、上述した形態例は、本発明を分かりやすく説明するために詳細に説明したものであり、必ずしも説明した全ての構成を備えるものに限定されるものではない。また、ある形態例の一部を他の形態例の構成に置き換えることが可能であり、また、ある形態例の構成に他の形態例の構成を加えることも可能である。また、各形態例の構成の一部について、他の構成を追加、削除又は置換することも可能である。
3…リアルタイム蛍光測定部
5…制御部
7…融解温度測定部
9…温度補正部
11…記憶部
15…リアルタイム蛍光測定機構
21…反応容器
22…回転盤
23…データ処理部
24…記憶・演算部
25…装置制御部
26…反応プレート
31…分注機構
32…反応容器搬送機構
33…試料架設ポジション
34…核酸抽出試薬架設ポジション
35…核酸増幅試薬架設ポジション
36…消耗品架設ポジション
37…消耗品廃棄穴
38…反応容器廃棄穴
100…ネットワーク情報データベース
101…情報管理装置
102…核酸増幅装置
103…サービス情報管理装置
Claims (15)
- 個別に温度制御可能な複数の温度制御ブロックと、
各温度制御ブロックによって温度管理される反応容器内の試料をリアルタイムで蛍光測定するリアルタイム蛍光測定部と、
各温度制御ブロックで温度管理される1又は複数の反応容器に分注された温度校正試料の基準融解温度を記憶する記憶部と、
各温度制御ブロックに対応する各反応容器に収容された温度校正試料の融解温度を測定融解温度として測定する融解温度測定部と、
各温度制御ブロックに対応する測定融解温度と前記基準融解温度とを比較し、各差分値に基づいて各温度制御ブロックの温度絶対値を補正する温度補正部と
を有することを特徴とする核酸増幅装置。 - 請求項1に記載の核酸増幅装置において、
前記温度補正部による各温度制御ブロックの温度絶対値の補正終了後、温度校正試料の融解温度を自動的に測定し、補正後の温度制御ブロックの温度制御精度を画面表示する制御部を有することを特徴とする核酸増幅装置。 - 請求項1に記載の核酸増幅装置において、
前記温度補正部による各温度制御ブロックの温度絶対値の補正終了後、各温度制御ブロックについて温度校正試料の融解温度を自動的に測定する処理と、
測定結果に基づいて各温度制御ブロックの温度絶対値を補正する処理と
を繰り返し実行する制御部を有する
ことを特徴とする核酸増幅装置。 - 請求項3に記載の核酸増幅装置において、
前記制御部は、全ての温度制御ブロックの温度制御精度が、予め設定された温度精度範囲内に入るまで前記補正する処理と前記測定する処理を繰り返す
ことを特徴とする核酸増幅装置。 - 請求項4に記載の核酸増幅装置において、
前記制御部は、予め設定された回数を繰り返しても、予め設定された温度精度範囲内に温度制御精度が入らない温度制御ブロックについて、温度制御異常を意味するアラームを画面表示する
ことを特徴とする核酸増幅装置。 - 請求項5に記載の核酸増幅装置において、
前記制御部は、温度制御異常を意味するアラームが画面表示された温度制御ブロックに対し、通常動作中もアラームの表示を継続する
ことを特徴とする核酸増幅装置。 - 請求項5に記載の核酸増幅装置において、
前記制御部は、温度制御異常を意味するアラームが画面表示された温度制御ブロックに対し、通常動作の使用対象から除外する
ことを特徴とする核酸増幅装置。 - 個別に温度制御可能な複数の温度制御ブロックと、各温度制御ブロックによって温度管理される反応容器内の試料をリアルタイムで蛍光測定するリアルタイム蛍光測定部と、各温度制御ブロックで温度管理される1つ又は複数の反応容器に分注された温度校正試料の基準融解温度を記憶する記憶部と、各温度制御ブロックに対応する各反応容器に収容された温度校正試料の融解温度を測定融解温度として測定する融解温度測定部と、各温度制御ブロックに対応する測定融解温度と前記基準融解温度とを比較し、各差分値に基づいて各温度制御ブロックの温度絶対値を補正する温度補正部とを有する核酸増幅装置と、
前記反応容器に検体又は試料を分注する分注機構と、
前記核酸増幅装置に前記反応容器を搬送し、前記複数の温度制御ブロックのうちいずれかに搬送する搬送機構と
を有することを特徴とする核酸分析装置。 - 請求項8に記載の核酸分析装置において、
前記温度補正部による各温度制御ブロックの温度絶対値の補正終了後、温度校正試料の融解温度を自動的に測定し、補正後の温度制御ブロックの温度制御精度を画面表示する制御部を有することを特徴とする核酸分析装置。 - 請求項8に記載の核酸分析装置において、
前記温度補正部による各温度制御ブロックの温度絶対値の補正終了後、各温度制御ブロックについて温度校正試料の融解温度を自動的に測定する処理と、
測定結果に基づいて各温度制御ブロックの温度絶対値を補正する処理と
を繰り返し実行する制御部を有する
ことを特徴とする核酸分析装置。 - 請求項9に記載の核酸分析装置において、
前記制御部は、全ての温度制御ブロックの温度制御精度が、予め設定された温度精度範囲内に入るまで前記補正する処理と前記測定する処理を繰り返す
ことを特徴とする核酸分析装置。 - 請求項10に記載の核酸分析装置において、
前記制御部は、予め設定された回数を繰り返しても、予め設定された温度精度範囲内に温度精度が入らない温度制御ブロックについて、温度制御異常を意味するアラームを画面表示する
ことを特徴とする核酸分析装置。 - 請求項12に記載の核酸分析装置において、
前記制御部は、温度制御異常を意味するアラームが画面表示された温度制御ブロックに対し、通常動作中もアラームの表示を継続する
ことを特徴とする核酸分析装置。 - 請求項12に記載の核酸分析装置において、
前記制御部は、温度制御異常を意味するアラームが画面表示された温度制御ブロックに対し、通常動作の使用対象から除外する
ことを特徴とする核酸分析装置。 - 請求項8に記載の核酸分析装置において、
前記核酸分析装置はネットワーク経由で前記温度校正試料の基準融解温度を取得し、他の核酸分析装置との間で前記基準融解温度を共有する
ことを特徴とする核酸分析装置。
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