Patent Publication Number: US-2023152273-A1

Title: Electrophoresis Device and Analysis Method

Description:
TECHNICAL FIELD 
     The present disclosure relates to an electrophoresis device and an analysis method. 
     BACKGROUND ART 
     As a method for analyzing the base sequence or the base length of a DNA, an electrophoresis method is known widely. As one of the analysis method using electrophoresis, there is capillary electrophoresis. The capillary electrophoresis is a technology where a fine tube called a capillary is filled with a separation medium such as acrylamide to perform electrophoresis. To be more specific, when a sample including a DNA is disposed at one end of the capillary and a high voltage is applied to both ends of the capillary in the state, the DNA that is a charged particle negatively charged moves to the positive pole side inside the capillary depending on own size namely the base length. Also, by measuring time required for the sample to migrate through a constant distance (from a sample poring end of the capillary to a signal detection unit in normal cases), the base length of the DNA can be analyzed. Each DNA is labelled by a fluorescent dye, and fluorescence is produced by irradiation of excitation light. The fluorescence is detected by a photodetector. 
     In an analysis of the DNA by capillary electrophoresis, there is a case of using plural fluorescent dyes for the purpose of speeding up the analysis. The plural fluorescent dyes respectively produce different fluorescence receiving irradiation of excitation light. A spectrum obtained by dispersing this fluorescence and acquiring the same on a photodetector is called a fluorescence spectrum. Although each fluorescent dye has a different fluorescence spectrum respectively, they are not sharp, and the fluorescence spectra of respective fluorescent dyes overlap with each other. Therefore, in a photodetector, when DNA fragments labelled by a different fluorescent dye have a fragment length of a same degree, a fluorescence spectrum obtained by a photodetector becomes a linear sum namely a weighted sum of fluorescence spectra of plural kinds of the fluorescent dye. In order to obtain the signal strength (fluorescence strength) of each fluorescent dye from this state, a linear coefficient namely a weighted value of a spectrum of each fluorescent dye configuring a spectrum can be obtained from the spectrum obtained by a photodetector. 
     In order to obtain this weighted value, each fluorescence spectrum should be known in advance. Each fluorescence spectrum is to be intrinsically determined unitarily by a fluorescent dye and a separation medium without depending on a device. However, in an actual device, the fluorescence spectrum changes due to various reasons. One that is known well among them is the positional relation of a capillary and a photodetector. Therefore, when the capillary is to be replaced, before a sample of an analysis object (will be hereinafter expressed as an “actual sample”) is subject to electrophoresis, an operation of obtaining beforehand a fluorescence spectrum in the device and the capillary is required. This operation is called “spectral calibration”. Also, when electrophoresis is performed for plural samples simultaneously using a capillary array where plural capillaries are arrayed, it is required to obtain the fluorescence spectrum for each capillary. 
     Here, an example of the spectral calibration related to a prior art will be explained. 
       FIG.  1    is a diffraction grating image (bottom) imaged in a photodetector of a multi-capillary electrophoresis device, and a drawing (top) showing the signal strength distribution of a capillary corresponding to the A-A′ direction of the diffraction grating image. The multi-capillary electrophoresis device separates fluorescence emitted from each fluorescent dye by irradiating laser light having a specific wavelength to the capillary in the wavelength direction by a diffraction grating, and detects the separated light by a photodetector such as a CCD to acquire a diffraction grating image. Also, the signal strength distribution (spectrum) of the diffraction grating image is acquired. 
     The bottom of  FIG.  1    is a diffraction grating image when laser light is irradiated to a capillary where four pieces of capillaries are arrayed, the vertical axis shows the sequence direction of the capillary, and the horizontal axis shows the wavelength direction. In the top of  FIG.  1   , the vertical axis shows the signal strength (brightness value (RFU)), and the horizontal axis shows the wavelength. Further, although  FIG.  1    shows an example of measuring a spectrum continuously (discretely for each pixel in fact) using a diffraction grating, it may be data which are obtained by sampling the spectrum described above with a wide wavelength interval. For example, as illustrated in the diffraction grating image of  FIG.  1   , only the signal strength in the twenty wavelengths λ(0) to λ(19) may be acquired for each capillary. Also, an arithmetic mean of the signal strength in the vicinity of each of the wavelengths λ(0) to λ(19) may be taken. 
       FIG.  2    is a flowchart showing a spectral calibration method of a prior art. 
     In step S 101 , an operator performs electrophoresis of a matrix standard. The matrix standard is a reagent for acquiring a fluorescence spectrum and obtaining a matrix described below. The matrix standard includes four kinds of DNA fragments with different length respectively labeled by different fluorescent dye. Information of the length or the order of the length of the DNA fragment corresponding to each fluorescent dye is known. 
       FIG.  3 A  is a drawing showing a waveform of a signal strength obtained by performing electrophoresis of the matrix standard, the vertical axis shows the signal strength, and the horizontal axis shows the time. In step S 101 , it is assumed to obtain fluorescent spectra of four kinds of the fluorescent dye (ROX, TMR, R 110 , and R6G), and  FIG.  3 A  shows a state of laying the signal strength waveform of each fluorescent dye on one graph. As shown in  FIG.  3 A , a sharp peak appears at a time corresponding to the length of the DNA fragment labeled by each fluorescent dye. Since the DNA fragment with different length is labeled respectively by a different fluorescent dye, each fluorescent dye produces light in isolation at each peak time (t0, t1, t2, and t3). Therefore, by acquiring a spectrum at the time when only a specific fluorescent dye produces light (t0, t1, t2, t3, and t4 in  FIG.  3 A ), the fluorescence spectrum of each fluorescent dye is obtained. 
     Returning to  FIG.  2   , in step S 102 , a computation control circuit of the multi-capillary electrophoresis device calculates the fluorescence strength from the spectrum of each time of the signal strength obtained in step S 101 . Processing of the present step may be performed for each scanning time, and may be performed after accumulating spectrum data of a portion of a constant time interval. 
     In step S 103 , the computation control circuit detects the peak time of the signal strength waveform of  FIG.  3 A . As described above, since the appearance order of the peak corresponding to the length of the DNA fragment labeled by each fluorescent dye is known, the kind of the fluorescent dye can be identified by the appearance time of the peak.  FIG.  3 A  shows the situations ROX produces light at time t0, TMR produces light at time t1, R 110  produces light at time t2, and R6G produces light at time t3 in isolation respectively. The spectrum of each time corresponds to each fluorescence spectrum. That is to say, each fluorescence spectrum is known by acquiring the spectrum of each peak time. 
       FIG.  3 B  is the fluorescence spectrum acquired from the signal strength waveform of  FIG.  3 A , the vertical axis shows the fluorescence strength, and the horizontal axis shows the wavelength. As shown in  FIG.  3 B , the computation control circuit acquires the fluorescence spectrum of each fluorescent dye based on the signal strength waveform. 
     Returning to  FIG.  2   , in step S 104 , the computation control circuit acquires a matrix M using each fluorescence spectrum. The mathematical expression 1 described below shows an example of the matrix M of a case of acquiring the signal strength in the twenty wavelengths λ(0) to λ(19). The element of the matrix M corresponds to the strength ratio of the signal strength of each fluorescent dye at each time and in each wavelength. This ratio is a rate relative to a maximum value among the wavelength of each fluorescent dye for example. For example, an element WX1 of the mathematical expression 1 is a ratio of the fluorescence strength of the fluorescent dye ROX at the time t0 and the wavelength λ(1). It means that as this values is larger, contribution to the fluorescence strength of the wavelength is stronger. The matrix M is used to obtain each fluorescence strength from the spectrum waveform obtained by the photodetector. 
     
       
         
           
             
               
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     The operation of step S 101  to S 104  described above is the spectral calibration. When there exist more than one of the capillaries, it is required to acquire the matrix M for each capillary. Also, it is required to perform the spectral calibration whenever the capillary is disposed, the component is replaced, and so on. 
     The matrix M obtained in the spectral calibration is also called a reference spectrum, and is identical to the fluorescence spectrum of an actual sample ideally. However, in practice, a deviation possibly occurs between the reference spectrum and the fluorescence spectrum of the actual sample. When the deviation occurs, a weighted value is not calculated correctly, and erroneous fluorescence strength is recorded. In a serious case, a pseudo peak appears at a peak time same as the time of the main peak. 
       FIG.  4    is a fluorescence spectrum when a pseudo peak appears. The pseudo peak possibly occurs by overlapping of the fluorescence spectrum of each color, and the effect by this overlapping is observed largely when a deviation occurs between the reference spectrum and the fluorescence spectrum of the actual sample. Also, when there are a plurality of the main peaks, this pseudo peak is observed in all of the main peaks. 
     The deviation between the reference spectrum and the fluorescence spectrum of the actual sample generally occurs due to the difference in the fluorescent dye and the electrophoresis condition between the time of the spectral calibration and the time of electrophoresis of the actual sample. That is to say, since the operator is required to repeat the spectral calibration whenever the fluorescent dye and the electrophoresis condition used for an actual sample are changed, the labor and the cost increase. 
     Patent Literature 1 discloses a gene analysis device that is “characterized by acquiring a reference fluorescence spectrum by using an allelic ladder and a size standard that is known information for a DNA fragment used in electrophoresis of an actual sample, and is characterized in that, on capillaries not using an allelic ladder, the spectral calibration is performed by detecting an amount of shift in the fluorescence spectrum of the size standard and calculating the fluorescence spectrum by shifting the reference fluorescence spectrum using the amount of shift” (refer to the abstract of the literature). Thus, since it is not required to perform electrophoresis using a special matrix standard, the spectral calibration can be achieved in a short time and at a low cost. 
     The size standard is a mixture of the known DNA fragments labeled by a specific fluorescent dye. The allelic ladder is a mixture of the known DNA fragments labeled by a fluorescent dye same as that of the actual sample. In an operation described in Patent Literature 1, the size standard is mixed for all samples at the time of electrophoresis. Also, the allelic ladder is analyzed by a capillary separate from that of the actual sample. 
     Citation List 
     Patent Literature 
     Patent Literature 1: JP-A No. 2014-117222 
     SUMMARY OF INVENTION 
     Technical Problem 
     However, in Patent Literature 1, the amount of shift of the fluorescence spectrum between the capillaries is calculated using a specific fluorescent dye, and such a case is not assumed that the amount of shift described above differs according to the fluorescent dye. Therefore, according to the fluorescent dye, there is a case that an appropriate reference spectrum is not obtained, a deviation occurs between the reference spectrum and the fluorescence spectrum of the actual sample, and a pseudo peak occurs. Also, in the example 3 of Patent Literature 1, an example of performing the spectral calibration for each capillary is cited. However, for that purpose, a peak formed of a fluorescent dye of a single color becomes necessary. Therefore, there is a case that a reference spectrum cannot be obtained in such sample that plural peaks overlap with each other. As a result, a deviation possibly occurs between the reference spectrum and the fluorescence spectrum of the actual sample. From the above, since the method of Patent Literature 1 is hardly applied to an optional fluorescent dye and an optional sample, it is required to repeat the spectral calibration whenever the electrophoresis condition and the fluorescent dye are changed. Therefore, the labor of the operator and the cost increase. 
     Therefore, the present disclosure provides an electrophoresis device and an analysis method reducing the labor of the operator and the cost. 
     Solution to Problem 
     In order to solve the problem described above, an electrophoresis device of the present disclosure includes an electrophoresis path of a sample, a dispersion element for dispersing light from the sample within the electrophoresis path, a photodetector for detecting the light dispersed by the dispersion element, and a computation unit for determining the spectrum of the light on the basis of a signal from the photodetector, and is characterized in that the computation unit corrects the spectrum using correction factors determined for each electrophoresis condition or fluorescent dye. 
     Also, another electrophoresis device of the present disclosure includes an electrophoresis path of a sample, a dispersion element for dispersing light from the sample within the electrophoresis path, a photodetector for detecting the light dispersed by the dispersion element, and a computation unit for calculating signal strength of the light on the basis of a signal of the photodetector, and is characterized in that the photodetector acquires the signal with a signal acquisition width that is set so that a correlation coefficient between spectra of plural fluorescent dyes becomes equal to or greater than a predetermined value. 
     Other features related to the present disclosure will be clarified by description of the present description and the attached drawings. Also, aspects of the present disclosure will be achieved and actualized by elements and combination of various elements, detailed description hereinbelow, and aspects of the attached claims. 
     Description of the present description is only a typical exemplification, and does not limit the claims or the application example of the present disclosure in any means. 
     Advantageous Effects of Invention 
     According to the present disclosure, it is not required to repeat the spectral calibration whenever the electrophoresis condition and the fluorescent dye are changed. As a result, the labor of the operator and the cost are reduced. Problems, configurations, and effects other than those described above will be clarified by explanation of embodiments described below. 
    
    
     
       BRIEF DESCRIPTION OF DRAWINGS 
         FIG.  1    is a drawing showing the signal strength of fluorescence detected by a multi-capillary electrophoresis device (top) and the wavelength (bottom). 
         FIG.  2    is a flowchart showing a conventional spectral calibration method. 
         FIG.  3 A  is a drawing for explaining a summary of spectral calibration related to a prior art. 
         FIG.  3 B  is a drawing for explaining a summary of spectral calibration related to a prior art. 
         FIG.  4    is a drawing for explaining a pseudo peak. 
         FIG.  5    is a schematic view showing a multi-capillary electrophoresis device related to a first embodiment. 
         FIG.  6    is a schematic view showing a configuration of an optical system within a constant temperature reservoir. 
         FIG.  7    is a flowchart showing a calculation method of a correction factor related to the first embodiment. 
         FIG.  8 A  is a drawing for explaining a summary of calculation of a matrix M′ in the first embodiment. 
         FIG.  8 B  is a drawing for explaining a summary of calculation of a matrix M′ in the first embodiment. 
         FIG.  9    is a flowchart showing an application method of a correction factor in electrophoresis of an actual sample. 
         FIG.  10    is a flowchart of an electrophoresis method of an actual sample. 
         FIG.  11    is a drawing for explaining gauss fitting. 
         FIG.  12    is a flowchart showing an analysis method of a sample related to a second embodiment. 
         FIG.  13    is a drawing showing a result of an experiment example 1. 
         FIG.  14    is a flowchart showing an analysis method of a sample related to a third embodiment. 
         FIG.  15 A  is a drawing showing a fluorescent dye used in an experiment example 2. 
         FIG.  15 B  is a drawing showing a result of the experiment example 2. 
         FIG.  16    is a flowchart showing an analysis method of a sample related to a fifth embodiment. 
         FIG.  17 A  is a fluorescence spectrum acquired in an experiment example 3. 
         FIG.  17 B  is a fluorescence spectrum acquired in a control experiment of the experiment example 3. 
         FIG.  18    is a flowchart showing an analysis method of a sample related to a sixth embodiment. 
     
    
    
     DESCRIPTION OF EMBODIMENTS 
      Embodiments will be hereinafter explained referring to the attached drawings. In the attached drawings, there is also a case of expressing an element having a same function by a same reference sign. Further, although the attached drawings show embodiments and implementation examples in line with the technical principle of the present disclosure, they are for the purpose of understanding of the present disclosure, and are not to be used by any means for interpreting the technology of the present disclosure in a limiting manner. Description of the present description is only a typical exemplification, and does not limit the claims or the application example of the present disclosure in any means. 
     In the present embodiments, although explanation thereof is made in detail to be sufficiently enough for a person with an ordinary skill in the art to implement the present disclosure, it is to be understood that other implementations and aspects are also possible and changing of the configuration and construction and replacement of various elements are possible without deviating from the scope and spirit of the technical thought of the present disclosure. Therefore, description below should not be interpreted to be limited to it. 
     First Embodiment 
     As described in Background Art, when the deviation occurs between the reference spectrum and the fluorescence spectrum of the actual sample, a true weighted value is not calculated, and erroneous fluorescence strength is recorded. This deviation occurs mainly because the spectrum changes mainly by denaturalization of the fluorescent dye. Denaturalization of the fluorescent dye occurs by improper pH, storage at an improper temperature, and excessive excitation of the dye. Further, denaturalization of the fluorescent dye possibly occurs also when a migration voltage differs between the time of the spectral calibration and the time of migration of the actual sample. In an electrophoresis device including plural capillaries, since the excitation light strength differs in each capillary, a deviation possibly occurs. It is to be noted also that the degree of denaturalization differs according to the fluorescent dye in each example cited above. Further, even when the actual sample is labeled by a fluorescent dye different from the matrix standard, a deviation occurs as a matter of course. 
      Therefore, in the first embodiment, explanation will be given on operation of a case a migration voltage differs between the time of the spectral calibration by an operator having purchased the multi-capillary electrophoresis device and the time of migration of the actual sample (correction of the fluorescence spectrum. Also, in the present description, there is a case that spectral calibration implemented by a manufacturer of a multi-capillary electrophoresis device before shipment of the device is referred to “the first spectral calibration”, and spectral calibration by an operator having purchased the multi-capillary electrophoresis device is referred to “the second spectral calibration”. 
     Configuration Example of Multi-Capillary Electrophoresis Device 
       FIG.  5    is a schematic view showing a configuration of a multi-capillary electrophoresis device  500  related to the first embodiment. As shown in  FIG.  5   , the multi-capillary electrophoresis device  500  includes a device body  501  and a control computer  502 . 
     The device body  501  includes a computation control circuit  503 , a photodetector  504 , a constant temperature reservoir  505 , a capillary array  506 , a light source  507 , a light irradiation unit  508 , a load header  509 , a negative pole buffer container  511 , a sample container  512 , a polymer cartridge  513 , a positive pole buffer container  514 , a positive pole  515 , a high tension power source  516 , an array header  517 , a transporter  518 , a syringe mechanism  520 , a heating/cooling mechanism  523 , and a diffraction grating  524 . 
     The device body  501  is connected to the control computer  502  in a communicatable manner. An operator can operate each unit included in the device body  501  by operating the control computer  502 . The control computer  502  receives data acquired by the device body  501  (such as a detection signal of the photodetector  504 ). The control computer  502  includes a display that displays the data having been received. Also, the control computer  502  may be incorporated into the device body  501 . 
     The computation control circuit  503  performs calculation processing of a measurement value (fluorescence strength) based on a detection signal of the photodetector  504 , and performs correction of the measurement value (fluorescence strength). Also, the computation control circuit  503  controls the device body  501  according to an input and a command from the control computer  502 . 
     The photodetector  504  is a light sensor detecting fluorescence generated by laser light as excitation light irradiated from the light source  507  to the capillary array  506 . For the light source  507 , liquid laser, gas laser, semiconductor laser, and the like can be used appropriately, and an LED also can be used alternatively. The light source  507  may be configured to irradiate excitation light from both sides of an array of the capillary array  506 , and may be configured to irradiate excitation light in time division. 
     The constant temperature reservoir  505  is a temperature control mechanism for controlling the temperature of the capillary array  506 . The constant temperature reservoir  505  is covered by a heat insulation material to keep the temperature within the reservoir constant, and the temperature is controlled by the heating/cooling mechanism  523 . Thus, the temperature of a major portion of the capillary array  506  can be maintained to a constant temperature of approximately 60° C. for example. 
     The capillary array  506  is configured by arraying plural capillaries  519  (electrophoresis paths) (four in an example of  FIG.  5   ). The capillary array  506  can be configured as a replacement member capable of being replaced appropriately by a new one when damage and qualitative deterioration have been confirmed. Also, the capillary array  506  can be replaced by a separate capillary array including a capillary with different number of piece and length according to measurement. 
     Each of the plural capillaries  519  configuring the capillary array  506  can be configured of a glass pipe with several tens to several hundreds µm of the inside diameter and several hundreds µm of the outside diameter. Also, in order to increase the strength, the surface of the glass pipe may be covered by a polyimide coat. However, at a position where laser light is irradiated and the vicinity thereof, the polyimide coat on the surface of the capillaries  519  is removed. The inside of the capillaries  519  is filled with a separation medium for separating DNA molecules in a biological sample (sample). Here, polyacrylamide-based separation gel (will be hereinafter expressed “polymer”) which is commercially available for electrophoresis use is to be used. 
     The light irradiation unit  508  is disposed in a part of the capillary array  506 . As described below, the light irradiation unit  508  is configured to be capable of causing the laser light (excitation light) from the light source  507  to enter the capillaries  519  of a plural number of piece commonly and introducing fluorescence produced from the capillaries  519  of a plural number of piece to the photodetector  504 . To be more specific, in order to irradiate laser light that is measurement light to a light irradiation portion arranged in the capillary array  506 , the light irradiation unit  508  includes a projection optical system such as an optical fiber and a lens. The diffraction grating  524  (dispersion element) disperses the light from the capillaries  519 , and causes the same to enter the photodetector  504 . 
     Although an example of detecting fluorescence from a fluorescent dye by irradiation of excitation light by the photodetector  504  is explained in the present disclosure, detected light is not limited to fluorescence, and may be absorbed light, produced light, and the like. 
     The load header  509  is arranged at an end of the capillary array  506 . The load header  509  functions as a negative pole to which a negative voltage for introducing the biological sample (sample) into the capillaries  519  is applied. At the other end of the capillary array  506 , the array header  517  is arranged, and the array header  517  bundles the plural capillaries  519  into one. Also, the array header  517  includes a sharp point portion  521  for insertion into the polymer cartridge  512  in the lower surface of the array header  517 . 
     The transporter  518  is configured to mount the negative pole buffer container  511 , the sample container  512 , the polymer cartridge  513 , and the positive pole buffer container  514  on the upper the surface of the transporter  518  and to transport them. As an example, the transporter  518  includes three motors and linear actuators, and can move in three axial directions of up and down, left and right, and front and rear. 
     The negative pole buffer container  511  and the positive pole buffer container  514  are containers holding a buffer for migration, and the sample container  512  is a container holding a sample of the measurement object (sample). 
     The polymer cartridge  513  is a container holding polymer for migration. The polymer cartridge  513  is sealed by a raw material with high plasticity such as rubber or silicone at an upper portion  522 , and is connected to the syringe mechanism  520  for filling the polymer and the transporter  518 . 
     Procedures in filling the polymer into the capillaries  519  from the polymer cartridge  513  are as per (1) to (3) below.
     (1) The transporter  518  is operated, and the array header  517  moves to the upper side of the polymer cartridge  513 .   (2) The sharp point portion  521  of the array header  517  penetrates the upper portion  522  of the polymer cartridge  513 . At this time, since the upper portion  522  of the polymer cartridge  513  having high plasticity encloses the sharp point portion  521  of the array header  517 , both are closely attached to each other, and the polymer cartridge  513  and the capillaries  519  are connected to each other in a sealed state.   (3) The syringe mechanism  520  pushes up the polymer of the inside of the polymer cartridge  513 , and pours the polymer into the capillaries  519 .   

     In the positive pole buffer container  514 , the positive pole  515  applying a positive voltage for migration is disposed so as to contact the buffer. The high tension power source  516  is connected between the positive pole  515  and the load header  509  as a negative pole. 
     The transporter  518  transports the negative pole buffer container  511  and the sample container  512  to a negative pole end  510  of the capillaries  519 . At this time, the positive pole buffer container  514  interlockingly moves to the sharp point portion  521  that corresponds to a positive pole end of the capillaries  519 . 
     The sample container  512  incorporates sample tubes of a number of piece same as that of the capillaries  519 . The operator dispenses the DNA to the sample tubes. 
     The computation control circuit  503  (computation unit) includes a measurement value computation unit  5032 , a correction factor computation unit  5033 , a correction factor database  5034 , and a correction unit  5035 . 
     The measurement value computation unit  5032  calculates a measurement value (fluorescence strength) based on a detection signal of the photodetector  504 . The correction factor computation unit  5033  calculates a correction factor for correcting the measurement value calculated by the measurement value computation unit  5032 . The correction factor database  5034  stores the correction factor calculated by the correction factor computation unit  5033 . Also, the correction unit  5035  calculates a corrected measurement value by applying the correction factor stored in the correction factor database  5034  to the measurement value of the measurement value computation unit  5032 . Computation processing of each unit of the computation control circuit  503  described above can be achieved by that a processor such as a CPU and an MPU for example performs a program. 
       FIG.  6    is a schematic view showing a configuration of an optical system within the constant temperature reservoir  505 . As shown in  FIG.  6   , the light irradiation unit  508  includes plural reflection mirrors  602  (two in  FIG.  6   ) and a condenser lens  603  as an example. The reflection mirror  602  changes the traveling direction of laser light  601  from the light source  507 . Also, the condenser lens  603  condenses the laser light to a light irradiation portion of the capillary array  506 . Thus, the laser light  601  enters the plural capillaries  519  in sequence. The fluorescent dye within each capillary  519  is excited by the laser light  601 , and produces information light (fluorescence having a wavelength depending on the sample). This information light is dispersed to a wavelength direction by the diffraction grating  524 . The information light having been dispersed is detected by the photodetector  504 . At this time, although the photodetector  504  can measure a spectrum continuously (discretely for each pixel in fact), in the present embodiment, as an example, only the signal strength in the twenty wavelengths λ(0) to λ(19) is to be acquired. 
     Thus, by observing the fluorescence strength of the fluorescence produced by entering of the laser light  601  by the photodetector  504 , an analysis of the DNA during electrophoresis is enabled. Electrophoresis means to separate a sample by a difference of movability depending on the property of a sample, the movability being given to a sample in a capillary  119  by an electric field action generated between the negative pole and positive pole buffers. Here, explanation will be given exemplifying a case where the sample is a DNA. 
     The DNA has a negative electric charge in the polymer by a phosphodiester bond which corresponds to a skeleton of a double helix. Therefore, the DNA moves to the positive pole side in a DNA electric field. At this time, since the polymer has a net-like structure, movability of the DNA depends on easiness in going through the net, namely the size of the DNA. A DNA having a short base length easily goes through the net-like structure and movability becomes high. Results of a DNA having a long base length are opposite. Since the DNA is labeled by a fluorescence substance (fluorescent body) beforehand, the photodetector  504  optically detects the DNA in sequence starting from a DNA having a short base length. Normally, the measuring time and the voltage application time are set matching a sample with the longest migration time. 
     Calculation Method for Correction Factor 
     As described above, the present embodiment proposes a correction method for the fluorescence spectrum of a case where the migration voltage differs between the time of the spectral calibration and the time of migration of the actual sample. The manufacturer of the multi-capillary electrophoresis device  500  obtains a correction factor for correcting the fluorescence spectrum acquired at the time of migration of an actual sample and registers the correction factor to the correction factor database  5034  of the computation control circuit  503  before shipment of the device. 
       FIG.  7    is a flowchart showing a calculation method for a correction factor. The calculation method for the correction factor will be summarized. First, in step S 1 , the manufacturer performs the spectral calibration using a matrix standard, and acquires a matrix M becoming a reference by the computation control circuit  503 . Next, in step S 2 , a matrix M′ used for correction is acquired by the computation control circuit  503 . Lastly, in step S 3 , a correction factor matrix K is acquired by the computation control circuit  503 . 
     Step S 1   
     In step S 1 , the manufacturer performs the spectral calibration using a matrix standard including a DNA fragment labeled by an optional fluorescent dye (the first spectral calibration). In the present embodiment, as an example, ROX, TMR, R 110 , and R6G are used as the fluorescent dye. The migration voltage should be made same as a migration voltage in the spectral calibration before migration of the actual sample described below (the second spectral calibration). In the present embodiment, although the migration voltage is made 15 kV as an example, the migration voltage is not limited to it. 
     The manufacturer registers the kind of the fluorescent dye and the migration voltage in the computation control circuit  503  by operating an input device of the control computer  502 . The measurement value computation unit  5032  is to obtain the matrix M with this condition. 
     Here, one of the problems to be solved in the present disclosure is that, when the migration voltage differs between the time of the spectral calibration by an operator and the time of migration of the actual sample, a deviation occurs between the reference spectrum and the fluorescence spectrum of the actual sample. The migration voltage affects the time required for electrophoresis and the separation capacity which is one of the important quality indicators in an analysis. Therefore, in using the multi-capillary electrophoresis device, an operator frequently changes the migration voltage of the actual sample according to the necessity. Also, whenever the migration voltage of the actual sample is changed, the operator is required to repeat the spectral calibration with a same migration voltage as that of the actual sample. 
     In order to solve this problem, the present embodiment proposes to perform the first spectral calibration with various migration voltages before shipment of the multi-capillary electrophoresis device, to quantify the deviation between the spectra found out there, and to thereby register such correction factor to minimize the deviation in the computation control circuit  503  beforehand. The correction factor is registered along with the information such as the fluorescent dye and the migration voltage having been used. 
     The operator having purchased the device selects an optional migration voltage out of those having been registered in the computation control circuit  503 , performs the second spectral calibration, and is enabled thereafter to make an actual sample to migrate with an optional migration voltage having been registered in the computation control circuit  503  in a similar manner. That is to say, even when the migration voltage of an actual sample may be changed by any number of times within a range registered in the computation control circuit  503 , the operator is not required to repeat the spectral calibration each time. 
     When the operation described above is assumed, in step S 1 , the manufacturer should perform migration of the matrix standard not only with 15 kV but also with plural voltages. Also, all of the matrix M having been acquired should be registered in the computation control circuit  503  along with the information of the migration voltage and the fluorescent dye. 
     The calculation method for the matrix M is as per one described above. 
     Step S 2   
     In step S 2 , the manufacturer makes the matrix standard to migrate with a fluorescent dye and a migration condition same as those of an actual sample. Here, the actual sample is to be labeled by a fluorescent dye same as that for the matrix standard having been used in step S 1  and is to be made to migrate with 7.5 kV. At this time, by operating an input device of the control computer  502 , the manufacturer registers the kind of the fluorescent dye and the migration voltage in the computation control circuit  503 . 
     As described above, since the matrix standard includes DNA fragments with different length having been labeled respectively by a different fluorescent dye, each fluorescent dye produces light in isolation at each peak time (t0′, t1′, t2′, and t3′). Also, since the appearance order of the peak time corresponding to each fluorescent dye is known, the kind of the fluorescent dye corresponding to each peak time can be identified. 
       FIG.  8 A  is a drawing showing a waveform of a signal strength obtained by performing electrophoresis of the matrix standard, the vertical axis shows the signal strength, and the horizontal axis shows the time. As shown in  FIG.  8 A , ROX produces light at the time t′0, TMR produces light at the time t′1, R 110  produces light at the time t′2, and R6G produces light at the time t′3 respectively in isolation. The spectrum of each time corresponds to the fluorescence spectrum of each fluorescent dye. Therefore, the computation control circuit  503  acquires the fluorescence spectrum of each fluorescent dye by acquiring the spectrum of each peak time. 
       FIG.  8 B  is the fluorescence spectrum acquired from the signal strength waveform of  FIG.  8 A , the vertical axis shows the fluorescence strength, and the horizontal axis shows the wavelength. 
     The measurement value computation unit  5032  calculates the matrix M′ using each fluorescence spectrum. The mathematical expression 2 described below shows an example of the matrix M′ of a case of acquiring the signal strength in the twenty wavelengths λ(0) to λ(19). The element of the matrix M′ corresponds to the strength ratio of each fluorescent dye at each peak time (t′0, t′1, t′2, and t′3) and each wavelength. For example, an element W’X1 of the mathematical expression 2 is a rate of the fluorescence strength of the fluorescent dye ROX at the time t′0 and the wavelength λ(1).  
     
       
         
           
             
               
                 M′ = 
                 
                   
                     
                       
                         
                           
                             W 
                             
                               ′ 
                               
                                 X 
                                 0 
                               
                             
                           
                         
                         
                           
                             W 
                             
                               ′ 
                               
                                 X 
                                 1 
                               
                             
                           
                         
                         
                           
                             W 
                             
                               ′ 
                               
                                 X 
                                 2 
                               
                             
                           
                         
                       
                       
                         
                           
                             W 
                             
                               ′ 
                               
                                 T 
                                 0 
                               
                             
                           
                         
                         
                           
                             W 
                             
                               ′ 
                               
                                 T 
                                 1 
                               
                             
                           
                         
                         
                           
                             W 
                             
                               ′ 
                               
                                 T 
                                 2 
                               
                             
                           
                         
                       
                       
                         
                           
                             W 
                             
                               ′ 
                               
                                 R 
                                 0 
                               
                             
                           
                         
                         
                           
                             W 
                             
                               ′ 
                               
                                 R 
                                 1 
                               
                             
                           
                         
                         
                           
                             W 
                             
                               ′ 
                               
                                 R 
                                 2 
                               
                             
                           
                         
                       
                       
                         
                           
                             W 
                             
                               ′ 
                               
                                 G 
                                 0 
                               
                             
                           
                         
                         
                           
                             W 
                             
                               ′ 
                               
                                 G 
                                 1 
                               
                             
                           
                         
                         
                           
                             W 
                             
                               ′ 
                               
                                 G 
                                 2 
                               
                             
                           
                         
                       
                     
                     • 
                     • 
                     • 
                     • 
                     • 
                     • 
                     • 
                     
                       
                         
                           
                             W 
                             
                               ′ 
                               
                                 X 
                                 18 
                               
                             
                           
                         
                         
                           
                             W 
                             
                               ′ 
                               
                                 X 
                                 19 
                               
                             
                           
                         
                       
                       
                         
                           
                             W 
                             
                               ′ 
                               
                                 T 
                                 18 
                               
                             
                           
                         
                         
                           
                             W 
                             
                               ′ 
                               
                                 T 
                                 19 
                               
                             
                           
                         
                       
                       
                         
                           
                             W 
                             
                               ′ 
                               
                                 R 
                                 18 
                               
                             
                           
                         
                         
                           
                             W 
                             
                               ′ 
                               
                                 R 
                                 19 
                               
                             
                           
                         
                       
                       
                         
                           
                             W 
                             
                               ′ 
                               
                                 G 
                                 18 
                               
                             
                           
                         
                         
                           
                             W 
                             
                               ′ 
                               
                                 G 
                                 19 
                               
                             
                           
                         
                       
                     
                   
                 
                   
               
             
             
               
                   
                   
                   
                   
                   
                   
                 
                   
                     
                       
                         X 
                         ⇒ 
                       
                     
                     
                       
                         ROX 
                       
                     
                   
                   
                     
                       
                         T 
                         ⇒ 
                       
                     
                     
                       
                         TMR 
                       
                     
                   
                   
                     
                       
                         R 
                         ⇒ 
                       
                     
                     
                       
                         R110 
                       
                     
                   
                   
                     
                       
                         G 
                         ⇒ 
                       
                     
                     
                       
                         R6G 
                       
                     
                   
                 
               
             
           
         
       
     
     Further, in step S 2  also, due to such reason as described in step S 1 , in an actual operation, the matrix standard is made to migrate with plural voltages including 7.5 kV, and all acquired matrices M′ are registered in the computation control circuit  503  along with the information such as the migration voltage and the fluorescent dye. 
     Step S 3   
     Returning to  FIG.  7   , in step S 3 , the measurement value computation unit  5032  transmits the matrices M and M′ having been calculated to the correction factor computation unit  5033 . The correction factor computation unit  5033  acquires the correction factor matrix K based on the matrices M and M′. In the fluorescent dye i and the wavelength j, the element of the correction factor matrix K is defined to be the element k(ij) of the correction factor matrix k(ij)=w′(ij)/w(ij). As described already, the fluorescent dye and the migration voltage used in step S 2  have been registered in the computation control circuit  503 . Therefore, k(ij) can be accumulated in the correction factor database  5034  along with the information of the migration condition and the fluorescent dye used for calculation. At this time, as described in steps S 1  and S 2 , when the matrices M and M′ have been acquired with plural migration voltages, the correction factor computation unit  5033  calculates the correction factor matrix K in all combinations thereof, and registers the same in the correction factor database  5034  along with the information of the migration voltage and the fluorescent dye. 
      Analysis Method by Electrophoresis of Actual Sample 
       FIG.  9    is a flowchart showing an application method of a correction factor in electrophoresis of an actual sample by an operator. 
     Step S 11   
     Steps S 1  to S 3  described above have already been completed at the time point when the operator purchased the multi-capillary electrophoresis device  500 . The operator only has to perform only operations of step S 11  and onward. Also, it is assumed that, at the time of the purchase (after step S 3 ), the capillaries were detached and attached for the purpose of transportation of the device, and the positional relation between the photodetector  504  and the capillaries  519  changed. That is to say, the device is in a state of requiring the spectral calibration again. 
     In step S 11 , the operator performs the spectral calibration using the matrix standard in a manner similar to step S 1 . For the sake of convenience, the spectral calibration performed by the operator is referred to “the second spectral calibration”. The migration voltage in the second spectral calibration can be selected optionally as far as it is a migration voltage registered in the correction factor database  5034 . In the present embodiment, as an example, migration is to be effected with 15 kV. Also, with respect to the fluorescent dye, it is assumed that the matrix standard has been labeled by ROX, TMR, R 110 , and R6G. The matrix M acquired by the measurement value computation unit  5032  in the second spectral calibration of step S 11  is made a matrix M(r). 
     Step S 12   
     In step S 12 , the operator performs migration of the actual sample. Although the actual sample is an unknown sample, the kind of the fluorescent dye and the migration voltage are assumed to be known. The migration condition of the actual sample is made 7.5 kV used in step S 2 . With respect to the fluorescent dye, it is assumed that the actual sample also has been labeled by ROX, TMR, R 110 , and R6G in a manner similar to the matrix standard. 
       FIG.  10    is a flowchart of an electrophoresis method of the actual sample in step S 12 . As shown in  FIG.  10   , the basic procedure of electrophoresis includes sample preparation (step S 121 ), analysis start (step S 122 ), separation medium filling (step S 123 ), preparatory migration (step S 124 ), sample introduction (step S 125 ), and migration analysis (step S 126 ). 
     Step S 121   
     In step S 121 , as sample of preparation before starting an analysis, the operator sets the sample and the reagent to the multi-capillary electrophoresis device  500 . To be more specific, first, the operator fills the negative pole buffer container  511  and the positive pole buffer container  514  shown in  FIG.  5    with a buffer solution which forms a part of an energizing path. For the buffer solution, a commercially available electrolyte fluid for electrophoresis can be used for example. Also, the operator dispenses the actual sample which is the analysis object into the well of the sample container  512 . The actual sample is a PCR product of a DNA for example. Also, the operator pours a separation medium for causing electrophoresis of the sample into the syringe mechanism  520 . For the separation medium, the polymer described above is to be used. Also, the operator replaces the capillary array  506  when deterioration of the capillaries  519  is presumed or when the length of the capillaries  519  is to be changed. 
     Step S 122   
     In step S 122 , by operating an input device of the control computer  502 , the operator registers the kind of the fluorescent dye and the migration voltage used for the actual sample to the computation control circuit  503 . Also, the operator inputs an instruction of analysis start to the control computer  502 . When the instruction of the analysis start is inputted, the control computer  502  transits the instruction to the device body  501 . Thus, the device body  501  starts in an analysis. 
     Step S 123   
     In step S 123 , the device body  501  starts polymer filling into the capillaries  519 . Polymer filling is a procedure of filling new polymer into the capillaries  519  to form a migration path. 
      In polymer filling in the present embodiment, first, the negative pole buffer container  511  is carried to right below the load header  509  by the transporter  518  shown in  FIG.  5    so as to be capable of receiving the spent polymer discharged from the negative pole end  510  of the capillaries  519 . Also, the syringe mechanism  520  is driven to fill the capillaries  519  with new polymer, and the spent polymer is disposed of. Lastly, in order to prevent the separation medium from being dried, the negative pole end  510  is immersed in the buffer solution within the negative pole buffer container  511 . 
     Step S 124   
     In step S 124 , the device body  501  performs preparatory migration. Preparatory migration is a procedure of applying a predetermined voltage to the polymer to achieve a state of the polymer suitable to electrophoresis. 
     In preparatory migration in the present embodiment, first, the negative pole end  510  is immersed in the buffer solution within the negative pole buffer container  511  by the transporter  518 , and the energizing path is formed. Also, by the high tension power source  516 , voltage of approximately several to several tens kV is applied to the polymer for several to several tens minutes to achieve a state of the polymer suitable to electrophoresis. Lastly, in order to prevent the polymer from being dried, the negative pole end  510  is immersed in the buffer solution within the negative pole buffer container  511 . 
     Step S 125   
     In step S 125 , the device body  501  introduces a sample component to the migration path. This step may be performed automatically, or may be performed by that a control signal is transmitted from the control computer  502  from time to time. 
     In sample introduction in the present embodiment, first, the negative pole end  510  is immersed in the sample held within the well of the sample container  512  by the transporter  518 . Thus, an energizing path is formed, and a state of enabling to introduce the sample component into the migration path is achieved. Also, a pulse voltage is applied to the energizing path by the high tension power source  518  to introduce the sample component to the migration path. Lastly, in order to prevent the polymer from being dried, the negative pole end  510  is immersed in the buffer solution within the negative pole buffer container  511 . 
     Step S 126   
     In step S 126 , the device body  501  performs migration analysis. In migration analysis, each sample component included in the sample is separated and analyzed by electrophoresis. 
     In migration analysis in the present embodiment, first, the negative pole end  510  is immersed in the buffer solution within the negative pole buffer container  511  by the transporter  518 , and the energization path is formed. Next, a high voltage of 7.5 kV is applied to the energization path by the high tension power source  516  to generate an electric field in the migration path. By the electric field having been generated, each sample component within the migration path moves to the light irradiation unit  508  at a speed depending on the property of each sample component. That is to say, the sample component is separated by the difference in the moving speed thereof. Also, the photodetector  504  effects detection in sequence from the sample component having reached the light irradiation unit  508 . 
     For example, when the sample includes many DNAs with different base length, a difference occurs in the moving speed by the base length thereof, and the DNA reaches the light irradiation unit  508  in sequence from a DNA with short base length. To each DNA, a fluorescent dye corresponding to an analysis object is bonded. When excitation light is irradiated to the light irradiation unit  508  from the light source  507 , information light (fluorescence having a wavelength depending on a sample) is produced from the sample and is discharged to the outside. This information light is dispersed in the wavelength direction by the diffraction grating  524 , and is detected by the photodetector  504 . An example of images detected by the photodetector  504  is  FIG.  1   . During migration analysis, in the photodetector  504 , this information light is detected at a constant temporal interval, and image data are transmitted to the computation control circuit  503 . Alternatively, in order to reduce the information amount to be transmitted, the photodetector  504  may transmit not the image data but brightness (signal strength) of only a part of regions in the image data. For example, the strength of only a wavelength position of a constant interval may be transmitted for each capillary. 
     As stated in the explanation of  FIG.  1   , in the present embodiment, out of the image data described above, only the signal strength data in the twenty wavelengths λ(0) to λ(19) are to be transmitted to the computation control circuit  503  for each capillary. The signal strength data expresses a spectrum of each DNA sample in each capillary, and this spectrum is stored in the measurement value computation unit  5032 . In the measurement value computation unit  5032 , the spectra of all of the capillaries  519  at all detection times during migration analysis described above are stored. Further, although the spectra of all detection times can be stored in the measurement value computation unit  5032 , when only a predetermine peak time is important for an operator, only the spectra of the vicinity of the predetermine time may be stored. 
     Step S 127   
     In step S 127 , when acquisition of the image data having been planned is completed, the device body  501  stops application of voltage, and migration analysis is finished. 
     The above is an example of processing of electrophoresis processing (step S 12 ) in  FIG.  9   . Also, steps S 123  to S 127  may be performed automatically by the device body  501 , and may be performed by transmission of a control signal from the control computer  502  from time to time. 
     Step S 13   
     Returning to  FIG.  9   , in step S 13 , the correction unit  5035  calls up a correction factor matrix K having combination of same migration voltage and fluorescent dye as that of the time of acquisition of the matrix M(r) and the actual sample of step S 12  from the correction factor database  5034 , and calculates a matrix M(r)k by multiplication of each element of the matrix M(r) and each element k(ij) of the matrix K. 
      Step S 14   
     In step S 14 , the correction unit  5035  calculates the fluorescence strength. To be more specific, the correction unit  5035  calculates the strength of each fluorescent dye from the image data obtained in electrophoresis processing (step S 12 ) described above. In the present step S 14 , the strength ratio of each fluorescent dye in the wavelengths λ(0) to λ(19) only has to be multiplied to the spectrum of each capillary  519  at each time and to be added. When this is expressed by a matrix, the result is as per the mathematical expression 3 below.  
     
       
         
           
             
               
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                 
                   
                     
                       
                         c 
                         = 
                         M 
                         
                           r 
                         
                         k f 
                       
                     
                   
                   
                     
                       
                         c 
                         = 
                         
                           
                               
                             
                               c 
                               X 
                             
                               
                               
                             
                               c 
                               T 
                             
                               
                               
                             
                               c 
                               R 
                             
                               
                               
                             
                               c 
                               G 
                             
                               
                           
                         
                       
                     
                   
                   
                     
                       
                         f 
                         = 
                         
                           
                             
                               f 
                               0 
                             
                               
                               
                             
                               f 
                               1 
                             
                               
                               
                             
                               f 
                               2 
                             
                               
                               
                             • 
                             • 
                             • 
                             • 
                             • 
                             • 
                               
                             
                               f 
                               
                                 18 
                               
                             
                               
                               
                             
                               f 
                               
                                 19 
                               
                             
                               
                           
                         
                       
                     
                   
                 
               
             
             
               
                 M 
                 
                   r 
                 
                 k 
                 = 
                 
                   
                     
                       
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 X 
                                 0 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     X 
                                     0 
                                   
                                 
                               
                             
                           
                         
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 X 
                                 1 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     X 
                                     2 
                                   
                                 
                               
                             
                           
                         
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 X 
                                 2 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     X 
                                     2 
                                   
                                 
                               
                             
                           
                         
                       
                       
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 T 
                                 0 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     T 
                                     0 
                                   
                                 
                               
                             
                           
                         
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 T 
                                 1 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     T 
                                     1 
                                   
                                 
                               
                             
                           
                         
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 T 
                                 2 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     T 
                                     2 
                                   
                                 
                               
                             
                           
                         
                       
                       
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 R 
                                 0 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     R 
                                     0 
                                   
                                 
                               
                             
                           
                         
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 R 
                                 1 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     R 
                                     1 
                                   
                                 
                               
                             
                           
                         
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 R 
                                 2 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     R 
                                     2 
                                   
                                 
                               
                             
                           
                         
                       
                       
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 G 
                                 0 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     G 
                                     0 
                                   
                                 
                               
                             
                           
                         
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 G 
                                 1 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     G 
                                     1 
                                   
                                 
                               
                             
                           
                         
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 G 
                                 2 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     G 
                                     2 
                                   
                                 
                               
                             
                           
                         
                       
                     
                     • 
                     • 
                     • 
                     • 
                     • 
                     • 
                     • 
                     
                       
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 X 
                                 18 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     X 
                                     18 
                                   
                                 
                               
                             
                           
                         
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 X 
                                 19 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     X 
                                     19 
                                   
                                 
                               
                             
                           
                         
                       
                       
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 T 
                                 18 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     T 
                                     18 
                                   
                                 
                               
                             
                           
                         
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 T 
                                 19 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     T 
                                     19 
                                   
                                 
                               
                             
                           
                         
                       
                       
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 R 
                                 18 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     R 
                                     18 
                                   
                                 
                               
                             
                           
                         
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 R 
                                 19 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     R 
                                     19 
                                   
                                 
                               
                             
                           
                         
                       
                       
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 G 
                                 18 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     G 
                                     18 
                                   
                                 
                               
                             
                           
                         
                         
                           
                             w 
                             
                               
                                 
                                   r 
                                 
                               
                               
                                 G 
                                 19 
                               
                             
                             
                               k 
                               
                                 
                                   
                                     G 
                                     19 
                                   
                                 
                               
                             
                           
                         
                       
                     
                   
                 
               
             
             
               
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                   
                 
                   
                     
                       
                         X 
                         ⇒ 
                       
                     
                     
                       
                         ROX 
                       
                     
                   
                   
                     
                       
                         T 
                         ⇒ 
                       
                     
                     
                       
                         TMR 
                       
                     
                   
                   
                     
                       
                         R 
                         ⇒ 
                       
                     
                     
                       
                         R110 
                       
                     
                   
                   
                     
                       
                         G 
                         ⇒ 
                       
                     
                     
                       
                         R6G 
                       
                     
                   
                 
               
             
           
         
       
     
     Here, the vector C expresses the fluorescence strength of each fluorescent dye having been used. Therefore, the elements Cx, C T , C R , and C G  of the vector C express the fluorescence strength of ROX, TMR, R 110 , and R6G respectively. The vector f expresses the signal strength observed by the photodetector  504 . The elements f 0  to f 19  of the vector f express the signal strength in the wavelengths λ(0) to λ(19) respectively. The elements f 0  to f 19  may be an arithmetic mean and the like of the signal strength of the vicinity of the wavelengths λ(0) to λ(19) respectively. 
      Also, in the measurement signal of each of the wavelengths λ(0) to λ(19) detected by the photodetector  504 , Raman scattering light from the polymer filled in the capillaries  519  is included as a base line signal in addition to a signal by the fluorescent dye. Therefore, when the vector f is to be calculated, it is required to remove this base line signal beforehand. 
     As an example of the removal method for the base line signal, the spectrum of the Raman scattering light is obtained before shipment of the device beforehand, and the spectrum is stored in the computation control circuit  503  as the base line signal. Also, the signal by a fluorescent dye is obtained by deducting the base line signal from the measurement signal at each time, and it can be made the vector f. Alternatively, the minimum value in the vicinity of each time can be made the base line signal value at the time. 
     In converting the measurement vector f into a fluorescence strength vector, the matrix M(r)k is used. 
     The correction unit  5035  calculates the fluorescence strength of each fluorescent dye from the measurement spectrum by the mathematical expression 3 described above. By performing this processing to the spectrum of each capillary  519  of each time, the time series data of the fluorescence strength of each capillary  519  can be obtained. These time series data of the fluorescence strength are hereinafter referred to a fluorescence strength waveform. 
     Step S 15   
     In step S 15 , the correction unit  5035  performs peak detection with respect to the fluorescence strength waveform described above. In peak detection, the center position of the peak (peak time), the height of the peak, and the width of the peak are mainly important. The center position of the peak corresponds to the DNA fragment length. The height of the peak is used for quality evaluation of the magnitude of the DNA density in the sample, and so on. The width of the peak is also important in evaluating the quality of the sample and the electrophoresis result. As one of the methods for estimating a peak parameter of such actual data, Gaussian fitting which is a known technology can be used. 
       FIG.  11    is a drawing showing a concept of the Gaussian fitting. As shown in  FIG.  11   , the Gaussian fitting is processing for calculating such parameter (average value µ, standard deviation σ, and maximum amplitude value A) that the Gauss function g approximates the actual data most with respect to the actual data in a constant interval. As an indicator expressing the degree of approximation of the actual data, the least square error of the actual data and the Gauss function value is used frequently. As a numerical value calculation method for minimizing this least square error, the parameter can be optimized using a method such as a Gauss-Newton method. Alternatively, such method that accuracy of a case where two or more peak waveforms are mixed, a case where the data in the vicinity of the peak are dissymmetric, and so on is improved may be applicable. Also, when the variance σ of the Gauss function g is determined, the full width at half maximum (FWHM) of it is obtained by an expression shown in  FIG.  11   . This value can be made the peak width. 
     Thus, the correction unit  5035  obtains the peak parameter with respect to the fluorescence strength waveform of all fluorescent dyes. At this time, when the width of the peak and the height of the peak do not fulfill a predetermined threshold condition, they may be removed from the peak. 
     By the operation described above, the signal strength of the actual sample obtained by migration of 7.5 kV is calculated accurately using the matrix M obtained by the migration voltage of 15 kV. Although a combination of predetermined migration voltage is exemplified in the present embodiment, in practice, the operator can optionally select the migration voltage of the second spectral calibration (step S 11 ) and the actual sample migration (step S 12 ) within a range having been registered in the correction factor database  5034 . 
     Technical Effects 
     As described above, in the first embodiment, before shipment of the multi-capillary electrophoresis device  500 , migration is performed in a same condition as the first spectral calibration and the actual sample with plural migration voltages, and the correction factor matrix K for correcting the deviation of the spectrum is acquired for every combination of the migration voltage and is registered in the correction factor database  5034  along with the information of the fluorescent dye. An operator having purchased the device can perform the second spectral calibration and migration of the actual sample with a combination of the optional migration voltage registered in the correction factor database  5034 . Also, even when the operator may change the voltage of the time of migration of the actual sample, the reference spectrum and the fluorescence spectrum of the actual sample do not deviate from each other, therefore even when the second spectral calibration is not repeated, a correct fluorescence strength can be acquired. 
     Second Embodiment 
     Although the matrix M′ is acquired using the matrix standard in the first embodiment, the second embodiment proposes a method for acquiring the matrix M′ using a known DNA sample. The known DNA sample is a PCR product of a DNA, a commercially available standard sample, and so on. In the present embodiment, all of the matrix standard, the known DNA sample, and the actual sample as an example are to be labeled by ROX, TMR, R 110 , and R6G. Also, the times (t0′, t1′, t2′ and t3′) at which each fluorescent dye produces light in isolation during migration of the known DNA sample are to be known. 
       FIG.  12    is a flowchart showing an analysis method of a sample related to the second embodiment. 
     In step S 21 , in a manner similar to step S 1 , the manufacturer performs spectral calibration using the matrix standard, and the measurement value computation unit  5032  acquires the matrix M. The migration voltage is made 15 kV. 
     In step S 22 , the manufacturer causes the known DNA sample to migrate. 
     In step S 23 , the measurement value computation unit  5032  acquires a spectrum at the time (t0′, t1′, t2′ and t3′) at which each fluorescent dye produces light in isolation, and creates the matrix M′ from the strength ratio of each fluorescent dye. The migration voltage is made 7.5 kV. 
     In step S 24 , in a manner similar to step S 3 , the correction factor computation unit  5033  calculates the correction factor matrix K based on the matrices M and M′. The correction factor matrix K is registered in the correction factor database  5034  along with the information of the migration voltage and the fluorescent dye. As described in step S 1  of the first embodiment, in an actual operation, steps S 21  and S 21  are performed with various migration voltages, and the plural matrices M and M′ are acquired. When migration is effected with plural voltages, all correction factor matrices K are registered. 
     In a manner similar to the first embodiment, steps S 21  to S 24  have been performed by the manufacturer side before shipment of the multi-capillary electrophoresis device  500 , and the correction factor matrix K has already been registered in the correction factor database  5034 . The work actually performed by the operator having purchased the device becomes the next step S 25  and onward. Here, it is assumed that, after step S 24 , the capillaries  519  have been detached and attached at the time of transportation, and the positional relation between the photodetector  504  and the capillaries  519  has changed. When detachment and attachment of the capillaries  519  have not been performed in step S 24  and onward, out of the matrices M obtained in step S 21 , one with a migration voltage same as that of migration of the actual sample (step S 21 ) can be selected to be made the matrix M(r)k described below. 
     In step S 25 , the operator performs the second spectral calibration in a manner similar to step S 11 , and the measurement value computation unit  5032  acquires the matrix M(r). Although the migration voltage in step S 25  is made 15 kV as an example, in an actual operation, an optional one can be selected out of the migration voltages having been registered in the correction factor database  5034 . 
     In step S 26 , in a manner similar to step S 12 , the operator performs migration of the actual sample. Although the migration voltage here is made 7.5 kV as an example, in actual practice, the migration voltage can be selected optionally from those having been registered in the correction factor database  5034 . 
      Since steps S 27  to S 29  are similar to steps S 13  to S 15  ( FIG.  9   ) explained in the first embodiment, explanation thereof will be omitted. 
     By the operation described above, even when the migration voltage differs between the time of the first spectral calibration (step S 21 ) and the time of migration of the actual sample (step S 25 ), the reference spectrum and the fluorescence spectrum of the actual sample do not deviate from each other, and the fluorescence strength of the actual sample is calculated accurately. Although a combination of predetermined migration voltage is exemplified here, in practice, the operator can optionally select the migration voltage of the second spectral calibration (step S 25 ) and the actual sample migration (step S 26 ) within a range having been registered in the correction factor database  5034 . 
     Technical Effect 
     As described above, in the second embodiment, in a manner similar to the first embodiment, the operator having purchased the device can perform the second spectral calibration and migration of the actual sample with a combination of optional migration voltages having been registered in the correction factor database  5034 . Also, even when the operator may change the voltage of the time of migration of the actual sample, the reference spectrum and the fluorescence spectrum of the actual sample do not deviate from each other, therefore even when the second spectral calibration is not repeated, a correct fluorescence strength can be acquired. 
     Experiment Example 1 
     The effect of the second embodiment was confirmed by a procedure described below. 
     Sample 
     As the matrix standard for the time of the first spectral calibration (step S 21 ), BigDye (Registered Trade Mark) Terminator v3.1 Matrix Standards (Dye Set Z) (made by Applied Biosystems) was used. For both of the known DNA sample (step S 22 ) and the actual sample (step S 26 ), 3500/3500×L Sequencing Standards, BigDye (Registered Trade Mark) Terminator v3.1 (made by Applied Biosystems) was used. With respect to all of the samples described above, ROX, TMR, R 110 , and R6G are used as the fluorescent dye. 
     Analysis Procedure 
     In the experiment example 1, as a verification of the second embodiment, steps S 21  to S 26  were performed in a manner described in steps S 1 , S 12 , S 2 , S 3 , S 11 , and S 12  respectively. The capillary length at the time of migration was 36 cm, the applied voltage at the time of pouring the sample was 1.6 kV, the applied voltage at the time of migration was 15 kV at the time of the first spectral calibration (step S 21 ), and the voltage at the time of migration of the known sample and migration of the actual sample was 7.5 kV. 
     Next, steps S 27  to S 29  were performed in a manner described in steps S 13  to S 15 . 
     As a control of the second embodiment, light intensity calculation and peak detection of the actual sample were performed not applying the correction factor matrix K but using the matrix M. Between the second embodiment and its control, the signal strength of the pseudo peak was compared. 
     Experiment Result 
       FIG.  13    is a drawing showing the result of the experiment example 1. In  FIG.  3   , the matrix M, the matrix M′, and the correction factor matrix K obtained by steps S 21 , S 23 , and S 24  are shown. 
     With respect to the graph in  FIG.  13   , the horizontal axis shows the peak time, and the vertical axis shows the fluorescence strength. Although the pseudo peak is confirmed in the control, it is obvious that the pseudo peak is reduced in the method of the second embodiment. 
     Third Embodiment 
     Although explanation was made on a case where the migration voltage was different between the time of the second spectral calibration and the time of migration of the actual sample was explained in the first and second embodiments, a case where the fluorescent dye is different will be explained in the third embodiment. In the present embodiment, as an example, the matrix standard used in the first spectral calibration is to be labeled by FAM, JOE, TMR, and CXR. Also, the actual sample is to be labeled by R6G, R 110 , TMR, and ROX. 
       FIG.  14    is a flowchart showing an analysis method of a sample related to the third embodiment. 
     In step S 31 , in a manner similar to step S 1 , the manufacturer performs the spectral calibration using the matrix standard, and the measurement value computation unit  5032  acquires the matrix M. However, for the sample, a matrix standard labeled by FAM, JOE, TMR, and CXR is used. 
     In step S 32 , in a manner similar to step S 1 , the manufacturer acquires the matrix M′ . However, for the sample, a matrix standard labeled by R6G, R 110 , TMR, and ROX is used. 
     In step S 33 , in a manner similar to step S 3 , the correction factor computation unit  5033  calculates the correction factor matrix K based on the matrices M and M′. The correction factor matrix K is registered in the correction factor database  5034  along with the information of the migration voltage and the fluorescent dye. As described in step S 1  of the first embodiment, in an actual operation, steps S 31  and S 32  are performed with a combination of various fluorescent dyes, and the plural matrices M and M′ are acquired. When migration is effected with a combination of plural fluorescent dyes, all correction factor matrices K are registered. 
     In a manner similar to the first embodiment, steps S 31  to S 33  have been performed by the manufacturer side before shipment of the multi-capillary electrophoresis device  500 , and the correction factor matrix K has already been registered in the correction factor database  5034 . The work actually performed by the operator having purchased the device becomes the next step S 34  and onward. Here, it is assumed that, after step S 33 , the capillaries  519  have been detached and attached at the time of transportation, and the positional relation between the photodetector  504  and the capillaries  519  has changed. When detachment and attachment of the capillaries  519  have not been performed in step S 33  and onward, out of the matrices M obtained in step S 31 , one with a fluorescent dye same as that of migration of the actual sample (step S 35 ) can be selected to be made the matrix M(r)k described below. 
     In step S 34 , the operator performs the second spectral calibration in a manner similar to step S 11 , and the measurement value computation unit  5032  acquires the matrix M(r). Although the fluorescent dye in step S 34  uses FAM, JOE, TMR, and CXR in the present embodiment, in an actual operation, an optional one can be selected out of the fluorescent dyes having been registered in the correction factor database  5034 . 
     In step S 35 , in a manner similar to step S 12 , the operator performs migration of the actual sample. As an example, the actual sample is to be labeled by R6G, R 110 , TMR, and ROX. However, in an actual operation, an optional one can be selected out of the fluorescent dyes having been registered in the correction factor database  5034 . 
     Since steps S 36  to S 38  are similar to steps S 13  to S 15  ( FIG.  9   ) explained in the first embodiment, explanation thereof will be omitted. 
     By the operation described above, even when the fluorescent dye may differ between the time of the spectral calibration (step S 31 ) and the time of migration of the actual sample (step S 35 ), the reference spectrum and the fluorescence spectrum of the actual sample do not deviate from each other, and the fluorescence strength of the actual sample is calculated accurately. Although a combination of predetermined fluorescent dye is exemplified here, in practice, the operator can optionally change the fluorescent dye of the second spectral calibration (step S 34 ) and the actual sample migration (step S 35 ) within a range having been registered in the correction factor database  5034 . 
     Technical Effect 
     As described above, in the third embodiment, before shipment of the multi-capillary electrophoresis device  500 , migration with a condition same as that of the first spectral calibration and the actual sample is performed using a sample labeled by a set of different fluorescent dyes, and the correction factor matrix K for correcting the deviation of the spectrum is acquired for each combination of the fluorescent dye and is registered in the correction factor database  5034 . The operator having purchased the device can perform the second spectral calibration and migration of the actual sample with a combination of optional fluorescent dyes having been registered in the correction factor database  5034 . Also, even when the operator may change the fluorescent dye of the time of migration of the actual sample, the reference spectrum and the fluorescence spectrum of the actual sample do not deviate from each other, therefore even when the second spectral calibration is not repeated, an accurate fluorescence strength can be acquired. 
     Experiment Example 2 
     The effect of the third embodiment was confirmed by a procedure described below. 
     Sample 
     As the matrix standard for the time of the first spectral calibration (step S 31 ), PowerPlex (Registered Trade Mark) 4C Matrix Standards (made by Promega Corporation) was used. For acquisition of the matrix M′ (step S 32 ), BigDye (Registered Trade Mark) Terminator v3.1 Matrix Standards (Dye Set Z) (made by Applied Biosystems) was used. For the actual sample (step S 35 ), 3500/3500×L Sequencing Standards, BigDye (Registered Trade Mark) Terminator v3.1 (made by Applied Biosystems) was used. 
       FIG.  15 A  is a drawing showing a fluorescent dye used in the experiment example 2. As shown in  FIG.  15 A , for the matrix standard (step S 31 ), FAM, JOE, TMR, and CXR are used as a fluorescent dye. Also, ROX, TMR, R 110 , and R6G are used for both of steps S 32  and S 35  as a fluorescent dye. 
     Analysis Procedure 
     In the experiment example 2, as a verification of the third embodiment, steps S 31 , S 32 , S 33 , and S 34  were performed in a manner described in steps S 1 , S 1 , S 3 , and S 11  respectively. The capillary length at the time of migration was 36 cm, the applied voltage at the time of pouring the sample was 1.6 kV, the applied voltage at the time of migration was 15 kV at all steps at the time of the first spectral calibration (step S 31 ). 
     Next, steps S 35  to S 38  were performed in a manner described in steps S 12  to S 15 . 
     As a control of the third embodiment, light intensity calculation and peak detection of the actual sample were performed not applying the correction factor matrix K but using the matrix M. Between the third embodiment and its control, the signal strength of the pseudo peak was compared. 
     Experiment Result 
       FIG.  15 B  is a drawing showing the result of the experiment example 2. In  FIG.  15 B , the matrix M, the matrix M′, and the correction factor matrix K obtained in steps S 31  to S 33  are shown. 
     With respect to the graph in  FIG.  15 B , the horizontal axis shows the peak time, and the vertical axis shows the fluorescence strength. Although the pseudo peak is confirmed in the control, it is obvious that the pseudo peak is reduced in the method of the third embodiment. 
     Fourth Embodiment 
     In the first embodiment, the correction factor matrix K obtained in a predetermined device was applied to data of the actual sample obtained in the same device. The fourth embodiment proposes a method for applying the correction factor matrix K obtained in a predetermined device to data of the actual sample obtained in a separate device. 
     In the present embodiment, as an example, explanation will be made exemplifying a case where the fluorescent dye is different in a similar manner to the third embodiment ( FIG.  14   ). As an example, the matrix standard used in the first spectral calibration (step S 31 ) is to be labeled by FAM, JOE, TMR, and CXR. Also, the actual sample is to be labeled by R6G, R 110 , TMR, and ROX. 
      The first spectral calibration (step S 31 ), acquisition of the matrix M′ (step S 32 ), and calculation of the correction factor matrix K (step S 33 ) are performed in a similar manner to the third embodiment by the manufacturer side with a predetermined multi-capillary electrophoresis device A. The device A transmits the correction factor matrix K to a different device (plural devices) through a network for example, and causes the correction factor matrix K to be registered in the correction factor database  5034  of each device. For example, the correction factor matrix K may be registered in all multi-capillary electrophoresis devices which are before shipment. 
     Step 34 and onward can be performed in an optional device where the correction factor matrix K same as that in the device A has been registered. 
     Technical Effect 
     As described above, in the fourth embodiment, the correction factor matrix K acquired using a predetermined multi-capillary electrophoresis device is also registered in other devices. Thus, since it is not required to measure the correction factor matrix K in each device, the cost and labor on the manufacturer side are reduced. 
     Fifth Embodiment 
     In the first embodiment, by multiplication of the correction factor matrix K to the matrix M(r) obtained in the second spectral calibration, deviation between the matrix M(r) and the fluorescence spectrum of the actual sample was prevented. The fifth embodiment proposes a method for preventing deviation by changing the wavelength width (signal acquisition width) of a signal detected by the photodetector. With respect to processing similar to that of the first embodiment, explanation will be omitted. 
       FIG.  16    is a flowchart showing an analysis method of a sample related to the fifth embodiment. 
     In the present embodiment, the photodetector  504  of the multi-capillary electrophoresis device  500  is to measure the signal strength in the twenty wavelengths λ(0) to λ(19) in sampling the data. Although the twenty wavelengths were cited here as only an example, in practice, an arithmetic mean of the signal strength in the vicinity of each of the wavelengths λ(0) to λ(19) may be taken. Also, the matrix standard is to be labeled by CXR, and the actual sample is to be labeled by ROX. The fluorescence spectra of these fluorescent dyes are known and do not agree to each other. 
     Since the photodetector  504  of the present embodiment detects the twenty wavelengths only, the fluorescence spectrum of CXR is expressed by a vector Vm formed of twenty pieces of elements, and the fluorescence spectrum of ROX is expressed by a vector Vs formed of twenty pieces of elements. 
     In step S 51 , the measurement value computation unit  5032  defines the twenty wavelengths (signal acquisition width) so that the correlation coefficient of the vector Vm and the vector Vs is maximized. At this time, a weight may be given to the local maximum or its vicinity of the spectrum. Further, if it is not an imposition in an actual practice, the correlation coefficient only has to be made sufficiently high, and is not necessarily required to be made a maximum value. That is to say, the signal acquisition width is defined so that the correlation coefficient becomes a predetermined value or more. 
     In step S 52 , in a similar manner to step S 1 , the operator performs the spectral calibration. At this time, the measurement value computation unit  5032  calculates a vector Vc formed of twenty pieces of elements. 
     In step S 53 , in a similar manner to step S 12 , the operator performs migration of the actual sample. Here, the spectrum f acquired by the measurement value computation unit  5032  expresses the signal strength observed by the photodetector  504 . Elements f0 to f19 thereof respectively express the signal strength in the wavelengths λ(0) to λ(19). 
     In step S 54 , the correction unit  5035  calculates the fluorescence strength. To be more specific, with respect to the spectrum of each of the capillaries  519  at each time, the strength ratio of each fluorescent dye in each of the wavelengths λ(0) to λ(19) only has to be multiplied and to be added up. Expression of it by a matrix is as per the mathematical expression 4 below.  
     
       
         
           
             
               
                 
                   
                     c=Vmf 
                   
                 
               
               
                 
                   
                     f= 
                     
                       
                         
                           f 
                           0 
                         
                           
                         
                           f 
                           1 
                         
                           
                         
                           f 
                           2 
                         
                         ⋅ 
                         ⋅ 
                         ⋅ 
                         ⋅ 
                         ⋅ 
                         ⋅ 
                           
                         
                           f 
                           
                             18 
                           
                         
                           
                         
                           f 
                           
                             19 
                           
                         
                       
                     
                   
                 
               
               
                 
                   
                     Vm= 
                     
                       
                         
                           w 
                           0 
                         
                           
                         
                           w 
                           1 
                         
                           
                         
                           w 
                           2 
                         
                           
                         ⋅ 
                         ⋅ 
                         ⋅ 
                         ⋅ 
                         ⋅ 
                         ⋅ 
                           
                         
                           w 
                           
                             18 
                           
                         
                           
                         
                           w 
                           
                             19 
                           
                         
                       
                     
                   
                 
               
             
           
         
       
     
     The vector c is a fluorescence strength spectrum. The vector f expresses the signal strength detected by the photodetector  504 . The elements f 0  to f 19  of it respectively express the signal strength in the wavelengths λ(0) to λ(19). 
     Also, as described in step S 14  of the first embodiment, in the measurement signal of each of the wavelengths λ(0) to λ(19) detected by the photodetector  504 , a Raman scattering light from the polymer filled within the capillary is included as a base line signal in addition to a signal by the fluorescent dye. Therefore, in calculating the vector f, it is required to remove this base line signal beforehand. Base line removal may be performed by a method described in step S 14 . 
     In step S 55 , the correction unit  5035  performs peak detection in a similar manner to step S 15 . 
     By the operation described above, even when the fluorescent dye may differ between the time of the spectral calibration (step S 1 ) and the time of migration of the actual sample (step S 12 ), the reference spectrum and the fluorescence spectrum of the actual sample do not deviate from each other, and the fluorescence strength of the actual sample is calculated accurately. 
     Technical Effect 
     As described above, in the fifth embodiment, the photodetector  504  detects light from the capillaries  519  with such wavelength that the correlation coefficient of the fluorescence spectra of the plural fluorescent dyes becomes large. Thus, since the matrix M(r)k is not required at the time of migration, the time required for the analysis can be shortened, and a burden on the computation control circuit  503  can be reduced. 
     Experiment Example 3 
     The effect of the fifth embodiment was confirmed by a procedure described below. 
     Device 
     The multi-capillary electrophoresis device  500  ( FIG.  5   ) explained in the first embodiment can be used. However, as an example in the present embodiment, the photodetector  504  is to detect the signal strength in the twenty wavelengths between 520 nm and 690 nm. In the present experiment example 3, the signal acquisition width for causing a mutual correlation coefficient of two spectra to become sufficiently high is to have been known. The twenty wavelengths expressed by a vector is made λ test . Also, as a control, a case of acquiring signals at equal intervals of 8.9 nm in a same section is assumed, and the wavelengths of the portion of these twenty pieces are expressed by a vector as λ ctrl . The mathematical expression 5 below shows elements of λ test  and λ ctrl .  
     
       
         
           
             
               
                 λ 
                 test = 
               
             
             
               
                 
                   
                     520 
                      529 538 547 556 565 574 583 602 627 634 640 646 653 659 665 
                   
                 
               
             
             
               
                 
                   
                     671 
                      678 684 690 
                   
                 
               
             
             
               
                 λ 
                 ctrl = 
               
             
             
               
                 
                   
                     520 
                      529 538 547 556 565 574 583 592 601 605 518 627 636 645 654 
                   
                 
               
             
             
               
                 
                   
                     663 
                      672 680 690 
                   
                 
               
             
           
         
       
     
     Sample 
     As the matrix standard for the time of the spectral calibration (step S 52 ), out of the four peaks included in PowerPlex (Registered Trade Mark) 4C Matrix Standards (made by Promega Corporation), one labeled by CXR was used. For migration of the actual sample (step S 53 ), out of the four peaks included in BigDye (Registered Trade Mark) Terminator v3.1 Matrix Standards (Dye Set Z) (made by Applied Biosystems), one labeled by ROX was used. 
     Analysis Procedure 
     In the experiment example 3, as a verification of the fifth embodiment, steps S 52  and S 53  were performed in a manner described in steps S 11  and S 12  respectively. The capillary length at the time of migration was 36 cm, the applied voltage at the time of pouring the sample was 1.6 kV, the applied voltage at the time of migration was 15 kV for both of the time of the spectral calibration (step S 52 ) and the time of migration of the actual sample (step S 53 ). 
     Experiment Result 
       FIG.  17 A  is a fluorescence spectrum acquired in the experiment example 3. In  FIG.  17 A , a fluorescence spectrum obtained with λ test  is shown. The mathematical expression 6 below shows the signal strength of the vector Vm and the vector Vs in λ test . As shown in the mathematical expression 6, the correlation coefficient (corr.) of the vector Vm and the vector Vs of a case of applying λ test  (the fifth) embodiment became 0.998.  
     
       
         
           
             
               
                 Vm =  
               
             
             
               
                 
                   
                     0 
                     .01 0 
                     .02 0 
                     .02 0 
                     .03 0 
                     .03 0 
                     .04 0 
                     .06 0 
                     .06 0 
                     .16 1 
                     .00 0 
                     .86 0 
                     .71 0 
                     .58 0 
                     .43 0 
                     .35 0 
                     .29 
                   
                 
               
             
             
               
                 
                   
                     0 
                     .24 0 
                     .21 0 
                     .19 0 
                     .17 
                   
                 
               
             
             
               
                 Vs = 
               
             
             
               
                 
                   
                     0 
                     .01 0 
                     .01 0 
                     .02 0 
                     .02 0 
                     .02 0 
                     .02 0 
                     .04 0 
                     .04 0 
                     .09 1 
                     .00 0 
                     .89 0 
                     .75 0 
                     .62 0 
                     .46 0 
                     .36 0 
                     .29 
                   
                 
               
             
             
               
                 
                   
                     0 
                     .24 0 
                     .21 0 
                     .19 0 
                     .17 
                   
                 
               
             
             
               
                 c 
                 o 
                 r 
                 r 
                 . 
                 = 
                 0.998 
               
             
           
         
       
     
       FIG.  17 B  is a fluorescence spectrum acquired in the control experiment of the experiment example 3. In  FIG.  17 B , a fluorescence spectrum obtained with λ ctrl  is shown. The mathematical expression 7 below shows the signal strength of the vector Vm and the vector Vs in λ ctrl . As shown in the mathematical expression 7, the correlation coefficient (corr.) of the vector Vm and the vector Vs of a case of λ ctrl  became 0.986.  
     
       
         
           
             
               
                 Vm = 
               
             
             
               
                 
                   
                     0 
                     .01 0 
                     .02 0 
                     .02 0 
                     .03 0 
                     .03 0 
                     .04 0 
                     .06 0 
                     .06 0 
                     .16 0 
                     .38 0 
                     .70 1 
                     .00 0 
                     .99 0 
                     .79 0 
                     .59 0 
                     .39 
                   
                 
               
             
             
               
                 
                   
                     0 
                     .30 0 
                     .23 0 
                     .20 0 
                     .17 
                   
                 
               
             
             
               
                 Vs = 
               
             
             
               
                 
                   
                     0 
                     .01 0 
                     .01 0 
                     .02 0 
                     .02 0 
                     .02 0 
                     .02 0 
                     .04 0 
                     .04 0 
                     .09 0 
                     .24 0 
                     .54 0 
                     .87 1 
                     .00 0 
                     .83 0 
                     .62 0 
                     .42 
                   
                 
               
             
             
               
                 
                   
                     0 
                     .30 0 
                     .23 0 
                     .20 0 
                     .17 
                   
                 
               
             
             
               
                 c 
                 o 
                 r 
                 r 
                 . 
                 = 
                 0.986 
               
             
           
         
       
     
     As is clear from the mathematical expressions 6 and 7, it is known that the mutual correlation coefficient became higher than that of the control (λ ctrl ) in applying λ test  namely the fifth embodiment. 
     Sixth Embodiment 
     In the first to the third embodiments, explanation was made on a case where the migration voltage or the fluorescent dye differed between the time of the second spectral calibration and the time of migration of the actual sample. In the sixth embodiment, a case where both of the migration voltage and the fluorescent dye differ will be explained. In the present embodiment, as an example, the matrix standard used in the first spectral calibration is to be labeled by FAM, JOE, TMR, and CXR. Also, the actual sample is to be labeled by R6G, R 110 , TMR, and ROX. The migration voltage of the time of the spectral calibration is made 15 kV, and the migration voltage of the time of migration of the actual sample is made 7.5 kV. 
       FIG.  18    is a flowchart showing an analysis method of a sample related to the sixth embodiment. 
     In step S 61 , in a similar manner to step S 1 , the manufacturer performs the spectral calibration using the matrix standard, and the measurement value computation unit  5032  acquires the matrix M. The migration voltage is made 15 kV. 
     In step S 62 , in a similar manner to step S 2 , the manufacturer acquires the matrix M′. However, for the sample, a matrix standard labeled by R6G, R 110 , TMR, and ROX is used. The migration voltage at this time is 7.5 kV. 
     In step S 63 , in a manner similar to step S 3 , the correction factor computation unit  5033  calculates the correction factor matrix K based on the matrices M and M′. The correction factor matrix K is registered in the correction factor database  5034  along with the information of the migration voltage and the fluorescent dye. As described in step S 1  of the first embodiment, in an actual operation, steps S 61  and S 62  are performed with various combinations of the migration voltage and the fluorescent dye, and the plural matrices M and M′ are acquired. When migration is effected with plural migration voltages and plural fluorescent dyes, all correction factor matrices K are registered. 
     In a manner similar to the first embodiment, steps S 61  to S 63  have been performed by the manufacturer side before shipment of the multi-capillary electrophoresis device  500 , and the correction factor matrix K has been registered already in the correction factor database  5034 . The work actually performed by the operator having purchased the device becomes the next step S 64  and onward. Here, it is assumed that, after step S 63 , the capillaries  519  have been detached and attached at the time of transportation, and the positional relation between the photodetector  504  and the capillaries  519  has changed. When detachment and attachment of the capillaries  519  have not been performed in step S 63  and onward, out of the matrices M obtained in step S 61 , one with a fluorescent dye which is the same as that of migration of the actual sample (step S 65 ) can be selected to be made the matrix M(r)k described below. 
     In step S 64 , the operator performs the second spectral calibration in a manner similar to step S 11 , and the measurement value computation unit  5032  acquires the matrix M(r). Although the migration voltage and the fluorescent dye in step S 64  can be made to be the same as those of the first spectral calibration (step S 61 ) as an example, in practice, optional ones can be selected out of those having been registered in the correction factor database  5034 . 
     In step S 65 , the operator performs migration of the actual sample. Although the migration voltage and the fluorescent dye used here are the same as those of the time of acquisition of the matrix M′ (step S 62 ) as an example, in practice, optional ones can be selected out of those having been registered in the correction factor database  5034 . 
     Since steps S 66  to S 68  are also similar to steps S 13  to S 15  ( FIG.  9   ) explained in the first embodiment, explanation thereof will be omitted. 
     By the operation described above, even when both of the fluorescent dye and the migration voltage to be used may differ between the time of the spectral calibration (step S 61 ) and the time of migration of the actual sample (step S 65 ), the reference spectrum and the fluorescence spectrum of the actual sample do not deviate from each other, and the fluorescence strength of the actual sample is calculated accurately. Although a combination of predetermined fluorescent dye and migration voltage was exemplified here, in practice, the operator can optionally change the migration voltage and the fluorescent dye of the second spectral calibration (step S 64 ) and the actual sample migration (step S 65 ) within a range having been registered in the correction factor database  5034 . 
     Technical Effect 
     As described above, in the sixth embodiment, before shipment of the multi-capillary electrophoresis device  500 , the first spectral calibration and migration of the actual sample are performed with different migration voltages using a sample labeled by a set of different fluorescent dyes, and the correction factor matrix K for correcting the deviation of the spectrum is acquired for each combination of the fluorescent dye and the migration voltage and is registered in the correction factor database  5034 . The operator having purchased the device can perform the second spectral calibration and migration of the actual sample with a combination of optional fluorescent dye and migration voltage having been registered in the correction factor database  5034 . Thus, according to the present embodiment, the degree of freedom of the fluorescent dye and the migration voltage used by an operator improves compared to the first to the third embodiments. 
     Seventh Embodiment 
     Although explanation was made on a case where the migration voltage or the fluorescent dye differed between the time of the second spectral calibration and the time of migration of the actual sample in the first to the third embodiments, a case where the chemical property or the composition of the polymer differs will be explained in the seventh embodiment. As described above, since the polymer is only an example of the separation medium, it is needless to mention that same operation can be applied to separation media other than the polymer. 
     Since the analysis method related to the seventh embodiment can be performed by a flow similar to that of the first embodiment for example, only the different points will be hereinafter explained. 
     In the seventh embodiment, as an example, the polymer used in the spectral calibration (steps S 1  and S 11 ) is to contain 4% of polyacrylamide. Also, the polymer used at the time of migration of the actual sample (steps S 2  and S 12 ) is to contain 7% of polyacrylamide. Both of the matrix standard and the actual sample are to be labeled by R6G, R 110 , TMR, and ROX, and the migration voltage for the both is to be made 15 kV. 
     As described in the first embodiment, in an actual operation, steps S 1  and S 2  are performed with combinations of various kinds of the polymer, and plural matrices M and M′ are acquired. The various kinds of the polymer mentioned here means a polymer containing polyacrylamide of various concentrations as an example. 
     In step S 3 , the correction factor computation unit  5033  calculates the correction factor matrix K based on the matrices M and M′ . The correction factor matrix K is registered in the correction factor database  5034  along with the information of the kind of the polymer and so on. When migration is effected using plural polymers, all correction factor matrices K are registered. 
     According to the method of the present embodiment, even when the composition of the polymer may differ between the time of the spectral calibration (step S 1 ) and the time of migration of the actual sample (step S 12 ), the reference spectrum and the fluorescence spectrum of the actual sample do not deviate from each other, and the fluorescence strength of the actual sample is calculated accurately. Although a combination of the predetermined composition was exemplified here, in practice, the operator can optionally change the chemical property of the polymer of the second spectral calibration (step S 11 ) and the actual sample migration (step S 12 ) within a range having been registered in the correction factor database  5034 . Further, the method of the present embodiment can be applied also to a case where the composition of the polymer differs. 
     Eighth Embodiment 
     Although explanation was made on a case where the migration voltage or the fluorescent dye differed between the time of the spectral calibration and the time of migration of the actual sample in the first to the third embodiments, a case where the length of the capillaries  519  differs will be explained in the eighth embodiment. 
     Since the analysis method related to the eighth embodiment can be performed by a flow similar to that of the first embodiment for example, the different points will be hereinafter explained. 
     In the eighth embodiment, as an example, the capillary length at the time of the spectral calibration (steps S 1  and S 11 ) is to be made 50 cm, and the capillary length at the time of migration of the actual sample (steps S 2  and S 12 ) is made 36 cm. 
     As described in the first embodiment, in an actual operation, steps S 1  and S 2  are performed with combinations of various capillary lengths, and plural matrices M and M′ are acquired. 
     In step S 3 , the correction factor computation unit  5033  calculates the correction factor matrix K based on the matrices M and M′ . The correction factor matrix K is registered in the correction factor database  5034  along with the information of the capillary length. When migration is effected with plural capillary lengths, all correction factor matrices K are registered. 
     According to the method of the present embodiment, even when the capillary length may differ between the time of the spectral calibration (step S 1 ) and the time of migration of the actual sample (step S 12 ), the reference spectrum and the fluorescence spectrum of the actual sample do not deviate from each other, and the fluorescence strength of the actual sample is calculated accurately. Although a combination of the predetermined length was exemplified here, in practice, the operator can optionally change the capillary length of the second spectral calibration (step S 11 ) and the actual sample migration (step S 12 ) within a range having been registered in the correction factor database  5034 . 
     Ninth Embodiment 
     Although explanation was made on a case where the migration voltage or the fluorescent dye differed between the time of the spectral calibration and the time of migration of the actual sample in the first to the third embodiments, a case where the composition or the chemical property of the positive pole buffer differs will be explained in the ninth embodiment. 
     Since the analysis method related to the ninth embodiment can be performed by a flow similar to that of the first embodiment for example, the different points will be hereinafter explained. 
     In the ninth embodiment, as an example, pH of the positive pole buffer used in the spectral calibration (steps S 1  and S 11 ) is to be 7.5. pH of the positive pole buffer at the time of migration of the actual sample (steps S 2  and S 12 ) is to be 8.0. 
     As described in the first embodiment, in an actual operation, steps S 1  and S 2  are performed with combinations of the positive pole buffer of various pH, and plural matrices M and M′ are acquired. 
     In step S 3 , the correction factor computation unit  5033  calculates the correction factor matrix K based on the matrices M and M′ . The correction factor matrix K is registered in the correction factor database  5034  along with the information of pH of the positive pole buffer. When migration is effected using the positive pole buffers of plural pH, all correction factor matrices K are registered. 
     According to the method of the present embodiment, even when pH of the positive pole buffer may differ between the time of the spectral calibration (step S 1 ) and the time of migration of the actual sample (step S 12 ), the reference spectrum and the fluorescence spectrum of the actual sample do not deviate from each other, and the fluorescence strength of the actual sample is calculated accurately. Although a combination of the predetermined pH of the positive pole buffer was exemplified here, in practice, the operator can optionally change pH of the positive pole buffer of the second spectral calibration (step S 11 ) and the actual sample migration (step S 12 ) within a range having been registered in the correction factor database  5034 . Further, the method of the present embodiment can be applied also to a case where the composition of the positive pole buffer differs. 
     Tenth Embodiment 
     Although explanation was made on a case where the chemical property of the positive pole buffer differed in the ninth embodiment, a case where the composition or the chemical property of a negative pole buffer differs will be explained in the tenth embodiment. 
     In the tenth embodiment, as an example, pH of the negative pole buffer used in the spectral calibration (steps S 1  and S 11 ) is to be 7.5. pH of the negative pole buffer at the time of migration of the actual sample (steps S 2  and S 12 ) is to be 8.0. Since other points are similar to the ninth embodiment, explanation thereof will be omitted. 
     According to the method of the present embodiment, even when pH of the negative pole buffer may differ between the time of the spectral calibration (step S 1 ) and the time of migration of the actual sample (step S 12 ), the reference spectrum and the fluorescence spectrum of the actual sample do not deviate from each other, and the fluorescence strength of the actual sample is calculated accurately. Although a combination of the predetermined pH was exemplified here, in practice, the operator can optionally change pH of the positive pole buffer of the second spectral calibration (step S 11 ) and the actual sample migration (step S 12 ) within a range having been registered in the correction factor database  5034 . Further, the method of the present embodiment can be applied also to a case where the composition of the negative pole buffer differs. 
     Eleventh Embodiment 
     Although explanation was made on a case where the chemical property of the positive pole buffer differed in the ninth embodiment and a case where the chemical property of the negative pole buffer differed in the tenth embodiment, a case where the chemical property or the composition of a sample solution differs will be explained in the eleventh embodiment. 
     Since the analysis method related to the eleventh embodiment can be performed by a flow similar to that of the first embodiment for example, the different points will be hereinafter explained. 
     In the eleventh embodiment, as an example, pH of a solution of the matrix standard used in the spectral calibration (steps S 1  and S 11 ) is to be 7.5. pH of a solution of the actual sample used in steps S 2  and S 12  is to be 8.0. 
     As described in the first embodiment, in an actual operation, steps S 1  and S 2  are performed with combinations of the samples of various pH, and plural matrices M and M′ are acquired. 
     In step S 3 , the correction factor computation unit  5033  calculates the correction factor matrix K based on the matrices M and M′ . The correction factor matrix K is registered in the correction factor database  5034  along with the information of pH of the sample solution. When migration is effected using the samples of plural pH, all correction factor matrices K are registered. 
     According to the method of the present embodiment, even when pH of the sample solution may differ between the time of the spectral calibration (step S 1 ) and the time of migration of the actual sample (step S 12 ), the reference spectrum and the fluorescence spectrum of the actual sample do not deviate from each other, and the fluorescence strength of the actual sample is calculated accurately. Although a combination of the sample solution of predetermined pH was exemplified here, in practice, the operator can optionally change pH of the sample solution of the second spectral calibration (step S 11 ) and the actual sample migration (step S 12 ) within a range having been registered in the correction factor database  5034 . Further, the method of the present embodiment can be applied also to a case where the composition of the sample solution differs. 
     Twelfth Embodiment 
     In the twelfth embodiment, explanation will be made on a case where the temperature of the constant temperature reservoir  505  differs. 
     Since the analysis method related to the twelfth embodiment can be performed by a flow similar to that of the first embodiment for example, the different points will be hereinafter explained. 
     In the twelfth embodiment, as an example, the temperature of the constant temperature reservoir  505  at the time of the spectral calibration (steps S 1  and S 11 ) is to be 42° C. Also, the temperature of the constant temperature reservoir  505  at the time of migration of the actual sample (steps S 2  and S 12 ) is to be 60° C. 
     As described in the first embodiment, in an actual operation, steps S 1  and S 2  are performed with combinations of various temperatures, and plural matrices M and M′ are acquired. 
     In step S 3 , the correction factor computation unit  5033  calculates the correction factor matrix K based on the matrices M and M′ . The correction factor matrix K is registered in the correction factor database  5034  along with the information of the temperature of the constant temperature reservoir  505 . When migration is effected with the temperature of the constant temperature reservoir  505  being kept at plural temperatures, all correction factor matrices K are registered. 
     According to the method of the present embodiment, even when the temperature of the constant temperature reservoir  505  may differ between the time of the spectral calibration (step S 1 ) and the time of migration of the actual sample (step S 12 ), the reference spectrum and the fluorescence spectrum of the actual sample do not deviate from each other, and the fluorescence strength of the actual sample is calculated accurately. Although a predetermined combination of the temperature of the constant temperature reservoir  505  was exemplified here, in practice, the operator can optionally change the temperature of the constant temperature reservoir  505  of the second spectral calibration (step S 11 ) and the actual sample migration (step S 12 ) within a range having been registered in the correction factor database  5034 . 
     On Manifestation 
     As an example of a method for confirming infringement of the present disclosure, verification described below can be cited. Explanation will be made based on  FIG.  9   . 
     In the device of the object, step S 11  (spectral calibration) is performed. Although the migration voltage is 15 kV at this time, in practice, the analysis is effected at such migration speed that the migration voltage becomes equivalent to 7.5 kV. The migration speed can be adjusted by adding a salt of an appropriate amount to the sample. Alternatively, while registering the migration voltage to be 15 kV in terms of the device, in practice, migration may be effected with the migration voltage of 7.5 kV. Thereafter, according to the method of the first embodiment, the actual sample is analyzed (steps S 13  to S 15 ). At this time, when the pseudo peak increases compared to the time of the first embodiment, it is highly possible that the device of the object has applied the correction factor determined for each migration voltage to the fluorescence spectrum of the matrix standard. 
     Also, such verification as described below is also possible. Explanation will be made based on  FIG.  14   . In this case, in step S 31 , a fluorescent dye different from the actual fluorescent dye is registered. When the pull-up increases compared to the third embodiment after the analysis of the actual sample (steps S 34  to S 36 ), it is highly possible that the correction factor determined for each fluorescent dye has been applied to the fluorescence spectrum of the matrix standard. 
     Modification 
     The present disclosure is not limited to the embodiments described above, and includes various modifications. For example, the embodiments described above were explained in detail for easy understanding of the present disclosure, and it is not necessarily required to include all configurations having been explained. Also, a part of an embodiment can be substituted by a configuration of other embodiments. Also, a configuration of an embodiment can be added with a configuration of other embodiments. Further, with respect to a part of the configuration of each embodiment, it is possible to add a part of a configuration of other embodiments, to be deleted, or to be substituted. 
     
       
         
           
               
             
               
                 Reference Signs List 
               
             
            
               
                   101 ... device body, 
               
               
                   102 ...control computer, 
               
               
                   103 ...computation control circuit 
               
               
                   104 ...photodetector, 
               
               
                   105 ...constant temperature reservoir, 
               
               
                   106 ...capillary array, 
               
               
                   107 ...light source, 
               
               
                   108 ...light irradiation unit, 
               
               
                   109 ...load header, 
               
               
                   110 ...negative pole end, 
               
               
                   111 ...negative pole buffer container 
               
               
                   112 ...sample container, 
               
               
                   113 ...polymer cartridge, 
               
               
                   114 ...positive pole buffer container, 
               
               
                   115 ...positive pole, 
               
               
                   116 ...D.C. power source, 
               
               
                   117 ...array header, 
               
               
                   118 ...transporter, 
               
               
                   119 ...capillary, 
               
               
                   120 ...syringe mechanism, 
               
               
                   121 ... sharp point portion, 
               
               
                   122 ...polymer cartridge upper portion, 
               
               
                   123 ...heating/cooling mechanism, 
               
               
                   201 ...laser light, 
               
               
                   202 ...reflection mirror, 
               
               
                   203 ...condenser lens