Abstract:
Provided is an imaging apparatus which can detect an occurrence of a vibration using imaged images without a new constituent such as a vibration sensor being added thereto, wherein when imaging a position reference image for checking positions of observation targets on the image and a plurality of luminance reference images for checking the spectral distributions of the observation targets, a first vibration detection unit detects a vibration for the position reference image, a mask image for checking the positions is created on the basis of the position reference image for which no vibration is detected, and the luminance reference image is compared with the mask image every time the luminance reference image is imaged, thereby to judge whether positions deviate or not, and in a case where the positions deviate, the position reference image is re-imaged, thereby eliminating the image imaged with a vibration occurring.

Description:
FIELD OF THE INVENTION  
       [0001]     The present invention relates to an imaging apparatus that stores acquired and processed images as well as analyzes the same. More particularly, the present invention relates to an imaging apparatus which eliminates vibrating components from the target images and performs a high reliability data analysis.  
       BACKGROUND OF THE INVENTION  
       [0002]     In recent years, an image processing technique using an imaging apparatus is applied in various fields, and there are many applications to such as a gene expression analyzer in medical fields. For example, there is a gene expression analyzer using a real time PCR method, a DNA micro array (also referred to as DNA chip), or a semiconductor nanocrystal.  
         [0003]     A description will be given hereinafter of a gene expression analyzer using a fluorescence microscope as a prior art imaging apparatus.  
         [0004]     Imaging targets by the prior art gene expression analyzer are beads having various spectral characteristics, each having a diameter of about 10 μm. A specific mRNA is combined with a bead having each spectral characteristic. The gene expression analyzer images beads and analyzes the spectral characteristics of each beads, and thereby identifies an mRNA that corresponds to the kind of the existing beads.  
         [0005]      FIG. 13  is a block diagram illustrating a prior art gene expression analyzer using a fluorescence microscope.  
         [0006]     In the prior art gene expression analyzer  600  shown in  FIG. 13 , there are provided a well plate  601  comprising a plurality of wells  602  for receiving a plurality of beads as observation targets, a well plate driving unit  603  for moving the well plate  601  in X and Y directions on a two-dimensional plane, a position reference imaging unit  630  for imaging silhouettes of the plurality of beads, a luminance reference imaging unit  640  for imaging luminance images of plural beads through plural optical filters each having a passing wavelength and different from each other, a CCD camera controller  611  for controlling a CCD camera  610 , and a CPU  620  which analyzes the images imaged by respective imaging units  630  and  640  as well as controls the whole apparatus  600 .  
         [0007]     More specifically, the position reference imaging unit  630  includes an LED  606  as reference light, an objective lens  605 , a z-axis driving unit  612  for moving the objective lens  605  in z-axis direction, a dichroic mirror  607  for reflecting a light of wavelength less than a predetermined value while passing a light of wavelength equal to or larger than the predetermined value, an imaging lens  609 , and a CCD camera  610 . A LED light from the LED  606  is applied to the plural beads as observation targets in the well  602 , and the obtained silhouette lights of the beads are enlarged by the objective lens  605  to pass through the dichroic mirror  607  and the bandpass filter  608 , and are collected by the imaging lens  609 . Then, the z-axis driving unit  612  drives the objective lens  605  to align the focus position of the objective lens  605 , and the CCD camera  610  images the silhouette lights to output a silhouette image as a two-dimensional image.  
         [0008]     The luminance reference imaging unit  640  includes an excitation light source  61 . 3 , an objective lens  605 , a z-axis driving unit  612 , a dichroic mirror  607 , a filter wheel  614  which holds plural bandpass filters  608  each passing only a predetermined wavelength band, a filter wheel driving unit  615  which rotatably drives the filter wheel  614 , an imaging lens  609 , and a CCD camera  610 . An excitation light from the excitation light source  613  is reflected by the dichroic mirror  607  and is applied to the plural beads as observation targets in the well  602  passing through the objective lens  605 . The light generated in accordance with the spectral characteristic of the respective beads in response to the applied light are enlarged by the objective lens  605 , and passes through the dichroic mirror  607  and the bandpass filter  608  to be collected by the imaging lens  609 . Meanwhile, the z-axis driving unit  612  drives the objective lens  605  so as to align the objective lens  605  in its focus position, and then the CCD camera  610  images the light which is emitted from the respective beads and is collected by the imaging lens  609 , to obtain a luminance image as a two-dimensional image.  
         [0009]     Further, the CPU  620  includes a controller  621  which controls the whole apparatus  600 , an analysis unit  622  which analyzes the two-dimensional image imaged by the CCD camera  610 , and a mask image creation unit  623  which creates mask image that shows the existing area of the beads as imaging targets on the basis of the position reference image.  
         [0010]     An operation of the prior art gene expression analyzer will be described.  FIG. 14  is a flowchart illustrating a series of operations for obtaining the spectral characteristics of beads as observation targets in the prior art gene expression analyzer.  
         [0011]     Initially, in step S 101 , position reference images for obtaining existing positions of plural beads as imaging targets are captured into the CPU  620  in the apparatus  600 . To be specific, the controller  621  in the CPU  620  controls the well plate driving unit  603  so as to move the well plate  601  receiving the observation targets to be positioned right above the objective lens  605 . Then, the controller  621  makes the LED  606  light to apply the LED light to the well  602 . The LED light becomes the silhouette light for the beads as observation targets in the well  602 , and the silhouette light is enlarged by the objective lens  605  and passes through the dichroic mirror  607  and the bandpass filter  608  to be collected by the imaging lens  609 , and then reaches the CCD camera  610 . The controller  621  instructs the z-axis driving unit  612  to align the objective lens  605  in its focus position so as to image the silhouette lights, and then instructs the CCD camera controller  611  to make the CCD camera  610  image the silhouette images of the plural beads as observation targets. Then, the analysis unit  622  in the CPU  620  binarizes the imaged silhouette images, and the binarized images are stored in the CPU  620  as position reference image for obtaining existing positions of the respective targets.  
         [0012]     In step S 102 , a plurality of images which have passed through the respective optical filters are captured into the CPU  620  as images for obtaining luminance values of the respective imaging targets. To be specific, the controller  621  initially makes the LED  606  unlighted and makes the excitation light source  613  apply an excitation light. The excitation light is a light of short wavelength such as a blue laser beam. When the excitation light is incident on the dichroic mirror  607 , due to the characteristic of the dichroic mirror  607  that it reflects a light of wavelength less than a predetermined value, the dichroic mirror  607  reflects the excitation light in the direction toward the objective lens  605 . The objective lens  605  focuses the light from the dichromic mirror  607  on the observation targets in the well  602 . The plural beads as observation targets existing in the well  602  present light emission patterns which respectively correspond to the spectral characteristics of the respective beads in response to the light applied from the objective lens  605 , and the lights emitted from the respective beads pass through the objective lens  605 , the dichroic mirror  607 , and the bandpass filter  608 , and further are collected by the imaging lens  609 , and then reach the CCD camera  610  similarly as described above for the silhouette lights. At this time, since the bandpass filter  608  only passes a specific wavelength band, only the light of the specific wavelength band among the light emitted from the observation targets reaches the CCD camera  610 . The controller  621  instructs the CCD camera controller  611  to make the CCD camera  610  image luminance images of only the specific wavelength bands having passed through the bandpass filter  608  among the light emitted from the observation targets. Then, the analysis unit  622  in the CPU  620  binarizes the imaged luminance images to be stored in the CPU  620  as luminance reference images.  
         [0013]     In step S 103 , it is confirmed whether a predetermined number of luminance reference images obtained as above are captured or not, and when it does not yet reach the predetermined number, the controller  621  controls the filter wheel driving unit  615  to rotate the filter wheel  614  and to set the bandpass filter  608  passing a different wavelength band in the light path. Then, after performing the same processing as described above, the CCD camera  610  images a luminance image of a specific wavelength band which has passed through the newly set bandpass filter  608  among the light emitted from the observation targets. The analysis unit  622  then binarizes the imaged luminance image to be stored in the CPU  620  as a new luminance reference image. This processing is repeated a predetermined number of times until for example eight pieces of luminance reference images are obtained, and luminance reference images of various wavelengths are obtained.  
         [0014]     After obtaining the position reference image and the luminance reference images as above, the processing transits to an analysis step of identifying the kind of the plural beads as observation targets using those reference image.  
         [0015]     Here, the beads appearing on the respective luminance reference images and the beads appearing on the position reference image should be located at the same positions. Accordingly, in the analysis step, the respective luminance values of the respective beads are obtained for each optical filter from the respective luminance reference images, to identify the kind of the beads is identified on the basis of the luminance values.  
         [0016]     Initially, in step S 104 , the mask image creation unit  623  in the CPU  620  creates a mask image indicating the bead presence areas using the captured position reference image.  
         [0017]      FIG. 15  is a diagram illustrating a mask image and luminance reference images. In  FIG. 15 , reference numeral  801  denotes a mask image showing bead presence areas, which is obtained by performing a masking processing that masks higher luminance portions at the center portions of the beads in the position reference image.  
         [0018]     In  FIG. 15 , reference numeral  701   a  denotes a first luminance reference image obtained after passing through the bandpass filter  608  that passes a light of 505 nm wavelength, the reference numeral  701   b  denotes a second luminance reference image obtained after passing through the bandpass filter that passes a light of 525 nm wavelength, and the reference numeral  701   c  denotes a third luminance reference image obtained after passing through the bandpass filter that passes a light of 545 nm wavelength.  
         [0019]     Here, 8 pieces of bandpass filters  608  that respectively passes the lights having wavelengths different from each other by 20 nm are used to obtain 8 pieces of luminance reference images in total.  
         [0020]      FIG. 15  shows only first three pieces among 8 pieces of luminance reference images.  
         [0021]     The areas B 1   m , B 2   m , and B 3   m  on the mask image  801  are bead presence areas where the beads B 1 , B 2 , and B 3  are present, respectively, and the areas B 1   a  to B 1   c , the areas (B 2   a  to B 2   c ) (do not appear in  FIG. 15 ), and the areas B 3   a  to B 3   c  on the first to third luminance reference images  701   a  to  701   c  are areas on the luminance reference images which correspond to the areas B 1   m , B 2   m , and B 3   m  on the mask image  801 , respectively.  
         [0022]     In steps S 105  to S 106 , assuming that the positions of the bead areas B 1   a  to B 1   c , B 2   a  to B 2   c , and B 3   a  to B 3   c  which are present on the respective luminance reference images are the same as the positions of the bead area B 1   m , B 2   m , and B 3   m  which are present on the mask image  801 , respectively, the analysis unit  622  in the CPU  620  obtains the respective luminance average values of the areas B 1   a  to B 1   c , B 2   a  to B 2   c , and B 3   a  to B 3   c  on the respective luminance reference images.  
         [0023]     Assuming, for example, that a luminance average value of the area B 1   a  on the first luminance reference image  701   a  is A, a luminance average value of the area B 1   b  on the second luminance reference image  701   b  is B, and a luminance average value of the area B 1   c  on the third luminance reference image  701   c  is C, in step S 107 , the luminance average values obtained as above are plotted to result in  FIG. 16 .  
         [0024]      FIG. 16  is a diagram illustrating the plotted luminance average values of the respective areas on the luminance reference images, which areas correspond to the three bead areas on the mask image.  
         [0025]     In  FIG. 16 , the abscissas indicates a wavelength transmitting through the bandpass filter while the ordinates indicates the luminance average value. The reference numeral  901  indicates a spectral curve indicating the characteristic of beads B 1 , obtained by plotting the 8 luminance average values of the areas on the first to eighth luminance reference images corresponding to the bead presence area B 1   m  and connecting the plot points. Reference numeral  902  indicates a spectral curve indicating the characteristic of beads B 2 , obtained by plotting the respective luminance average values of the areas on the luminance reference images corresponding to the bead presence area B 2   m  and connecting the plot points. Reference numeral  903  indicates a spectral curve indicating the characteristic of beads B 3 , obtained by plotting the respective luminance average values of the areas on the luminance reference images corresponding to the bead presence area B 3   m  and connecting the plot points.  
         [0026]     In step S 108 , the analysis unit  622  analyzes the spectral characteristics of the respective beads B 1  to B 3  on the basis of the respective spectral curves  901  to  903  thus obtained and identifies the kinds of the beads, respectively.  
         [0027]     In the conventional method, as shown in  FIG. 17 ( a ), assuming that, when the mask image is overlaid on the first luminance reference image, the bead areas B 1   m  to B 3   m  on the mask image and the bead areas B 1   a  to B 3   a  on the first luminance reference image are located at the same positions, respectively, luminance average values of the respective areas on the luminance reference images corresponding to the bead presence areas B 1   m  to B 3   m  on the mask image are obtained and the kinds of the beads are respectively identified on the basis of the average values.  
         [0028]     In this conventional method, however, there is no means for detecting vibrations, and even when vibrations occur while the luminance reference image is being imaged and thereby the position of the beads on the luminance reference image is changed, the processing is performed similarly as described above. Therefore, when the mask image is overlay-displayed on the luminance reference image and a deviation has occurred between the beads presence areas B 1   m  to B 3   m  on the mask image and the bead areas on the luminance reference images as shown in  FIG. 17 ( b ), the analysis unit  622  cannot obtain luminance average values of the respective beads on the respective luminance reference images correctly, and thereby the kinds of the beads cannot be properly identified.  
         [0029]     FIGS.  17 ( a ) and  17 ( b ) are diagrams illustrating relationships between the beads areas present in the mask image and the bead areas present in the first luminance reference image, wherein  FIG. 17 ( a ) shows a case where the mask image is overlaid on the luminance reference image in its display when no vibrations occur and  FIG. 17 ( b ) shows a case where the mask image is overlaid on the luminance reference image in its display when vibrations occur.  
         [0030]     To solve this problem, it may be conceived to mount a vibration detection sensor close to the imaging target and to judge whether the positions of the beads has deviated or not before or during imaging the respective luminance reference images employing the technique disclosed in the Japanese Published Patent Application No. Hei.5-130545.  
         [0031]     However, when the vibration detection method as disclosed in the above patent reference is employed, it is necessary to provide a vibration sensor in the apparatus  600 , which results in an increase in cost as well as necessitating a large amount of labor for interrelating the vibrations and the deviations in the beads positions.  
         [0032]     This comes from that as long as a detailed analysis is not made as to the amplitude and the direction of the vibrations, a mounting position of a sensor, and relationships between the vibrations and the sensor positions, it is difficult to properly relate the allowable range for the sensor output and the allowable range for the actual beads position deviations.  
       SUMMARY OF THE INVENTION  
       [0033]     The present invention is directed to solving the above described problems and has for its object to provide an imaging apparatus that can detect an occurrence of vibrations, and can perform data analysis with high accuracy, without providing a new constituent such as a vibration sensor in the apparatus.  
         [0034]     Other objects and advantages of the invention will become apparent from the detailed description that follows. The detailed description and specific embodiments described are provided only for illustration since various additions and modifications within the spirit and scope of the invention will be apparent to those of skill in the art from the detailed description.  
         [0035]     In order to solve the above-described problems, according to a 1st aspect of the present invention, there is provided an imaging apparatus which comprises: a position reference image creation unit for applying reference light to plural imaging targets each having the same shape, and creating silhouettes of the imaging targets, thereby to obtain a position reference image utilized for obtaining positions at which the respective imaging targets are present; a luminance reference image creation unit for applying an excitation light to the imaging targets, and creating luminance images of the imaging targets for respective optical filters each having a predetermined passing wavelength band, thereby to obtain plural luminance reference images utilized for obtaining luminance of respective imaging targets for each optical filter; a first vibration detection unit for detecting at least one among a change amount of the number or the area and a shape change of the imaging targets on the position reference image, and judging whether vibrations have occurred in the respective imaging targets during imaging the imaging target by the position reference image creation unit on the basis of the detected result; a mask image creation unit for creating a mask image that shows areas where the imaging targets are present from the position reference image for which the first vibration detection unit has detected no vibrations; and a second vibration detection unit for overlay-displaying the mask image on the respective luminance reference images, detecting whether or not the imaging targets are present outside the existing areas of the respective imaging targets, and judging whether or not vibrations have occurred in the respective imaging targets during imaging the imaging target by the luminance reference image creation unit on the basis of the detected result.  
         [0036]     Therefore, it can be detected whether or not a vibration occurred during imaging using an image obtained by the apparatus.  
         [0037]     According to a 2nd aspect of the present invention, in the imaging apparatus of the 1st aspect, the first vibration detection unit includes a shape change detection unit for obtaining a characteristic amount indicating a shape change of each imaging target in the position reference image, the characteristic amount detected by the first vibration detection unit is compared with a predetermined threshold value for the characteristic amount, and when among the plural imaging targets in the position reference image, there are present a predetermined number of or a predetermined rate of imaging targets that have the characteristic amounts larger than the threshold value, it is judged that vibrations have occurred.  
         [0038]     Therefore, it can be detected whether or not a vibration occurred when the silhouettes of the imaging targets were imaged using the obtained position reference image.  
         [0039]     According to a 3rd aspect of the present invention, in the imaging apparatus of the 2nd aspect, the characteristic amount for the imaging target having a spherical shape is the largest diameter of the imaging target.  
         [0040]     Therefore, a shape of the imaging target can be easily obtained.  
         [0041]     According to a 4th aspect of the present invention, in the imaging apparatus of the 2nd aspect, the characteristic amount for the imaging target that has a spherical shape and comprises a substance of a high light transparency is the largest diameter of a high luminance portion of the imaging target.  
         [0042]     Therefore, a shape of the imaging target can be more properly and easily obtained.  
         [0043]     According to a 5th aspect of the present invention, in the imaging apparatus of the 1st aspect, the mask image creation unit creates a mask image by pasting areas corresponding to the imaging targets and their outer circumference areas in the position reference image for which imaging target areas no vibrations are detected by the first vibration detection unit.  
         [0044]     Therefore, it can be easily detected whether or not a vibration occurred before or when the luminance image of the imaging target was imaged using the created mask image.  
         [0045]     According to a 6th aspect of the present invention, in the imaging apparatus of the 1st aspect, the second vibration detection unit overlay-displays the mask image on the luminance reference images, and compares the luminance values of the pixels that are located outside the existing areas of the imaging targets in the luminance reference images with a predetermined threshold value for the luminance value, and when among the pixels located outside the existing areas of the imaging targets, there are present a predetermined number of or a predetermined rate of pixels that have the luminance values larger than the threshold value, it is judged that vibrations have occurred.  
         [0046]     Therefore, it can be more reliably detected whether or not a vibration occurred during imaging using the mask image.  
         [0047]     According to a 7th aspect of the present invention, in the imaging apparatus of the 1st aspect, the second vibration detection unit overlay-displays the mask image on the luminance reference images, and compares the luminance values of the pixels that are located close to the outer circumferences of the existing areas of the imaging targets in the luminance reference images with a predetermined threshold value for the luminance value, and when among the pixels that are located close to the circumferences of the existing areas of the imaging targets, there are present a predetermined number of or a predetermined rate of pixels having higher luminance values than the threshold value, it is judged that vibrations have occurred.  
         [0048]     Therefore, it can be detected in a shorter time whether or not a vibration occurred before or when a luminance image was imaged since the targets to be detected are reduced.  
         [0049]     According to an 8th aspect of the present invention, in the imaging apparatus of the 1st aspect, when vibrations are detected by either the first vibration detection unit or the second vibration detection unit, all of the position reference image and the respective luminance reference images are newly captured.  
         [0050]     Therefore, an image for which a vibration is detected can be securely eliminated, and as a result reliability of the data analyzed by the apparatus can be substantially improved.  
         [0051]     According to a 9th aspect of the present invention, in the imaging apparatus of the 1st aspect, when vibrations are detected by either the first vibration detection unit or the second vibration detection unit, the luminance reference images for which vibrations are detected as well as the position reference image for obtaining the existing positions of the imaging targets on the luminance reference images for which vibrations are detected are newly captured. Therefore, a time for eliminating the images for which a vibration is detected and obtaining all the images can be shortened. 
     
    
     BRIEF DESCRIPTION OF THE DRAWINGS  
       [0052]      FIG. 1  is a diagram illustrating a construction of a gene expression analyzer using a fluorescence microscope according to the present invention.  
         [0053]      FIG. 2  is a diagram illustrating a flow chart of image capture steps according to a first embodiment of the present invention.  
         [0054]      FIG. 3  shows an image obtained when no vibration occurred while the position reference image imaging unit was performing imaging according to the first embodiment of the present invention wherein  FIG. 3 ( a ) shows a silhouette image,  FIG. 3 ( b ) is an enlarged view of a bead shown in  FIG. 3 ( a ), and  FIG. 3 ( c ) shows a position reference image.  
         [0055]      FIG. 4  shows an image obtained when a strong vibration occurred while the position reference image imaging unit was performing imaging according to the first embodiment of the present invention wherein  FIG. 4 ( a ) shows a silhouette image, and  FIG. 4 ( b ) is a diagram showing that one of the beads shown in  FIG. 4 ( a ) is binarized.  
         [0056]      FIG. 5  shows an image obtained when a gentle vibration occurred while the position reference image imaging unit was performing imaging according to the first embodiment of the present invention wherein  FIG. 5 ( a ) shows a silhouette image,  FIG. 5 ( b ) is a diagram showing that one of the beads shown in  FIG. 5 ( a ) is binzrized, and  FIG. 5 ( c ) is a diagram illustrating only a higher luminance portion of the bead shown in  FIG. 5 ( b ).  
         [0057]      FIG. 6  is a diagram illustrating a flow chart of a first vibration detection steps according to the first embodiment of the present invention.  
         [0058]      FIG. 7  is a diagram for explaining a method for creating a mask image according to the first embodiment of the present invention wherein  FIG. 7 ( a ) shows a position reference image,  FIG. 7 ( b ) is a diagram illustrating an intermediate process of creating the mask image, and  FIG. 7 ( c ) shows the mask image obtained on the basis of the image shown in  FIG. 7 ( a ).  
         [0059]      FIG. 8  is a diagram illustrating a mask image and a plurality of luminance reference images according to the first embodiment of the present invention.  
         [0060]      FIG. 9  is a diagram illustrating a relationship between a mask image and a first luminance reference image according to the first embodiment of the present invention, wherein  FIG. 9 ( a ) is a diagram showing that the mask image is overlaid on the luminance reference image in a case where no vibration occurred, and  FIG. 9 ( b ) is a diagram showing that the mask image is overlaid on the luminance reference image in a case where a vibration occurred.  
         [0061]      FIG. 10  is a diagram illustrating a flow chart of a second vibration detection steps according to the first embodiment of the present invention.  
         [0062]      FIG. 11  shows a histogram which is created on the basis of a first luminance reference image according to the first embodiment of the present invention.  
         [0063]      FIG. 12  is a diagram illustrating a flow chart of the image capturing steps according to a second embodiment of the present invention.  
         [0064]      FIG. 13  is a diagram illustrating a construction of a prior art gene expression analyzer using a fluorescence microscope.  
         [0065]      FIG. 14  is a flow chart illustrating a series of operations performed by the prior art gene expression analyzer.  
         [0066]      FIG. 15  is a diagram illustrating a mask image and a plurality of luminance reference images for the prior art.  
         [0067]      FIG. 16  is a diagram illustrating plotted luminance average values.  
         [0068]      FIG. 17  is a diagram illustrating a relationship between a mask image and a first luminance reference image for the prior art wherein  FIG. 17 ( a ) is a diagram showing that the mask image is overlaid on the luminance reference image in a case where no vibration occurred, and  FIG. 17 ( b ) is a diagram showing that the mask image is overlaid on the luminance reference image in a case where a vibration occurred. 
     
    
     DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT  
       [0069]     Hereinafter, embodiments of an imaging apparatus according to the present invention will be described in detail with reference to the drawings.  
       Embodiment 1  
       [0070]     The imaging apparatus according a first embodiment detects whether or not a vibration occurs before or while an image is imaged on the basis of the imaged images. Here, in the first embodiment, a gene expression analyzer which analyzes spectral characteristics of beads which are present in the imaged image and identifies the kinds of the beads, thereby identifying mRNAs corresponding to the kinds of the beads, respectively, is taken as an example of an imaging apparatus as described in the background of the invention.  
         [0071]     Further, imaging targets are beads having various spectral characteristics, each of which has a diameter of about 10 μm.  
         [0072]      FIG. 1  is a diagram illustrating a construction of the gene expression analyzer according to the first embodiment.  
         [0073]     In  FIG. 1 , the gene expression analyzer  100  according to the first embodiment comprises: a well plate  101  formed by a plurality of wells  102  into which a plurality of beads as observation targets are injected; a well plate driving unit  103  which moves the well plate  101  in the X and Y directions on the two-dimensional plane; a position reference image imaging unit  130  which images silhouettes of the plurality of beads as imaging targets; a luminance reference image imaging unit  140  which images luminance images of the beads through a plurality of optical filters each having a passing wavelength band different from each other, respectively; a CCD camera controller  111  which controls a CCD camera  110 ; and a CPU  120 . The CPU  120  is provided with a first vibration detection unit  124  which detects whether or not a vibration occurred when the position reference image was being imaged and a second vibration detection unit  125  which detects whether or not a vibration occurred before or when the luminance reference image was imaged in addition to a controller  121  which controls the whole apparatus  100 , an analysis unit  122  which analyzes images imaged by the CCD camera  110 , and a mask image creation unit  123  which creates a mask image indicating bead presence areas on the basis of the position reference image.  
         [0074]     Then, in  FIG. 1 , the portions identical or corresponding to those shown for the prior art apparatus  600  in  FIG. 11  are denoted by the reference numerals identical or corresponding to those designated for the prior art apparatus.  
         [0075]     Hereinafter, a description will be given in detail. The position reference image imaging unit  130  in the apparatus  100  includes a LED  106  as reference light, an objective lens  105 , a z-axis driving unit  112 ; a dichroic mirror  107 ; an imaging lens  109 , and a CCD camera  110 . A LED light from the LED  106  is applied to the plurality of beads as observation targets in the well  102 , and the obtained silhouette lights of the beads are enlarged by the objective lens  105  and then pass through the dichroic mirror  107  and the bandpass filter  108 , and are collected by the imaging lens  109 . At this time, the objective lens  105  is aligned with the focus position by means of the z-axis driving unit  112 , and the CCD camera  110  images the silhouette lights to obtain a silhouette image which is transformed into a two-dimensional image.  
         [0076]     The luminance reference image imaging unit  140  in the apparatus  100  includes an excitation light source  113 , an objective lens  105 , a z-axis driving unit  112 , a dichroic mirror  107 , a filter wheel  114 , a filter wheel driving unit  115 , an imaging lens  109 , and a CCD camera  110 . An excitation light from the excitation light source  113  is applied to the plurality of beads as observation targets in the well  102  through the dichroic mirror  107  and the objective lens  105 , and the lights emitted from the respective beads by the applied light are enlarged by the objective lens  105 , and pass through the dichroic mirror  107  and the bandpass filter  108  and are collected by the imaging lens  109 . At this time, the z-axis driving unit  112  moves the objective lens  105  to align the objective lens  105  with the focus position, and thereafter the CCD camera  110  images the lights emitted from the respective beads, which lights are collected by the imaging lens  109 , to obtain a luminance image which is transformed into a two-dimensional image.  
         [0077]     Next, an operation will be described.  
         [0078]     The gene expression analyzer  100  according to the first embodiment images the plurality of beads as observation targets and captures a position reference image and a plurality of luminance reference images and then analyzes the spectral characteristics of the beads using the captured images as described above for the prior art apparatus  600  (refer to  FIG. 14 ).  
         [0079]     The process of the analysis steps according to the first embodiment is the same as the process of the analysis steps for the prior art apparatus, and here a process of image capture steps of capturing images will be described in detail.  
         [0080]      FIG. 2  is a flow chart illustrating a series of flows for the image capture steps performed by the gene expression analyzer according to the first embodiment.  
         [0081]     Initially, in step S 1 , a position reference image for obtaining positions where the plurality of beads as imaging targets are present is captured into the CPU  120  in the apparatus  100 . To be specific, the controller  121  in the CPU  120  initially controls the well plate driving unit  103  so as to move the well plate  101  so that the well  102  into which the observation targets have been injected is positioned right above the objective lens  105 . Then, the controller  121  lights the LED  106  so as to apply the LED light to the well  102 . The LED light becomes silhouette lights of the beads as observation targets in the well  102 , and the silhouette lights are enlarged by the objective lens  105 , and pass through the dichroic mirror  107  and the bandpass filter  108 , and further are collected by the imaging lens  109 , and reach the CCD camera  110 . The controller  121  instructs the Z axis driving unit  112  to align the objective lens  105  with the focus position to image the silhouette lights, and thereafter instructs the CCD camera controller  111  to make the CCD camera  110  image the silhouette image of the plurality of beads. Then, the analysis unit  122  in the CPU  120  binarizes the imaged silhouette image. The binarized image is stored in the CPU  120  as the position reference image for obtaining positions where the respective imaging targets are present.  
         [0082]      FIG. 3 ( a ) is a diagram illustrating an imaged silhouette image,  FIG. 3 ( b ) is an enlarged view of a bead which is present in the silhouette image shown in  FIG. 3 ( a ), and  FIG. 3 ( c ) is a diagram illustrating a position reference image obtained on the basis of the silhouette image shown in  FIG. 3 ( a ). Then, while some hundreds of beads appear on an actual silhouette image,  FIG. 3  shows only five pieces of beads for simplicity.  
         [0083]     As shown in  FIG. 3 ( b ) a bead has a much lighter area at its center. This is because the bead is made of translucent acryl and the bead functions as a lens in the case of the LED light being applied, and the light is focused on the center part, and the center part becomes very light while the periphery of the bead becomes very dark since the light is bent and does not reach the periphery.  
         [0084]     Next, in step S 2 , the first vibration detection unit  124  in the CPU  120  detects whether or not a vibration occurred when the position reference image imaging unit  130  imaged a silhouette image.  
         [0085]     Hereinafter, a vibration detection method performed by the first vibration detection unit  124  will be described.  FIG. 6  is a flow chart illustrating a flow of the first vibration detection steps.  
         [0086]     In step S 21 , the number and area change detection unit  124   a  in the first vibration detection unit  124  initially detects the number of beads or each bead area on the position reference image, and in step S 22  compares the number of beads or each bead area which is detected with a previously held threshold value.  
         [0087]     Then, in a case where it is judged in the step S 22  that the number of beads or each bead area is less than the threshold value, it is judged that a strong vibration occurred when the silhouette image was imaged, and the step shifts to step S 1  shown in  FIG. 2 , and a silhouette image is re-imaged again.  
         [0088]     Here, why the vibration can be detected by detecting the number of beads or each bead area appearing on the position reference image will be described.  
         [0089]      FIG. 4 ( a ) is a diagram illustrating a silhouette image obtained in a case where a strong vibration occurred when the silhouette image was imaged, and  FIG. 4 ( b ) is a diagram showing that the bead B 5  shown in  FIG. 4 ( a ) is binarized. A silhouette image  201  imaged when the strong vibration occurred during imaging substantially blurs as shown in  FIG. 4 ( a ), and as a result, the number of beads are substantially reduced (the number is reduced from 5 to 3 in  FIG. 4 ), or each area of the bead parts is reduced as shown in  FIG. 4 ( b ).  
         [0090]     By utilizing this phenomenon, the number and area change detection unit  124   a  detects the number of beads and each bead area on the position reference image, and compares the number of beads or each bead area with a predetermined threshold value, thereby detecting whether or not a strong vibration occurred during imaging.  
         [0091]     Next, a method for calculating a threshold value to be previously held in the number and area change detection unit  124   a  will be described.  
         [0092]     The number N of beads to be injected into a well  102  is previously set, and the beads are uniformly distributed in the well  102 . Accordingly, when the well area in the well  102  is S W  and the well area on the position reference image is S M , the number Nx of beads on the position reference image is obtained as Nx=(S W /S M )×N and a value having this as an upper limit is set as a threshold value for the number of beads. As this threshold value is closer to the upper limit value, the smaller vibration can be detected.  
         [0093]     On the other hand, since each bead has the same shape, each bead area is calculated on the basis of the threshold value for the number of beads and the calculated area is set as a threshold value for the bead area. For example, the threshold value for the number of beads and the threshold value for each bead area on the position reference image  201  are set to 200 pieces and 80 pixels, respectively.  
         [0094]     These threshold values are previously held, and in a case where the number of beads on the position reference image  201 , which is detected by the number and area change detection unit  124   a , is less than 200 pieces, it is judged that a strong vibration occurred during imaging. Otherwise, in a case where a bead area having the largest area of the respective bead areas on the position reference image  201 , which is detected by the number and area change detection unit  124   a , is less than 80 pixels, it is judged that a strong vibration occurred during imaging. Of course, all the bead areas on the position reference image  201  are calculated on the basis of the threshold value for the number of beads and the threshold value for each bead area, and the calculated value may be held as a threshold value for the bead area. In this case, when the bead area detected by the number and area change detection unit  124   a  is less than 200 pieces×80 pixels=16000 pixels, it is judged that a strong vibration occurred.  
         [0095]     On the other hand, in a case where it is judged in the step S 22  that the value detected by the number and area change detection unit  124   a  is equal to or larger than the threshold value, the shape change detection unit  124   b  in the first vibration detection unit  124  subsequently detects a characteristic amount of a bead which is present on the position reference image in step S 23 , and compares the characteristic amount with the threshold value which is previously held in the shape change detection unit  124   b  in step S 24 .  
         [0096]     Then, when it is judged in the step S 24  that the characteristic amount of the bead is larger than the threshold value, it is judged that a gentle vibration occurred while the silhouette image was imaged, and the step shifts to step S 1  shown in  FIG. 2 , and a silhouette image is re-imaged again.  
         [0097]      FIG. 5 ( a ) is a diagram illustrating a silhouette image obtained when a gentle vibration occurred during imaging,  FIG. 5 ( b ) is an enlarged view of a bead B 5  present on the position reference image, which is obtained by binarizing the bead B 5  shown in  FIG. 5 ( a ), and  FIG. 5 ( c ) is a diagram illustrating a part having a higher luminance which is present at the center part of the bead B 5 .  
         [0098]     While the bead which is present in the silhouette image imaged when a gentle vibration occurred during imaging has its shape distorted a little and becomes elliptical as shown in  FIG. 5 ( a ), the number of beads remains unchanged and each bead area changes just a little, and thereby the vibration detection method performed by the number and area change detection unit  124   a  described above cannot be applied.  
         [0099]     However, in a case where a gentle vibration occurred, the bead in the silhouette image has its shape distorted as described above, and therefore the shape change detection unit  124   b  utilizes this phenomenon to detect a change in bead shape and detects whether or not a gentle vibration occurred during imaging.  
         [0100]     Hereinafter, a method for detecting a change in bead shape performed by the shape change detection unit  124   b  will be described.  
         [0101]     The shape change detection unit  124   b  obtains a change in shape of the bead which is present on the position reference image by detecting whether or not the shape of the bead is elliptical. Accordingly, the largest bead diameter is detected as a value indicating a shape of the bead on the position reference image (hereinafter, referred to as “characteristic amount”), and when the largest diameter is larger than the previously held threshold value, it is judged that the vibration occurred.  
         [0102]     Then, as shown in  FIG. 5 ( b ), in a case where the bead has a higher light transmittancy, the light is focused on the center part of the bead which functions as a lens and the center part has a much higher luminance value and keeps a stable shape also after binarized, and therefore the largest diameter of the shape of this part having the higher luminance is set as the characteristic amount.  
         [0103]     Then, a value whose lower limit is a diameter of a bead in a normal state is set as the previously held threshold value. As the threshold value is closer to the lower limit, the smaller vibration can be detected. Then, the judgement as to whether the shape is elliptical or not can be also made in a method in which a length of the major axis and a length of the minor axis of the bead are obtained and in a case where the ratio therebetween is other than 1, the bead shape is judged as ellipse.  
         [0104]     Hereinafter, a method for obtaining a characteristic amount in the case of the bead having a higher light transmittancy will be described.  
         [0105]     In  FIG. 5   c , the reference numeral  220  indicates a shape of a white area in a part of a bead, which has the higher luminance. For the shape  220 , a coordinate of the rightmost pixel  211  is (Xmax, Ya), a coordinate of the leftmost pixel  212  is (Xmin, Yb), a coordinate of the uppermost pixel  213  is (Xa, Ymax), and a coordinate of the lowermost pixel  214  is (Xb, Ymin).  
         [0106]     At this time, a width  215  is obtained as (Xmax−Xmin), and the height  216  is obtained as (Ymax−Ymin). In order to obtain a major axis of the ellipse, the width (Xmax−Xmin) is compared with the height (Ymax−Ymin), and when the width (Xmax−Xmin) is larger, L=(Xmax−Xmin)  2 +(Ya−Yb)  2 , while when the height (Ymax−Ymin) is larger, L=(Xa−Xb) 2 +(Ymax−Ymin) 2 .  
         [0107]     The value of L obtained here is the square of the length  217  and is close to the square of the major axis  218  of the ellipse.  
         [0108]     Then, the value of L is used only for comparison as to magnitude, and there is no need to obtain the square root thereof and the value L is used as it is.  
         [0109]     The values of Ls of all the beads which are present on the position reference image are obtained as described above and thereafter an average value thereof is obtained as a value of LA.  
         [0110]     At this time, a threshold value to be previously held in the shape change detection unit  124   b  is a value Lmax whose lower limit is the average value LA of the diameters of all the beads which are present on the position reference image in the case of no vibration having occurred.  
         [0111]     On the other hand, in a case where no change in bead shape is detected by the shape change detection unit  124   b  in the step S 24 , it is judged that no vibration occurred and the step proceeds to the next step S 3  shown in  FIG. 2 .  
         [0112]     In step S 3 , a mask image is created on the basis of the position reference image.  
         [0113]     Hereinafter, a mask image will be described.  FIG. 7 ( a ) is a diagram illustrating a position reference image,  FIG. 7 ( b ) is a diagram illustrating an intermediate process of creating a mask image on the basis of the image shown in  FIG. 7 ( a ), and  FIG. 7 ( c ) is a diagram illustrating the created mask image.  
         [0114]     The mask image indicates presence positions of beads which are present on the position reference image and is obtained by making the position reference image shown in  FIG. 7 ( a ). To be specific, as shown in  FIG. 7 ( b ), the center parts of the beads having the higher luminances are initially masked and thereafter the processing is performed so that the outer circumferences of the respective beads B 1 , B 2  and B 3  become slightly larger, thereby obtaining the mask image as shown in  FIG. 7 ( c ). Then, in the first embodiment, the processing is performed so that the outer circumference of each of the beads B 1 , B 2  and B 3  becomes one pixel larger.  
         [0115]     Then, in step S 4 , a plurality of luminance reference images for obtaining the respective average luminance values of the imaging targets which have already passed through the respective optical filters are captured into the CPU  120 . To be specific, the controller  121  in the CPU  120  initially has the LED  106  unlighted and makes the excitation light source  113  apply an excitation light. The dichroic mirror  107  reflects the excitation light in the direction of the objective lens  105 . The objective lens  105  focuses the light from the dichromic mirror  107  on a plurality of beads as the observation targets in the well  102 . The plurality of beads which are present in the well  102  indicate light emission patterns corresponding to the spectral characteristics by the light which is applied from the objective lens  105 , respectively, and the lights emitted from the respective beads pass through the objective lens  105 , the dichroic mirror  107  and the bandpass filter  108 , and further are collected by the imaging lens  109 , and reach the CCD camera  110  as described for the bead silhouette lights obtained by applying the LED light to the beads. At this time, since the bandpass filter  108  has the characteristic that it passes only a specific wavelength band, only the light of the specific wavelength band emitted from the beads reach the CCD camera  110 . In this state, the controller  121  instructs the CCD camera controller  111  to make the CCD camera  110  image a luminance image of only the specific wavelength band which passes through the bandpass filter  108  among the lights emitted from the beads. Then, the analysis unit  122  in the CPU  120  binarizes the imaged luminance image and the binarized image is stored as a luminance reference image in the CPU  120 .  FIG. 8  is a diagram illustrating the mask image and the luminance reference image.  
         [0116]     A luminance reference image is captured into the CPU  120  in the step S 4 , and thereafter in step S 5  the second vibration detection unit  125  detects whether or not a vibration occurred when the luminance image to be captured was imaged using the mask image created in the step S 3 .  
         [0117]     Hereinafter, a vibration detection method for the luminance reference image performed by the second vibration detection unit  125  will be described.  
         [0118]     When a vibration is detected for the luminance reference image, the position reference image  210  imaged with no vibration occurring has been already stored in the CPU  120  and the mask image  401  indicating the bead presence positions in the image has been created on the basis of the position reference image.  
         [0119]     As described above, the outer circumferences of the bead areas B 1   m , B 2   m , and B 3   m  on the mask image  401  are slightly larger than the outer circumferences of the beads B 1 , B 2  and B 3  which are present on the position reference image, respectively.  
         [0120]     Here, since the processing is performed so that each of the outer circumferences of the bead areas B 1   m , B 2   m , and B 3   m  is one pixel larger than each of the outer circumferences of the beads B 1 , B 2  and B 3 , when no vibration occurred when the luminance image was imaged, in a case where the mask image  401  is overlay-displayed on the luminance reference image  301 , the masked bead areas B 1   m , B 2   m  and B 3   m  on the mask image  401  should be present around the outer circumferences of the bead areas B 1   a  to B 3   a  on the luminance reference image  301   a , respectively, as shown in  FIG. 9 ( a ).  
         [0121]      FIG. 9  is a diagram illustrating a relationship between the bead areas on the mask image and the bead areas on the first luminance reference image, wherein  FIG. 9 ( a ) shows that the mask image is overlay-displayed on the luminance reference image in a case where no vibration occurred while  FIG. 9 ( b ) shows that the mask image is overlay-displayed on the luminance reference image in a case where a vibration occurred before or during imaging.  
         [0122]     In  FIG. 9 , regions B 1   a , B 2   a , and B 3   a  are regions that are displayed by an emission from the bead B 1 , B 2 , and B 3  by irradiating an excitation light to the beads B 1 , B 2 , and B 3 , respectively. Regions B 1   m , B 2   m , and B 3   m  are regions displayed by overlay-displaying the mask area of the beads B 1 , B 2 , and B 3  on the luminance reference image, respectively.  
         [0123]     In a case where no vibration occurred before or when the luminance reference image was imaged, since no deviation in bead position between the position reference image and the luminance reference image is generated, the respective bead areas B 1   a , B 2   a , and B 3   a  on the luminance reference image  301   a  are located inside the respective bead areas B 1   m , B 2   m , and B 3   m  which are obtained by overlay-displaying the mask image on the luminance reference image as shown in  FIG. 9 ( a ). Accordingly, in this case, no part having a higher luminance is present outside the bead areas B 1   m , B 2   m  and B 3   m  which are obtained by overlay-displaying the mask image on the luminance reference image  301   a.    
         [0124]     On the other hand, when a vibration occurred before or when the luminance reference image was imaged, deviations in the bead positions between the position reference image and the luminance reference image are generated, the respective bead areas B 1   a , B 2   a , and B 3   a  on the luminance reference image  301   a  extrudes outside the respective bead areas B 1   m , B 2   m , and B 3   m  which are obtained by overlay-displaying the mask image on the luminance reference image  301   a  as shown in  FIG. 9 ( b ). Accordingly, in this case, parts having the higher luminances are present outside the bead areas B 1   m , B 2   m  and B 3   m  which are obtained by overlay-displaying the mask image on the luminance reference image  301   a.    
         [0125]     The second vibration detection unit  125  utilizes the above described phenomenon to detect a vibration for the luminance reference image.  FIG. 10  is a flow chart illustrating a flow of the second vibration detection steps.  
         [0126]     Initially, in step S 51 , the luminance distribution in the first luminance reference image  301   a  is checked. In the luminance reference image  301   a  of the first embodiment, the background part which occupies a larger part of the whole area indicates a lower luminance while only the excited areas B 1   a , B 2   a  and B 3   a  where the beads are present indicate higher luminances.  
         [0127]     Accordingly, in order to check the luminance distribution in the luminance reference image, for each gradation, the number of pixels indicating the gradation is obtained on the basis of the luminance reference image  301   a  to create a histogram.  FIG. 11  shows a histogram created on the basis of the luminance reference image.  
         [0128]     Then, in step S 52 , the histogram created in the step S 51  is searched from the low luminance side to obtain a luminance value IL at which the number of pixels positioned on the low luminance side totals to 5% of all the pixels, and further, the histogram is searched from the high luminance side to obtain a luminance value IH at which the number of pixels positioned on the high luminance side totals to 0.5% of all the pixels. Here, the ratio of the total number of pixels is different between the low luminance side and the high luminance side because the distribution on the low luminance side is dense while the distribution on the high luminance side is thin.  
         [0129]     Then, the luminance values IL and IH obtained as described above are substituted into the (formula 1) shown as below to obtain a boundary value TH. 
 
Boundary value  TH =( IL+IH )/2  (formula 1) 
 
         [0130]     Then, in step S 53 , the whole luminance reference image  301   a  is searched to count the number of pixels having the higher luminances than the boundary value TH obtained as described above among the pixels positioned outside the bead areas B 1   m , B 2   m , and B 3   m  in the mask image overlay-displayed.  
         [0131]     In step S 54 , the number of pixels having the higher luminance than the boundary value TH obtained in the step S 53  is compared with a threshold value previously held in the second vibration detection unit  125 , and in a case where the number of pixels having the higher luminance than the boundary value TH is larger than the threshold value, it is judged that a vibration occurred during imaging, and the step shifts to step S 1  shown in the  FIG. 2 , and the position reference image is also re-imaged again.  
         [0132]     Then, in this case, the total number of pixels positioned outside the bead areas B 1   m , B 2   m , and B 3   m  in the mask image which is overlay-displayed on the luminance reference image  301   a  is obtained, and in a case where a ratio of the number of pixels having the higher luminance than the boundary value TH obtained in the step S 53  to the total number of pixels exceeds a predetermined ratio, it may be judged that a vibration occurred.  
         [0133]     Further, while all the pixels positioned outside the bead areas B 1   m , B 2   m , and B 3   m  in the mask image which is overlay-displayed on the luminance reference image  301   a  are searched in the above described method, in a case where the expected vibration is not so strong and the amount of deviation of the beads due to the vibration is approximately lower than a diameter of the bead, it can be detected whether or not the vibration occurred by searching only the pixels close to the outer circumferences of the bead areas B 1   m , B 2   m , and B 3   m  in the mask image which is overlay-displayed on the luminance reference image for shortening processing time.  
         [0134]     That is, the luminances of the pixels bordering the outer circumferences of the respective bead areas B 1   m , B 2   m , and B 3   m  in the mask image which is overlay-displayed on the luminance reference image are checked to count the number of pixels indicating the higher luminances than the boundary value TH. Then, in a case where the total number of pixels counted, or a ratio of the total number of pixels counted to the total number of pixels whose luminances are checked is larger than a predetermined threshold value, it may be judged that a vibration occurred.  
         [0135]     On the other hand, in a case where it is judged in the step S 54  that the number of pixels having the higher luminances than the boundary value TH obtained in the step S 53  is equal to or lower than the threshold value, it is judged that no vibration occurred, and the step shifts to the next step S 6  shown in  FIG. 2 .  
         [0136]     In step S 6 , it is judged whether or not a predetermined number of luminance reference images are captured into the CPU  120 , and in a case where the predetermined numbers have not been captured yet, the step shifts to the step S 4  shown in  FIG. 2 , and the next luminance image is imaged and it is detected whether or not a vibration occurred before or when the luminance image was imaged using the mask image  401 .  
         [0137]     On the other hand, in a case where it is judged in the step S 6  that the predetermined number of luminance reference images are captured, the image capture step is completed.  
         [0138]     As described above, in the first embodiment, the CPU  120  contains a first vibration detection unit  124  which detects whether or not a vibration occurred when the position reference image imaging unit  130  imaged a silhouette image, and a second vibration detection unit  125  which detects whether or not a vibration occurred when the luminance reference image imaging unit  140  imaged a luminance image. The first vibration detection unit  124  uses the imaged position reference image to detect whether or not a vibration occurred in imaging, and further the second vibration detection unit  125  uses a mask image indicating bead presence positions which is created on the basis of the position reference image for which no vibration is detected to detect whether or not a vibration occurred in imaging every time a luminance reference image is imaged. In a case where the respective vibration detection units  124  and  125  detect vibrations, all the images are re-imaged, and thereby images imaged when the vibration occurred can be securely and easily eliminated, thereby enabling the apparatus to make data analysis with substantially high reliability.  
         [0139]     Then, while in the first embodiment the background portion of the luminance reference image has a lower luminance and the areas in which beads are excited have higher luminances, even when the relation of the luminnaces is reverse, the vibration can be detected in a like manner.  
       Embodiment 2  
       [0140]     In the first embodiment, when a vibration is detected for the luminance reference image even after some luminance reference images among a predetermined number of luminance reference images are obtained, all the images are re-obtained. On the other hand, in the second embodiment, when the vibration is detected for the luminance reference image after some luminance reference images among a predetermined number of luminance reference images are obtained, not all the images are not re-obtained, but images imaged before the vibration is detected are held as they are, the position reference image which is used for obtaining luminance values of the luminance reference image for which the vibration is detected is obtained and the luminance reference image for which the vibration is detected is re-imaged again.  
         [0141]     The vibration detection method according to the second embodiment is applied to a case where an amount of deviation of the bead is lower than the diameter of the bead when the mask image is overlay-displayed on the luminance reference image. Accordingly, it is assumed that mechanical countermeasures are taken for the imaging apparatus according to the second embodiment and thereby the vibration during imaging is substantially reduced.  
         [0142]     In the second embodiment, a gene expression analyzer which has the constituents similar to those described for the first embodiment, and analyzes the spectral characteristics of the beads which are present in the imaged image to identify the kinds of the beads and identifies mRNAs corresponding to the kinds of the beads is taken as an example of an imaging apparatus. Further the imaging targets are beads having various spectral characteristics, each of which has a diameter of about 10 μm.  
         [0143]     Hereinafter, an operation will be described.  
         [0144]     The gene expression analyzer of the second embodiment images beads as observation targets and captures a position reference image and a plurality of luminance reference images and thereafter analyzes the spectral characteristics of the beads using the captured images as described for the prior art apparatus  600 . (refer to  FIG. 14 )  
         [0145]     The process of analysis steps according to the second embodiment is the same as the process of analysis steps for the prior art apparatus, and therefore the process of image capture steps of capturing images will be described in detail here.  
         [0146]      FIG. 12  is a flow chart illustrating a series of flows of the image capture steps of the gene expression analyzer according to the second embodiment.  
         [0147]     Initially, in step S 1 , a position reference image for obtaining positions where a plurality of beads as imaging targets are present is captured into the CPU  120  in the apparatus  100 . To be specific, the controller  121  in the CPU  120  initially controls the well plate driving unit  103  so as to move the well plate  101  so that the well  102  into which the observation targets have been injected is positioned right above the objective lens  105 . Then, the LED  106  is lighted and silhouette lights of the plurality of beads as observation targets in the well  102  pass through the objective lens  105 , the dichroic mirror  107 , the bandpass filter  108 , and the imaging lens  109 , and the CCD camera  110  images a silhouette image. Then, the position reference image A obtained by binarizing the silhouette image by the analysis unit  122  in the CPU  120  is stored in the CPU  120 .  
         [0148]     Next, in step S 2 , the first vibration detection unit  124  detects whether or not a vibration occurred when the position reference image imaging unit  130  imaged the silhouette image with using the same method as that described for the first embodiment with reference to  FIG. 6 . Then, while in the first embodiment the number and area change detection unit  124   a  detects a strong vibration on the basis of the change in the number of beads or the respective bead areas, which beads are present on the position reference image, and thereafter the shape change detection unit  124   b  detects change in bead shape to detect a gentle vibration, only the shape change detection unit  124   b  may detect the vibration which occurred when the image was imaged in the second embodiment since it is assumed that the vibration is substantially reduced.  
         [0149]     Then, in a case where a vibration is detected in the step S 2 , the step shifts to the step S 1 , and the position reference image A is re-obtained again.  
         [0150]     On the other hand, in a case where no vibration is detected in the step S 2 , the step shifts to the next step S 3  and a mask image A indicating bead presence positions on the image is created on the basis of the position reference image A in the same method as described for the first embodiment.  
         [0151]     Thereinafter, in step S 4 , the first luminance reference image is captured. Then, in step S 5 , the second vibration detection unit  125  detects a vibration for the luminance reference image obtained in the step S 4  using the mask image A created in the step S 3  in the same method as described for the first embodiment.  
         [0152]     In a case where no vibration is detected in the step S 5 , the step shifts to the next step S 6 , and it is judged whether or not a predetermined number of luminance reference images are captured into the CPU  120 , and in a case where the predetermined number of luminance reference images have not been captured yet, the step shifts to the step S 4 , and the steps S 4  to S 6  are repeated until the predetermined number of luminance reference images can be obtained.  
         [0153]     Here, for example, it is assumed that a vibration is detected in the step S 5  after the sixth luminance reference image is captured into the CPU  120 . In this case, the step shifts to step S 7 , and the position reference image A′ for the luminance reference image for which the vibration is detected is obtained again.  
         [0154]     Then, in step S 8 , the first vibration detection unit  124  detects whether or not a vibration occurred when the image was imaged using the position reference image A′ in the same method as performed in the step S 2 .  
         [0155]     In a case where no vibration is detected in the step S 8 , the step shifts to the next step S 9 , and a mask image A′ is created on the basis of the position reference image A′.  
         [0156]     On the other hand, in a case where a vibration is detected in the step S 8 , the step shifts to the step S 7  and the position reference image A′ is re-obtained again.  
         [0157]     Next, in step S 10 , the sixth luminance reference image is captured again. Then, in step S 11 , the second vibration detection unit  125  detects a vibration for the sixth luminance reference image using the mask image A′ created in the step S 9  in the same method as described for the first embodiment. Then, in a case where no vibration is detected in the step S 1 , the step shifts to step S 12 , and it is judged whether the remaining luminance reference images are captured into the CPU  120  or not, and in a case where the remaining luminance reference images have not been captured yet, the steps S 10  to S 12  are repeated until all the remaining luminance reference images are obtained.  
         [0158]     The images obtained in the method described above are the position reference image A, the position reference image A′ and 8 pieces of luminance reference images, and the bead areas on the respective first to fifth luminance reference images are obtained on the basis of the bead areas on the mask image A, and the bead areas on the respective sixth to eighth luminance reference images are obtained on the basis of the bead areas on the mask image A′.  
         [0159]     In the second embodiment, it is assumed that the amount of deviation of the bead due to the vibration is approximately lower than the diameter of the bead as described above, and therefore the correspondence relationship of the bead between the mask image A and the mask image A′ can be easily obtained by utilizing the fact that the respective beads are overlapped.  
         [0160]     Accordingly, in order to create the spectral distribution chart as shown in  FIG. 16 , the average of the luminances of each area on each of the first to fifth luminance reference images, which area corresponds to each bead area on the mask image A and the average of the luminances of each area on each of the sixth to eighth luminance reference images, which area corresponds to each bead area on the mask image A′ being overlapped on each bead area on the mask image A may be obtained to plot all the luminance averages.  
         [0161]     The spectral characteristics of the respective beads can be obtained on the basis of the spectral curve created as described above, thereby enabling the kinds of the beads to be identified according thereto.  
         [0162]     Further, after the sixth luminance reference image is obtained as described above, for example, in a case where a vibration is detected for the seventh luminance reference image, the step shifts to step S 7 , and the position reference image A″ for the seventh luminance reference image for which the vibration is detected is obtained again, and a mask image A″ is created on the basis of the position reference image A″. When the analysis is made, the position reference image A, the position reference image A′, the position reference image A″, and 8 pieces of luminance reference images are used, and each bead area on each of the first to fifth luminance reference images is obtained on the basis of each bead area on the mask image A, each bead area on the sixth luminance reference image is obtained on the basis of each bead-area on the mask image A′, and each bead area on each of the seventh to eighth luminance reference images is obtained on the basis of each bead area on the mask image A″. Further, in order to create the spectral distribution chart shown in  FIG. 16 , the respective luminance averages of the bead areas on the first to fifth luminance reference images, which areas correspond to the bead areas on the mask image A, the respective luminance averages of the bead areas on the sixth luminance reference image, which areas correspond to the bead areas on the mask image A′ which overlap on the bead areas on the mask image A, and the respective luminance averages of the bead areas on the seventh to eighth luminance reference images, which areas correspond to the bead areas on the mask image A″ which overlap on the bead areas on the mask image A′ may be obtained to plot all the luminance averages.  
         [0163]     As described above, in the second embodiment, the CPU  120  contains a first vibration detection unit  124  which detects whether or not a vibration occurred when the position reference image imaging unit  130  imaged a silhouette image and a second vibration detection unit  125  which detects whether or not a vibration occurred when the luminance reference image imaging unit  140  imaged luminance images. The position reference image A is imaged and thereafter the first vibration detection unit  124  detects whether or not a vibration occurred during imaging using the imaged position reference image A, and further the respective luminance reference images are imaged and thereafter the second vibration detection unit  125  detects whether or not a vibration occurred during imaging using a mask image A indicating bead presence positions on the image, which mask image A is created on the basis of the position reference image A for which no vibration is detected. In a case where a plurality of luminance reference images are already imaged, and then a vibration is detected for the luminance reference image which is subsequently imaged, a position reference image A′ which is other than the position reference image A is obtained again, and it is detected whether or not a vibration occurred for the luminance reference image which is obtained again after the vibration is detected using the mask image A′ created on the basis of the position reference image A′, and therefore the image imaged with the vibration occurring can be eliminated, thereby enabling the apparatus to make data analysis with extremely high reliability. Further, the time for obtaining all the images can be shortened.  
         [0164]     The imaging apparatus according to the present invention is useful as a bioanalyzer such as a gene expression analyzer using a fluorescence microscope, which observes the spectral characteristics using a plurality of filter images since reliable data analysis can be made by preventing the positions of the filter images from deviating from each other due to the influence of vibration.