Abstract:
An optical microscope includes a light source irradiating a sample with an illuminating light; a mirror having a variable reflection surface for reflecting the illuminating light; a correction table storing data of plural shapes of the reflection surface which correspond to changes in a focal position and an aberration; and a controller selecting from the plural shapes of the reflection surface a reflection surface suitable for corrections of the focal position and aberration, and controlling the mirror so that the selected reflection surface is formed.

Description:
CROSS-REFERENCE TO RELATED APPLICATIONS  
       [0001]     This application is based upon and claims the benefit of priority from Japanese Patent Application No. 2005-004025, filed Jan. 11, 2005, the entire contents of which are incorporated herein by reference.  
       BACKGROUND OF THE INVENTION  
       [0002]     1. Field of the Invention  
         [0003]     The present invention relates to an optical microscope that allows an observation of a light emitted by a sample with an illuminating light from a light source irradiating the sample.  
         [0004]     2. Description of the Related Art  
         [0005]     Conventionally, an optical microscope has been widely used, by which a light emitted by a sample is observed with an illuminating light irradiating the sample. Generally, to obtain a clear image of a sample, i.e., an image on which an objective lens is in focus, it is necessary to keep an objective lens and the sample located at a certain distance therebetween. However, a thermal drift or a variety in thickness of a cover glass covering the sample would cause changes in the distance between the objective lens and the sample, resulting in changes in a focal position and an aberration, thus making it impossible to obtain a clear image. To solve this inconvenience, a mechanical adjustment has been generally employed for corrections of the focal position and aberration. However, since a correction collar for the mechanical adjustment is attached to the objective lens, manipulation of this correction collar is so uneasy that a lot of efforts are necessary, especially when the thickness of the cover glass varies. As a technique to correct the focal position and the aberration without using the mechanical adjustment, an optical microscope using an adaptive optical unit for modulating a wave surface of an incoming light is disclosed (See U.S. Pat. No. 6,771,417, for example).  
       SUMMARY OF THE INVENTION  
       [0006]     An optical microscope according to one aspect of the present invention includes a light source irradiating a sample with an illuminating light; a mirror having a variable reflection surface for reflecting the illuminating light; a correction table storing data of plural shapes of the reflection surface which correspond to changes in a focal position and an aberration; and a controller selecting from the plural shapes of the reflection surface a reflection surface suitable for corrections of the focal position and aberration, and controlling the mirror so that the selected reflection surface is formed.  
         [0007]     The above and other objects, features, advantages and technical and industrial significance of this invention will be better understood by reading the following detailed description of presently preferred embodiments of the invention, when considered in connection with the accompanying drawings. 
     
    
     BRIEF DESCRIPTION OF THE DRAWINGS  
       [0008]      FIG. 1  is a block diagram showing a schematic structure of an optical microscope according to a first embodiment of the present invention;  
         [0009]      FIG. 2  is a flowchart showing an operation procedure for corrections of a focal position and an aberration performed by a controller according to the first embodiment of the present invention;  
         [0010]      FIG. 3  is a schematic diagram showing a table structure stored in a correction table according to the first embodiment of the present invention;  
         [0011]      FIG. 4  shows detailed contents of an objective lens table according to the first embodiment of the present invention;  
         [0012]      FIG. 5  is a block diagram showing a schematic structure of an optical microscope according to a second embodiment of the present invention;  
         [0013]      FIG. 6  is a schematic diagram showing a table structure stored in a correction table according to the second embodiment of the present invention; and  
         [0014]      FIG. 7  is a flowchart showing an operation procedure for correction of a focal position and an aberration according to the second embodiment of the present invention 
     
    
     DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS  
       [0015]      FIG. 1  is a block diagram showing a schematic structure of an optical microscope  1  according to a first embodiment of the present invention. As shown in  FIG. 1 , the optical microscope  1  includes a light source  10  that emits an illuminating light; a collector lens that converts an illuminating light to a parallel light; a condenser lens  12  that focuses a parallel-converted illuminating light on a sample  13 ; an objective lens  15   a  that receives, via a cover glass  14 , a light emitted by the sample  13  on which the illuminating light is focused; a revolver  16  that stores plural objective lenses  15   a  and  15   b ; a half mirror  17  that splits the parallel illuminating light converted by the objective lens  15   a ; an adaptive mirror  18  that variably forms its reflection surface to reflect the light having passed through the half mirror  17 ; an imaging optical system  19  that focuses the light reflected by the adaptive mirror  18 ; a CCD camera  20  that obtains an observation image via an optical electronic conversion of the light focused by the imaging optical system  19 ; a controller  21  that controls the adaptive mirror  18  based on the light volume of the observation image obtained by the CCD camera  20  to change the shape of the reflection surface; and a correction table  22  that stores a plurality of voltage patterns to be applied to the adaptive mirror  18 . The controller  21  not only controls an arrangement of the objective lenses  15   a  and  15   b  by rotating the revolver  16 , but also reads the voltage pattern stored in the correction table  22 , and applies the read voltage pattern to the adaptive mirror  18 .  
         [0016]     In general, the adaptive mirror  18  includes at least one electrode, and an application of voltage to the electrode allows a change in the shape of the reflection surface. With such a change in the shape of the reflection surface, a focal position of the objective lens  15   a  changes. As a result, when the focal position changes due to an occurrence of a thermal drift or when an aberration changes due to a change in the thickness of the cover glass  14 , the changed focal position and aberration can be corrected by changing the shape of the reflection surface of the adaptive mirror  18 .  
         [0017]     With reference to  FIGS. 2 and 3 , an operation procedure of corrections of the focal position and aberration performed by the controller  21  will be described.  FIG. 2  is a flowchart showing the operation procedure of the controller  21 . First, the controller  21  performs an initial setting, which is a normal setting for optical system of the optical microscope  1  (step S 101 ). After the initial setting, the controller  21  starts an observation of the sample  13 . When an out-of-focus blurred image is obtained during the observation (step S 102 ), the controller  21  selects, from the correction table  22 , an objective lens table corresponding to the objective lens  15  currently used by the optical microscope  1  (step S 103 ).  
         [0018]      FIG. 3  is a schematic diagram showing a table structure stored in the correction table  22 . As shown in  FIG. 3 , the correction table  22  contains objective lens tables A 1  to A 1  each of which contains voltage patterns B 10  to Bm 0  each corresponding to a thermal drift factor. Each of those voltage patterns B 10  to Bm 0  contains voltage patterns B 11  to B 1   n , B 21  to B 2   n , . . . , Bm 1  to Bmn, each corresponding to a cover glass factor.  
         [0019]     The voltage patterns B 10  to Bm 0  each corresponding to the thermal drift factor represent voltage patterns used for corrections of the changed focal position and aberration when the focal position and aberration changes due to the thermal drift factor. The voltage patterns B 11  to B 1   n , B 21  to B 2   n , . . . , Bm 1  to Bmn each corresponding to the cover glass factor represent voltage patterns used for corrections of the changed focal position and aberration when the focal position and aberration changes due to a cover glass factor, i.e., a variety in the thickness of the cover glass  14 .  
         [0020]     Generally, the degree of changes in the focal position and aberration due to the thermal drift factor is higher than that due to the cover glass factor. Accordingly, the degree of changes in a voltage value corresponding to the thermal drift factor becomes higher than that corresponding to the cover glass factor.  
         [0021]     After selecting an objective lens table corresponding to the objective lens currently used in the optical microscope  1 , the controller  21  selects voltage patterns B 10  to Bm 0  that correspond to the thermal drift factor and stored in the selected objective lens table, and sequentially applies them to the adaptive mirror  18  (step S 104 ). After that, the controller  21  selects the voltage pattern where maximum light volume is obtained among the voltage patterns B 10  to Bm 0  (step S 105 ). When the voltage pattern B 10  is selected, for example, the controller  21  selects the voltage patterns B 11  to B 1   n  corresponding to the cover glass factor and sequentially applies them to the adaptive mirror  18  (step S 106 ). Next, the voltage pattern where maximum light volume is obtained is selected among the voltage patterns B 11  to B 1   n  (step S 107 ). When the voltage pattern B 11  is selected, for example, the controller  21  applies the voltage pattern B 11  to the adaptive mirror  18  (step S 108 ). Here ends the correction operation for the focal position and aberration.  
         [0022]     Here, detailed contents of an objective lens table A 1  will be explained with reference to  FIG. 4 . When the adaptive mirror  18  is divided into four, thereby having, four electrodes (first to fourth electrodes), the objective lens table A 1  has electrode tables A 11  to A 14  which correspond to four electrodes respectively. In a sheet A 11 , a plurality of voltage values applied to the first electrode are stored, and voltage values corresponding to the thermal drift factor and voltage values corresponding to the cover glass factor are stored in a matrix format.  
         [0023]     The voltage values corresponding to the thermal drift factor are stored in the sheet from 150.0 V to 155.0 V at 1 V intervals. The voltage values corresponding to the cover glass factor are stored from 150.1 V to 155.9 V at 0.1 V intervals. In each of sheets A 12  to A 14 , voltage values are stored in the same format as shown in the sheet A 11 . However, voltage values in each sheet are not necessarily the same.  
         [0024]     The voltage values applied to each of the first to fourth electrodes by the controller  21  can be calculated in advance by simulation calculation or the like. It is generally known that the shape of the reflection surface corresponding to the thermal drift factor is a curved surface which approximates in a low order function, whereas the shape of the reflection surface corresponding to the cover glass factor is a curved surface which approximates in a higher order function. The combination of the thermal drift factor and the cover glass factor provides the shape of the reflection surface expressed by a low order function overlapped with a high order function. Consequently, the correction table  22  in which such a combination is stored in advance enables a handling of various changes in a focal position and an aberration.  
         [0025]     The number of electrodes of the adaptive mirror  18  is not limited to four. However, it is necessary to prepare the same number of electrode tables as electrodes of the adaptive mirror  18  to be used.  
         [0026]     In the first embodiment, when an out-of-focus blurred image is obtained, the controller  21  selects the voltage patterns stored in the correction table  22 , and applies them sequentially to the adaptive mirror  18 , to perform quick and secure corrections of the focal position and aberration.  
         [0027]     In the first embodiment, when the light volume received by the CCD camera  20  is maximum, the corrections of the focal position and aberration end. Alternatively, the corrections of the focal position and aberration may end when the light volume received by the CCD camera  20  reaches a predetermined value. The CCD camera  20  may be replaced by an optical receiver such as an optical line sensor and a photo detector. If an observation using an incident-light illumination or fluorescence is desired, the light source of the illuminating light should be installed so that the adaptive mirror  18  is in front of the light source (closer side to the CCD camera  20  than to the adaptive mirror  18  in  FIG. 1 ) and thus the illuminating light passes through the adaptive mirror  18 .  
         [0028]     In the first embodiment, the voltage pattern corresponding to the cover glass factor is selected after the selection of the voltage pattern corresponding to the thermal drift factor. Alternatively, all voltage patterns may be sequentially selected, regardless of the differentiation between the thermal drift factor and the cover glass factor.  
         [0029]     An optical microscope according to a second embodiment of the present invention will next be described. In the first embodiment, the corrections of the focal position and aberration are conducted by using the adaptive mirror  18  in the optical microscope  1  categorized as a transmitting illumination type. On the other hand, in the second embodiment, corrections of the focal position and aberration are conducted by using an adaptive mirror in an optical microscope categorized as a confocal scanning type.  
         [0030]      FIG. 5  is a block diagram showing a schematic structure of the optical microscope  2  according to the second embodiment. As shown in  FIG. 5 , the optical microscope  2  includes a laser source  31  that emits a laser beam; a collimating optical system  32  that converts the laser beam to a parallel light; a dichroic mirror  33  that reflects the parallel-converted laser beam; a reflection mirror  34  that reflects the laser beam reflected by the dichroic mirror  33 ; an adaptive mirror  35  that variably forms its reflection surface to reflect the laser beam reflected by the reflection mirror; a scanner  36  that reflects the laser beam reflected by the adaptive mirror  35  into a fluorescence sample  40 , thereby to scan the fluorescence sample  40 ; a projection lens  37  that focuses the irradiating laser beam from the scanner  36 ; an intermediate imaging lens  38  that converts the laser beam focused by the projection lens  37  to a parallel light; an objective lens  39  that focuses the laser beam parallel-converted by the intermediate imaging lens  38  on the fluorescence sample  40 ; an imaging lens  41  that focuses fluorescence emitted by the fluorescence sample  40 ; a confocal pinhole  42  that is located in a conjugating position with a focal position of the objective lens  39 ; a photo-multiplier tube (PMT)  43  that performs a photoelectric conversion of fluorescence passed through the confocal pinhole  42 ; a controller  44  that obtains a fluorescence image of the fluorescence sample  40  using an electrical current converted by the PMT  43 , and controls a shape of the reflection surface of the adaptive mirror  35 ; and a correction table  45  that stores a plurality of voltage patterns to be applied to the adaptive mirror  35 .  
         [0031]     In this confocal scanning optical microscope  2 , when a thermal drift occurs due to a heat generation of the laser source  31  or a power supply (not shown) or the like, or when the thickness of the cover glass (not shown) covering the fluorescence sample  40  is different from that of a standard cover glass, the controller  44  obtains an out-of-focus blurred fluorescence image.  
         [0032]     When such an out-of-focus blurred fluorescence image is obtained, the controller  44  applies the voltage pattern stored in the correction table  45  to the adaptive mirror  35  sequentially. The controller  44  then varies the shape of the reflection surface of the adaptive mirror  35  to perform corrections of the focal position and aberration based on the light volume of the obtained fluorescence image. In other words, the controller  44  corrects the focal position and aberration by applying to the adaptive mirror  35  the voltage pattern where maximum light volume is obtained.  
         [0033]     In the second embodiment, the adaptive mirror  35  and the correction table  45  are used in the confocal scanning optical microscope  2  to correct the focal position and aberration. The correction table  45 , as shown in  FIG. 6 , may have a structure in which voltage patterns corresponding to the laser wavelength of the laser source  31  is added to the correction table  22  described in the first embodiment. Such a structure allows a correction of an aberration due to a laser wavelength factor.  
         [0034]      FIG. 7  is a flowchart showing an operation procedure performed by the controller  44 . The operation procedure of the controller  44  (steps S 201  to S 203 ) corresponds to the operation procedure described in the first embodiment (steps S 101  to S 103 ). After these steps, the controller  44  selects a wavelength table corresponding to the laser beam in use (step S 204 ). Following operation procedure after step  204  (steps S 205  to S 209 ) corresponds to the operation procedure described in the first embodiment (steps S 104  to S 108 ).  
         [0035]     In the first and second embodiments, the controller ( 21 ,  44 ) is configured to apply to the adaptive mirror ( 18 ,  35 ) the voltage patterns stored in the correction table ( 22 ,  45 ), based on the light volume of the obtained image. Alternatively, in each of the first and the second embodiments, the voltage pattern may be applied to the adaptive mirror ( 18 ,  35 ) based on one of an electric signal and an electric current output by the CCD camera  20  or the PTM  43 , without involving the controller ( 21 ,  44 ).  
         [0036]     With such a simple structure, quick and secure corrections of at least one of the focal position and the aberration can be realized.  
         [0037]     Additional advantages and modifications will readily occur to those skilled in the art. Therefore, the invention in its broader aspects is not limited to the specific details and representative embodiments shown and described herein. Accordingly, various modifications may be made without departing from the spirit or scope of the general inventive concept as defined by the appended claims and their equivalents.