Abstract:
Provided is a microscope having an immersion objective lens, a nozzle, and a liquid supplying mechanism. The immersion objective lens condenses light from a sample through liquid. The nozzle supplies the liquid to an upper surface of the immersion objective lens. The liquid supplying mechanism cooperates with one of a lens moving mechanism that moves the immersion objective lens and a sample moving mechanism that maintains and moves the sample, and moves the nozzle relative to the immersion objective lens to supply the liquid.

Description:
CROSS-REFERENCE TO RELATED APPLICATIONS  
       [0001]     This application is based upon and claims the benefit of priority from Japanese Patent Application No. 2005-128268, filed Apr. 26, 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 a microscope apparatus provided with an immersion objective lens.  
         [0004]     2. Description of the Related Art  
         [0005]     Function analysis of genes is widely conducted through experiments on cultured cells; one of such experiments is performed by time-lapse observation in which cultured cells are intermittently photographed for a prolonged time period. The cultured cells, i.e., living cells are generally damaged by photostimulation. Hence, to minimize the damages to the living cells, an objective lens with a high numerical aperture (NA) is employed in the observation of the cultured cell because such a lens can capture more fluorescence with a smaller amount of exciting light.  
         [0006]     Suitable objective lenses with a high NA for such observation are immersion objective lenses, which are employed together with high-refractive index liquid which fills up a space between the immersion objective lens and a sample to be observed. Japanese translation of PCT international application No. 2004-531765 proposes a liquid feeder which supplies liquid to the immersion objective lens. The proposed liquid feeder supplies liquid to the immersion objective lens through an outlet of a feeding unit arranged near a side of an exit lens of the immersion objective lens. Since the outlet of the feeding unit is located close to the exit lens of the immersion objective lens, a simply-structured liquid feeder can supply the liquid without the need of a moving mechanism for the feeding unit.  
         [0007]     The above-described arrangement, however, in which the outlet of the feeding unit is located near the side of the exit lens, is disadvantageous in that a large amount of liquid is required for filling up a gap between the exit lens and the sample. The immersion objective lenses can be classified into two groups depending on the types of employed liquid; namely, the water immersion objective lenses that employ water and oil immersion objective lenses that employ oil. On the one hand, the use of the oil immersion tends to accompany increase in experiment cost, since the oil employed for the oil immersion is expensive, and a larger amount of oil needs to be supplied than is required simply for observation. On the other hand, the use of the water immersion also accompanies increase in experiment cost, since a container with a large capacity is required for storage of a larger amount of liquid than is necessary, and a high-performance pump needs to be provided for reduction of time required for the liquid feed.  
         [0008]     A necessary amount of supplied liquid can be minimized with the use of a nozzle and a pump that are generally used for supply of determinate quantity of liquid. For the minimization of the amount of supplied liquid, however, the liquid must be fed from substantially directly above a targeted portion, i.e., the immersion objective lens. At a time of the observation, the nozzle has to be removed from a moving range of the immersion objective lens, and hence an additional mechanism for nozzle retraction and a driving unit for the mechanism are necessary, which implies a further increase in equipment cost.  
       SUMMARY OF THE INVENTION  
       [0009]     A microscope apparatus according to one aspect of the present invention includes an immersion objective lens that condenses light from a sample through a liquid; a nozzle that supplies the liquid to an upper surface of the immersion objective lens; and a liquid feeding mechanism that cooperates with one of a lens moving mechanism that moves the immersion objective lens and a sample moving mechanism that holds and moves the sample, and moves the nozzle relative to the immersion objective lens to supply the liquid.  
         [0010]     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  
       [0011]      FIG. 1  is a schematic diagram of a microscope apparatus according to a first embodiment of the present invention;  
         [0012]      FIG. 2  is a detailed diagram of a mechanical unit of a liquid feeder in the microscope apparatus shown in  FIG. 1 ;  
         [0013]      FIG. 3  is a flowchart of a liquid feed process according to the first embodiment of the present invention;  
         [0014]      FIG. 4  shows main parts of a microscope apparatus according to a second embodiment of the present invention;  
         [0015]      FIG. 5  is an enlarged view of a nozzle shown in  FIG. 4  and a surrounding portion thereof;  
         [0016]      FIG. 6  shows main parts of a microscope apparatus according to a third embodiment of the present invention; and  
         [0017]      FIG. 7  schematically shows an objective lens of  FIG. 6  and a surrounding portion thereof as viewed from above. 
     
    
     DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS  
       [0018]     Exemplary embodiments of the present invention will be described in detail below with reference to the accompanying drawings.  
         [0019]     A microscope apparatus according to a first embodiment of the present invention is shown in  FIG. 1 . The microscope apparatus of  FIG. 1  includes a culture section  101  for maintenance of a proper environment, and a microscope section  102  for observation. The culture section  101  and the microscope section  102  each have a heat insulating layer  112  which blocks heat transfer to/from outside, and a heater  103  arranged in contact with an inner wall of the heat insulating layer  112 , so that temperature of each of the culture section  101  and the microscope section  102  can be maintained at a constant level. At a portion where the culture section  101  and the microscope section  102  are joined, an elastic sealing member  108  is provided to secure air-tightness of the culture section  101 .  
         [0020]     The culture section  101  further includes a temperature sensor  104 , a moisturizing pad  105 , a CO 2  sensor  106 , and an electromagnetic valve  107 . The microscope section  102  has a controller  109  which controls the heater  103  according to signals supplied from the temperature sensor  104 , the electromagnetic valve  107  according to signals supplied from the CO 2  sensor  106 , thereby maintaining the culture section  101  in which a cultured cell  110  is placed generally at a temperature of 37° C., a CO 2  concentration of 5%, and a relative humidity of at least 95% so as to maintain an activity of the cultured cell  110 . Further, the culture section  101  includes a transmitted light source  111  which is arranged on a top surface thereof to allow for an observation with transmitted light.  
         [0021]     The microscope section  102  further includes an immersion objective lens  113 , a focusing unit  114  which moves the immersion objective lens  113  up and down in a vertical direction, a stage  115  on which a sample container  121  housing the cultured cell  110  is placed, an imaging lens  116  which focuses parallel light rays on the immersion objective lens  113 , an incident light source  117  which illuminates the sample through the immersion objective lens  113 , a fluorescent filter  118 , and a charge coupled device (CCD) camera  119 . The stage  115  has a linearly moving unit and a rotationally moving unit and is able to two-dimensionally move the sample container  121  in a plane perpendicular to an optical axis of the immersion objective lens  113  relative to the immersion objective lens  113 . The transmitted light source  111  attached to the culture section  101  is employed for a morphological observation which aims at capturing an overall image of the cultured cell  110 , while the incident light source  117  and the fluorescent filter  118  are employed for fluorescent observation of a specific site of the cultured cell  110  with the use of fluorescent dye, fluorescent protein, or the like.  
         [0022]     The stage  115  protrudes from the microscope section  102  toward the culture section  101  with an elastic sealing member  120  placed between a bottom surface of the culture section  101  and the stage  115  so that humidity inside the culture section  101  does not leak out to the microscope section  102 . A function of the sealing member  120  can alternatively be realized by a gap between the stage  115  and the bottom surface of the culture section  101  when the gap is set to approximately submillimeter. The controller  109  is also connected to and controls the transmitted light source  111 , the focusing unit  114 , the stage  115 , the incident light source  117 , and the CCD camera  119 .  
         [0023]     The space between the immersion objective lens  113  and the sample container  121  is filled with liquid such as water or oil. Since the liquid is not supported by a specific holding unit, the liquid may adhere to the sample containers  121  when plural sample containers  121  are used for observation, or may expand due to change in observation position in the sample container  121  even when the single sample container  121  is employed for observation. In such cases, the amount of liquid may become insufficient and the refill of the liquid may become necessary.  
         [0024]     The liquid feeder is largely divided into a mechanical unit and a liquid delivery unit. The liquid delivery unit has a liquid feeding unit and a liquid discharging unit. The liquid feeding unit includes a feeding tank  122  which stores the liquid, a feeding pump  123  which delivers the liquid, a rotating arm  124  which rotates around an axis parallel to the optical axis of the immersion objective lens  113 , and a nozzle  125  which is fixed to the rotating arm  124 . The nozzle  125  and the rotating arm  124  are penetrated by a thin hole inside, and the rotating arm  124 , the feeding pump  123 , and the feeding tank  122  are connected with each other by a tube  126  made of silicon or the like. The liquid discharging unit includes a waste liquid saucer  127  in which the liquid drops off from the immersion objective lens  113  is stored, a discharge pump  128  which serves to discharge a fixed amount of stored liquid, and a discharge tank  129  which stores the liquid. The waste liquid saucer  127 , the discharge pump  128 , and the discharge tank  129  are connected with each other by a tube  126  similar to the tube in the liquid feeding unit. The tube  126  extends from the feeding pump  123  to the rotating arm  124  along the inner wall of the microscope  102 .  
         [0025]      FIG. 2  is a detailed diagram of the mechanical unit of the liquid feeder. A solid line in  FIG. 2  shows a liquid feeding state where the immersion objective lens  113  is in a lower position, and a two-dot chain line in  FIG. 2  shows an observation state in which the immersion objective lens  113  is in an upper position. The mechanical unit includes a cam member  130  provided in the focusing unit  114 , and the rotating arm  124  arranged at an upper portion of the microscope section  102 . The cam member  130  has a cam surface inclined toward a vertical moving direction of the immersion objective lens  113 . The cam member  130  triggers an operation to drive the rotating arm  124 . The rotating arm  124  is rotatably supported by a bearing  131  on an inner side surface of the upper surface of the microscope section  102 , and protrudes toward the culture section  101  from the microscope section  102 . The nozzle  125  is fixed to the rotating arm  124  so that the nozzle  125  is arranged perpendicular to the rotating axis of the rotating arm  124 .  
         [0026]     As shown in  FIG. 1 , an elastic sealing member  132  is placed between the rotating arm  124  and the microscope section  102 . In  FIG. 2 , the rotating arm  124  is provided with a rotating pin  133  which contacts with the cam member  130  within the moving range of the focusing unit  114 . The rotating pin  133  is arranged perpendicular to the rotation axis of the rotating arm  124 . On an inner side of the microscope section  102 , a restricting pin  134  is attached. The restricting pin  134  contacts with the rotating pin  133  and restricts the rotation of the rotating arm  124 . To bring the rotating pin  133  into contact with the restricting pin  134 , a hook  135  attached inside the upper portion of the microscope section  102  is connected to the rotating pin  133  by an elastic spring member  136 .  
         [0027]     In  FIG. 1 , the sample container  121  is fitted into a depressed portion  115   a  on the stage  115 , and fixed by a plate-like elastic metal fixing member  137 . When the cultured cell  110  is observed, the immersion objective lens  113  is located at an upper position within the moving range of the focusing unit  114 . In the observation state, the rotating arm  124  is brought into contact with the restricting pin  134  due to the force from the spring member  136  connected to the rotating arm  124 . In this state, the rotating pin  133  of the rotating arm  124  is not in contact with the cam member  130  attached to the focusing unit  114 .  
         [0028]     When an observation target changes to the cultured cell  110  contained in the other sample container  121  placed on the stage  115 , the immersion objective lens  113  is lowered by a significant degree by the focusing unit  114  so that the stage  115  does not interfere with the immersion objective lens  113 . Then, the cam member  130  pushes the rotating pin  133  to cause the rotation of the rotating arm  124  against the tensile force of the spring member  136 , whereby the nozzle  125  attached to the rotating arm  124  is placed near and above a top lens of the immersion objective lens  113 . Thus, the nozzle  125  moves in conjunction with the movement of the immersion objective lens  113  caused by the focusing unit  114 .  
         [0029]     Since the objective lens is positioned to an accuracy of submicrometer (μm), even a small amount of external force can easily cause defocusing and the proper repositioning of the objective lens is difficult. In the present embodiment, however, the focusing unit  114  rotates the rotating arm  124  not to a focus position required for an image pick-up, and hence, the application of external force to the focusing unit  114  would not cause defocusing. In addition, since the temperature of the liquid reaches the same level as the temperature of the microscope section  102  while passing through the tube  126  running along the inner wall of the microscope section  102 , in other words, the temperature of the liquid becomes the same as the temperature of the immersion objective lens  113  to which the liquid is delivered, there would be no defocusing caused by the temperature change in the immersion objective lens  113 . Still in addition, since the nozzle  125  is placed near and above the top lens of the immersion objective lens  113 , the liquid can be supplied through the nozzle  125  by a minimum amount required for the observation. Still in addition, since the rotation of the nozzle  125  can be realized without the need of a dedicated driving unit, the microscope can be manufactured by low cost.  
         [0030]     The timing of liquid feed will be described below with reference to the flowchart of  FIG. 3 .  
         [0031]     On power-up of the microscope apparatus, the feeding unit starts control (Step S 1 ).  
         [0032]     The necessity of liquid feed to the immersion objective lens  113  is evaluated based on following three Conditions 1 to 3 (Step S 2 ).  
         [0033]     Condition 1: The liquid, particularly the water, of the immersion objective lens  113  decreases by evaporation. The necessity of the liquid feed is determined based on the time elapsed since last liquid feed. When the time elapsed since the last liquid feed exceeds a predetermined time period, a liquid feed operation starts. Here, an optimal value is set in advance as a value of the predetermined time period depending on the composition of the liquid, for example, depending on whether the liquid is water or oil.  
         [0034]     Condition 2: The liquid of the immersion objective lens  113  adheres to the bottom of the container while transferred from one container to another, and the amount available gradually decreases. The amount of liquid decrease is determined based on the number of transfers among containers. When the number of transfers exceeds a predetermined number, the liquid feed operation starts.  
         [0035]     Condition 3: When the point of observation changes in the single container, the container is moved. Then, the liquid of the immersion objective lens  113  adheres to the bottom of the container as if the liquid is applied thereto, and the available amount of liquid decreases. The amount of decreased liquid is determined based on the distance the container moves. When the moving distance exceeds a predetermined distance, the liquid feed operation starts.  
         [0036]     In the liquid feed operation, the focusing unit  114  is first lowered so that the nozzle  125  is placed over the immersion objective lens  113  (Step S 3 ).  
         [0037]     The feeding pump  123  and the discharge pump  128  are operated for a predetermined time period (Step S 4 ). The operation time of the feeding pump  123  may be set so that a slightly larger amount of liquid is supplied than is necessary in order to prevent supply shortage. For example, if the required amount is 0.2 cc, the operation time is set so that 0.3 cc liquid is supplied.  
         [0038]     When the observation point needs to be changed, the stage  115  is moved (Step S 5 ).  
         [0039]     It is decided whether the operation of the feeding pump  123  and the discharge pump  128 , and the moving operation of the stage  115  have been finished (Step S 6 ).  
         [0040]     Then, the focusing unit  114  is moved to a predetermined position (Step S 7 ).  
         [0041]     Finally, latency T2 is set by a timer, and after the latency T2 passes, the feed operation ends (Step S 8 ). The latency T2 is set to eliminate a small temperature difference between the liquid and the immersion objective lens  113 , and serves to prevent defocusing from being caused by the temperature difference which induces extension of material used in the immersion objective lens  113 .  
         [0042]     According to the above-described control manner, the liquid feed to the immersion objective lens  113  is performed simultaneously with the two-dimensional movement of the immersion objective lens  113  relative to the optical axis. Hence, the above manner can shorten the time interval between observations compared with a control manner in which steps are conducted sequentially in series. In addition, since the latency T2 is provided in Step S 8  prior to the observation, defocusing of the immersion objective lens  113  can be prevented from happening by the temperature change, whereby an image with no blurring can be taken.  
         [0043]     In the first embodiment, the microscope apparatus provided with the culture section is described. The liquid feeder of the present embodiment, however, can be applied to a general manual microscope.  
         [0044]     A microscope apparatus according to a second embodiment has a liquid feed-related portion shown in  FIG. 4  which is different from the microscope apparatus of the first embodiment. The parts not shown in  FIG. 4  are the same as those in the first embodiment. The microscope apparatus according to the second embodiment includes a culture section  200  and a microscope section  201 . The microscope section  201  includes a linearly moving stage  202  which is attached to an upper surface thereof and which is movable in one axial direction. The linearly moving stage  202  is further attached to a rotary stage  203  which is rotatable. The linearly moving stage  202  has a sealing portion  202   a  which protrudes toward the culture section  200  from the microscope section  201 . Further, the rotary stage  203  has a tray connecting portion  203   a  which protrudes toward the culture section  200  from the microscope section  201 .  
         [0045]     The tray connecting portion  203   a  has a male dovetail  203   b , whereas a tray  205  has a female dovetail  205   a  which has such a shape that it is engaged with the male dovetail  203   b . The sealing portion  202   a  has a nozzle  206  for feeding the liquid to the immersion objective lens  204 . An elastic sealing member  207  is arranged between the sealing portion  202   a  and the upper surface of the microscope section  201  to prevent the leakage of humidity generated in the culture section  200  to the microscope section  201 . The function of the sealing member  207  may alternatively be realized by setting a gap between the sealing portion  202   a  and the upper surface of the microscope section  201  to approximately submillimeter.  
         [0046]     A general glass bottom dish employed as the sample container  208  has an outer diameter of approximately 35 mm and an observation range of approximately 10 mm in diameter. The sample container  208  is placed on the tray  205  on a circle of approximately 70 mm in radius around a rotation axis Y of the rotary stage  203 . The moving direction of the linearly moving stage  202  is parallel to a straight line which is perpendicular to the rotation axis Y of the rotary stage  203  and an optical axis X of the immersion objective lens  204 . The linear stage  202  and the rotary stage  203  can cooperate to move the sample container  208  placed on the tray  205  two-dimensionally within a plane perpendicular to the optical axis of the immersion objective lens  204 . Since the sample container  208  has an observation range of 10 mm in diameter, the moving range of the linear stage  202  is required to be approximately 10 mm for observation. Thus, the observation range of the sample container  208  can be observed by the CCD camera. Since the rotation angle of the rotary stage  203  relative to the imaging range of the CCD camera is sufficiently small, an operator can regard the movements as movements in two perpendicular directions during operation.  
         [0047]     The linear stage  202  of the present embodiment has a moving range which is larger than the necessary range 10 mm for observation by 20 mm. The additional moving range of 20 mm is set based on the half length, i.e., 15 mm of the outermost diameter 30 mm of the immersion objective lens  204 . With the moving range of 30 mm, the nozzle  206  placed outside the immersion objective lens  204  can be arranged near and above a top lens of the immersion objective lens  204  within the observation range of the sample container  208 .  
         [0048]     As shown in  FIG. 5 , the sealing portion  202   a  has a depressed portion  202   b  for the attachment of the nozzle  206 . The sealing portion  202   a  has a depressed portion  202   b  and a small hole  202   c  which communicates with inside the microscope section  201  as shown in  FIG. 4 . The small hole  202   c  is connected to a tube  209  which is connected to the feeding pump and the feeding tank though not shown in  FIG. 4 .  
         [0049]     As shown in  FIG. 5 , the nozzle  206  has a protruding portion  206   a  which has such a shape that it is engaged with the depressed portion  202   b  of the sealing portion  202   a , and is provided with a small hole  206   b  which runs from the protruding portion  206   a  to the bottom surface near the nozzle tip. Further, an O ring  210  is inserted into a groove  206   c  provided in a cylindrical portion of the protruding portion  206   a . Further, the nozzle  206  has a flange  206   d  slightly larger than the protruding portion  206   a , and the flange  206   d  serves as a contact surface to the sealing portion  202   a  of the nozzle  206 . The nozzle  206  is fixed to the sealing portion  202   a  by the elasticity of the O ring  210 . Therefore, when the controller of the microscope apparatus commits a processing error, or when the operator exerts external force on the microscope apparatus, e.g., at cleaning, the nozzle can be easily removed so that the damage to the microscope apparatus and contamination inside the microscope apparatus can be prevented.  
         [0050]     A control flow will be described. First, the necessity of the liquid feed is determined. When the liquid feed is necessary, the focusing unit  211  is first lowered for the prevention of interference between the tray  205  and the immersion objective lens  204 . Thereafter, two operations are performed in parallel. One is an operation of rotational movement of the rotary stage  203  at the exchange of the sample container  208  or the change in the observation position; another is the liquid feed operation. In the rotational movement operation, the rotary stage  203  is rotated after the liquid feed operation so that the next observation point is located on the optical axis X of the immersion objective  204 . Further, in the liquid feed operation, the nozzle  206  is arranged near and above the top lens of the immersion objective lens  204  by the linearly moving stage  202 . Then the liquid is supplied and thereafter the nozzle  206  is returned back to a substantially original position by the linearly moving stage  202 . At the return of the nozzle  206 , the nozzle  206  is positioned so that the next observation point is located on the optical axis X of the immersion objective lens  204 . When the two operations are finished, the focusing unit  211  is raised to the focusing position of the immersion objective lens  204 . After the latency for the correction of temperature difference passes, the observation starts.  
         [0051]     In the above-described control, the liquid feed operation starts after the immersion objective lens  204  is lowered by the focusing unit  211 , and the tray  205  is moved by the linear stage  202  and the rotary stage  203 . Therefore, the tray  205  is not interfered by the immersion objective lens  204 . Still further, since the liquid feed operation and the movement to the next observation point are performed in parallel, the latency can be shortened.  
         [0052]     In the above-described control, when the shape of the tray  205  is considered and the movement of the focusing unit  211  is made faster than the movement of the linear stage  202 , the interference between the tray  205  and the immersion objective lens  204  can be prevented and the latency in the control system can be shortened.  
         [0053]     In the present embodiment, the enlargement of the moving range of the linear stage and the addition of nozzle to the sealing portion of the linear stage make the liquid feed possible. Hence, an additional driving unit is not necessary, whereby the minimum liquid feed can be realized with a fewer number of parts than the parts added in the first embodiment.  
         [0054]      FIG. 6  shows a microscope apparatus according to a third embodiment, in particular a portion related with the liquid feed and is different from the parts in the first embodiment. Those not shown in  FIG. 6  are the same as those in the first embodiment. The microscope apparatus of the present embodiment is similar to the microscope apparatus according to the second embodiment in many points, and in  FIG. 6 , the similar parts to the parts in the microscope apparatus according to the second embodiment are designated by the same reference characters.  FIG. 7  shows the objective lens in  FIG. 6  and the surrounding portion thereof viewed from above.  
         [0055]     As shown in  FIG. 6 , a focusing unit  300  has a rotating-type revolver  302  which serves to switch among plural objective lenses. Two immersion objective lenses  301   a  and  301   b  can be attached to the revolver  302 , which moves the immersion objective lenses  301   a  and  301   b  by rotation. The revolver  302  is designed so as to be able to position the immersion objective lens at intervals of 90 degrees. In a plane perpendicular to the optical axis X in the optical system of the microscope section  201  in which the objective lens is arranged for observation in  FIG. 7 , the tip of the nozzle  303  is arranged on an axis Z which is parallel to the optical axis X and passing through a point 90 degrees off from the optical axis X and which is on the circle around the center of rotation of the revolver  302  and passing through the optical axis X.  
         [0056]     Though two objective lenses can be attached to the revolver  302  in the present embodiment, objective lenses of a number “n” may be attached to the revolver  302 . In this case, the revolver  302  is designed so as to be able to position the objective lenses at intervals of θ degrees (here, θ=360÷2n). The nozzle  303  is arranged on the circle around the center of rotation of the revolver  302  and passing through the optical axis X. The arranged position of the nozzle  303  is θ (here, θ=360÷2n) degrees off from the optical axis X.  
         [0057]     When one of the immersion objective lenses  301   a  and  301   b  is arranged on the optical axis X for observation, the other is not placed on the axis Z. The immersion objective lens located on the optical axis X is raised at the time of observation, so that the working distance (WD) between the immersion objective lens and the sample container  208  is approximately 0.1 mm. Further, when one of the immersion objective lenses  301   a  and  301   b  is placed on the axis Z for the liquid feed, the other is not placed on the optical axis X, and hence, it is not necessary to raise the focusing unit  300 . Therefore, one of the top lenses of the immersion objective lenses  301   a  and  301   b  can be placed near and below the nozzle  303 , which can then supply the liquid to the immersion objective lens from above.  
         [0058]     The control flow will be described. First, the necessity of the liquid feed is decided. When the liquid feed is necessary, the focusing unit  300  is lowered for the prevention of the interference between the tray  205  and the immersion objective lens  301   a . Thereafter, two operations are performed in parallel. One is the movement of the rotary stage  203  and the linear stage  202  at the exchange of the sample container  208  and the changes in observation point, and the other is the liquid feed operation. In the liquid feed operation, the immersion objective lens  301   a  is placed on the axis Z by the rotation of the revolver  302 . Then, the top lens of the immersion objective lens  301   a  is placed near and below the nozzle  303 . Sequentially, the liquid is supplied, and the immersion objective lens  301   a  is put back on the optical axis X by the rotation of the revolver  302 . Thereafter, the focusing unit  300  is raised to the observation position. After the elapse of the latency for the temperature difference correction, the observation begins.  
         [0059]     In the present embodiment, the immersion objective lens is placed below the fixed nozzle  303  by the revolver  302  which serves as a switching unit between the immersion objective lenses  301   a  and  301   b . Thus, the liquid feed of minimum amount can be realized in a still simpler configuration than the configuration of the second embodiment.  
         [0060]     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.