JP4388298B2 - Microscope system - Google Patents

Description

[0001]
BACKGROUND OF THE INVENTION
The present invention relates to a microscope system including, for example, an electric focusing device that adjusts a relative distance between an objective lens and a specimen to perform autofocusing on the specimen.
[0002]
[Prior art]
Currently, there are microscopes that can observe a fine specimen and record an observation image of the specimen as video image data. This microscope is widely used for, for example, research in the biological field and inspection processes in the industrial field. When this microscope is used, the focusing operation is usually performed by adjusting the focus on the specimen by operating the focusing handle. However, when the focal depth is shallow and the focus range is narrow like a high-magnification objective lens, considerable skill is required to quickly perform the focusing operation, and the workability is poor.
[0003]
Such poor workability has an adverse effect on observer fatigue and a reduction in production efficiency in the industrial field. Particularly during routine operations such as inspection processes, it is very important to reduce the inspection time by quickly performing the focusing operation.
[0004]
Various autofocus (hereinafter referred to as AF) devices for microscopes that can automatically perform the focusing operation from such circumstances have been proposed, and many proposals for improving these have been made.
[0005]
The AF apparatus in the industrial field not only improves the operability and throughput described above, but also leaks defects in each layer and the line width between patterns for a specimen having a step shape such as a semiconductor wafer formed in multiple layers. Therefore, there is a need to detect and measure, and to measure a minute step shape on a specimen with high accuracy, and an AF apparatus having performance suitable for such inspection and measurement has been proposed.
[0006]
In such an AF apparatus in the industrial field, light such as an infrared laser is projected onto the specimen for reasons such as compatibility with the specimen and shortening of the AF time, and the state of the reflected light from the specimen is detected and focused. There are many so-called active AF systems that perform operations.
[0007]
On the other hand, in an AF apparatus in the biological field, a more accurate focus position is required, and a transmission type specimen having a low reflectance that is impossible in the active method is used. The so-called passive AF method that performs the operation is the mainstream.
[0008]
In recent years, not only in the industrial and biological fields, the number of cases in which the drive of the stage on which the specimen is placed in the plane (XY plane) direction is motorized and used in combination with AF to automate and speed up specimen inspection. ing. For example, in the biological field, a so-called telepathology technique for inspecting a pathological tissue from a remote place is applicable, and in the industrial field, the above-described automatic defect inspection apparatus for a semiconductor wafer is applicable.
[0009]
In a microscope system equipped with such an AF device, the focus is shifted by AF accompanying movement in the XY plane, that is, when the specimen is moved in the XY plane, the specimen is reflected at a specific position, for example, active AF. In a range where the rate is extremely low, or in the case of passive AF, where the contrast is extremely low, the AF operation fails in the sample area where focusing is difficult in each AF method, and the focus position is shifted.
[0010]
When such a focus shift occurs, an accurate specimen cannot be inspected and an inspection error occurs, or the observer needs to manually focus, leading to a decrease in throughput.
[0011]
In order to alleviate such a problem, for example, Japanese Patent Laid-Open No. 2001-91846 sequentially stores XYZ positions when AF is successful, and when AF fails at the current position, It is described that the focus shift due to AF failure is minimized by reading the position and moving the stage to the Z position.
[0012]
[Problems to be solved by the invention]
However, if the specimen surface is tilted, if the previous AF is performed at a position far from the current position, the Z position for focusing is completely different between the position where the previous AF was performed and the current position, Even if the stage is moved to the Z position when the previous AF was successful, the focus position will be greatly shifted at the current position.
[0013]
In Japanese Patent Laid-Open No. 2001-91846, a reference sample is placed on a stage as an initial setting, AF is performed at each position on the reference sample, and its focus position (Z position) is stored in advance. It is disclosed that the position is used to correct the Z position of the stage as the specimen moves in the XY plane.
[0014]
However, in the correction of the focus position, only correction of the tilt of the stage itself is taken into consideration, and no countermeasure is taken against the tilt of the sample surface placed on the stage. In addition, it takes a lot of time to perform the initial setting.
[0015]
Therefore, an object of the present invention is to provide a microscope system capable of stably obtaining an image of a specimen by suppressing focus deviation with respect to the specimen as much as possible even when AF does not operate in the specimen.
[0016]
[Means for Solving the Problems]
  The present inventionThe first aspect ofMakes it possible to move the stage on which the specimen is placed in a plane direction perpendicular to the optical axis of the objective lens, and to adjust the distance between the stage and the objective lens relative to each other to automatically focus the specimen. In the microscope system, the position data indicating coordinates in the plane direction of the stage when the autofocus is performed on the specimen and the focus data of the distance between the specimen and the objective lens are stored. When the position storage means and the autofocus is not performed on the specimen,The position data of the coordinates closest to the coordinates that are not autofocused are retrieved from the position storage means, and the focus data corresponding to the retrieved position data is used.A microscope system comprising: a focus position correcting unit that changes a relative distance between the stage and the objective lens..
[0017]
  In addition, the present inventionSecond aspect ofIsThe stage on which the specimen is placed can be moved in a plane direction perpendicular to the optical axis of the objective lens, and the distance between the stage and the objective lens is relatively adjusted to perform autofocus on the specimen. In a microscope system to perform, a position memory that stores position data indicating coordinates in a plane direction of the stage when the specimen is autofocused, and focus data of a distance between the specimen and the objective lens Means for calculating the inclination of the sample surface based on the position data and the focus data at at least two coordinates stored in the position storage means; and When the focus is not performed, the position data at the coordinates where the autofocus is not performed and the sample inclination calculation means And a focus position correcting unit that predicts a focus position based on the calculated inclination of the sample and changes a relative distance between the stage and the objective lens based on the focus position. Characterized byIt is a microscope system.
[0018]
  The present inventionThe third aspect of the present invention includes an autofocus execution feasibility judging means for judging whether or not the autofocus at each coordinate is possible for the sample. It is a microscope system of an aspect.
  According to a fourth aspect of the present invention, the stage on which the sample is placed can be moved in a plane direction perpendicular to the optical axis of the objective lens, and the distance between the stage and the objective lens is set. In a microscope system that performs real-time autofocusing that adjusts relatively and always follows the focus position with respect to the specimen, the stage moves each time the stage moves in the plane direction reaches a preset amount of movement. Position data in the plane direction and focus data of the distance between the specimen and the objective lens, and at least two autofocus positions stored in the position memory Sample inclination calculation means for calculating the inclination of the sample surface based on the position data and the focus data, and for the sample When the focus position cannot be tracked, a focus position is predicted based on the tilt of the sample surface calculated by the sample tilt calculating means within a range where the focus position cannot be tracked, and the stage and the stage are determined according to the focus position. A microscope system comprising: a focus position correcting unit that moves relative to the objective lens.
[0019]
DETAILED DESCRIPTION OF THE INVENTION
Hereinafter, a first embodiment of the present invention will be described with reference to the drawings.
[0020]
FIG. 1 is a configuration diagram of a microscope system. A plurality of objective lenses 2 are attached to a revolver body 1 constituting an electric revolver. The revolver body 1 is rotated by driving a revolver motor 3, and an objective lens 2 having an arbitrary magnification is inserted into the optical path.
[0021]
A revolver hole (hereinafter abbreviated as “revo hole”) position detection unit 4 detects which hole position of the revolver body 1 is attached with the objective lens 2 and sends a detection signal to the control unit 5.
[0022]
The control unit 5 receives a selection signal for the objective lens 2 from the operation unit 6 and sends a signal for inserting the objective lens 2 instructed by the selection signal into the optical path to the revolver motor drive unit 7. Have
[0023]
The revolver motor drive unit 7 inputs a signal from the control unit 5, drives the revolver motor 3 to rotate the revolver body 1, and inserts the instructed objective lens 2 into the optical path.
[0024]
As shown in FIG. 2, the control unit 5 is connected to a CPU main body 8 via a data bus 9, a ROM 10, a RAM 11 and an I / O port 12. Among these, the ROM 10 stores a program for controlling the microscope system. The RAM 11 is, for example, a volatile memory that stores data necessary for controlling the microscope system. The I / O port 12 inputs and outputs control signals. Further, the control unit 5 is provided with peripheral circuits such as an oscillator and an address decoder which are necessary for controlling the CPU main body 8 although not shown. Control of each peripheral device is performed via the I / O port 12 and the data bus 9.
[0025]
An XY stage 14 is provided on the Z stage 13. A sample S is placed on the XY stage 14. The Z stage 13 is movable in the Z direction (optical axis direction) and moves up and down in the optical axis direction by driving the focusing motor 15. The XY stage 14 can move in the XY plane perpendicular to the optical axis direction, and moves two-dimensionally in the XY plane by driving the XY stage motor 16.
[0026]
A focusing motor drive unit 17 is connected to the focusing motor 15, and an XY stage motor driving unit 18 is connected to the XY stage motor 16. These motors 15, 16 are driven by the motor driving units 17, 18, respectively. To do. The focusing motor driving unit 17 and the XY stage motor driving unit 18 are connected to the I / O port 12 of the control unit 5 and input control signals from the control unit 5.
[0027]
The operation unit 6 performs operation instructions for the microscope system, and includes an AF start switch, a focus setting switch, and an XY movement switch.
[0028]
On the other hand, the reference light source 19 is used for AF, and outputs laser light having a wavelength in the visible light wavelength region such as an infrared laser. The reference light source 19 is pulse-lit by the laser driving unit 20 and the intensity of the emitted laser light is controlled.
[0029]
The laser light emitted from the reference light source 19 is shaped into parallel light by the collimator lens 21 and half the diameter of the light beam is cut by the light projecting side stopper 22. The cut laser light is incident on a polarization beam splitter (hereinafter referred to as PBS) 23, where only the P-polarized component is reflected.
[0030]
The laser beam of the P-polarized component reflected by the PBS 23 is once condensed by the condenser lens group 24 and enters the chromatic aberration lens group 25. The chromatic aberration lens group 25 is moved in the optical axis direction (arrow A direction) when the chromatic aberration lens driving motor 27 is driven by the chromatic aberration lens driving unit 26. The chromatic aberration can be corrected by moving the chromatic aberration lens group 25 in the optical axis direction. The chromatic aberration lens driving unit 26 drives the chromatic aberration lens driving motor 27 in accordance with a control signal sent from the control unit 5.
[0031]
The laser beam that has passed through the chromatic aberration lens group 25 passes through the λ / 4 plate 28, is polarized by 45 °, and enters the dichroic mirror 29. The dichroic mirror 29 reflects a laser beam only in the infrared region. The laser light beam reflected by the dichroic mirror 29 is condensed by the objective lens 2 and irradiated onto the sample S as a spot-shaped image.
[0032]
The light beam reflected by the sample S travels through the optical path opposite to the optical path of laser beam irradiation to the sample S, that is, travels through the objective lens 2 and the dichroic mirror 29 and enters the λ / 4 plate 28, where it is polarized by 45 °. Thus, the s-polarized light component is switched. The light beam polarized by the λ / 4 plate 28 travels through the color correction lens group 25 and the condenser lens group 24 and enters the PBS 23. Since the light beam incident on the PBS 23 is an S-polarized component, it passes through the PBS 23 as it is, and is further imaged on the light receiving sensor 31 by the condenser lens group 30.
[0033]
The light receiving sensor 31 is a two-divided photodiode, and is divided into light receiving regions of an A range and a B range by the dividing line 31a. The light receiving sensor 31 is arranged with the dividing line 31a aligned on the optical axis.
[0034]
As shown in FIGS. 3A and 3B, the spot of the light beam formed on the light receiving sensor 31 has a narrow spot area when it is at the focus position (when the sample S is at the focus position). Therefore, the light receiving sensor 31 outputs a sensor signal indicating a high light intensity. When the spot is above the focus position (front pin position) as shown in FIGS. 4A and 4B, the light intensity distribution is biased toward the A range. The sensor signal shown is output. 5A and 5B, when the spot is on the lower side (rear pin position) from the focus position, the light intensity distribution is biased toward the B range, and the light receiving sensor 31 has such a light intensity distribution. A sensor signal indicating is output.
[0035]
The signal processing unit 32 receives the sensor signal output from the light receiving sensor 31, divides the sensor signal into signal components (A range signal and B range signal) of the A range and the B range, and these A range signals. A total of each light intensity for each B range signal is calculated, and the A range which is symmetrical with respect to the focus position with the horizontal axis as shown in FIG. Each curve of the signal and the B range signal is detected, and these curves are sent to the controller 5.
[0036]
The controller 5 inputs an A range signal and a B range signal, and from these signals,
(AB) / (A + B) (1)
6b is obtained, and a control signal for controlling the Z stage 13 in the vertical direction is sent to the focusing motor drive unit 17 so that the incident light intensity in this curve becomes the zero cross point. Then, focus operation is performed.
[0037]
As described above, the active AF optical system is configured by controlling the lighting of the light source 19 that emits laser light and detecting the reflected light when the sample S is irradiated.
[0038]
On the other hand, an illumination light source 33 for observing the specimen S is provided. The illumination light emitted from the illumination light source 33 passes through the lens 34, is reflected by the half mirror 35, and irradiates the sample S from above. Reflected light from the specimen S passes through the objective lens 2, passes through the half mirror 35, then passes through the dichroic mirror 29, and becomes observation light.
[0039]
An encoder 36 is provided for detecting the amount of movement when the Z stage 13 is moved up and down in the optical axis direction using a manual focusing mechanism (not shown). The output pulses of the encoder 36 are counted by a pulse counter 37, and this count value is sent to the control unit 5.
[0040]
As shown in FIG. 7, the control unit 5 includes a position storage unit 38 and a focus position correction unit 39. Among these, the position storage unit 38 includes position data (XY coordinates) in the plane direction of the XY stage 14 when AF is performed on the sample S, and focus data of the distance between the sample S and the objective lens 2, that is, the Z stage. For example, the RAM 11 has a function of storing the 13 positions in the Z direction.
[0041]
If the AF is not performed on the sample S, the focus position correcting unit 39Not AFThe RAM 11 has a function of retrieving the XY coordinate closest to the XY coordinate of the XY stage 14 from the RAM 11 and moving the Z stage 13 in the optical axis direction according to the position in the Z direction corresponding to the retrieved XY coordinate.
[0042]
Next, the AF operation of the apparatus configured as described above will be described.
[0043]
For example, as shown in FIG. 8, the observer has a plurality of observation positions Q with respect to the surface of the semiconductor wafer S._A, Q_B, Q_CThe semiconductor wafer S is inspected while focusing on.
[0044]
First, the observer must observe the observation position Q._AThen, the AF start switch of the operation unit 6 is pressed to perform the first AF operation. That is, the laser light emitted from the reference light source 19 is shaped into parallel light by the collimator lens 21, and half of the diameter of the light beam is cut by the light projection side stopper 22, and this cut laser light is incident on the PBS 23. Only the P-polarized component is reflected. The laser beam of the P-polarized component reflected by the PBS 23 is once condensed by the condenser lens group 24 and enters the chromatic aberration lens group 25. The laser beam that has passed through the chromatic aberration lens group 25 passes through the λ / 4 plate 28. The laser beam only in the infrared region that is transmitted and polarized by 45 ° and incident on the dichroic mirror 29 and reflected by the dichroic mirror 29 is condensed by the objective lens 2 and irradiated onto the sample S as a spot-shaped image. .
[0045]
The light beam reflected by the sample S enters the PBS 23 through the optical path opposite to the optical path of the laser beam irradiation to the sample S, passes through the PBS 23 as it is, and is coupled onto the light receiving sensor 31 by the condenser lens group 30. Imaged.
[0046]
In this light receiving sensor 31, the spot of the focused light beam is at the focus position shown in FIGS. 3 (a) and 3 (b) and from the focus position shown in FIGS. 4 (a) and 4 (b) to the upper side (front pin position). There are cases where there are cases where there is a lower position (rear pin position) than the focus positions shown in FIGS. 5 (a) and 5 (b), and each outputs a sensor signal indicating a light intensity distribution.
[0047]
The signal processing unit 32 receives the sensor signal output from the light receiving sensor 31, divides the sensor signal into an A range signal and a B range signal, and sums up each light intensity for each of the A range signal and the B range signal. 6A is detected, and each curve of the A range signal and the B range signal that are symmetrical with respect to the focus position with the horizontal axis as the vertical direction (defocus) of the Z stage 13 as shown in FIG. 6A is detected. These curves are sent to the controller 5.
[0048]
The controller 5 inputs the A range signal and the B range signal, calculates the above equation (1) from these signals, obtains a curve as shown in FIG. 6B, and the incident light intensity in this curve is the zero cross point. A control signal for controlling the Z stage 13 in the Z direction is sent to the focusing motor drive unit 17 so that the focusing operation is performed.
[0049]
Here, the observation position Q_AIf the AF operation is successful, the above-described manual adjustment of the focus position and the switch operation for storing the focus position are not necessary, and after the AF operation is completed, the position storage unit 38 of the control unit 5 displays the current (observation position Q_A) The Z position of the Z stage 13 is the first observation position Q._AFocus position (Z_A) And this focus position (Z_A) Observation position Q_AXY coordinates (X_A, Y_A), That is, the focus position (X_A, Y_A, Z_A) Is stored in the RAM 11.
[0050]
On the other hand, when the focus is not normally achieved, that is, when the AF operation is unsuccessful due to factors such as the shape of the sample S or the reflectance, the control unit 5 displays an AF failure on a display unit (not shown) and notifies the observer. And urge manual focusing.
[0051]
The observer moves the Z stage 13 in the optical axis direction using the encoder 36 or a manual focusing mechanism (not shown), moves the specimen S to the focus position, and then presses the focus setting switch of the operation unit 6. In response to the pressing operation of the focus setting switch, the position storage unit 38 of the control unit 5 automatically detects the observation position Q._AFocus position (X_A, Y_A, Z_A) Is stored in the RAM 11.
[0052]
Next, the observation position Q_AThe observer who has finished observing the sample S moves the XY stage using the XY movement switch of the operation unit 6 to observe the observation position Q._BIs moved into the field of view, and the AF operation is started. When this AF operation is successful, the position storage unit 38 of the control unit 5 automatically detects the observation position Q as in the first time._BFocus position (X_B, Y_B, Z_B) Is stored in the RAM 11.
[0053]
On the other hand, when the AF operation is unsuccessful, the position storage unit 38 of the control unit 5 sets the focus position Z when the AF operation or manual focusing is performed for the first time._AIs read from the RAM 11, and the focus position Z_AThen, the Z stage 13 is moved in the Z direction to prevent the focus from being greatly blurred due to AF failure.
[0054]
Next, the observer moves the XY stage by using the XY movement switch of the operation unit 6 to observe the observation position Q._CIs moved into the field of view, and the AF operation is started. When this AF operation is successful, the position storage unit 38 of the control unit 5 automatically performs the observation position Q as described above._CFocus position (X_C, Y_C, Z_C) Is stored in the RAM 11.
[0055]
On the other hand, when the AF operation is unsuccessful, the focus position correction unit 39 of the control unit 5 determines the current (observation position Q_C) Coordinates (X_C, Y_C) And the coordinates of the observation positions (X_N, Y_N) And read these coordinates (X_N, Y_N) Current coordinates (X_C, Y_C) Closest observation position, ie
(X_C-X_N)²+(Y _C -Y _N ) ²... (2)
The focus position (X_N, Y_N, Z_N) And select the Z position Z at this focus position._NThe Z stage 13 is moved.
[0056]
That is, in the observation of the semiconductor wafer S shown in FIG._A, Q_BIf the AF operation is successful at the observation position Q_CObservation position Q for_AAnd observation position Q_BAnd the observation position Q_AThan observation position Q_BIs the observation position Q_CSince the focus position correction unit 39 is close to the observation position Q,_BFocus position Z_BSearch for this focus position Z_BIs sent to the focusing motor drive unit 17. As a result, the Z stage 13 moves the focus position Z_BMove to.
[0057]
Observation position Q_BIf the AF operation fails, the Z stage 13 moves the focus position Z_AMove to.
[0058]
Thus, in the first embodiment, an arbitrary observation position Q_n(Eg Q_C), The observation position Q at which the AF operation was successful in the past._A, Q_B, ..., the closest observation position Q_A, Q_B, ..., (eg Q_B) Focus position (X_B, Y_B, Z_B), The Z stage 13 is moved in the Z direction. Therefore, even if the AF operation fails, the influence of the focus shift due to the tilt of the sample S can be avoided as much as possible, and the sample S is located at the Z position that is closest to the focus position. XY stage 14 can be moved in the X and Y directions to obtain a specimen image in focus that is close to the specimen S almost accurately.
[0059]
Since the focus position of the sample S often depends mainly on the inclination of the sample S, the closer the observation position where the AF operation has failed to the observation position where the AF operation has been successful in the past, the closer the Z position of the Z stage 13 becomes. The focus position moved by the position correction is close to the focus position when the original AF operation is successful, and an almost accurate focus position can be obtained. In other words, a plurality of observation positions Q on the same specimen S_A, Q_B, ..., Q_NAs the number of successful AF operations increases, more accurate Z position correction can be performed. Then, the focus position (X_B, Y_B, Z_B) Is stored in the RAM 11, the specimen S is observed at random positions, and even if the AF operation fails, it is possible to obtain a specimen image in focus that is almost exactly accurate. it can.
[0060]
Therefore, for example, in the defect inspection of a semiconductor wafer, many observation positions Q on the same semiconductor wafer surface._NTherefore, the Z position correction when the AF operation fails according to the present invention is effective.
[0061]
Next, a second embodiment of the present invention will be described. The same parts as those in FIG. 1 are denoted by the same reference numerals, and detailed description thereof is omitted.
[0062]
Compared with the first embodiment, the microscope system of the present invention is provided with a sample inclination calculation unit 40 and a focus position correction unit 41 in the control unit 5 as shown in the functional block diagram of FIG.
[0063]
The sample inclination calculation unit 40 has a function of calculating the inclination of the sample S plane based on the XY coordinates of the XY stage 14 and the Z-direction position of the Z stage 13 at at least two autofocus positions stored in the RAM 11. .
[0064]
When the AF operation is not performed on the sample S, the focus position correction unit 41 is based on the XY coordinates of the XY stage 14 on which the AF operation is not performed and the inclination of the sample S surface calculated by the sample inclination calculation unit 40. And has a function of moving the Z stage 13 in the Z direction according to the focus position.
[0065]
Next, the AF operation of the apparatus configured as described above will be described.
[0066]
An observer, for example, as shown in FIG._A, Q_B, Q_C, Q_DThe semiconductor wafer S is inspected while focusing on.
[0067]
First, the observer must observe the observation position Q._AThen, the AF start switch of the operation unit 6 is pressed to perform the first AF operation. Here, the observation position Q_AIf the AF operation is successful, the above-described manual adjustment of the focus position and the switch operation at the time of storing the focus position are not necessary, and after the AF operation is completed, the position storage unit 38 of the control unit 5 stores the current Z stage 13. Z position of the first observation position Q_AFocus position (Z_A) And this focus position (Z_A) Observation position Q_AXY coordinates (X_A, Y_A), That is, the focus position (X_A, Y_A, Z_A) Is stored in the RAM 11.
[0068]
On the other hand, if the AF operation fails due to a failure in focusing normally, the control unit 5 displays AF failure on a display unit (not shown) to notify the observer, and prompts manual focusing. The observer moves the Z stage 13 in the Z direction by using the encoder 36 or a manual focusing mechanism (not shown), moves the sample S to the focus position, and then presses the focus setting switch of the operation unit 6. In response to the pressing operation of the focus setting switch, the position storage unit 38 of the control unit 5 automatically detects the observation position Q._AFocus position (X_A, Y_A, Z_A) Is stored in the RAM 11.
[0069]
Next, the observer moves the XY stage by using the XY movement switch of the operation unit 6 to observe the observation position Q._BIs moved into the field of view, and the AF operation is started. When this AF operation is successful, the position storage unit 38 of the control unit 5 automatically detects the observation position Q as in the first time._BFocus position (X_B, Y_B, Z_B) Is stored in the RAM 11.
[0070]
Further, the sample inclination calculation unit 40 of the control unit 5 is currently (observation position Q_BZ position of the Z stage 13, that is, the focus position Z_BAnd observation position Q_ACoordinates (X_A, Y_A) Focus position Z_ADifference ΔZ from_AB
ΔZ_AB= (Z_B-Z_A(3)
And the difference ΔZ_ABObservation position Q_ATo Q_BIs stored in the RAM 11 as a change in the focus position accompanying the movement to.
[0071]
On the other hand, when the AF operation is unsuccessful, the position storage unit 38 of the control unit 5 determines that the focus position Z_BAnd focus position Z_ADifference ΔZ from_ABThe focus position Z when the AF operation or manual focus is performed for the first time without calculating_AIs read from the RAM 11, and the focus position Z_AThe Z stage 13 is moved in the Z direction.
[0072]
Next, the observer moves the XY stage by using the XY movement switch of the operation unit 6 to observe the observation position Q._CIs moved into the field of view, and the AF operation is started. When this AF operation is successful, the position storage unit 38 of the control unit 5 automatically performs the observation position Q as described above._CFocus position (X_C, Y_C, Z_C) Is stored in the RAM 11.
[0073]
Further, the sample inclination calculation unit 40 of the control unit 5 is currently (observation position Q_CZ position of the Z stage 13, that is, the focus position Z_CAnd observation position Q_BCoordinates (X_B, Y_B) Focus position Z_BDifference ΔZ from_BC
ΔZ_BC= (Z_C-Z_B(4)
And the difference ΔZ_BCObservation position Q_BTo Q_CIs stored in the RAM 11 as a change in the focus position accompanying the movement to.
[0074]
On the other hand, when the AF operation is unsuccessful, the position storage unit 38 of the control unit 5 determines that the focus position Z_BAnd focus position Z_CDifference ΔZ from_BCWithout calculating the current position (observation position Q_C) The AF operation success position closest to the position, for example, the observation position Q_BIf the AF operation is successful, the observation position Q_BIf it fails, the observation position Q_AFocus position Z_BOr Z_AThe Z stage 13 is moved.
[0075]
By the way, in each of the operations so far, each observation position Q_B, Q_CIf the AF operation is successful, each observation position Q stored in the RAM 11 is_B, Q_CThe inclination when the sample S is a plane can be calculated from the XY coordinates and the shift of the focus position.
[0076]
That is, if the inclination in the X-axis direction is α and the inclination in the Y-axis direction is β,
α ・ ΔX_AB+ Β · ΔY_AB= ΔZ_AB                 ... (5)
α ・ ΔX_BC+ Β · ΔY_BC= ΔZ_BC                 ... (6)
ΔX in the two equations_AB, ΔY_AB, ΔZ_AB, ΔX_BC, ΔY_BC, ΔZ_BCEach difference ΔX stored in the RAM 11_AB, ΔY_AB, ΔZ_AB, ΔX_BC, ΔY_BC, ΔZ_BCBy substituting this value, the inclination α in the X-axis direction and the inclination β in the Y-axis direction are obtained.
[0077]
These differences ΔX_AB, ΔY_AB, ΔX_BC, ΔY_BCAs shown in FIG._AAnd Q_B, Q_BAnd Q_C, The movement amount of the XY stage 14 in the X-axis direction and the Y-axis direction, and the observation position Q by the sample inclination calculation unit 40 of the control unit 5_ATo Q_B, Q_BTo Q_CAnd is stored in the RAM 11 for each movement.
[0078]
Therefore, the focus position correction unit 41 calculates the above equations (5) and (6) to calculate the inclination of the plane in the current sample S, that is, the inclination in the X-axis direction α and the inclination in the Y-axis direction β. As a result, the focus position shift ΔZ accompanying the movement (ΔX, ΔY) in any XY axis direction can be reduced.
ΔZ = α · ΔX + β · ΔY (7)
It is possible to approximate and predict by the following equation.
[0079]
Next, the observer moves the XY stage by using the XY movement switch of the operation unit 6 to observe the observation position Q._DIs moved into the field of view, and the AF operation is started. When this AF operation is successful, the position storage unit 38 of the control unit 5 automatically performs the observation position Q as described above._DFocus position (X_D, Y_D, Z_D) In the RAM 11 and the current (observation position Q)_DZ position Z of Z stage 13)_DAnd observation position Q_CCoordinates (X_C, Y_C) Focus position Z_CDifference ΔZ from_CD(= Z_D-Z_C) And the observation position Q_CTo Q_DIs stored in the RAM 11 as a change in the focus position accompanying the movement to.
[0080]
On the other hand, when the AF operation fails, the focus position correction unit 41 of the control unit 5 moves the movement amount ΔX of the XY plane._CD, ΔY_CDTo calculate the above equation (7)
ΔZ_CD= Α ・ ΔX_CD+ Β · ΔY_CD                 (8)
Observation position Q_CAnd Q_DIn focus position ΔZ between_CDThis deviation ΔZ_CDAccordingly, a control signal for moving the Z stage 13 in the Z direction is sent to the focusing motor drive unit 17. As a result, the Z stage 13 moves ΔZ in the Z direction._CDJust move.
[0081]
As described above, in the second embodiment, the sample inclination calculation unit 40 calculates the inclination of the sample S surface based on the XY coordinates and the focus position Z of at least two autofocus positions stored in the RAM 11. When the AF operation is not performed on the sample S, the focus position is determined based on the XY coordinates of the XY stage 14 that is not AF-operated by the focus position correction unit 41 and the inclination of the sample S surface calculated by the sample inclination calculation unit 40. Since the Z stage 13 is moved in accordance with this focus position, an arbitrary observation position Q_N(X_N, Y_N), The amount of focus position correction when the AF operation fails can be set to a value that takes into account the inclination of the sample S, whereby the Z stage when the AF operation fails on the sample S surface having a uniform gradient. The Z position of 13 can be moved to a position closest to the focus position without being affected by the inclination of the sample S surface.
[0082]
Further, as in the first embodiment, a plurality of observation positions Q on the same specimen S are used._A, Q_B, ..., Q_NAs the number of successful AF operations increases, the inclination of the sample S can be predicted more accurately.
[0083]
Next, a third embodiment of the present invention will be described. The same parts as those in FIG. 1 are denoted by the same reference numerals, and detailed description thereof is omitted.
[0084]
The microscope system of the present invention is different from the second embodiment in that the AF operation mode and the focus position correction unit 42 are provided in the control unit 5 as shown in the functional block diagram of FIG. Different.
[0085]
The mode of the AF operation will be described. In the second embodiment, the observation position Q is_N(X_N, Y_N) And focus position Z_NAcquisition and storage of the observation position Q_N(X_N, Y_N) After the AF operation is started and the success / failure of the AF operation is determined. That is, each observation position Q_N(X_N, Y_N), An AF operation is started, and each observation position Q is different._N(X_N, Y_N) Is premised on that no AF operation is performed. This form of AF operation is called so-called one-shot AF, and each observation position Q_N(X_N, Y_N) AF operation performed only once.
[0086]
In contrast, each observation position Q_N(X_N, Y_NThe AF operation that performs the AF operation even when moving to () and always follows the focus position is referred to as real-time AF, and is frequently used to obtain a focused image continuously.
[0087]
In the case of real-time AF, the observation position Q of the specimen S as in the one-shot AF described in the second embodiment._N(X_N, Y_N) And focus position Z_NThe AF start command, which is the timing for acquiring the_AOnly after each observation position Q_N(X_N, Y_N) Is not emitted.
[0088]
In the real-time AF operation, when the focus position with respect to the specimen S cannot be obtained (AF failure), the reflectance of the specimen S is mainly low. In this case, the focus position tracking operation is not performed, and the Z stage 13 remains stopped in the Z direction, and waits until the reflectance of the sample S exceeds the predetermined value and the focus position can be tracked again. It is normal to become.
[0089]
For example, as shown in FIG. 12, when real-time AF is started from the observation position a on the surface of the specimen S and the observation position is moved along the observation locus K by the movement of the XY stage 14 in the XY direction, for example, reflection If the focus position cannot be detected within the low-rate range G and the focus position cannot be tracked, the Z stage 13 stops moving from the position b in contact with the range G in the Z direction and exits from the range G. The focus position tracking operation is resumed from c. When the Z stage 13 moves within the range G, the Z position remains at the Z position at the position b.
[0090]
Therefore, if the sample S has a gradient, the image of the sample S gradually blurs within the range G where real-time AF cannot be performed.
[0091]
For this reason, if the focus position cannot be tracked with respect to the sample S, the focus position correcting unit 42 can tilt the surface of the sample S calculated by the sample tilt calculating unit 40 within the range G where the focus position cannot be tracked. The focus position is predicted on the basis of the above, and the Z stage 13 is moved in the Z direction according to the focus position.
[0092]
By providing the focus position correction unit 42, the Z position of the Z stage 13 that is closest to the focus position is calculated within the range G where real-time AF cannot be performed, and as the observation position moves with respect to the specimen S, the Z position is moved as needed. By correcting the Z position, it is possible to acquire a sample image with the focus position being suppressed as much as possible until the position c at which the real-time AF operation can be performed again.
[0093]
Next, the AF operation of the apparatus configured as described above will be described.
[0094]
The focus position correction unit 42 divides into a plurality of grid-like areas according to the lengths ΔX and ΔY (for example, about 1 mm) with respect to the XY axes as indicated by broken lines in FIG. The lengths ΔX and ΔY can be set to arbitrary values by the operation unit 6.
[0095]
When the observer starts the real-time AF operation from the position a on the sample S and moves the observation position along the observation locus K, the movement amount in the X direction becomes ΔX or more, or the movement amount in the Y direction. When the value becomes ΔY or more, the focus position correction unit 42 acquires the coordinates (X, Y) and the focus position Z of the current observation position, and stores these coordinates (X, Y) and the focus position Z in the RAM 11. Remember. For example, on the specimen S shown in FIG.₁~ Q₇Each coordinate (X₁, Y₁) To (X₇, Y₇) And focus position Z₁~ Z₇Is stored in the RAM 11.
[0096]
Similar to the second embodiment, the sample inclination calculation unit 40 has at least two AF positions stored in the RAM 11, for example, each observation position Q in FIG.₁~ Q₇XY coordinates (X₁, Y₁) To (X₇, Y₇) And focus position Z₁~ Z₇Based on the above, the inclination of the sample S surface is calculated.
[0097]
Thus, until the observation position passes through the observation position b in FIG. 13 and enters the range G where the real-time AF operation cannot be performed, the correction formula for the focus position Z accompanying the movement of the sample S in the XY direction, that is, the above-described formula The focus position shift ΔZ is obtained by calculating Expression (7).
[0098]
Accordingly, the focus position correction unit 42 calculates the above expression (7) from the movement amounts ΔX and ΔY of the XY plane to obtain the focus position shift ΔZ on the observation locus K of the observation position, and the Z stage 13 according to the shift ΔZ. Is sent to the focusing motor drive unit 17 in the Z direction. Thereby, the Z stage 13 moves at any time in the Z direction by ΔZ.
[0099]
As a result, the focus position can be tracked even in the range G where the real-time AF operation cannot be performed.
[0100]
As described above, in the third embodiment, when the real-time AF operation is performed, if the focus position cannot be tracked with respect to the sample S, the inclination of the surface of the sample S is within the range G where the focus position cannot be tracked. The focus position is predicted based on the Z position, and the Z stage 13 is moved in the Z direction in accordance with the focus position. 13 can be controlled in the Z direction and can automatically switch to the normal real-time AF operation at the position c where the real-time AF operation is possible again, and cannot be inspected due to out-of-focus, and manual focus adjustment, AF operation You can avoid unnecessary operations such as resuming the image, and always obtain a focused sample image. .
[0101]
The present invention is not limited to the first to third embodiments described above, and various modifications can be made without departing from the scope of the invention in the implementation stage.
[0102]
Furthermore, the above embodiments include inventions at various stages, and various inventions can be extracted by appropriately combining a plurality of disclosed constituent requirements. For example, even if some constituent elements are deleted from all the constituent elements shown in the embodiment, the problem described in the column of the problem to be solved by the invention can be solved, and is described in the column of the effect of the invention. If the effect is obtained, a configuration from which this configuration requirement is deleted can be extracted as an invention.
[0103]
For example, in the first to third embodiments, the distance between the specimen S and the objective lens 2 is the Z stage 13 being moved. However, the present invention is not limited to this. For example, the revolver 1 to which the objective lens 2 is attached is Z. The method may be changed to a method in which the Z stage 13 and the revolver 1 are moved in the Z direction.
[0104]
In the first to third embodiments, the so-called active AF in which the sample S is irradiated with laser light and the AF operation is performed based on the state of the reflected light has been described. The present invention can also be applied to so-called passive AF that detects with a line sensor or the like and performs an AF operation based on the contrast value of image data acquired thereby, and has the same effect as the first to third embodiments. An effect can be obtained.
[0105]
Further, whether or not the AF operation is possible, for example, switching of the AF or Z position correction operation at the positions b and c in FIG. 13 is performed by the laser light incident on the light receiving sensor 31 as shown in FIGS. For example, in the case of a passive AF such as a contrast method, the contrast of the image of the sample S detected by a CCD line sensor or the like is realized by providing a predetermined threshold value for the intensity (A + B). If the total value is used, it is possible to determine whether or not AF is possible as in the active AF.
[0106]
A microscope system having an AF function can be applied to various types of microscopes such as an epi-illumination type, a transmission type microscope, and a laser scanning type.
[0107]
Further, in the first to third embodiments, the case where the sample S has an inclination has been described. However, one or both of the Z stage 13 and the XY stage 14 have an inclination, and the sample S is inclined. Even in this case, it is possible to obtain a specimen image stably while suppressing the focus shift with respect to the specimen S as much as possible. In this case, since the inclination of the Z stage 13 or the XY stage 14 is always constant, a plurality of focus positions (X_A, Y_A, Z_A) May be stored in the RAM 11 to form a database, and the focus position deviation may be corrected using this database.
[0108]
【The invention's effect】
As described above in detail, according to the present invention, it is possible to provide a microscope system that can stably obtain an image of a specimen by suppressing focus deviation with respect to the specimen as much as possible even when AF does not operate in the specimen. .
[Brief description of the drawings]
FIG. 1 is a configuration diagram showing a first embodiment of a microscope system according to the present invention.
FIG. 2 is a configuration diagram of a control unit in the first embodiment of the microscope system according to the present invention.
FIG. 3 shows a microscope system according to the first embodiment of the present invention.focusThe figure which shows the light intensity distribution on the light reception sensor in the case of a position.
FIG. 4 is a diagram showing a light intensity distribution on a light receiving sensor in the case of a front pin position in the first embodiment of the microscope system according to the present invention.
FIG. 5 is a diagram showing a light intensity distribution on a light receiving sensor in the case of a rear pin position in the first embodiment of the microscope system according to the present invention.
FIG. 6 is a diagram illustrating the operation of the microscope system according to the first embodiment of the present invention.focusThe figure for demonstrating operation | movement.
FIG. 7 is a block diagram showing functions of a controller in the first embodiment of the microscope system according to the present invention.
FIG. 8 is a diagram for explaining an AF operation of the first embodiment of the microscope system according to the present invention.
FIG. 9 is a block diagram showing functions of a controller in the second embodiment of the microscope system according to the present invention.
FIG. 10 is a diagram for explaining an AF operation of the second embodiment of the microscope system according to the present invention.
FIG. 11 is a block diagram showing functions of a controller in a third embodiment of a microscope system according to the present invention.
FIG. 12 is a diagram for explaining a normal real-time AF tracking operation;
FIG. 13 is a diagram for explaining an AF operation of the third embodiment of the microscope system according to the present invention.
[Explanation of symbols]
1: Revolver body
2: Objective lens
3: Revolver motor
4: Rebo hole position detector
5: Control section
6: Operation unit
7: Revolver motor drive
8: CPU body
9: Data bus
10: ROM
11: RAM
12: I / O port
13: Z stage
14: XY stage
15: Focusing motor
16: Motor for XY stage
17: Focusing motor drive
18: XY stage motor drive
19: Reference light source
20: Laser drive unit
21: Collimator lens
22: Light emission side stopper
23: Polarizing beam splitter (PBS)
24: Condensing lens group
25: Chromatic aberration lens group
26: Chromatic aberration lens driving unit
27: Motor for driving chromatic aberration lens
28: λ / 4 plate
29: Dichroic mirror
30: Condensing lens group
31: Light receiving sensor
32: Signal processor
33: Light source for illumination
34: Lens
35: Half mirror
36: Encoder
37: Pulse counter
38: Position storage unit
39: Focus position correction unit
40: Sample inclination calculation unit
41, 42: Focus position correction unit
S: Specimen

Claims (4)

The stage on which the specimen is placed can be moved in a plane direction perpendicular to the optical axis of the objective lens, and the distance between the stage and the objective lens is relatively adjusted to perform autofocus on the specimen. In the microscope system,
Position storage means for storing position data indicating coordinates in a plane direction of the stage when the specimen is autofocused, and focus data of a distance between the specimen and the objective lens;
When the autofocus is not performed on the sample, the position data of the coordinates closest to the coordinates that are not autofocused is retrieved from the position storage means, and the focus data corresponding to the retrieved position data is used. a focus position correction means for changing the relative distance between the stage and the objective lens Te,
A microscope system comprising: The stage on which the specimen is placed can be moved in a plane direction perpendicular to the optical axis of the objective lens, and the distance between the stage and the objective lens is relatively adjusted to automatically focus the specimen. In the microscope system that performs
Position data indicating coordinates in the plane direction of the stage when the autofocus is performed on the specimen;
Position storage means for storing focus data of a distance between the specimen and the objective lens;
Sample inclination calculating means for calculating the inclination of the sample surface based on the position data and the focus data at at least two coordinates stored in the position storage means;
When the autofocus is not performed on the sample, a focus position is predicted based on the position data at the coordinates where the autofocus is not performed and the tilt of the sample calculated by the sample tilt calculating unit. A focus position correcting means for changing a relative distance between the stage and the objective lens based on a position;
A microscope system characterized by comprising: 3. The microscope system according to claim 1, further comprising: an autofocus execution enable / disable determining unit configured to determine whether or not the autofocus at each coordinate is possible for the sample. The stage on which the specimen is placed can be moved in a plane direction perpendicular to the optical axis of the objective lens, and the distance between the stage and the objective lens is relatively adjusted to always move the stage relative to the specimen. In a microscope system that performs real-time autofocus that follows the focus position
Each time the amount of movement of the stage in the plane direction reaches a predetermined amount of movement, position data of the stage in the plane direction and focus data of the distance between the specimen and the objective lens are obtained. Position storage means for storing;
Sample inclination calculation means for calculating the inclination of the sample surface based on the position data and the focus data of at least two autofocus positions stored in the position storage means;
When the focus position cannot be tracked with respect to the sample, a focus position is predicted based on the tilt of the sample surface calculated by the sample tilt calculation means within a range where the focus position cannot be tracked, and the focus position is determined according to the focus position. Focus position correcting means for relatively moving a stage and the objective lens;
A microscope system comprising:

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