CN110799879A

CN110799879A - Microscope and method for microscopic examination of a sample under variable mechanical parameters

Info

Publication number: CN110799879A
Application number: CN201880043531.5A
Authority: CN
Inventors: 亚历山大·盖杜克; 德特勒夫·海因; 帕夫洛斯·伊利奥普洛斯
Original assignee: Carl Zeiss Microscopy GmbH
Current assignee: Carl Zeiss Microscopy GmbH
Priority date: 2017-06-29
Filing date: 2018-05-11
Publication date: 2020-02-14
Also published as: DE102017114562A1; WO2019001821A1

Abstract

The invention relates to a microscope and to a method for microscopically examining a specimen, wherein a mechanical parameter (theta) is variable in the microscope. In one step of the method, a first value (theta) of the mechanical parameter (theta) to be set is selected_Target). An electrically controllable actuating element (04) is actuated in order to set a first value (theta) of a mechanical variable (theta) to be set_Target). Adjusted first value (theta) of the mechanical parameter (theta)_Target) In this case, a first microscopic image of the specimen is taken. In another step, mechanical parameters are selectedA second value (theta) of (theta) to be adjusted_Target). Actuating an electrically controllable actuating element (04) in order to set a second value (theta) of the mechanical variable (theta) to be set_Target). The second value (theta) adjusted for the mechanical parameter (theta)_Target) In this case, a second microscopic image of the specimen is taken.

Description

Microscope and method for microscopic examination of a sample under variable mechanical parameters

Technical Field

The invention relates to a microscope and to a method for microscopically examining a specimen, wherein mechanical parameters of the microscope are variable. The mechanical parameter is preferably formed by an inclination angle which is variable in at least one spatial direction between the objective of the microscope and the stage of the microscope. Such an inclination angle indicates the inclination between the objective lens and the stage, also called tilt.

Background

JP 2001059599 a2 and JP 2010102344 a2 show a pendulum arm for a digital microscope. The swing arm frame comprises a swing arm which can swing around a horizontal rotation axis.

DE 102013005999 a1 and DE 102013222295 a1 disclose a pendulum arm support for a digital microscope, in which the pendulum arm movement can be blocked by a high-torque electromagnetic brake arranged about the axis of rotation. The jam can be released by releasing the high torque electromagnetic brake for a continuous time while the key is depressed. The key is arranged on the swing arm so that it can be easily depressed with at least one finger of the same hand.

US 6,642,686B 1 shows a controllable arm with which, in particular, medical operations can be supported. The rotatable arm may for example carry a microscope as instrument.

Disclosure of Invention

Starting from the prior art, the object of the invention is to expand the scope of application of microscopes in which mechanical parameters, such as the tilt angle between the objective and the stage, are variable.

The solution of the invention to the above object is a method according to claim 1 and a microscope according to the accompanying claim 15.

The method according to the invention is used for the microscopic examination of samples with a microscope, in particular with a digital microscope. The electronic image conversion is preferably carried out in a digital microscope, wherein the captured image is further processed into the form of digital data and displayed on an electronic image reproduction device. The microscope comprises preferably at least one objective and preferably an image sensor for converting an image directly or indirectly imaged from the objective onto the image sensor.

The mechanical parameters of the microscope are variable. The mechanical parameter is preferably formed by the position, angle or distance with respect to the sample, with respect to the stage, with respect to the image sensor, with respect to the objective assembly, with respect to the filter and/or with respect to the microscope illumination light source. In this way, mechanical parameters can be formed, for example, by the height and/or lateral position of the stage, the distance from the image sensor or zoom adjustment. Alternatively, the mechanical parameter is preferably formed by the position, angle or distance of an adjusting element for adjusting a physical variable of the microscope. It may be, for example, a rotatable adjusting element, with which, for example, the wavelength, the brightness at the microscope or the polarization or the phase in the illumination or detection channel of the microscope may be adjusted.

The mechanical parameter is preferably formed by an inclination angle which is variable in at least one spatial direction between the objective of the microscope and the stage of the microscope. In a microscope, the tilt angle between the objective lens and the specimen-carrying stage of the microscope is variable. The stage is used to arrange the sample. The tilt angle is defined between the optical axis of the objective lens and a vertical line on the stage. The tilt angle indicates the tilt between the objective lens and the stage, also called tilt. The tilt axis is formed by a tilted rotation axis oriented perpendicular to the optical axis. The tilt angle is defined in at least one spatial direction, but may also be defined by two values in a spherical coordinate system, for example.

The method comprises the step of selecting a first value of the mechanical parameter to be adjusted. The first value of the mechanical parameter to be set defines the quantity of the mechanical parameter which is to be set. In a further step, an electrically controllable electromechanical actuating element is actuated in order to set a first value of the mechanical variable to be set. The actuating element is used to set the mechanical parameter to a first value to be set. In a further step, a first microscopic image of the sample is taken with the aid of the microscope while adjusting the first value of the mechanical parameter.

In a further step, a second value of the mechanical parameter to be set is selected. The second value is different from the first value. The second value of the mechanical parameter to be set defines the amount of the angle of inclination which should be set. In a further step, an electrically controllable electromechanical actuating element is actuated in order to set a second value of the mechanical variable to be set. In a further step, a second microscopic image of the sample is taken with the aid of the microscope while adjusting the second value of the mechanical parameter.

A particular advantage of the method according to the invention is that microscopic images are taken at defined values of the mechanical parameter, for example at defined values of the inclination angle, respectively, in order to be able to microscopically examine the sample further on this basis. The method according to the invention allows, for example, an improved further examination of low-contrast samples. By the method according to the invention, the operator is supported when adjusting the defined values of the mechanical parameter, so that the effort for checking the sample at different values of the mechanical parameter is greatly reduced and is not prone to errors.

A preferred embodiment of the method according to the invention further comprises the step of executing after the activation of the electrically controllable actuating element, respectively. In this step, a focused sample is realized; in particular the sample is focused again, since by changing the mechanical parameters, for example by changing the tilt angle or by changing the height of the stage, it is no longer necessary to focus the sample through the objective. Here, alternatively or additionally, the specimen in the image to be recorded is centered again, for which purpose the stage is displaced in its height and/or laterally.

In a first preferred embodiment, the actuating element is formed by an actuator which can be driven by an electric motor and by means of which the mechanical parameters can be set and adjusted. By energizing the electric motor, the mechanical parameters can be adjusted.

In a second preferred embodiment, the mechanical parameters can be changed manually. The operator of the microscope adjusts the mechanical parameters with his strength, for which purpose he preferably has to first actuate the unlocking device. The actuating element is formed by an electrically controllable setting device with which the mechanical parameters can be set. The setting device is preferably formed by an electrically controllable mechanical brake or a high-torque electromagnetic brake. In this second preferred embodiment, the step of activating the electrically controllable actuating element comprises in each case the step of measuring a time profile in which the value of the mechanical parameter is changed manually. For this purpose, an angle sensor or a distance sensor is preferably used. In a further sub-step, a previously determined time profile predicting the manual change of the value of the mechanical parameter will result in a point in time of the respective value of the mechanical parameter to be adjusted. Accordingly, the electrically actuable setting device is actuated at a previously determined point in time in order to ensure that the respectively to be set value of the mechanical parameter, for example the respectively to be set value of the tilt angle, is actually achieved and, for example, the dependency of the accuracy on the angle size or on the distance between the objective and the sample is eliminated. By actuating the setting device, which can be electrically actuated, the mechanical parameter is fixed so that it can no longer be changed manually.

In the embodiment in which the mechanical parameter is formed by the tilt angle, a previously determined time curve predicting a manual change of the value of the tilt angle will result in a value Θ of the tilt angle, which is to be adjusted in each case_TargetThe step of time point of (a) preferably comprises a plurality of sub-steps. In a sub-step, the initial value Θ of the tilt angle is measured_InitialIn a further step, the angular velocity ω and the angular acceleration α are determined from a previously determined time profile of the manual change of the value of the inclination angle_Target＝Θ_Initial+ω·t+1/2·α·t²Predicting a manual change of the value of the tilt angle by means of tWill result in the time points of the respective values of the tilt angle to be adjusted.

The previously determined time curve predicting the manual change of the value of the mechanical parameter is preferably repeated continuously at the point in time at which the respective to-be-adjusted value of the mechanical parameter would result until the value of the mechanical parameter has been manually changed to the respective to-be-adjusted value of the mechanical parameter. By such successive repetition, highly accurate prediction is realized. Accordingly, in the embodiment in which the mechanical parameter is formed by the inclination angle, a previously determined time profile, which preferably continuously predicts repeated manual changes to the value of the inclination angle, will result in the respective value Θ of the inclination angle to be adjusted_TargetUntil the value of the tilt angle has been manually changed to the value theta of the tilt angle, respectively to be adjusted_Target。

In the embodiment in which the mechanical parameter is formed by the inclination angle, a time profile for the purpose of manually changing the value of the inclination angle determined in advance will result in the value Θ of the inclination angle to be respectively adjusted_TargetPreferably also taking into account the mass distribution on the microscope and/or the force exerted by manually changing the value of the tilt angle. In this way, it can be precisely predicted that a previously determined time profile of a manual change of the value of the tilt angle will result in a value Θ of the tilt angle which is to be set in each case_TargetThe time point of (a).

In embodiments in which the mechanical parameter is formed by the tilt angle, the objective is preferably fastened to the swing arm of the microscope. The rocker arm may be a component of a rocker arm stand. The swing arm may also be referred to as a tilt arm. The arm is pivotable, whereby the angle of inclination between the objective lens and the stage is variable.

Alternatively or additionally, the object table is preferably rotatable. The object table is preferably rotatable about an axis arranged parallel to the plane of extension of the object table or particularly preferably lies in the plane of extension of the object table. By rotating the objective lens about this axis, the tilt angle between the objective lens and the stage is variable. The object table is also rotatable about an axis oriented perpendicular to the extension plane of the object table. Rotation about this axis results in a change in the tilt angle as the sample on the carriage also extends vertically.

In a particularly preferred embodiment, the method comprises a further step, in which a feedback is generated to the operator if the value of the mechanical parameter has changed by a predefined amount and/or a value of the mechanical parameter has been adjusted to be respectively adjusted. In this way, the operator can quickly check whether the respective values of the mechanical parameters to be set have been reached, thus facilitating the operation of the microscope. Furthermore, if the value of the mechanical parameter has changed by a predefined amount; for example, if the value of the tilt angle has changed by, for example, 5 °, the operator is always able to obtain feedback. The feedback is preferably visually, audibly, and/or tactilely perceptible.

The feedback to the operator is particularly preferably perceptible in the sense of touch and is preferably generated using an electrically actuable setting device. For this purpose, in the embodiment in which the mechanical parameter is formed by the tilt angle, the change in the value of the tilt angle, i.e. in particular the swiveling movement of the swing arm or the rotation of the rotatable stage, is preferably interrupted briefly as it changes over time, which the operator can perceive so that the operator realizes that the value of the tilt angle has changed by a predefined amount. The interruption of the value of the inclination angle over time is generated in particular by an electrically actuable setting device in the form of a brake interrupting the swiveling movement of the swiveling arm or the rotation of the rotatable object table in a very short continuous time.

Alternatively or additionally, the feedback to the operator is acoustically perceptible and is preferably generated using a buzzer or similar sound signal transmitter. By means of the acoustic signal, the operator knows that the value of the machine parameter has changed by a predefined amount or that a value of the machine parameter to be set in each case has been set.

In a preferred embodiment of the method according to the invention, the step of selecting the respective values of the mechanical parameter to be set is carried out manually by an operator. The operator specifies two or more values to be set for the machine parameter, which he preferably enters via the user interface.

In a further preferred embodiment of the method according to the invention, the respective values of the mechanical parameter to be set are selected from a plurality of stored values. Preferably, during the execution of the method of the invention, the stored values are chosen for a sample of a previous microscopic examination and subsequently stored. In this way, the same values of the mechanical parameters are selected for the two samples to be microscopically examined and corresponding microscopic images are acquired, whereby an extended evaluation can be carried out, for example, by comparison. The stored values preferably represent pre-programmed or pre-adjusted values accessed during the execution of the method.

In a further preferred embodiment of the method according to the invention, the selection of the respectively to be adjusted value of the mechanical parameter is effected by incrementing the previously selected value of the mechanical parameter by an increment. Thus, in embodiments where the mechanical parameter is formed by the tilt angle, the sample is subjected to scanning microscopy at the tilt angle. The first value of the mechanical parameter represents a starting value.

It is particularly preferred to select and adjust not only two values of the mechanical parameter, but also a plurality of values of the mechanical parameter. Accordingly, the method preferably comprises a step in which a further value of the mechanical parameter to be set, which differs from the other values of the mechanical parameter, is selected. In a further step, an electrically controllable actuating element is actuated in order to set a further value of the mechanical variable to be set. In a further step, a further microscopic image of the sample is taken with a further value of the mechanical parameter being adjusted. The steps are preferably repeated a plurality of times for other values of the mechanical parameter.

The method according to the invention is preferably configured for microscopic examination of at least two samples. In this case, the same first value of the mechanical parameter, the same second value of the mechanical parameter and, if necessary, the same further values of the mechanical parameter are respectively selected and adjusted for taking a microscopic image of the first specimen and for taking a microscopic image of the second specimen. In this way, microscopic images are taken at the same value of the mechanical parameter for at least two samples.

In a further preferred embodiment of the method according to the invention, different magnifications of the objective and/or different objectives are selected for the acquisition of the first and second and, if appropriate, further microscopic images of the specimen. This can match the spatial extent of the sample or allow microscopic examination of a particular region of the sample at higher magnification.

A preferred embodiment of the method according to the invention comprises the further step in which a 2.5-dimensional reconstruction or a 3-dimensional reconstruction of the sample is generated from the plurality of microscopic images. Preferably, a plurality of values of the mechanical parameter, preferably formed by the tilt angle, are selected and adjusted in order to take a corresponding plurality of microscopic images and to generate a 2.5-dimensional or 3-dimensional reconstruction of the sample. Depth information can also be obtained from the microscopic images, since the sample is microscopically examined at different values of the tilt angle or the sample distance.

When taking microscopic images of the sample at various different tilt angles, one or more areas of the sample can be covered. Therefore, it is preferred to take further microscopic images of the sample at further adjusted tilt angles in order to take those sample regions that cover further tilt angles.

The microscope according to the invention is used for microscopic examination of a sample. It preferably comprises an objective lens for optically imaging the sample. The microscope also preferably includes a stage for arranging the sample. The mechanical parameters of the microscope are variable. Preferably, as a mechanical parameter, the tilt angle between the objective lens and the stage is variable. The microscope further comprises an electrically controllable manipulation element for adjusting the mechanical parameter value and a control unit configured for implementing the method according to the invention. The control unit is preferably configured for implementing preferred embodiments of the method according to the invention. The microscope preferably also has the features described in connection with the method according to the invention.

Drawings

Further details and refinements of the invention are derived from the following description of preferred embodiments of the invention with reference to the drawing. In the figure:

fig. 1 shows a first preferred embodiment of a microscope according to the invention;

fig. 2 shows a second preferred embodiment of a microscope according to the invention;

FIG. 3 shows two states of the microscope shown in FIG. 1 during the execution of the method according to the invention; and

fig. 4 shows two states of the microscope shown in fig. 2 during the execution of the method according to the invention.

Detailed Description

Fig. 1 shows a first preferred embodiment of a microscope according to the invention. The microscope comprises a base 01 on which a stage 02 is fastened. The swing arm 03 is rotatably supported on the base 01, and can be swung with respect to the base 01 by means of an electric actuator 04 so that it is also tilted with respect to the stage 02 and the specimen 06 located thereon (see fig. 2). An objective lens 07 and an optical module 08 are fastened to the swing arm 03. The swing of the swing arm 03 can be unlocked by the unlocking key 09.

As an alternative to the electric actuator 04, an electrically actuable brake 11 (see fig. 2) can be used on the swing arm 03, which brake can block the swing of the swing arm 03 relative to the base 01 and the object table 02 by braking. The operator must then apply a torque to swing the swing arm 03. The electrically actuable brake 11 (shown in fig. 2) of the pivot arm 03 is actuated such that, once the desired pivot angle value is reached, the pivot arm 03 cannot be pivoted further manually.

Fig. 2 shows a second preferred embodiment of a microscope according to the invention, which, like the embodiment shown in fig. 1, comprises a stage 02, an objective lens 07 and an optical module 08. In contrast to the embodiment shown in fig. 1, the objective lens 07 and the optical module 08 are not fastened to a pivotable pivot arm, but rather are arranged fixedly. In other words, the stage 02 is rotatable. The rotational movement of the object carrier 02 can be braked by the electrically actuable brake 11, so that the rotatable object carrier 02 can be fixed. The embodiment shown in fig. 2 comprises a further objective lens 12 and a further optical module 13 arranged below the object table 02.

Fig. 3 shows two states of the microscope shown in fig. 1 during the execution of the method according to the invention. The left part of fig. 3 shows the microscope in an initial state, in which the swing arm 03 is not tilted, so that the tilt angle Θ is equal to zero. According to the method, a value theta of the tilt angle theta to be adjusted is selected_Target。

FIG. 3The right-hand part of (a) shows the microscope at the value to be adjusted theta for adjusting the tilt angle theta according to the method_TargetThe adjustment can be effected by actuating the unlocking key 09. The pivot arm 03 has been pivoted by driving the electrodynamic actuator 04 to the value Θ of the angle of inclination Θ_{At present}. Continuing to drive the electrodynamic actuator 04 until the value Θ of the inclination angle Θ to be set is reached_Target。

Fig. 4 shows two states of the microscope shown in fig. 2 during the execution of the method according to the invention. The left part of fig. 4 shows the microscope in an initial state, in which the stage 02 is not tilted, so that the tilt angle Θ is equal to zero. According to the method, a value theta of the tilt angle theta to be adjusted is selected_Target。

The right-hand part of fig. 3 shows the microscope at the value Θ to be adjusted for adjusting the tilt angle Θ according to this method_TargetThe status of the period. The object table 02 is rotated manually, wherein the object table 02 has been rotated to a value Θ of the inclination angle Θ_{At present}. According to this method, the rotational movement of the object table 02 is measured and the value Θ to be set up which reaches the inclination angle Θ is predicted_TargetThe time point of (a). At this point in time, the electrically actuated brake 11 is actuated, so that no further rotational movement of the object table 02 is possible.

List of reference numerals

01 base

02 objective table

03 swing arm

04 electric actuator

05 -

06 sample

07 Objective lens

08 optical module

09 unlocking key

10 -

11 electrically operable brake

12 additional objective lens

13 additional optical module

Claims

1. Method for microscopic examination of a sample (06) with a microscope, wherein on the microscope a mechanical parameter (Θ) is variable, and wherein the method comprises at least the following steps:

-selecting a first value (Θ) of said mechanical parameter (Θ) to be adjusted_Target)；

-actuating an electrically controllable actuating element (04; 11) in order to set a first value (Θ) of the mechanical parameter (Θ) to be set_Target)；

-an adjusted first value (Θ) of said mechanical parameter (Θ)_Target) Taking a first microscopic image of the sample (06);

-selecting a second value (Θ) of the mechanical parameter (Θ) to be adjusted_Target)；

-actuating the electrically controllable actuating element (04; 11) in order to set a second value (Θ) of the mechanical parameter (Θ) to be set_Target) (ii) a And

-adjusting the second value (Θ) at the mechanical parameter (Θ)_Target) In case (2), a second microscopic image of the sample (06) is taken.

2. Method according to claim 1, characterized in that it comprises further steps which are respectively carried out after the actuation of the electrically controllable steering elements (04; 11) and which comprise focusing the sample (06).

3. Method according to claim 1 or 2, characterized in that the actuating element is formed by an actuator (04) which can be driven by an electric motor and by means of which the mechanical parameter (Θ) can be adjusted.

4. Method according to any one of claims 1 to 3, characterized in that the mechanical parameter (Θ) is manually variable and the operating element is formed by a setting device (11) by means of which the mechanical parameter (Θ) can be set, and in that the step of actuating the electrically controllable operating element (11) comprises the following sub-steps, respectively:

-measuring the value (Θ) of said mechanical parameter (Θ)_{At present}) Time profile of the manually-induced change of (a);

-predicting the value (Θ) of said mechanical parameter (Θ)_{At present}) Will result in a value (Θ) of the mechanical parameter (Θ) which is to be set in each case_Target) The time point of (a); and

-electrically controlling the electrically operable setting device (11) at a previously determined point in time.

5. Method according to claim 4, characterized in that said mechanical parameter is formed by an inclination angle (Θ) and in that the value (Θ) of said inclination angle (Θ) is predicted_{At present}) Will result in a value (Θ) of the tilt angle (Θ) which is to be adjusted in each case_Target) The step of time point of (a) comprises the substeps of:

-measuring an initial value Θ of said tilt angle_Initial；

-by the value (Θ) for said tilt angle (Θ)_{At present}) To determine the angular velocity omega and the angular acceleration α, and

by equation Θ solved for time t_Target＝Θ_Initial+ω·t+1/2·α·t²Starting from this, the value (Θ) of the tilt angle (Θ) is predicted_{At present}) Will result in a value (Θ) of the tilt angle, which is to be set in each case_Target) The time point of (a).

6. Method according to claim 4 or 5, characterized in that the prediction of the value (Θ) of the mechanical parameter (Θ) is repeated continuously_{At present}) Will result in a value (Θ) of the mechanical parameter (Θ) which is to be set in each case_Target) Until the manual change of the value of the mechanical parameter (Θ) has resulted in a value (Θ) of the mechanical parameter (Θ) to be adjusted, respectively_Target)。

7. The method according to any one of claims 1 to 6, characterized in that the mechanical parameter is formed by the inclination angle (Θ) between the objective (07) of the microscope and the stage (02) of the microscope; wherein the objective (07) is fastened to a pivotable rocker arm (03), or wherein the object table (02) is rotatable about an axis parallel to the extension plane of the object table (02), whereby the inclination angle (Θ) between the objective (07) and the object table (02) can be changed.

8. The method according to any one of claims 1 to 7, characterized in that it further comprises the steps of:

-when the value of the mechanical parameter (Θ) has changed by a predefined amount and/or a value (Θ) of the mechanical parameter (Θ) to be adjusted in each case has been adjusted_Target) When this occurs, feedback is generated to the operator.

9. Method according to claim 8, characterized in that the feedback is haptically perceptible and is generated with an electromagnetically manipulatable setting device (11).

10. Method according to any one of claims 1 to 9, characterized in that the values (Θ) of the mechanical parameter (Θ) to be respectively adjusted are chosen by incrementing a previously chosen value of the mechanical parameter (Θ) by an increment_Target)。

11. The method according to any one of claims 1 to 10, characterized in that it further comprises the following further steps repeated a plurality of times:

-selecting other values (Θ) of said mechanical parameter (Θ) with said mechanical parameter (Θ)_Target) Different further values (Θ) to be set_Target)；

-actuating the electrically controllable actuating element (04; 11) in order to set a further value (Θ) of the mechanical variable (Θ) to be set_Target) (ii) a And

-adjusted further values (Θ) of the mechanical parameter (Θ)_Target) In case (2), a further microscopic image of the sample (06) is taken.

12. The method according to any one of claims 1 to 11, characterized in that different magnifications of the objective lens are selected and/or different objective lenses are selected for taking a first microscopic image of the sample (06) and for taking a second microscopic image of the sample (06).

13. The method according to any one of claims 1 to 12, characterized in that the method is configured for microscopic examination of at least two samples (06), wherein identical first values (Θ) of the mechanical parameter (Θ) are respectively selected and adjusted_Target) A second value (Θ) identical to said mechanical parameter (Θ)_Target) For taking a microscopic image of the first sample (06) and for taking a microscopic image of the second sample (06).

14. The method according to any one of claims 1 to 13, characterized in that it further comprises the steps of:

-generating a 2.5-dimensional or 3-dimensional rendering of the sample (06) from a plurality of microscopic images.

15. Microscope for microscopic examination of a sample (06), wherein a mechanical parameter (Θ) is variable, comprising:

-an electrically controllable actuating element (04; 11) for setting the value of the mechanical parameter (Θ); and

-a control unit configured for implementing the method according to any one of claims 1 to 14.