Light microscope with automatic focusing

1 . A method for automatically focusing a microscope, comprising:

an initial coarse focusing in which a coarse focus setting is determined, followed by a subsequent fine focusing in which a fine focus setting is determined based on the coarse focus setting,

the initial coarse focusing comprising:

determining the coarse focus setting at a lateral xy-location of a lateral edge or corner of a cover slip, a microscope slide, a well of a multiwell plate, or a sample vessel assessed as a best-suited lateral xy-location for a focus determination based on predetermined criteria out of a plurality of assessed lateral xy-locations of lateral edges or corners of the cover slip, the microscope slide, the well of the multiwell plate, or the sample vessel in an overview image, wherein the coarse focus setting is determined at the best-suited lateral xy-location identified in the overview image;

the subsequent fine focusing comprising:

determining the fine focus setting at the sample by recording image data at the sample with the coarse focus setting and modifying the coarse focus setting to enhance image sharpness; and

after determining the fine focus setting, recording at least one sample image using the fine focus setting.

2 . The method of claim 1 ,

wherein an actuating device performs a relative movement between the sample and a detection beam path with the result that the detection beam path is directed at the best-suited lateral xy-location, and subsequently the coarse focus setting is determined.

3 . The method of claim 1 ,

wherein the initial coarse focusing uses a segmentation algorithm or detection algorithm via which boundaries of objects are ascertained in the overview image, wherein the segmentation algorithm or detection algorithm is based on a machine learning algorithm.

4 . The method of claim 1 ,

wherein a position of one of a plurality of boundary portions is selected as the best-suited lateral xy-location, in dependence on an alignment of the respective boundary portion relative to an illumination direction used in the subsequent determination of the coarse focus setting.

5 . The method of claim 1 , further comprising

providing a learning mode to a user, in which:

reference overview images of reference samples are represented,

a marking tool is provided for the user with which the user can mark one or more lateral edges in each reference overview image,

a machine learning algorithm establishes, based on the reference overview images and the boundaries marked by the user, criteria with which the best-suited lateral xy-location is ascertained for determining the coarse focus setting.

6 . The method of claim 1 ,

wherein a plurality of lateral xy-locations of a plurality of edges and at least one corner are ascertained in the overview image, and the at least one corner is used as the location where the coarse focus setting is determined.

7 . The method of claim 1 ,

wherein the initial coarse focusing includes ascertaining boundaries of at least two objects that are not the sample itself in the overview image, wherein at least one of the at least two objects is a sticker, a printed area or a mark on a microscope slide, and the boundary of the at least one of the at least two objects is or comprises a periphery of the sticker, of the printed area or of the mark.

8 . The method of claim 1 ,

wherein determining the coarse focus setting comprises at least:

recording a plurality of microscope images of different height focusing at the best-suited lateral xy-location;

determining a respective image sharpness quality factor of the microscope images;

determining that microscope image or those microscope images having the maximum image sharpness quality factor;

establishing the coarse focus setting based on the microscope image/microscope images having the maximum image sharpness quality factor.

9 . The method of claim 8 ,

wherein, as part of the determination of the coarse focus setting, a cover slip thickness is also determined, wherein a lateral cover slip edge is ascertained in the overview image,

wherein during the determination of the microscope images with a maximum image sharpness quality factor, two microscope images with a local maximum in the image sharpness quality factor are ascertained,

wherein a cover slip thickness is ascertained from a difference between height values of the two microscope images.

10 . The method of claim 1 ,

wherein as part of the determination of the coarse focus setting, a microscope slide inclination is also ascertained, wherein at least three locations of a boundary are ascertained on which then in each case a local focus setting that indicates a height position of the respective location is ascertained,

wherein the microscope slide inclination is ascertained based on the at least three height positions.

11 . The method of claim 10 , further comprising

selecting the locations of the boundary that are used for ascertaining the microscope slide inclination such that the sample is located laterally between these locations of the boundary.

12 . The method of claim 10 ,

wherein the number of locations of the boundary used for determining the microscope slide inclination is selected in dependence on a lateral distance between said locations.

13 . The method of claim 10 , further comprising

using the ascertained microscope slide inclination for the fine focus setting in subsequent sample examinations, wherein for different lateral positions of the sample a respective fine focus setting is adjusted in accordance with the microscope slide inclination.

14 . The method of claim 1 ,

wherein, as part of the determination of the coarse focus setting, an orientation alignment is ascertained by

ascertaining in the overview image at least one cover slip periphery and one microscope slide boundary as boundaries,

ascertaining the coarse focus setting and thus a height position in each case for the cover slip periphery and the microscope slide boundary,

ascertaining, using a comparison of the height positions of the cover slip periphery and of the microscope slide boundary, whether the cover slip is located above or below the microscope slide.

15 . The method of claim 14 ,

wherein the ascertained microscope slide inclination is taken into account for the ascertainment of the orientation alignment by calculating a height offset based on the microscope slide inclination for the height positions to be compared.

16 . The method of claim 1 , further comprising

ascertaining at least one cover slip periphery and one microscope slide boundary as boundaries in the overview image,

determining a focus setting and thus a height position in each case for the cover slip periphery and the microscope slide boundary, and

setting the coarse focus setting for subsequent sample examinations to a height position, wherein the height position is between the determined height positions of the cover slip periphery and the microscope slide boundary.

17 . A computer program product comprising instructions that, upon execution by a computing unit of a light microscope, cause the latter to carry out the method of claim 1 .

18 . A light microscope with automatic focusing, comprising:

an electronic control and evaluation device, which is configured to perform an initial coarse focusing in which a coarse focus setting is determined, followed by a subsequent fine focusing in which a fine focus setting is determined based on the coarse focus setting;

wherein the electronic control and evaluation device is configured to:

determine the coarse focus setting at a lateral xy-location of a lateral edge or corner of one of: a cover slip, a microscope slide, a well of a multiwell plate, or a sample vessel assessed as a best-suited lateral xy-location for a focus determination based on predetermined criteria out of a plurality of assessed lateral xy-locations of lateral edges or corners of the cover slip, the microwave slide, the well of the multiwell plate, or the sample vessel in an overview image, wherein the coarse focus setting is determined at the best-suited lateral xy-location identified in the overview image;

determine the fine focus setting at the sample by recording image data at the sample with the coarse focus setting and modifying the coarse focus setting to enhance image sharpness; and

after determination of the fine focus setting, record at least one sample image using the fine focus setting.

19 . The light microscope of claim 18 ,

wherein, as part of the determination of the coarse focus setting, the electronic control and evaluation device is configured to ascertain a microscope slide inclination, wherein at least three locations of a boundary are ascertained on which then in each case a local focus setting that indicates a height position of the respective location is ascertained, and wherein the microscope slide inclination is ascertained based on the at least three height positions.

20 . A method for automatically focusing a microscope, comprising:

an initial coarse focusing in which a coarse focus setting is determined, followed by a subsequent fine focusing in which a fine focus setting is determined based on the coarse focus setting,

the initial coarse focusing comprising:

determining the coarse focus setting at a lateral xy-location of a text or an edge or corner of a text field on a microscope slide or sample vessel assessed as best-suited for a focus determination based on predetermined criteria out of a plurality of assessed lateral xy-locations of text or lateral edges or corners of the text field on the microscope slide or the sample vessel in an overview image, wherein the coarse focus setting is determined at the best-suited lateral xy-location identified in the overview image;

the subsequent fine focusing comprising:

determining the fine focus setting at the sample by recording image data at the sample with the coarse focus setting and modifying the coarse focus setting to enhance image sharpness; and

after determination of the fine focus setting, recording sample images at xy-locations of the sample using the fine focus setting.

21 . A method for automatically focusing a microscope, comprising:

an initial coarse focusing in which a coarse focus setting is determined at an xy-location outside a sample, followed by a subsequent fine focusing in which a fine focus setting is determined at the sample based on the coarse focus setting, the initial coarse focusing comprising:

analyzing an overview image showing the sample and an environment to ascertain a plurality of lateral xy-locations of a lateral edge or corner of one of: a cover slip, a microscope slide, a well of a multiwell plate, a sample holder, a sample vessel, or another barrier which physically limits a specimen area;

determining, for each lateral xy-location of the plurality of lateral xy-locations, a suitability for determination of the coarse focus setting based on predetermined criteria;

selecting at least one best-suited lateral xy-location out of the plurality of lateral xy-locations, wherein the at least one best-suited lateral xy-location identifies a part of the overview image which is laterally distanced from the sample; and

determining the coarse focus setting at the selected at least one best-suited lateral xy-location of the lateral edge or corner of the cover slip, microscope slide, well, sample holder, sample vessel, or the another barrier which physically limits the specimen area, while disregarding other regions recorded in the overview image for the determining of the coarse focus setting;

the subsequent fine focusing comprising:

determining the fine focus setting at the sample by recording image data at the sample with the coarse focus setting and modifying the coarse focus setting to enhance image sharpness; and

recording at least one sample image with the fine focus setting; and

providing a learning mode to a user, in which:

reference overview images of reference samples are represented, a marking tool is provided for the user with which the user can mark one or more lateral edges in each reference overview image, a machine learning algorithm establishes, based on the reference overview images and the boundaries marked by the user, criteria with which the at least one best-suited lateral xy-location of the lateral edge or corner is ascertained for determining the coarse focus setting; and

wherein as part of the determination of the coarse focus setting, a microscope slide inclination is also ascertained, wherein at least three locations of a boundary are ascertained on which then in each case a local focus setting that indicates a height position of the respective location is ascertained, wherein the microscope slide inclination is ascertained based on the at least three height positions.