Real-time autofocus scanning

Patent metadata

Field	Value
Publication number	US-11422351-B2
Application number	US-202017105251-A
Country	US
Kind code	B2
Filing date	Nov 25, 2020
Priority date	Sep 29, 2017
Publication date	Aug 23, 2022
Grant date	Aug 23, 2022

Abstract

Real-time autofocus. In an embodiment, a scanning apparatus includes an imaging sensor, a focusing sensor, an objective lens, and processor(s) configured to analyze image data captured by the imaging and focusing sensors, and move the objective lens. Real-time autofocus during scanning of a sample is achieved by determining a true-Z value for the objective lens for a point on a sample and for each of a plurality of regions on the sample. The true-Z values and/or surfaces calculated therefrom are used to determine a predicted-Z value for an unscanned region of the sample. The objective lens is adjusted to the predicted-Z value at the beginning of the unscanned region. After scanning the region, a true-Z value is determined for the region and compared to the predicted-Z value. A rescan of the region is initiated if the comparison exceeds a predetermined threshold.

First claim

What is claimed is:

1. A digital scanning apparatus comprising:
an objective lens;
at least one sensor positioned in an optical path of the objective lens and configured to sense a field of view of the objective lens; and
at least one processor configured to:
adjust a height of the objective lens during scanning of a sample, and
for each of a plurality of regions of the sample to be scanned:
determine a predicted objective lens distance Z value for the region based on a stored objective lens distance Z value that is associated with a location that is nearest to the location of the region than any other location that is associated with any stored objective lens distance Z value, wherein each stored objective lens distance Z value represents a distance between the objective lens and the sample that provides best focus at the associated location,
at a start of scanning the region, adjust a distance between the objective lens and the sample based on the predicted objective lens distance Z value,
scan the region to acquire a buffer of image data,
based on the buffer of image data, determining an objective lens distance Z value that represents a distance between the objective lens and the sample that provides best focus for the region after scanning the region, and
store the objective lens distance Z value, determined for the region, in association with a location on the sample.

2. The apparatus of claim 1 , wherein the at least one processor of the digital scanning apparatus is further configured to, for each of the plurality of regions of the sample to be scanned:
after determining the objective lens distance Z value for the region, determine a difference between the objective lens distance Z value for the region and the predicted objective lens distance Z value for the region; and,
when the difference exceeds a predetermined threshold, initiating a rescan of the region.

3. The apparatus of claim 2 , wherein the at least one processor of the digital scanning apparatus is further configured to, after scanning the plurality of regions:
determine whether or not the difference between the objective lens distance Z value and the predicted objective lens distance Z value exceeds the predetermined threshold for a predetermined percentage of the scanned plurality of regions; and,
when determining that the difference between the objective lens distance Z value and the predicted objective lens distance Z value exceeds the predetermined threshold for the predetermined percentage of the scanned plurality of regions, initiate a rescan of the entire sample.

4. The apparatus of claim 2 , wherein the predetermined threshold is within a range of 0.5 microns to 0.9 microns.

5. The apparatus of claim 1 , wherein the at least one processor of the digital scanning apparatus is further configured to, when determining an objective lens distance Z value that represents a distance between the objective lens and the sample that provides best focus for a respective region, determine a distance between the objective lens and the sample that is associated with a peak contrast value from the respective buffer, wherein the at least one sensor comprises a focusing sensor with one or more linear arrays, wherein each of the one or more linear arrays is tilted such that each pixel in the linear array is in a different image plane, and wherein the at least one processor of the digital scanning apparatus is further configured to, when determining a distance between the objective lens and the sample that is associated with a peak contrast value from the respective buffer:
for each of a plurality of columns of pixels in the respective buffer, calculate an average contrast value;
identify the peak contrast value as a highest one of the determined average contrast values; and
determine the distance between the objective lens and the sample that is associated with the column of pixels for which the highest average contrast value was calculated,
wherein each average contrast value is an average contrast ratio value.

6. The apparatus of claim 1 , wherein:
the at least one processor of the digital scanning apparatus is further configured to calculate a global sample surface using all of the stored objective lens distance Z values, wherein the predicted objective lens distance Z value for each of the plurality of regions is further based on the global sample surface,
the at least one processor of the digital scanning apparatus is further configured to, after scanning each of the plurality of regions, recalculate the global sample surface using the objective lens distance Z value determined for that region, and
the at least one processor of the digital scanning apparatus is further configured to continually update the global sample surface as new objective lens distance Z values are determined and stored, to reflect all stored objective lens distance Z values for the sample.

7. The apparatus of claim 1 , wherein the at least one processor of the digital scanning apparatus is further configured to, for each of one or more of the plurality of regions, calculate a local sample surface using objective lens distance Z values determined for at least a subset of neighboring regions of the sample, wherein the predicted objective lens distance Z value for the region is further based on the local sample surface, wherein the at least one processor of the digital scanning apparatus is further configured to, for each of the plurality of regions:
when an objective lens distance Z value has not been calculated for any neighboring regions, determine the predicted objective lens distance Z value for the region based on the stored objective lens distance Z value that is associated with the location that is nearest to the location of the region than any other location that is associated with any stored objective lens distance Z value, and not based on any local sample surface; and,
when an objective lens distance Z value has been calculated for neighboring regions such that a local sample surface is calculated, determine the predicted objective lens distance Z value for the region further based on the local sample surface.

8. The apparatus of claim 1 , wherein the at least one processor of the digital scanning apparatus is further configured to, prior to scanning the plurality of regions:
select at least one macro-focus point on the sample;
acquire image data comprising the macro-focus point at a plurality of image planes corresponding to a plurality of distances between the objective lens and the sample;
determine a objective lens distance Z value that represents a distance between the objective lens and the sample that provides best focus for the macro-focus point; and
store the objective lens distance Z value, determined for the macro-focus point, in association with a location of the macro-focus point on the sample.

9. The apparatus of claim 8 , wherein the at least one processor of the digital scanning apparatus is further configured to, when selecting at least one macro-focus point on the sample, select the at least one macro-focus point to be within a predetermined range from an edge of the sample.

10. The apparatus of claim 9 , wherein the at least one processor of the digital scanning apparatus is further configured to, when selecting at least one macro-focus point on the sample, select the at least one macro-focus point to be within the predetermined range from a line representing a maximum length across the sample, wherein the predetermine range is a radius or a range of the at least one sensor.

11. The apparatus of claim 8 , wherein the plurality of distances comprise both a closest and farthest possible distance between th

Assignees

• Leica Biosystems Imaging Inc

Inventors

• Chen Leng-Chun
• Olson Allen
• Zou Yunlu
• Najmabadi Peyman
• Crandall Greg

Classifications

• H04N25/711

  Time delay and integration [TDI] registers; TDI shift registers · CPC title

• G06T2207/10064

  Fluorescence image · CPC title

• G06T2207/10061

  from scanning electron microscope · CPC title

• G06T7/50

  Depth or shape recovery · CPC title

• G02B21/244 Primary

  using image analysis techniques · CPC title