US-20260126636-A1 - OBJECTIVE LENS UNIT FOR A MICROSCOPE, MICROSCOPE COMPRISING THE OBJECTIVE LENS UNIT, AND METHOD OF SETTING THE OBJECT PLANE OF A MICROSCOPE

US20260126636A1US 20260126636 A1US20260126636 A1US 20260126636A1US-20260126636-A1

Abstract

An objective lens unit for a microscope includes at least one optically detectable target on at least one optical component of the objective lens unit which has a well-defined position relative to a distal front end of an enclosure of the objective lens unit or a distal end of the microscope, respectively. An adjustable lens of the objective lens unit is adjusted to sharply image at least one of the optically detectable target(s), such as on a sensor of a camera attached to the microscope. Thus, a setting of the adjustable lens for a reference object plane is determined. The adjustable lens may subsequently be adjusted to shift the object plane a distance into a distal direction to set the object plane at an intended location distal from the distal front end of the enclosure or the distal end of the microscope, respectively, in a sample volume.

Inventors

Sandro Link
Rudolf Gati

Assignees

METTLER-TOLEDO GMBH

Dates

Publication Date: 20260507
Application Date: 20251020
Priority Date: 20241104

Claims (15)

1 . An objective lens unit for a microscope, the objective lens unit comprising: an enclosure, the enclosure having a distal front end; a front end optical aperture of the objective lens unit provided in the distal front end of the enclosure; an objective lens system arranged inside the enclosure and proximal from the front end optical aperture of the objective lens unit, wherein the objective lens system is configured to collect light received through the front end optical aperture of the objective lens unit, and wherein the objective lens system comprises an adjustable lens; and at least one optical component distal from the adjustable lens, wherein at least one optically detectable target is provided on the at least one optical component.
2 . The objective lens unit of claim 1 , wherein: the adjustable lens is a motorless automated adjustable lens.
3 . The objective lens unit of claim 2 , wherein: the motorless automated adjustable lens comprises one of a tuneable lens and a lens axially displaceable along the optical axis of the objective lens unit by a piezo actuator.
4 . The objective lens unit of claim 1 , wherein: the at least one optically detectable target is provided on a plane surface extending perpendicular to the optical axis of the objective lens unit.
5 . The objective lens unit of claim 1 , wherein: an immersion lens is provided distal from the adjustable lens and configured to receive light through the front end optical aperture.
6 . The objective lens unit of claim 1 , wherein: at least one of the at least one optically detectable target is provided on a most distal component of the objective lens unit.
7 . The objective lens unit of claim 6 , wherein: the most distal optical component of the objective lens unit is one of an immersion lens and a window closing the front end optical aperture.
8 . The objective lens unit of claim 1 , wherein: at least one of the at least one optically detectable target is 3-dimensionally shaped.
9 . The objective lens unit of claim 1 , wherein: the objective lens unit comprises at least one first optically detectable target and at least one second optically detectable target; and the at least one first optically detectable target and the at least one second optically detectable target are provided at different axial positions.
10 . The objective lens unit of claim 1 , wherein: the enclosure comprises a cap and a sleeve; the objective lens system is provided inside the sleeve; the cap comprises a lateral sheath, a front wall, a rear port, and the at least one optical component; the front end optical aperture of the objective lens unit is provided in the front wall of the cap; the at least one optically detectable target is provided on the at least one optical component of the cap; and the sleeve is at least partially received inside the cap or the cap is at least partially received inside the sleeve.
11 . A cap configured for the objective lens unit of claim 10 , wherein: the cap comprises a rear port, a front wall, and a lateral sheath extending axially from the front wall; the front end optical aperture is formed in the front wall; the lateral sheath is either configured to receive a sleeve therein and the sleeve is insertable in an axial direction through the rear port of the cap or is configured to be received in the sleeve and the lateral sheath is insertable in an axial direction in the sleeve; the cap comprises the at least one optical component and a most distal one of the at least one optical component closes the front end optical aperture; and the at least one optically detectable target is provided on at least one of the optical components.
12 . The cap of claim 11 , wherein: at least one of the at least one optically detectable target is provided on the most distal one of the at least one optical component.
13 . A microscope comprising the objective lens unit of claim 1 and an optical sensor, wherein the optical sensor is functionally coupled to the objective lens unit to receive light transmitted from a front end of the objective lens unit and through the objective lens system.
14 . A method for setting an object plane of a microscope, wherein: the microscope comprises at least one optical sensor and at least one adjustable lens; the at least one adjustable lens is configured to axially shift the object plane of the microscope relative to a distal end of the microscope, wherein the object plane is a plane in which objects are sharply imaged on the at least one optical sensor; at least one optically detectable target is provided in an area imaged on the optical sensor and at a determined axial position relative to the distal end of the microscope; and the method comprising the steps of: defining an intended object plane position relative to the distal end of the microscope, located at an intended distance distal from the distal end of the microscope; adjusting the at least one adjustable lens until the at least one optically detectable target is sharply imaged on the optical sensor; and adjusting the at least one adjustable lens to axially move the object plane along the optical axis of the microscope to the intended object plane position.
15 . The method according to claim 14 , wherein: two focussing targets are used which are provided at different axial positions, in an area imaged on the optical sensor and at a known axial distance from each other; the two focussing targets are provided by either: at least one of the at least one optically detectable target being 3-dimensionally shaped and comprising, at a known axial distance from each other, at least two optically distinct features, each of these features forming one of the at least two focussing targets; and/or at least one first optically detectable target at a first axial position and at least one second optically detectable target at a second axial position, whereby the first axial position and the second axial position are at a known axial distance from each other, each of the first and the second optically detectable targets forming one of the two focussing targets; the method comprising the steps of: performing a first calibration adjustment by adjusting the at least one adjustable lens until a first one of the focussing targets is sharply imaged on the optical sensor; performing a second calibration adjustment by adjusting the at least one adjustable lens until a second one of the focussing targets is sharply imaged on the optical sensor; determining an adjustment calibration difference as a magnitude of adjustable lens adjustment applied for shifting the object plane from the first focussing target to the second focussing target; determining a required adjustable lens adjustment magnitude required to set the object plane to the intended object plane position from the: adjustment calibration difference; the axial distance between the first one of the focussing targets and the second one of the focussing targets; and the axial distance between the first or the second one of the focussing targets and the intended object plane position; and adjusting the at least one adjustable lens by applying the required adjustable lens adjustment magnitude to set the object plane to the intended object plane position.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of European Application No. 24210649.0 filed Nov. 4, 2024, and European Application No. 25163723.7 filed Mar. 14, 2025, the disclosures of each of which are hereby incorporated by reference as if fully restated herein. TECHNICAL FIELD The herein claimed subject matter relates generally to instruments and methods applicable for microscopy. More in particular, it relates to the subject matter set forth in the claims. BACKGROUND In certain applications of microscopy, it is required to set the object plane of the microscope, i.e., a plane in which objects are sharply imaged, a certain distance in front of, or distal from, a front end or front end optical aperture of the microscope, without an object to be referenced present during the object plane setting. This may typically be the case in, while not limited to, in-line and in-situ microscopy, in particular when the objects to be observed move through a sample volume. In said case, there is, for instance, no stationary object which could be focused on. One typical, while again non-limiting, instance is cytometry, in particular image cytometry of bioprocesses. Bioprocesses are processes which use complete living, biological cells or their components such as for example bacteria, enzymes or chloroplasts, to obtain desired products. A bioprocess is typically done in a bioreactor, i.e., a process vessel which is preferably either reusable and therefore sterilizable tank or a single-use bag. The biological cells being part of the bioprocesses are dispensed in a liquid medium, establishing thereby the suitable environment for the desired process. Typically, the medium comprises for example nutrients for the cells in question as well as gases needed by them such as O2 and CO2. In most cases, it is important to ensure that no cells other than the ones involved in the bioprocess are present in the bioreactor, implying that it is preferred to maintain a sterile barrier between the process and the outside as long as possible thereby minimizing the number and duration of occasions where a contamination with unwanted cells can happen. Often, the bioprocess uses a means to mix the cells and the medium constantly. Examples of such means are stirrers arranged on the inside of the bioreactor or shakers which move the whole bioreactor. Therefore, in general, the cells suspended in the media are moving inside the bioreactor. In many cases, the medium is closely monitored to ensure that the desired conditions are maintained. For this monitoring, there are today different optical, opto-chemical and electrochemical sensors available and in use which can measure for example the pH-value or the amount of dissolved oxygen in-line or in-situ in a reliable manner. There is, however, currently no similar reliable measurement device available to monitor the cells directly. In-line or in-situ measurements are—in the case of a process, in particular a bioprocess—measurements which take place directly in the reactor respectively in the bioreactor or in a tube transporting the fluid to be monitored. In particular, the fluid to be monitored is the liquid of the bioprocess comprising the medium and the biological cells. Cytometry is applied to characterize living or dead biological cells, and preferably for an image cytometer using the invention at hand the cells to be characterized are provided in a liquid. For instance, density, size and morphology of biological cells may be determined by microscopy. In the absence of reliable and quantitative in-line measurement methods, bioengineers have to take samples to perform cytometry with off-line measurement devices. This requires taking a sample volume of the bioreactor within certain intervals, which increases the risk of contamination. In addition, it is time-consuming and due to the sparse interval, little statistics and no live information about cytometry is available to the operator. An in-line cytometer, characterizing the biological cells in the, typically liquid, environment in which they are cultivated, would drastically reduce the complexity compared to off-line methods and would allow the process controller to monitor the cell count and cell characteristics, such as the ones named above, close to real-time. In certain applications, the sample volume in which objects are sharply imaged may be optically confined: In a first dimension, the sample volume can be optically confined by the depth of field of the microscope. Preferably, it is the objective lens unit which determines the depth of field of the microscope. In the two dimensions perpendicular to the first dimension it is the area imaged which confines the sample volume optically: the area is determined for instance, by the field of view of the microscope, defined for example by the size of the optical sensor attached to the microscope or by an aperture arranged in the optical path, for example the front end op