Systems, methods, and apparatus for immersion media coating and lens cleaning

Abstract

An imaging system configured for automated application and / or removal of immersion medium can include: (i) a sample stage; (ii) an imaging assembly disposed on a first side of the sample stage and having an immersion objective configured to be selectively aligned with an optical axis of the imaging system; and (iii) an applicator positioned to selectively interact with a lens surface of the immersion objective to deposit or remove immersion medium.

Description

Technical Field

[0001] This disclosure generally relates to microscopes. More specifically, this disclosure relates to systems, methods, and apparatus for applying and cleaning immersion media on objectives. Background Technology

[0002] Microscopy involves observing small objects, often microscopic. Traditional microscopes incorporate lens systems to magnify and thus allow observation of these small objects. In optical microscopes, the lens system guides a magnified image of the small object to the microscope's eyepiece, while in digital microscopes, the image is focused onto an image sensor. In either case, optical microscopes are typically used to capture images of microscopic objects. At lower magnifications, a wide field of view (at lower resolution) allows for rapid navigation within or between samples, and once a point of interest is identified, a higher-resolution objective can be moved into the optical axis, allowing for more detailed observation and imaging of that point of interest. Objectives with high numerical apertures provide high resolution but are limited by the refractive index of air unless an immersion medium with a refractive index greater than air is placed between the sample and the objective. Therefore, immersion media are typically used in optical microscopy applications to obtain high-resolution images of the sample.

[0003] While immersion media can be manually applied to objectives, the application of immersion media to high-resolution objectives has become a bottleneck for system productivity and efficiency with the advent of automated, high-throughput imaging systems. Objectives in many automated imaging systems are difficult to access quickly and are often confined to small spaces. Previous efforts to automate the application of immersion media to objectives have encountered numerous problems and limitations. For example, due to space constraints and the dynamic movement of components within automated imaging systems, existing systems can typically only effectively apply immersion media to a single objective. Furthermore, existing systems include additional motors or actuators for moving and / or flipping the immersion applicator into and out of its operable configuration, increasing the mechanical and operational complexity of these systems. If additional or different immersion objectives are to be used during automated imaging, existing systems require reconfiguration of the immersion media applicator and / or objectives within the system, or in some cases, the addition of multiple applicators.

[0004] Furthermore, existing systems fail to provide an automated and efficient method for cleaning immersion media from immersion objectives. Prolonged exposure to some immersion media (such as cedarwood oil) can negatively impact the lifespan or functionality of objectives. Even for non-corrosive immersion media whose properties remain relatively unchanged over time or upon exposure to light, best practices for optimal imaging system performance include removing the immersion media from the lens immediately after use or between applications. Existing systems fail to address this additional challenge in the field of automated imaging systems utilizing optical microscopy methods.

[0005] Therefore, there are many shortcomings and problems in the field of automated optical microscopy, and there is a great need for systems, methods and apparatus that can conveniently and automatically apply immersion media to objectives in automated systems and / or automatically clean or remove immersion media from objectives, especially when using automated optical microscopes. Summary of the Invention

[0006] The various embodiments disclosed herein relate to apparatus, methods, and systems for applying immersion media and cleaning lenses.

[0007] A first aspect of the disclosed embodiments provides an imaging system configured for automatically applying and / or removing an immersion medium. The imaging system includes: (i) a sample stage; (ii) an imaging assembly disposed on a first side of the sample stage and having an immersion objective configured to selectively align with the optical axis of the imaging system; and (iii) an applicator positioned to selectively interact with the lens surface of the immersion objective to deposit or remove the immersion medium.

[0008] In one aspect, the applicator includes an immersion medium nozzle. The immersion medium nozzle can be configured to dispense a bubble-free immersion medium and can include a bubble sensor configured to detect the presence of bubbles at the immersion medium nozzle or in an upstream line supplying the immersion medium to the nozzle. The applicator may additionally or alternatively include a liquid sensor for detecting the presence of immersion medium at the immersion medium nozzle. The liquid sensor may include an optical sensor, a multimeter for measuring the resistance at the lens surface of an immersion objective, or a capacitor sensor.

[0009] In one aspect, the imaging system further includes a hose connecting the applicator to an immersion medium reservoir, and may also include a pump associated with the hose, the pump being configured to dispense immersion medium from the immersion medium reservoir via the applicator. For example, the pump may be configured to dispense a desired volume of immersion medium based on operating time and / or the number of operating cycles.

[0010] In one aspect, the applicator is positioned on a first side of the sample stage and can be integrated into and translate together with the sample stage. Such a sample stage can be a motorized xy stage configured to position the applicator adjacent to the lens surface of the immersion objective, such that dispensing the immersion medium from the applicator causes the immersion medium to deposit onto the lens surface of the immersion objective.

[0011] In one aspect, the imaging system is an inverted microscope, wherein the imaging component is positioned below the sample stage, and a first side of the sample stage is the bottom of the sample stage, such that the applicator is oriented on the bottom of the sample stage toward the immersion objective. Alternatively, the imaging system can be an upright microscope, wherein the imaging component is positioned above the sample stage, and a first side of the sample stage is the top of the sample stage, such that the applicator is oriented on the top of the sample stage toward the immersion objective.

[0012] In one aspect, the imaging system further includes a wiping material configured to clean and / or remove the immersion medium from the lens surface of the immersion objective. The wiping material may contain a cleaning agent and, in some cases, may be an applicator. In this case, the imaging system may further include a displacement element operatively connected to the wiping material and configured to selectively move the wiping material. The selective movement of the wiping material may be, or may include, linear or back-and-forth movement, movement within a single plane, or rotational movement. The displacement element may be, for example, a solenoid that provides a similar vibrational amplitude to the wiping material, or may otherwise be associated with a mechanism for moving the sample stage. Additionally or alternatively, the imaging system includes a computing system configured to generate a map of the wiping material, track portions of the wiping material previously used for cleaning the immersion objective, and guide the movement of the wiping material during subsequent cleaning operations to interact with the immersion objective in clean or unused areas of the wiping material.

[0013] In one aspect, the applicator is a suction device configured to remove immersion medium from the lens surface of an immersion objective. Such an applicator can be positioned in a fixed location within the housing of the imaging system. The imaging assembly may include a lens slider or turntable on which the immersion objective is mounted and selectively positioned below the applicator to receive immersion medium from the applicator onto the lens surface of the immersion objective. Such an exemplary imaging system may additionally include a wiping agent (e.g., containing a cleaning agent) configured to clean and / or remove immersion medium from the lens surface of the immersion objective.

[0014] In one aspect, the applicator is a wiping material, and the imaging system may additionally include a vibrating element operatively connected to and configured to selectively vibrate the wiping material. A lens slider or turntable may be selectively positioned below the wiping material to remove the immersion medium from the lens surface of the immersion objective.

[0015] In one aspect, the imaging system includes a second immersion objective, and the applicator is further configured to selectively interact with the corresponding lens surface of the second immersion objective.

[0016] This disclosure further includes a method for automatically applying an immersion medium to an immersion objective. In one aspect, the method includes: obtaining an imaging system as disclosed herein, positioning a sample stage relative to the immersion objective such that an applicator associated with the imaging system is adjacent to the lens surface of the immersion objective, and dispensing an immersion medium from the applicator onto the lens surface of the immersion objective.

[0017] In one aspect, a method for automatically applying an immersion medium to an immersion objective includes: obtaining an imaging system as disclosed herein; positioning the immersion objective relative to an applicator such that the applicator is adjacent to the lens surface of the immersion objective; and dispensing the immersion medium from the applicator onto the lens surface of the immersion objective. In some aspects, the method of dispensing the immersion medium may additionally include dispensing an air-free immersion medium.

[0018] This disclosure further includes a method for automatically removing an immersion medium from the lens surface of an immersion objective. In one aspect, the method includes: obtaining the imaging system disclosed herein, which includes a wiping or suction device; positioning a sample stage relative to the immersion objective such that an applicator is adjacent to the lens surface of the immersion objective; and removing the immersion medium from the lens surface of the immersion objective via the wiping or suction device.

[0019] Embodiments of this disclosure further include a kit for automatically dispensing immersion media. In one aspect, the kit includes an immersion media reservoir configured to hold a volume of immersion media, an applicator (such as a nozzle) fluidly connected to the immersion media reservoir via an immersion media hose, and a miniature pump operable to move the immersion media from the immersion media reservoir through the immersion media hose and to the nozzle for dispensing.

[0020] In one aspect, the kit includes computer-executable instructions that, when executed by one or more processors of a computer system, cause the applicator to automatically dispense the immersion medium. In another aspect, the computer-executable instructions, when executed by a processor of a computer system, cause a wiping or suction device to remove the immersion medium from the lens surface of the immersion objective, or otherwise clean the lens surface of the immersion objective via a wiping or suction device.

[0021] In one aspect, the kit further includes a liquid sensor configured to detect the presence of immersion medium at the nozzle, a check valve associated with the immersion medium hose to prevent pumped immersion medium from flowing back into the immersion medium reservoir when not being pumped, a level indicator associated with the immersion medium reservoir, and / or one or more of a waste reservoir and a waste level indicator associated with the waste reservoir.

[0022] In one respect, the kit's operable parts can be converted into automated optical microscopes after manufacturing.

[0023] In one aspect, an exemplary kit includes a nozzle associated with an immersion medium hose for fluid connection to an immersion medium reservoir and a miniature pump operable to move immersion medium from the immersion medium reservoir through the immersion medium hose and to the nozzle for dispensing at an immersion objective. Such an exemplary kit may include computer-executable instructions, when executed by one or more processors of a computer system, to cause the nozzle to automatically dispense immersion medium onto one or more immersion medium lenses at user-selected times and / or intervals. In one aspect, the kit further includes a wiping or suction device to remove immersion medium from the lens surface of the immersion objective, or otherwise clean the lens surface of the immersion objective via the wiping or suction device. The computer-executable instructions, when executed by a processor of a computer system, may further cause the wiping or suction device to remove (or otherwise clean) the immersion medium from the lens surface of the immersion objective. In one aspect, the kit may additionally include a liquid sensor configured to detect the presence of immersion medium at the nozzle, and a check valve associated with the immersion medium hose to prevent pumped immersion medium from flowing back into the immersion medium reservoir when not being pumped.

[0024] The purpose of this summary is to introduce, in a simplified form, the selection of concepts further described below in the detailed description. This summary is not intended to identify key or essential features of the claimed subject matter, nor is it intended to be used as an indication of the scope of the claimed subject matter.

[0025] Additional features and advantages of this disclosure will be set forth in the description which follows, and some of the description will be obvious, or may be learned by practice of this disclosure. The features and advantages of this disclosure may be realized and obtained by means of the instruments and combinations particularly pointed out in the appended claims. These and other features of this disclosure will become more fully apparent from the following description and the appended claims, or may be learned by practice of this disclosure as set forth below. Attached Figure Description

[0026] In order to describe the manner in which the above and other advantages and features of this disclosure can be obtained, a more specific description of the disclosure briefly described above will be presented by reference to specific embodiments thereof shown in the accompanying drawings. It should be understood that these drawings depict only typical embodiments of the disclosure and are therefore not to be construed as limiting the scope of the disclosure. The disclosure will be described and explained with additional features and details using the accompanying drawings, wherein:

[0027] Figure 1A This illustrates a typical upright microscope known in the prior art;

[0028] Figure 1B This illustrates a typical inverted microscope known in the prior art;

[0029] Figure 2 Example embodiments of systems incorporating features disclosed or contemplated herein are shown;

[0030] Figure 3 A plan view of a longitudinal cross-section of an exemplary imaging system according to an embodiment of the present disclosure is shown;

[0031] Figure 4A A schematic diagram of an exemplary immersion medium applicator associated with an imaging system having an objective lens mounted on an objective lens slider, according to an embodiment of the present disclosure, is shown.

[0032] Figure 4B A schematic diagram of an exemplary immersion medium applicator associated with an imaging system having an objective lens mounted on an objective lens stage, according to an embodiment of the present disclosure, is shown.

[0033] Figure 5A A top perspective view of the interior portion of the stage housing and the associated objective lens of an exemplary imaging system incorporating an immersion medium applicator into a microscope stage, according to an embodiment of the present disclosure, is shown.

[0034] Figure 5B This is a top perspective view of an exemplary sample holder arm configured for mounting on the xy stage of an imaging system according to an embodiment of the present disclosure, the sample holder arm having an immersion medium applicator incorporated on the side of the arm shown.

[0035] Figure 5C A schematic diagram of an exemplary nozzle and immersion medium sensor according to an embodiment of the present disclosure is shown;

[0036] Figure 6A This is a schematic diagram of an exemplary immersion medium applicator associated with a stage assembly according to an embodiment of the present disclosure, and is shown in a configuration in which the immersion medium is applied from a nozzle integrated into the stage to a desired objective lens associated with an inverted microscope system.

[0037] Figure 6B According to embodiments of this disclosure Figure 6A A schematic diagram of an exemplary immersion medium applicator, and shown in an imaging configuration in which the applied immersion medium is disposed between the objective lens and the bottom surface of a porous plate held by the stage assembly;

[0038] Figure 7 This is a schematic diagram of an exemplary immersion medium applicator associated with a stage assembly according to an embodiment of the present disclosure, and is shown in a configuration in which an immersion medium can be applied from a nozzle integrated / associated with the stage and applied to a desired objective lens of an upright microscope.

[0039] Figure 8 A schematic diagram of an exemplary lens cleaning apparatus according to an embodiment of the present disclosure is shown;

[0040] Figure 9 A schematic diagram of another exemplary lens cleaning device according to an embodiment of the present disclosure is shown; and

[0041] Figure 10 A schematic diagram of an exemplary immersion medium applicator and lens cleaning apparatus according to embodiments of the present disclosure is shown. Detailed Implementation

[0042] Before describing the various embodiments of this disclosure in detail, it should be understood that this disclosure is not limited to the parameters of the systems, methods, apparatus, products, and / or processes of particular examples, which can naturally vary. Therefore, while certain embodiments of this disclosure will be described in detail with reference to specific configurations, parameters, components, elements, etc., the descriptions are exemplary and should not be construed as limiting the scope of the claimed invention. Furthermore, the terminology used herein is for the purpose of describing embodiments and is not necessarily intended to limit the scope of the claimed invention.

[0043] Furthermore, it should be understood that, for any given component or embodiment described herein, any possible candidates or alternatives listed for said component may generally be used alone or in combination with each other, unless otherwise implied or expressly understood or stated. Additionally, it should be understood that, unless otherwise implied or expressly understood or stated, any list of such candidates or alternatives is exemplary only and not limiting.

[0044] Furthermore, unless otherwise indicated, the numbers used in the specification and claims to express quantities, composition, distances, or other measures should be understood to be modified by the term “about,” as defined herein. Therefore, unless otherwise indicated, the numerical parameters set forth in the specification and appended claims are approximations that may vary depending on the desired characteristics sought to be obtained from the subject matter presented herein. To a minimum, and without attempting to limit the application of the equivalence principle to the scope of the claims, each numerical parameter should be interpreted at least according to the number of significant figures reported and by applying general rounding techniques. Although the numerical ranges and parameters that articulate the broad scope of the subject matter presented herein are approximations, the values ​​set forth in particular examples are reported as precisely as possible. However, any numerical value inherently contains some error that is necessarily due to the standard deviation present in its corresponding test measurement.

[0045] The headings and subheadings used herein are for organizational purposes only and are not intended to limit the scope of this specification or claims.

[0046] Overview of Imaging Systems and Methods

[0047] Figure 1A and Figure 1BAn upright optical microscope 10 is shown. Figure 1A ) and inverted optical microscope 20 ( Figure 1B The prior art embodiments of the optical microscopes 10 and 20 shown include a sample stage 30 and an objective lens 40 aligned with the optical axis of the respective microscope 10 or 20. Each optical microscope 10 or 20 is associated with multiple objectives on a turntable for tissue imaging, the turntable being rotatable to align different objectives with the optical axis of the microscope. Both optical microscopes are capable of imaging samples using both incident and projected illumination, but... Figure 1A Orthogonal optical microscope 10 and Figure 1B One of the main differences between the inverted optical microscopes 20 is the position of the objective lens relative to the sample stage. Figure 1A The upright optical microscope 10 includes an objective stage positioned above the sample stage, while Figure 1B In the inverted optical microscope 20, the corresponding objective stage is positioned below the sample stage. This largely affects the type of sample observed with high-resolution objectives using each microscope.

[0048] For example, when observing samples using immersion objectives of upright or inverted optical microscopes, it is advantageous to directly observe the desired focal plane of the sample through the immersion medium without intervening air space and / or to minimize the different types of materials through which light passes before being received by the microscope's optical system. Therefore, samples within microplates are generally best observed / imaged using an inverted optical microscope, while cross-sectional tissue samples on slides with coverslips are best observed using an upright optical microscope.

[0049] Regardless of sample type or viewing angle, upright and inverted microscopes are connected via a common thread, and using immersion objectives on these platforms traditionally requires manually applying the immersion medium to the objective. For example, when applying the immersion medium to an inverted optical microscope (e.g., Figure 1B When applying the objective lens to an inverted optical microscope (20), the user can manually lower the objective stage (or partially rotate the stage to access the objective), apply the immersion medium, and return the objective to the focal plane, where the immersion medium bridges the gap between the objective surface and the glass surface that separates the immersion medium from the sample. Similarly, when applying the immersion medium to image the sample at high resolution on an upright microscope, the stage can be partially rotated to access the axially aligned high-resolution immersion objective below which the immersion medium can be deposited (or other suitable imaging surface between the sample and the objective). Rotating the immersion objective above the sample to an axially aligned position will cause the deposited immersion medium to form an immersion medium layer between the coverslip and the objective.

[0050] As illustrated above, manually applying immersion medium to observe a single sample is a time-consuming process and is best suited for open access to the objective stage. In automated imaging systems, each sample can be imaged at multiple different levels of resolution, which may require repeated changes between multiple objectives during the imaging process. For example, if such an imaging process requires the tandem use of two different immersion objectives, current systems lack an efficient and effective solution for applying and / or maintaining an appropriate volume of immersion medium on each objective between rotations. Furthermore, continuous or excessive application of immersion medium can cause the objectives to become greasy and / or may negatively impact the consistency and quality of the acquired images. Current systems also lack the capability for automated removal and / or cleaning of the immersion medium from the objectives.

[0051] The embodiments of this disclosure advantageously provide systems and methods for automatically applying and cleaning immersion media from objectives, and have particularly beneficial effects when applied to automated imaging systems. For example, in some embodiments disclosed herein, an immersion media applicator is integrated within the sample stage of an automated system. By moving the sample stage above the desired immersion objective such that the applicator's outlet nozzle is oriented above the lens surface, where it can discharge discrete volumes of immersion media, the immersion media can be applied to the desired objective. When integrated into the sample stage itself, very little additional space (if any) is required within the stage assembly area to implement the applicator. The immersion media reservoir associated with the nozzle can be housed outside the sample stage assembly area and pulled there via a flexible hose. In this way, embodiments of this disclosure advantageously allow the retrofitting of most automated imaging systems without affecting the functionality or mobility of existing components, and can advantageously reduce the mechanical complexity (and associated costs) associated with existing systems that require additional motors or flip mounts to move the nozzle on top of the objective. Furthermore, embodiments of this disclosure advantageously allow the application of an immersion medium into any objective lens within an automated imaging system that can be used to image a sample held by a sample stage, regardless of whether the objective lens is held by a turntable or an objective lens slider.

[0052] In some embodiments of this disclosure, the immersion medium applicator comprises a static hose and a nozzle extending into the stage assembly area of ​​the automated imaging system. Alternatively, by positioning the sample stage above a fixed objective lens for immersion medium application, the objective lens is moved to the applicator (e.g., via an objective lens slider), in which case the immersion medium can be applied directly to the desired objective lens. After automatically receiving the immersion medium, the objective lens can be repositioned onto the sample for high-resolution imaging. Additional features and beneficial effects of the disclosed system are provided herein with reference to the embodiments disclosed in the accompanying drawings.

[0053] For example, Figure 2This is a general schematic diagram of an exemplary system 100 incorporating features disclosed or contemplated herein. At the heart of system 100 is an imaging system 102, in which samples such as biological cells are imaged and analyzed. The exemplary imaging system 102 includes, but is not limited to, an optical microscope assembly 104 and a computing device 110. Within the optical microscope assembly 104 is an image sensor (e.g., any CCD or CMOS sensor array or chip, or other sensors as known in the art), configured to capture image data from a sample located within the field of view of the image sensor.

[0054] like Figure 2 As shown, the stage housing 106 can be mounted on or otherwise associated with the optical microscope assembly 104 to facilitate positioning the sample 108 in alignment with the optical system of the optical microscope assembly 104. The sample can be contained within or mounted on any sample receiving device, including, for example, a microscope slide 108a, a multi-well plate (e.g., Figure 2 The 96-well plate 108b shown is an example of this. Therefore, the stage housing 106 may contain one or more light sources to illuminate the sample 108, which may be, for example, white light or light of a defined wavelength. It should be understood that in some embodiments, the light source is contained within the microscope assembly 104. In embodiments where the light emitter contains a fluorophore, the light source may contain a fluorophore excitation light source. For example, the stage housing 106 may contain a light engine comprising multiple light-emitting diodes (LEDs) or lasers configured to emit white light and / or an excitation wavelength for exciting fluorophores within the sample 108. Additionally or alternatively, the stage housing 106 may contain filters for filtering the excitation and emission light, such as multi-position dichroic filters and / or multi-position emission filters.

[0055] As a general working example, a multi-well plate containing the sample can be positioned within the stage housing 106 such that the desired sample aperture is optically aligned with the optical system of the associated optical microscope assembly 104 containing the desired objective. The objective can be switched to a lower or higher resolution objective (e.g., by rotating the associated turntable or repositioning the objective via an objective slider) and the sample can be illuminated by a white light source.

[0056] As another example, the fluorophore excitation source can be automatically or manually directed to provide light with multiple bandwidths from violet (e.g., 380 nm) to near-infrared (e.g., at least 700 nm) and is designed to excite fluorophores such as, for example, cyan fluorescent protein (CFP) and far-infrared (i.e., near-infrared) fluorophores. Example bandwidths with appropriate excitation filters (e.g., excitation filter wheel selection driven by computing device 110) may include, but are not limited to, violet (e.g., 380–410nm LED and 386 / 23nm excitation filter), blue (e.g., 420–455nm LED and 438 / 24nm excitation filter), cyan (e.g., 460–490nm LED and 485 / 20nm excitation filter), green (e.g., 535–600nm LED and 549 / 15nm excitation filter), red (e.g., 620–750nm LED and 650 / 13nm excitation filter), and near-infrared (e.g., 700nm-IR LED & 740 / 13nm excitation filter). When it is desired to increase the brightness of red and scarlet dyes, the two green / excitation filter combinations listed above can optionally be provided via, for example, a mechanical flipper. Alternatively, a laser emitting any desired excitation bandwidth and / or wavelength can be used to utilize, replace, or supplement other LED bandwidths.

[0057] Additionally or alternatively, the stage housing 106 may include a stage assembly and positioning mechanisms configured to hold and selectively move the sample for objective observation aligned with the remainder of the optical system within the optical microscope assembly 104. It should be understood that the stage assembly may be configured to move in any of the three dimensions, as known in the art. For example, the stage assembly may be configured to move laterally (e.g., in x, y planes parallel to the associated objective surface) to position different portions of the sample within the field of view. Additionally or alternatively, the stage assembly may be configured to move in the z-direction using any mechanism known in the art (e.g., between parallel x, y planes each positioned at different distances from the objective surface), such as, for example, using a stepper motor and screw / nut combination to provide progressive movement of the sample toward / away from the objective.

[0058] The stage assembly can be moved to position the desired sample within the focal plane of the optical microscope assembly 104, and after image data is captured at the optical microscope assembly 104, the data can be viewed, analyzed, and / or stored in the associated computing device 110. Therefore, the embodiments disclosed or contemplated herein may include or utilize a dedicated or general-purpose computer containing computer hardware (such as, for example, one or more processors), as discussed in more detail below. Embodiments may also include physical media and other computer-readable media for carrying or storing computer-executable instructions and / or data structures. Such computer-readable media may be any available media accessible by a general-purpose or dedicated computer system. A computer-readable medium storing computer-executable instructions is a physical storage medium. A computer-readable medium carrying computer-executable instructions is a transmission medium. Therefore, by way of example and not limitation, embodiments may include at least two distinctly different kinds of computer-readable media: computer storage media and transmission media.

[0059] Computer storage media include RAM, ROM, EEPROM, CD-ROM or other optical disk storage devices, magnetic disk storage devices or other magnetic storage devices, or any other medium that can be used to store desired program code in the form of computer-executable instructions or data structures and that can be accessed by a general-purpose or special-purpose computer. A “network” is defined as one or more data links that enable the transmission of electronic data between computer systems and / or modules and / or other electronic devices. When information is transmitted or provided to a computer via a network or other communication connection (hardwired, wireless, or a combination of hardwired and wireless), the computer appropriately regards the connection as a transmission medium. Transmission media may include networks and / or data links that can be used to carry data or desired program code in the form of computer-executable instructions or data structures and that can be accessed by a general-purpose or special-purpose computer. Combinations of the foregoing should also be included within the scope of computer-readable media.

[0060] Furthermore, upon arrival at various computer system components, program code in the form of computer-executable instructions or data structures can be automatically transferred from the transmission medium to the computer storage medium (and vice versa). For example, computer-executable instructions or data structures received via a network or data link can be buffered in the RAM within a network interface module (e.g., a "NIC") and then ultimately transferred to the computer system RAM and / or a less volatile computer storage medium on the computer system. Therefore, it should be understood that computer storage media can be included within computer system components that also (or even primarily) utilize the transmission medium.

[0061] Computer-executable instructions include, for example, instructions and data that cause a general-purpose computer, special-purpose computer, or special-purpose processing device to perform a function or a set of functions. Computer-executable instructions can be, for example, binary, intermediate format instructions (such as assembly language), or even source code. Although the subject matter has been described in language specific to structural features and / or methodological actions, it should be understood that the subject matter defined in the appended claims is not necessarily limited to the features or actions described above. Rather, the described features and actions are disclosed as examples of the claims.

[0062] Those skilled in the art will understand that embodiments can be practiced in network computing environments with many types of computer system configurations, including personal computers, desktop computers, laptop computers, message processors, handheld devices, multiprocessor systems, microprocessor-based or programmable consumer electronics, network PCs, minicomputers, mainframe computers, tablets, smartphones, routers, switches, etc. Embodiments can also be practiced in distributed system environments where both local and remote computer systems perform tasks via network links (via wired data links, wireless data links, or a combination of wired and wireless data links). In a distributed system environment, program modules can be located in both local and remote memory storage devices. A program module of one entity can be located and / or run in another entity's data center or "cloud".

[0063] Continue to refer to Figure 2 In system 100, computing device 110 can also serve as a controller for system 100, and, alone or in combination with optical microscope assembly 104, for performing actions such as determining the number and resolution of images acquired for each sample, the imaging path employed, objectives for imaging various aspects of the sample, automatically applying immersion medium to the objectives, and / or automatically cleaning the immersion medium from the objectives. The computing device can also be used for image analysis and / or storing images and data acquired by optical microscope assembly 104. Computing device 110 may include a general-purpose or special-purpose computer or server as defined herein, or any other computerized device. As is known in the art, computing device 110 may communicate directly with optical microscope assembly 104 or via a network. In some embodiments, computing device 110 is integrated into optical microscope assembly 104. In some embodiments, computing device is integrated within optical microscope assembly.

[0064] System 100 may also include a user display device 112 to display results and / or system configuration. The optical microscope assembly 104 and / or computing device 110 may communicate directly or indirectly with the user display device 112 to program and / or control automated imaging methods, which may include, for example, automatically applying an immersion medium to the appropriate immersion objective before and / or during the imaging method.

[0065] In one embodiment, one or more method steps described herein are executed as a software application. However, the embodiments are not limited thereto, and the method steps may also be executed in firmware, hardware, or a combination of firmware, hardware, and / or software. Furthermore, the steps of the method may be present, in whole or in part, on the optical microscope assembly 104, the computing device 110, and / or other computing devices.

[0066] The operating environment of the apparatus for the system may include or utilize a processing system having one or more microprocessors and system memory. According to the practice of those skilled in the art of computer programming, unless otherwise indicated, the embodiments are described below with reference to the actions and symbolic representations of operations or instructions performed by the processing system. Such actions and operations or instructions are referred to as “computer-executed,” “CPU-executed,” or “processor-executed.”

[0067] The various embodiments disclosed herein relate to apparatus, methods, and systems for applying immersion media and cleaning lenses. Such embodiments advantageously improve microscopy systems by enabling the automatic application of immersion media to objectives before and / or during automated imaging operations. These embodiments may additionally or alternatively provide automated lens cleaning systems that advantageously remove immersion media and / or clean objectives at any time before, during, and / or after imaging a sample using immersion media, thereby improving the consistency and / or quality of images acquired by the relevant imaging system and reducing the likelihood of damage to objectives due to prolonged or repeated exposure to immersion media when not in use. The various embodiments can also be readily incorporated into existing imaging systems (e.g., at the original equipment manufacturer and / or as a modification of an already manufactured imaging system) without significantly impeding the movement and / or operation of system components, and advantageously, the small footprint of the disclosed immersion media applicator and the disclosed lens cleaner—besides their position and operation within the imaging system—enhances the utilization of objectives and imaging modalities.

[0068] Furthermore, the embodiments disclosed herein can improve the efficiency and convenience of applying and cleaning immersion media from and from objectives housed within automated imaging systems that may be difficult or time-consuming to access. Therefore, the embodiments disclosed herein provide researchers and other imaging system operators with additional flexibility when planning or implementing automated (e.g., high-throughput) image acquisition processes. For example, the immersion media applicator disclosed herein can be configured to easily apply immersion media to any number (e.g., some or all) of objectives while maintaining or returning the desired field of view with extremely high precision and accuracy. The embodiments disclosed herein can also advantageously reduce focus drift because the immersion media is in the same incubator / controlled environment as the objectives prior to its application.

[0069] Longer image capture events (e.g., hours or days) can also be achieved with less time spent maintaining the immersion medium interface between the objective and the sample, and it is advantageous to apply and / or clean the immersion medium on multiple objectives throughout the longer image capture event, reducing the amount or duration of imaging interruptions.

[0070] Exemplary immersion media applicators and systems

[0071] Now for reference Figure 3 , showed Figure 2 An exemplary embodiment of the imaging system 102 disclosed herein. The internal platform design of the exemplary system 102 is in... Figure 3 The image is shown as a cross-sectional side view. Typically, the imaging system 102 integrates the components needed to position a sample (e.g., a porous sample plate containing biological cells) for automated imaging.

[0072] The stage housing 106 includes a stage assembly 114 mounted in a manner that cooperates optically and mechanically with components constituting the microscope assembly 104. The stage assembly 114 typically includes a stage 116 on which a sample 108 can be positioned, and may include stage positioning mechanisms for selectively moving the stage in the xy-plane to observe the sample positioned thereon, as known in the art. In some embodiments, the stage housing is a stage assembly. Thus, as used herein, “stage housing” is intended to encompass a microscope sample stage for holding and / or positioning a sample to be imaged. The term may also be used to describe additional features associated with the stage, including, for example, elements for controlling environmental conditions around the sample, such as heating elements, cooling elements, fans, gas sensors / inlets (e.g., oxygen, carbon dioxide, etc.), vacuum, compressors, or any one or more other elements or physical housings associated with or coupled to the sample stage.

[0073] In the depicted embodiment, microscope assembly 104 houses an inverted microscope for sifting samples on sample plate 108b from below. The microscope includes objective assembly 118 containing multiple objectives, as known in the art, to obtain magnified views of the sample. In one embodiment, one or more standard objectives comprise one or more immersion objectives. Example standard objectives include 2X / 0.08NA, 4X / 0.16NA, 10X / 0.4NA, 20X / 0.45NA, 20X / 0.7NA, and 40X / 0.6NA objectives. Any one or more of the above standard objectives may be contained on a turntable or lens slider with any one or more of the following immersion objectives: 40X / 1.3NA, 50X / 0.95NA, 60X / 1.25NA, 100X / 1.25NA, 100X / 1.28NA, 100X / 1.3NA, and 100X / 1.4NA objectives. It should be understood that, depending on the operator’s preference and / or application, any number or combination of objectives (including other magnification levels and objective types known in the art) may also be used in the embodiments of this disclosure.

[0074] The microscope also includes a focusing drive mechanism 120 mechanically coupled to the microscope objective assembly 118. The objective assembly 118 can be moved up and down relative to the stage assembly 114 via the focusing drive mechanism 120 to align and focus any objective of the microscope objective assembly 118 onto biological cells disposed within the sample plate 108b. The focusing drive mechanism 120 may be an autofocus mechanism, although this is not required. The focusing drive mechanism 120 may be configured with a stepper motor and screw / nut combination that reduces backflash to provide, for example, a resolution as low as 0.006 μm / microstep to support the microscope objectives configured in the imaging system 102.

[0075] The stage assembly 114 further includes an immersion medium applicator 122 associated with the stage 116 for applying immersion medium to the objectives in the objective assembly 118. As an example embodiment illustrating the objective operation of the imaging system 102, the objective assembly 118 can be configured in a customized manner to provide multiple positions that enable the querying of cells within the sample plate 108b. The focusing drive mechanism 120 can automatically and reliably switch between objectives. When on a turntable, the objectives in this arrangement can, but do not necessarily, be positioned 60 degrees apart, which allows the primary objective to focus on the sample plate 108b without the other objectives interfering with the stage, sample plate 108b, or other components within the imaging system 102.

[0076] To change the objective lens, the focusing drive mechanism 120 can descend below the stage assembly 114, rotate to the next objective lens position, and then push the objective lens upward to the appropriate focusing height. For enhanced system safety, mechanical limit switches can be used to position the turntable, while one or more optical switches can be used to confirm that the objective lens position has been correctly switched. Furthermore, each optical position can be held in place by precisely machined mechanical pawls on the turntable.

[0077] When switching to an immersion objective that requires the application of an immersion medium before imaging, the focusing drive mechanism 120 lowers the objective below the stage. The stage assembly positions the applicator 122 above the objective and applies a predetermined volume of immersion medium to the immersion objective. The stage assembly can also reposition the sample 108 relative to the objective assembly 118 in the observation position before applying the immersion medium. The focusing drive mechanism 120 then positions the immersion objective at the appropriate focusing height for imaging.

[0078] The microscope assembly 104 also includes various known components for generating and / or recording images of the sample. These components may include, but are not limited to, an image sensor 124 (e.g., a monochrome CCD or CMOS camera or sensor), a light source 126 (e.g., a light engine comprising multiple LEDs), filters for filtering excitation and emission light (e.g., a multi-position dichroic filter wheel 128 and a multi-position emission filter wheel 130), and light guiding devices for guiding light through the microscope assembly (e.g., a tube lens 132 and a folding mirror 134). One or more of the aforementioned components are typically controlled by a computing device 110 to allow for automated imaging.

[0079] Microscope assembly 104 allows for both incident (or reflected) light microscopy and transmitted light microscopy. In incident illumination, light (e.g., white light) generated by light source 126 is projected along optical path 136 through optical components, where it is focused onto and illuminates sample 108. Reflected light is received at the objective and returns along reflected light path 138 to image sensor 124. Alternatively, transmitted white light may be generated by transmitted light assembly 140 for bright-field imaging. Light generated by transmitted light assembly 140 passes through the sample and is received at the objective of microscope assembly 104. The light travels along optical path 138 through assembly 104 until it is received at image sensor 124.

[0080] Although this discussion focuses on the use of inverted microscope configurations, it should be understood that upright microscope configurations can be used alternatively for screening from above the sample.

[0081] Immersion media applicators can be implemented in various ways. For example, Figure 4A and Figure 4BAn exemplary applicator assembly 200 according to embodiments of the present disclosure is shown for use in applying an immersion medium to an objective lens in an automated imaging system. The applicator assembly 200 may include a nozzle 202 for dispensing the immersion medium, fluidly coupled via a hose 206 to a reservoir 204 of the immersion medium. The immersion medium can be pumped from the reservoir 204 to the nozzle 202 via a line pump 208 configured to deliver a predetermined, small volume of immersion medium (e.g., 1-50 μL) in each cycle. It should be understood that the pump 208 can be calibrated and / or operable to handle various types and viscosities of immersion media. In some embodiments, such as Figure 4A In the embodiment shown, the applicator assembly 200 further includes a one-way check valve 210 to prevent the pumped immersion medium from flowing back down the hose 206 into the reservoir 204 between application cycles. Valve 210 can also be used to maintain pressure in the hose 206, thereby maintaining the immersion medium at the nozzle 202 between application cycles and / or preventing air bubbles from forming within the hose.

[0082] Therefore, the applicator assembly 200 can be configured to dispense an immersion medium free of air bubbles. In some embodiments, the applicator assembly includes a sensor 212 located at a distal end of the assembly 200. The sensor 212 may be a bubble sensor for detecting the presence of air bubbles at the immersion medium nozzle 202 or in the upstream hose 206 supplying the immersion medium to the nozzle 202. Additionally or alternatively, the sensor 212 may be a liquid sensor for detecting the presence of immersion medium at the nozzle 202. For example, the sensor 212 may be any of a capacitive sensor, an optical sensor, or a multimeter measuring the resistance at the dispensing tip of the nozzle 202. In operation, the sensor 212 may initially record whether liquid is present at the nozzle 202. If present, the pump 208 may be activated for a predetermined number of cycles and / or time periods to deliver a known volume of immersion medium (e.g., based on the type of pump, the diameter of the nozzle and hose, and the type / viscosity of the immersion medium being dispensed). Alternatively, if the sensor 212 does not record liquid at the nozzle 202, the pump 208 may be activated until the sensor 212 indicates the presence of liquid. As described above, once the sensor 212 records the liquid at the nozzle 202, the pump 208 can be activated for the required time and / or number of cycles to dispense the desired volume of immersion medium.

[0083] In some embodiments, pumping the immersion medium through the hose until the sensor records liquid can be performed by positioning the nozzle above the waste reservoir 214, thereby preventing any immersion medium from being accidentally discharged into the interior of the microscope assembly. Similarly, in embodiments where the applicator assembly includes a bubble sensor, the system is operable to clear the tubing into the waste reservoir 214 to ensure bubble-free tubing.

[0084] Continue to refer to Figure 4Aand Figure 4B For example, the volume of the submerged medium in the reservoir 204 can be monitored by a level detector 215a, which is operable to alert the relevant computing system and / or user that the volume of the submerged medium in the reservoir 204 has reached or decreased below a predefined lower threshold. In some embodiments, if the level detector 215a indicates that the volume of the submerged medium in the reservoir 204 has reached or decreased below a predefined lower threshold, the system can stop any current and / or additional automatic imaging until the submerged medium reservoir 204 is replenished above the predetermined lower threshold (e.g., as confirmed by the level detector 215a).

[0085] Similarly, waste reservoir 214 may be associated with a level detector 215b, operable to monitor and / or identify when the volume of waste within waste reservoir 214 meets or exceeds a predetermined upper limit threshold. Level detector 215b can alert the user and / or associated computing system that the waste reservoir is “full” and needs to be emptied of its liquid contents, and in some embodiments, can prevent the applicator from dispensing immersion medium into the waste reservoir until level detector 215b indicates that the volume of immersion medium within waste reservoir 214 has dropped below the upper limit threshold. This advantageously prevents immersion medium from overflowing from waste reservoir 214, thereby advantageously protecting components from potential damage due to exposure to the immersion medium.

[0086] It should be understood that the applicator assembly may be equipped with software or other computer-executable instruction sets for configuring a computing system to operate and / or communicate with liquid sensors, level detectors, nozzles, valves, and / or pumps. In this way, the applicator assembly can be integrated with an automated imaging system, thereby allowing the appropriate application of immersion media onto a desired objective lens. In some embodiments, the applicator assembly itself may include the operability necessary for communication with discrete components of the assembly and between discrete parts of the assembly to achieve any and / or all functions and operations of the assembly (or any individual part thereof), as disclosed herein.

[0087] In some embodiments, the applicator assembly is stationary within the associated automated imaging assembly. For example, the nozzle 202 of the applicator assembly 200 may be positioned along the path of the objective lens slider 216 (e.g., as shown in the image). Figure 4A (As indicated by arrow A in the diagram), allowing a volume of immersion medium 218 to be applied to any of the objectives 220a, 220b, 220c by selectively positioning the desired objective below nozzle 202. In some embodiments, the associated focusing drive mechanism (217) can control the z-direction of the objective slider 216 or the individual objectives held by the objective slider 216 (e.g., as shown by arrow A in the diagram), making it possible to apply a volume of immersion medium 218 to any of the objectives 220a, 220b, 220c by selectively positioning the desired objective below nozzle 202. Figure 4B(As indicated by arrow B in the image) to ensure that the desired objective is below nozzle 202 and / or to lift the objective to the nozzle to obtain the immersion medium when moving toward or away from it.

[0088] Continue to refer to Figure 4B The objective slider 216 may additionally include objectives 220a, 220b, 220c positioned on a turntable to allow them to rotate (e.g., as indicated by arrow C). In such an embodiment, the slider 216 may be positioned laterally and vertically to receive a quantity of immersion medium from the nozzle 202, and may rotate the objectives to the nozzle such that the objective receiving the immersion medium is positioned closest to the nozzle compared to another objective on the turntable.

[0089] While having a fixed nozzle position can advantageously reduce the risk of tangling or breaking of the operable hose, the repositioning accuracy of a moving objective is generally worse than that of a moving sample stage, resulting in unexpected shifts in the image field of view. Therefore, in some embodiments, the nozzle and hose can be integrated into the sample stage itself.

[0090] For example, such as Figures 5A to 5C As shown, the applicator assembly 300 may include a nozzle 302 integrated with the stage 304, such that the opening of the nozzle 302 faces the back of the stage 304 in the direction of the objective lens 308 (i.e., in an inverted microscope arrangement). A flexible tube 306 may be located within a channel formed within the sample stage 304 and guided to the exterior (not shown) of the assembly containing the immersion medium reservoir. In this configuration, the flexible tube 306 may have some additional slack to accommodate movement of the sample stage within the stage housing during sample reading and / or application of the immersion medium to the objective lens.

[0091] For example, such as Figure 5A As shown, a channel is formed within the sample stage 304 and provides an inconspicuous, defined path for the hose 306. In this configuration, the hose is advantageously protected from tangling or accidental damage when the user places or removes the sample for imaging. The nozzle 302 is positioned on the sample stage 304 such that the immersion lens 308 can access it within the normal operating / movement parameters of the sample stage and / or lens slider (or similar lens positioning device). In some embodiments, the nozzle 302 is positioned such that it can interact with the lens surface of each immersion lens on the associated lens slider (or other lens positioning device within the imaging system).

[0092] It should be understood that Figure 5AThe location of nozzle 302 may additionally include space for a wiping or suction device operable to clean the lens surface of the immersed lenses. As described above, the wiping or suction device is preferably positioned on the sample stage such that it can approach the lens surface of each immersed lens on the associated lens slider (or similar lens positioning device, such as a lens turntable) using normal operating / movement parameters of the imaging system. In some embodiments, a second channel is formed in the stage and may be fitted with one or more hoses for adding or removing washing solution from the immersed objectives.

[0093] Figure 5B Another embodiment of an applicator assembly 300 with a hose 306 integrated within a sample stage 304 is shown. A dedicated sample stage can be configured to retrofit an existing imaging system or can be incorporated into it during the original manufacturing process. As shown, the hose connects to an applicator (e.g., nozzle 302) for dispensing immersion media to the immersion objective. Figure 5B The modified sample stage 304 can be configured to dispense immersion medium onto an objective lens located below the stage (e.g., by protruding nozzles through holes formed in the bottom surface of the sample stage). Alternatively, the applicator can be oriented to dispense immersion medium onto an overhead immersion objective lens (e.g., by positioning the nozzles in the direction of the opening of a channel formed in the sample stage).

[0094] As by Figure 5B As further illustrated by the embodiments herein, the applicator assembly disclosed herein may include a wider diameter portion 307 of a hose 306 upstream of the applicator (e.g., nozzle 302) to prevent or reduce bubble formation while dispensing the immersion medium. The wider diameter portion 307 advantageously allows any air within the hose to be trapped therein and not to advance through the hose to the nozzle, which could easily cause bubbles or improper medium application. Furthermore, in some embodiments, such as Figure 5B The embodiment shown allows the wider diameter portion to be positioned for easy observation by the user or operator of the imaging system. Therefore, the operator can monitor for trapped air or other contaminants in the hose and take appropriate measures without affecting the experiment.

[0095] For details, please refer to the following: Figure 5C Components of an exemplary immersion medium applicator system are shown in a close-up view. For clarity, the portion of the stage 304 incorporating the applicator has been removed from the view. However, it should be understood that the immersion medium applicator system disclosed herein can be additionally mounted on the stage (similar to...). Figure 5C (As shown), rather than being integrally integrated into the stage. However, as... Figure 5CAs shown, an exemplary immersion medium applicator system may include a nozzle 302 oriented in the direction of the objective lens—in this case, facing the bottom surface of the stage 304—such that the nozzle 302 can selectively engage the objective lens of an inverted microscope system. The immersion medium applicator system may additionally include a sensor 303 for detecting the presence of immersion medium at the nozzle 302. In the exemplary embodiment shown, the sensor 303 includes a test lead electrically connected to a PCB 305 configured to measure the resistance between the nozzle and the test lead. Simultaneous contact of the immersion medium with both the (conductive) nozzle and the test lead reduces the resistance, indicating that the immersion medium is being dispensed from the nozzle. It should be understood that... Figure 5C The sensor 303 shown is illustrative, and other liquid sensors may be used within the scope of this disclosure.

[0096] In some embodiments, the objective lens may be positioned on a turntable that can rotate various lenses into the optical path of the microscope assembly. The objective lens on the turntable can be positioned in the correct focal plane by a focusing drive mechanism, but may be stationary relative to lateral movement (e.g., in the x and y directions). In such embodiments, the sample stage may be responsible for positioning the applicator nozzle in the correct xy coordinates to apply the immersion medium onto the objective lens. It should be understood that by limiting the movement of the objective lens, the repositioning accuracy of the sample field of view can be maximized compared to the repositioning accuracy when moving the objective lens.

[0097] In some embodiments, one or both of the stage and objective lens may be translated in the xy plane to orient the immersion medium applicator at a position where the immersion medium can be properly dispensed onto the desired objective lens. For example, as Figure 6A and Figure 6B As shown, a porous plate 310 containing a sample is held by a stage 304. The stage 304 includes an immersion medium applicator formed therein, with a nozzle 302 exposed on its underside, oriented toward the objective lens. In one embodiment, the stage 304 is moved (e.g., as...). Figure 6A (As indicated by arrow D in the diagram) to position the nozzle 302 above the desired immersion objective 312, as shown. Figure 6A As shown. A predetermined volume 314 of immersion medium is dispensed from nozzle 302 and applied to immersion objective lens 312. Stage 304 can then be repositioned to the desired sample for imaging (e.g., such as...). Figure 6B (As indicated by arrow D in the diagram), where the immersion medium forms an immersion layer between the immersion objective 312 and the sample aperture 316.

[0098] Additionally or alternatively, the objective lens can be moved relative to the sample stage to retrieve the immersion medium and return to an approximate field of view for imaging. For example, as Figure 6AAs shown, the objective slider 318 can be moved below the nozzle 302 of the immersion medium applicator (e.g., as indicated by arrow E) to receive a predetermined volume of immersion medium on the desired immersion objective 312. The objective slider 318 can then be repositioned to the sample aperture 316 (e.g., as indicated by arrow E). Figure 6B (As indicated by arrow E in the diagram) to position the immersion objective 312 at an approximate field of view for high-resolution imaging. Figure 6B As shown, the aligned immersion objective 312 can be moved to a position for observing the sample 316 via a z-motor driven objective holder 317. In some embodiments, both the stage and the objective lens move relative to each other during immersion medium application and / or objective lens and sample repositioning for imaging.

[0099] It should be understood that the aforementioned movement of the stage and / or lens can be achieved automatically, and the operator does not need to physically interact with the objective lens. Therefore, embodiments of this disclosure advantageously enable the automatic application of immersion medium to one or more objectives in an automated imaging system.

[0100] Continue to refer to Figure 6A and Figure 6B The system may additionally include a waste reservoir 313, positioned (e.g., on the objective lens slider or at a fixed location within the imaging system) such that an immersion media applicator can dispense media or dispense it into the reservoir 313. The reservoir may be equipped with a funnel or the like to guide the dispensed media into the reservoir 313. In some embodiments, the applicator may dispense a cleaning solution for cleaning the immersion objective lens and may additionally be equipped with a wiping or suction device for removing the dispensed cleaning solution from the lens surface. The waste reservoir 313 may be used to receive disposable wiping and / or may be connected to a suction device to receive aspirated media / washing solution from the lens surface of the immersion objective lens and / or directly from the applicator / suction device. In some embodiments, the waste reservoir 313 may be associated with a fill sensor 315 for monitoring the waste level in the reservoir 313 and may signal or otherwise indicate when the reservoir should be emptied.

[0101] It should be further understood that, although Figure 4A , Figure 4B , Figure 6A and Figure 6B The objective lens is shown as being oriented for an inverted microscope, but the embodiments disclosed herein can also be configured for use with an upright microscope system in which the objective lens is oriented above the sample to be observed.

[0102] An example system of this kind is Figure 7 As shown in the schematic diagram. The system includes an objective lens holder 320 on which an objective lens 322 is disposed. In some embodiments, such as Figure 7In the illustrated embodiment, the objective lens holder 320 includes a plurality of objectives disposed thereon. The objective lens holder 320 may be stationary relative to the stage housing 324, or in some embodiments, the objective lens holder 320 may be movable relative to the stage housing 324 in one or more of the x, y, and / or z directions. In some embodiments, the stage housing 324 may be laterally movable relative to the objective lens holder 320 (e.g., in the xy plane indicated by at least arrow F) and may additionally be movable relative to the objective lens holder 320 in the z direction (e.g., in the z plane indicated by at least arrow G). Thus, the stage housing 324 may be moved to selectively position the applicator nozzle 326 below the desired immersion objective (e.g., objective lens 322). The nozzle 326 can then be used with, for example, a tubular micropump and / or a liquid sensor (e.g., as mentioned above). Figure 4A and Figure 4B The desired volume of immersion medium 328 is dispensed onto the immersion objective lens (as shown and discussed) or through any other embodiments disclosed and / or conceived within the scope of this specification. The stage housing 324 can then be repositioned relative to the objective lens 322 such that the immersion medium contacts the imaging surface and forms an immersion layer therebetween for immersion imaging of the sample. Figure 7 As shown, the glass cover glass 330 covering the sample loaded onto the microscope slide 332 can be the desired imaging surface of the upright objective lens configuration shown.

[0103] Exemplary self-cleaning mirror system

[0104] In addition to the foregoing, embodiments of this disclosure further include systems for removing immersion media from objectives in automated imaging systems and / or for cleaning objectives in automated imaging systems. For example, Figure 8 A schematic diagram of an exemplary lens cleaning apparatus 400 is shown, comprising a wipe 402 configured to clean and / or remove immersion medium 404 from the lens surface of an immersion objective 406. As shown, the wipe 402 is associated with a stage 408 and can be positioned relative to the objective as described above with respect to nozzle positioning in embodiments having an immersion medium applicator. In some embodiments, the wipe may be dried lens paper for absorbing immersion medium from the lens. In some embodiments, the wipe contains a cleaning agent suitable for cleaning lens surfaces, as known in the art.

[0105] In some embodiments, the topology of the wipe is mapped and tracked by a computing system, such that clean or unused areas of the wipe are used to clean each subsequent objective. During the cleaning process, the objective can move linearly back and forth or rotate within a plane. In some embodiments, the wipe is associated with a displacement element (such as a solenoid) that provides a vibrational amplitude for the wipe's movement. For example, this movement can be achieved by vibrating or moving the sample stage.

[0106] This disclosure envisions alternative cleaning systems. As Figure 8 As a supplement or alternative to the wiping materials disclosed herein, the embodiments disclosed herein may include other exemplary lens cleaning devices. For example, Figure 9 A schematic diagram 410 shows a suction device 412 configured to remove the immersion medium 414 from the lens surface of the immersion objective 416. The suction device can be located in a fixed position within the imaging system housing, or as... Figure 9 As shown, the suction device 412 can be mounted on the sample stage 418 and can be positioned relative to the objective lens, as described above regarding nozzle positioning in an embodiment with an immersion medium applicator.

[0107] In some embodiments, the applicator and cleaning system can be combined into a single and / or collaborative system. For example... Figure 10 An exemplary dispensing and lens cleaning system is illustrated, comprising an immersion medium dispensing nozzle 420 and a suction device 422. As discussed herein, the dispensing nozzle 420 may be coupled to an immersion medium reservoir and operable to dispense immersion medium onto an immersion objective lens 424. Additionally or alternatively, the dispensing nozzle 420 may be coupled to a lens cleaning agent reservoir and operable to dispense cleaning agent onto an immersion objective lens 424. The suction device 422 may be positioned opposite the dispensing nozzle 420 (e.g., as shown in the image). Figure 10 (as shown), but it should be understood that the suction device 422 can be positioned at other locations on the associated stage 426 relative to the dispensing nozzle 420. When as shown Figure 10 As shown in the positioning diagram, the immersion medium and / or lens cleaner can be used to clean the immersion objective 424 by dispensing immersion medium and / or lens cleaner from nozzle 420 while removing the dispensed medium and / or cleaner flowing on the lens surface via suction device 422. This also advantageously allows for the removal of air bubbles from the immersion medium hose without moving nozzle 420 to a waste reservoir to clean the line. Alternatively, the line can be cleaned at the immersion objective. Any air bubbles within the line and dispensed onto the lens surface can be removed by suction device 422 and / or by an additional flow of immersion medium / cleaner on the lens surface, which is subsequently removed from the lens surface by suction device 422.

[0108] It should be understood that, in some embodiments, Figure 10 The nozzle 420 can be interchanged with or replaced by the wiping material (e.g., similar to...). Figure 8The wiping material 402. This advantageously allows the cleaning system to first remove any immersion medium from the lens surface via the suction device 422, and then clean and / or polish the lens surface with the wiping material, which in some embodiments may contain a cleaning agent. In this way, the wiping material can be used more times or for a longer period of time because it is soiled by less immersion medium (e.g., in the case of an oil-based immersion medium) and / or the cleaning agent is less diluted by the immersion medium absorbed into the wiping material (e.g., in the case of a water-based immersion medium).

[0109] List of abbreviations for defined terms

[0110] To aid in understanding the scope and content of this written description and the appended claims, several selected terms are defined directly below. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure pertains.

[0111] As used herein, the terms “approximately,” “about,” and “generally” refer to a quantity or condition that is close to a specific statement of whether the desired function or result is still being performed. For example, the terms “approximately,” “about,” and “generally” may refer to a quantity or condition that deviates from a specific statement by less than 10%, or less than 5%, or less than 1%, or less than 0.1%, or less than 0.01%.

[0112] It should be understood that the term "immersion medium" includes any natural or synthetic medium having a high refractive index (e.g., greater than 1.3, preferably greater than 1.5) suitable for improving the resolving power (i.e., numerical aperture) of high-resolution objectives. The term "immersion medium" is understood to include water or any transparent oil having the desired viscosity and optical properties for a given microscopic application.

[0113] As used herein, the term "immersion objective" is intended to include those objectives having a numerical aperture greater than 1 (i.e., the refractive index of air) and benefiting from or requiring the use of an immersion medium to obtain optimal performance. "Immersion objective" is understood herein to be synonymous with "high-resolution objective" or other objectives in the disclosed imaging system that automatically receives an immersion medium.

[0114] As used herein, the term "stage housing" includes a stage and / or stage assembly mounted in optical and mechanical cooperation with components constituting a microscope assembly. A "stage assembly" may be a stage on which a sample can be positioned, and may additionally include stage positioning mechanisms, as known in the art, for selectively moving the stage in the xy-plane to observe the sample positioned thereon. As used herein, the term "stage housing" may also be used to describe additional features associated with the stage or stage assembly, including, for example, elements for controlling environmental conditions around the stage and / or the mounted sample, such as heating elements, cooling elements, fans, gas sensors / inlets (e.g., oxygen, carbon dioxide, etc.), vacuum, compressors, or any one or more other elements or physical housings associated with or coupled to the sample stage.

[0115] Various aspects of this disclosure, including apparatus, systems, and methods, may be illustrated by reference to one or more embodiments or implementations that are essentially exemplary. As used herein, the term "exemplary" means "serving as an example, illustration, or description" and should not necessarily be construed as preferred or advantageous compared to other embodiments disclosed herein. Furthermore, references to this disclosure or "implementation" of the invention include specific references to one or more embodiments thereof, and such references are intended to provide illustrative examples rather than to limit the scope of the invention, the scope of which is indicated by the appended claims rather than the following description.

[0116] As used in this specification, words appearing in the singular encompass their plural counterparts, and words appearing in the plural encompass their singular counterparts, unless otherwise implied or explicitly stated. Therefore, it should be noted that, unless the context explicitly states otherwise, the singular forms “a,” “an,” and “the” as used in this specification and the appended claims include plural references. For example, a reference to a singular reference (e.g., “widget”) includes one, two, or more references, unless otherwise implied or explicitly stated. Similarly, unless the content and / or context clearly specify otherwise, a reference to multiple references should be interpreted as including a single reference and / or multiple references. For example, a reference to a plural form of reference (e.g., “widget”) does not necessarily require multiple such references. Rather, it should be understood that, unless otherwise stated, one or more references are contemplated herein, independent of the inferred number of references.

[0117] As used herein, directional terms such as “top,” “bottom,” “left,” “right,” “upper,” “lower,” “upper part,” “lower part,” “proximal end,” “farthest end,” “adjacent,” etc., are used only to indicate relative directions and are not intended to otherwise limit the scope of this disclosure and / or the claimed invention.

[0118] in conclusion

[0119] The terminology and expressions used herein are descriptive and not limiting, and their use is not intended to exclude any equivalents of the features or portions thereof shown and described, but rather to recognize that various modifications are possible within the scope of the claimed invention. Therefore, it should be understood that while the invention has been particularly disclosed through preferred embodiments, exemplary embodiments, and optional features, modifications and alterations to the concepts disclosed herein can be made by those skilled in the art, and such modifications and alterations are considered to be within the scope of the invention as defined by the appended claims. The specific embodiments provided herein are examples of useful embodiments of the invention, and various changes and / or modifications to the inventive features described herein and additional applications that will conceive of and possess by those skilled in the art without departing from the spirit and scope of the invention as defined by the claims are considered to be within the scope of this disclosure.

[0120] It should also be understood that systems, apparatuses, products, kits, methods, and / or processes according to certain embodiments of this disclosure may include, incorporate, or otherwise include the properties or features (e.g., components, members, elements, parts, and / or portions) described in other embodiments disclosed and / or described herein. Therefore, various features of certain embodiments may be compatible with, combined with, included in, and / or incorporated into other embodiments of this disclosure. Consequently, the disclosure of certain features with respect to a particular embodiment of this disclosure should not be construed as limiting the application or inclusion of said features to that particular embodiment. Rather, it should be understood that other embodiments may also include said features, members, elements, parts, and / or portions without necessarily departing from the scope of this disclosure.

[0121] Furthermore, unless a feature is described as another feature that needs to be combined with it, any feature herein may be combined with any other feature of the same or different embodiments disclosed herein. Moreover, to avoid obscuring aspects of the exemplary embodiments, various well-known aspects of illustrative systems, methods, devices, and the like are not described in particular detail herein. However, such aspects are contemplated herein.

[0122] All references listed in this application are incorporated herein by reference in their entirety, without any inconsistency with the disclosure herein. It will be apparent to those skilled in the art that methods, apparatuses, apparatus elements, materials, procedures, and techniques other than those specifically described herein can be applied to the practice of the invention as broadly disclosed herein without the need for excessive experimentation. All functional equivalents known in the art of the methods, apparatuses, apparatus elements, materials, procedures, and techniques specifically described herein are intended to be covered by this invention.

[0123] When groups of materials, compositions, components, or compounds are disclosed herein, it should be understood that all individual members of those groups and all their subgroups are disclosed separately. When Markush groups or other groupings are used herein, all individual members of said group and all possible combinations and subcombinations of said group are intended to be individually included in this disclosure. Unless otherwise stated, every combination of formulations or components described or exemplified herein can be used to practice the invention. Whenever a range, such as a temperature range, time range, or composition range, is given in the specification, all intermediate ranges and subranges, as well as all individual values ​​contained within the given range, are intended to be included in this disclosure.

[0124] All changes falling within the equivalent meaning and scope of the claims shall be covered within the scope of the claims.

Claims

1. An imaging system configured for automatically applying and / or removing an immersion medium, comprising: Sample stage; An imaging assembly is disposed on a first side of the sample stage and includes an immersion objective configured to be selectively aligned with the optical axis of the imaging system. as well as An applicator, positioned to selectively interact with the lens surface of the immersion objective to deposit or remove the immersion medium, wherein the applicator is integrated or mounted into the sample stage, and the applicator translates together with the sample stage. The applicator includes an immersion medium nozzle fluidly connected to an immersion medium reservoir via an immersion medium hose and operable to move immersion medium from the reservoir to the nozzle for dispensing, and the nozzle is connected to a lens cleaner reservoir and operable to move lens cleaner from the reservoir to the nozzle for dispensing. The imaging system further includes: A liquid sensor, configured to detect the presence of an immersion medium at the immersion medium nozzle; and A pump, associated with and configured to dispense immersion medium from the immersion medium reservoir through the immersion medium hose and to the immersion medium nozzle. Specifically, when the liquid sensor does not detect the immersion medium at the immersion medium nozzle, the pump is operated until the liquid sensor detects the immersion medium at the immersion medium nozzle. Wherein, after the liquid sensor detects the immersion medium at the immersion medium nozzle, the pump is operated to dispense a predetermined volume of immersion medium through the immersion medium nozzle.

2. The imaging system of claim 1, wherein the immersion medium nozzle is configured to dispense an immersion medium free of air bubbles.

3. The imaging system of claim 2, further comprising a bubble sensor configured to detect the presence of bubbles at the immersion medium nozzle or in an upstream pipeline supplying immersion medium to the immersion medium nozzle.

4. The imaging system of claim 1, wherein the liquid sensor comprises an optical sensor or a multimeter for measuring the resistance at the nozzle.

5. The imaging system of claim 1, wherein the liquid sensor comprises a capacitor sensor.

6. The imaging system of claim 1, wherein the pump is configured to dispense a desired volume of immersion medium based on operating time and / or the number of operating cycles.

7. The imaging system according to any one of claims 1 to 6, wherein the applicator is disposed on the first side of the sample stage.

8. The imaging system of claim 1, wherein the sample stage is an electric xy stage and is configured to position the applicator adjacent to the lens surface of the immersion objective, such that dispensing the immersion medium from the applicator causes the immersion medium to deposit onto the lens surface of the immersion objective.

9. The imaging system according to any one of claims 1 to 6 and 8, wherein the imaging system comprises an inverted microscope, wherein the imaging component is positioned below the sample stage, and the first side of the sample stage is the bottom of the sample stage, such that the applicator is oriented on the bottom of the sample stage toward the immersion objective.

10. The imaging system according to any one of claims 1 to 6, 8, wherein the imaging system comprises an upright microscope, wherein the imaging component is positioned above the sample stage, and the first side of the sample stage is the top of the sample stage, such that the applicator is oriented on the top of the sample stage toward the immersion objective.

11. The imaging system according to any one of claims 1 to 6 and 8, comprising a wiping material configured to clean and / or remove an immersion medium from the lens surface of the immersion objective, the wiping material containing a cleaning agent.

12. The imaging system of claim 11, wherein the applicator comprises the wiping material.

13. The imaging system of claim 11, further comprising a displacement element operatively connected to the wiping material and configured to selectively move the wiping material.

14. The imaging system of claim 13, wherein the selective movement of the wiping object comprises one or more of linear or back-and-forth movement, movement within a single plane, or rotational movement.

15. The imaging system of claim 13 or claim 14, wherein the translocation element is a solenoid that provides a similar vibration amplitude to the wiping material.

16. The imaging system of claim 11, further comprising a computing system configured to generate a map of the wiping material, track portions of the wiping material previously used to clean the immersion objective, and guide movement of the wiping material during subsequent cleaning operations to interact with the immersion objective in clean or unused areas of the wiping material.

17. The imaging system according to any one of claims 1 to 6 and 8, wherein the applicator includes a suction device configured to remove the immersion medium from the lens surface of the immersion objective.

18. The imaging system according to any one of claims 1 to 6, wherein the applicator is disposed at a fixed position within the housing of the imaging system.

19. The imaging system of claim 18, wherein the imaging component includes a lens slider or a turntable, and the immersion objective is mounted on the lens slider or turntable.

20. The imaging system of claim 19, wherein the lens slider or turntable is selectively positioned below the applicator to receive immersion medium from the applicator onto the lens surface of the immersion objective.

21. The imaging system of claim 18, comprising a wiping material configured to clean and / or remove an immersion medium from the lens surface of the immersion objective, the wiping material containing a cleaning agent, and wherein the imaging assembly includes a lens slider or turntable, the immersion objective being mounted on the lens slider or turntable.

22. The imaging system of claim 21, wherein the applicator comprises the wiping material.

23. The imaging system of claim 21 or claim 22, further comprising a vibration element operatively connected to the swab and configured to selectively vibrate the swab.

24. The imaging system of claim 21 or claim 22, wherein the lens slider or turntable is selectively positioned below the wiping material to remove the immersion medium from the lens surface of the immersion objective.

25. The imaging system of claim 18, wherein the applicator includes a suction device configured to remove the immersion medium from the lens surface of the immersion objective.

26. The imaging system according to any one of claims 1 to 6, 8, 12 to 14, 16, 19 to 22, 25, further comprising a second immersion objective, wherein the applicator is additionally configured to selectively interact with a corresponding lens surface of the second immersion objective.

27. A method for automatically applying an immersion medium to an immersion objective lens, comprising: Obtain the imaging system according to any one of claims 1 to 16; The sample stage is positioned relative to the immersion objective such that the applicator is adjacent to the lens surface of the immersion objective; as well as The immersion medium is dispensed from the applicator onto the lens surface of the immersion objective.

28. A method for automatically applying an immersion medium to an immersion objective lens, comprising: Obtain the imaging system according to any one of claims 18 to 26; The immersion objective is positioned relative to the applicator such that the applicator is adjacent to the lens surface of the immersion objective; as well as The immersion medium is dispensed from the applicator onto the lens surface of the immersion objective.

29. The method of claim 27 or claim 28, wherein the dispensed immersion medium is free of air bubbles.

30. The method of claim 28, further comprising returning the immersion objective to the observation position.

31. A method for automatically removing an immersion medium from the lens surface of an immersion objective lens, comprising: Obtain the imaging system according to any one of claims 1 to 17; The sample stage is positioned relative to the immersion objective such that the applicator is adjacent to the lens surface of the immersion objective; as well as The immersion medium is removed from the lens surface of the immersion objective via the applicator.

32. The method of claim 27 or claim 31, further comprising returning the sample stage to the observation position.

33. A kit for automatically dispensing immersion media, comprising: An immersion medium reservoir configured to hold a certain volume of immersion medium; A lens cleaner reservoir, the lens cleaner reservoir being configured to hold a certain volume of lens cleaner; A nozzle is fluidly connected to the immersion medium reservoir via an immersion medium hose, and the nozzle is connected to the lens cleaner reservoir and operable to move lens cleaner from the lens cleaner reservoir to the nozzle for dispensing; as well as A micropump operable to move immersion medium from the immersion medium reservoir through the immersion medium hose and to the nozzle for dispensing. The kit is used in conjunction with an imaging system, wherein the nozzle is integrated into or mounted into the sample stage of the imaging system, and the nozzle and the sample stage translate together. The kit further includes: A liquid sensor configured to detect the presence of an immersion medium at the nozzle. Specifically, when the liquid sensor does not detect the immersion medium at the nozzle, the micropump is operated until the liquid sensor detects the immersion medium at the nozzle. Wherein, after the liquid sensor detects the immersion medium at the nozzle, the micro pump is operated to dispense a predetermined volume of immersion medium through the nozzle.

34. The kit of claim 33, further comprising computer-executable instructions that, when executed by one or more processors of a computer system, cause the computer system to automatically allocate immersion medium.

35. The kit of claim 33 or claim 34, further comprising a check valve associated with the immersion medium hose to prevent pumped immersion medium from flowing back into the immersion medium reservoir when not being pumped.

36. The kit of claim 33 or claim 34, further comprising a level indicator associated with the immersion medium reservoir.

37. The kit of claim 33 or claim 34, further comprising a waste reservoir and a waste level indicator associated with said waste reservoir.

38. The kit according to claim 33 or claim 34, wherein the kit is operable to be converted into an automated optical microscope after manufacturing.

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