Microscope for examining a sample

Abstract

To provide a microscope system configured to feed a reagent to a biological sample for observation, which prevents deterioration of the sample and reagent.SOLUTION: The present invention relates to a microscope for microscopic examination of a sample 120, comprising: a microscope housing 102 enclosing an illumination optical system 118, a microscope stage 116, and an imaging optical system 124; an integrated sample chamber 106 located within the microscope housing 102; and an integrated reagent chamber 150 located within the microscope housing 102 and configured to supply a reagent to the sample 120.SELECTED DRAWING: Figure 1

Description

Technical Field

[0001] The concept of the present invention is directed to a microscope for microscopic examination of a sample, particularly a microscope for examining a sample disposed in a sample chamber to which a reagent is supplied to the sample before or during examination of the sample.

Background Art

[0002] Particularly in the field of microscopic examination of living samples such as cells, it is common to supply a reagent to the sample before microscopic examination of the sample. Reagents in the sense of the present application include all fluids, particularly liquids containing or not containing chemical or biological active substances for inducing a biological or chemical reaction with the sample or simply for preserving the sample, such as water, hormones or drugs in a suitable carrier substance, water-soluble or lipid-soluble or other substances. Usually, the sample itself is placed in a multi-well plate on a microscope table, in a Petri dish, in a microfluidic system, etc. A pump is used to supply the reagent to the sample. A syringe pump is often used for the injection process, and a peristaltic pump is used for continuously transporting the reagent, such as supplying the reagent to the sample and at the same time removing the reagent from the sample. On the other hand, manual pipetting can also be performed.

[0003] For storing the reagent, various types of containers such as Falcon tubes, Eppendorf tubes, Schott flasks are used. The containers are usually placed outside the microscope housing and sometimes placed in a temperature-controlled box. One or more pump systems are used to transport the reagent from the container to the sample, and the pump system is usually placed either outside the microscope housing or inside the microscope housing. The temperature-controlled box for storing the reagent container usually includes a heating element or an ice bath section.

[0004] In many cases, only small amounts of reagents need to be injected into biological samples, and these reagents must be stored in light-shielded containers at a temperature lower than the ambient temperature, for example, 4-8°C. This is especially true when injecting irritants such as hormones or drugs in 3D cell culture tests. Supplying reagents through long supply lines makes it difficult to maintain a constant temperature and protect the reagents from light as they reach the sample. Furthermore, a large amount of fluid is required to compensate for the dead volume inside the tubing, increasing the cost of the chemicals used.

[0005] On the other hand, in organ-on-a-chip (organ-on-a-chip) testing, for example, more reagents need to be exchanged. Injecting large quantities of reagents and coordinating peristaltic pump systems is often cumbersome, and because the temperature of the injected reagent differs from that of the sample being tested, temperature-dependent measurement artifacts may occur, or the sample may die back during testing. Therefore, ideally, the temperature of the injected reagent should be the same as that of the sample. [Overview of the project] [Problems that the invention aims to solve]

[0006] Given the aforementioned shortcomings, improvements are needed in the solution for supplying reagents to samples being examined under a microscope. [Means for solving the problem]

[0007] A conceptual embodiment of the present invention provides a microscope for examining a sample, as described in claim 1. The microscope comprises a microscope housing surrounding an illumination optical system, a microscope stage, and an imaging optical system, and further comprises an integrated sample chamber located within the microscope housing. Furthermore, an integrated reagent chamber configured for supplying reagents to the sample is located within the same microscope housing.

[0008] The concept of the present invention therefore allows for the provision of a microscope that includes an integrated reagent chamber located within the same microscope housing as an integrated sample chamber. This solution minimizes the distance between the reagent container and the sample. At the same time, the reagents can be transported and stored in a light-shielded state and with minimal temperature fluctuations. Another advantage is that reagents can be supplied to the sample before and / or during the examination without the user needing to access the sample.

[0009] It should be noted that the reagent chamber according to the concept of the present invention is configured to supply reagents to a sample. This means that the necessary reagent containers (which may be buffer containers) and, preferably, the associated pump system and corresponding reagent supply lines can be installed in the reagent chamber or are part of the reagent chamber.

[0010] In a preferred embodiment, the reagent chamber is itself light-shielded and / or thermally insulated. In this embodiment, the influence of the ambient atmosphere on the reagents inside the reagent chamber is minimized.

[0011] There are two options for integrating a reagent chamber into the microscope housing: the reagent chamber can be located outside or inside the sample chamber.

[0012] When the reagent chamber is located outside the sample chamber, the reagent chamber includes a connecting tube that connects the reagent chamber and the sample chamber. In this embodiment, since the reagent chamber and the sample chamber are separated, it is very preferable to thermally insulate the reagent chamber, including the connecting tube. In a preferred embodiment, the reagent chamber includes at least one reagent container and a pump system that delivers the reagent from the reagent container through at least one reagent supply line to a sample placed in the sample chamber. At least one supply line and / or any reagent removal line extends through the connecting tube.

[0013] In a preferred embodiment, the connecting tube includes a plug at at least one of its ends for (at least) partially closing the corresponding tube end. The plug is inserted, for example, into the end of the connecting tube facing the sample chamber. The plug is formed so that any supply / removal lines can still extend through the plug by corresponding through-holes. Such a plug serves to define the position of the supply / removal lines, provide high thermal insulation, and prevent the atmosphere of the sample chamber from entering the reagent chamber.

[0014] According to the second option, the reagent chamber is located within the sample chamber. This embodiment is particularly preferred when the sample chamber provides sufficient space to accommodate the reagent chamber. In most cases, especially for biological samples, the sample chamber is heated, i.e., the temperature and composition of the heating atmosphere within the sample chamber are controlled. In such cases, the temperature of the uninsulated reagent chamber approaches the temperature of the heated sample chamber, so it may not be necessary to use a thermally insulated reagent chamber. However, it is preferable that the reagent chamber has an opening that is at least partially closed by a plug. As already described in relation to the first option, the plug can serve the purpose of determining the specified location of any supply / removal lines. Furthermore, it can increase the insulation efficiency of the insulated reagent chamber. Finally, it prevents the heating atmosphere from entering the inside of the reagent chamber.

[0015] It should be noted that in some cases, an upper stage chamber may be used, which is positioned on the microscope stage within the sample chamber and configured to receive the sample. In such cases, only a very small volume of the upper stage chamber is kept warm. In these cases, a reagent supply line must be connected to the upper stage chamber so that reagents are supplied to the sample inside the upper stage chamber. Similarly, a reagent removal line must be connected to the upper stage chamber so that reagents can be removed from the sample, for example, by returning them to the reagent chamber.

[0016] In another preferred embodiment of the microscope, an integrated sample chamber is formed by an isolated housing within the microscope housing. This housing forming the sample chamber includes a lid that provides direct access to the microscope stage for placing a sample within the sample chamber. Furthermore, an integrated reagent chamber may be formed by another isolated housing within the microscope housing, this further housing including another door that provides direct access to at least one reagent container and / or pump system. By opening the door, the user can refill the reagent container, replace the reagent container, replace or adjust the pump system, connect reagent supply / removal lines, and so on.

[0017] In another preferred embodiment, the housing forming the sample chamber is configured such that the sample chamber is sealed when the lid is closed. This is particularly advantageous when the reagent chamber is located inside the sample chamber and when the sample chamber is kept warm. By using an uninsulated reagent chamber, the temperatures in the sample chamber and the reagent chamber are averaged out so that there is no temperature difference between the reagent and the sample.

[0018] In another preferred embodiment, the housing forming the sample chamber surrounds the microscope's illumination optical system and at least partially surrounds the microscope stage. Such a configuration corresponds to an inverted transmission microscope. Alternatively, another housing may surround the microscope's imaging optical system.

[0019] In a preferred embodiment, the reagent chamber is configured to be temperature-controlled. Temperature control of the reagent chamber is particularly useful when the reagent chamber is thermally insulated. This allows the reagent chamber to be maintained at a predetermined temperature so that any reagents in the container, supply line, or pump can be kept at the desired temperature. Generally, it is convenient to configure the housing, i.e., the portion surrounding the sample chamber, reagent chamber, and imaging optical system, to be air-conditioned or heated and / or temperature-controlled.

[0020] Regarding the possible temperature control of the sample chamber, it is preferable that the housing forming the sample chamber includes an interface for connecting an external temperature control unit to the sample chamber. This allows control of the environmental conditions inside the sample chamber by connecting the external temperature control unit to the interface. Typically, the temperature control unit includes a connection for supplying H2O, a connection for discharging H2O and / or the temperature control atmosphere, a connection for supplying N2 and / or O2, and a connection for supplying CO2 into the sample chamber. A desired temperature control atmosphere can be achieved by controlling at least one of the flow rate, temperature, and content of H2O and / or CO2 and / or N2 and / or O2 supplied to the temperature control atmosphere inside the sample chamber.

[0021] Temperature control of the reagent chamber can be achieved by at least one of a cooling unit, a heating unit, and a heat exchange unit located within the reagent chamber. Preferably, a Peltier element can be used as the cooling and heating unit. On the other hand, the cooling unit can simply be a cooling pack placed within the reagent chamber. Alternatively, a heat exchanger can be used to heat or cool the atmosphere inside the reagent chamber with respect to the heat transfer medium within the heat exchanger.

[0022] In another embodiment, the reagent chamber is connected to a temperature control unit configured to control the temperature inside the reagent chamber. The temperature control unit may be located outside the reagent chamber, for example, next to it, using an atmosphere connecting tube for exchanging atmosphere between the temperature control unit and the reagent chamber. For this purpose, it is preferable that the temperature control unit includes at least one fan for circulating a heated or cooled atmosphere between the temperature control unit and the reagent chamber. In this case as well, a cooling unit, a heating unit and / or a heat exchange unit may be provided within the temperature control unit to achieve a desired temperature of the atmosphere exchanged between the temperature control unit and the reagent chamber.

[0023] As described above, in a preferred embodiment, the reagent chamber is configured to be able to install at least one reagent container therein. During operation, the microscope includes a reagent chamber containing at least one reagent container. Further, the reagent chamber is preferably configured to be able to connect at least one of a reagent supply line and a reagent removal line to one of the at least one reagent containers. Also in this case, during operation of the microscope, the reagent chamber includes at least one of a reagent supply line and a reagent removal line connected to the reagent container. If there are a plurality of reagent containers, there may be a plurality of supply / removal lines.

[0024] In a preferred embodiment, the plug in the connection tube in the first option, or the plug at the opening of the reagent chamber in the second option, is configured to guide the reagent supply line and / or the reagent removal line through the plug.

[0025] Further, the reagent chamber preferably includes a pump system for transporting the reagent through the reagent supply line and / or the removal line. The pump system may include a syringe pump and / or a peristaltic pump.

[0026] Finally, the reagent chamber preferably includes a door that provides access to the reagent chamber.

[0027] It should be noted that the features of the above examples and embodiments, as well as the examples and embodiments further described below, can be combined, in whole or in part, with other examples and embodiments that are not explicitly mentioned herein but are part of the present disclosure.

Brief Description of the Drawings

[0028] [Figure 1] FIG. schematically shows a first embodiment of a microscope including a reagent chamber arranged outside the sample chamber of the microscope. [Figure 2] FIG. schematically shows a further embodiment of a microscope including a reagent chamber arranged inside the sample chamber of the microscope. [Figure 3]This figure schematically shows a further embodiment of a microscope that includes a reagent chamber located outside the sample chamber of the microscope. [Figure 4] This figure schematically shows a further embodiment of a microscope that includes a reagent chamber located outside the sample chamber of the microscope. [Figure 5] This figure schematically shows a further embodiment of a microscope, including a reagent chamber and a temperature control unit, both located outside the sample chamber of the microscope. [Figure 6] This figure schematically shows a further embodiment of a microscope, including a reagent chamber and a temperature control unit, both located outside the sample chamber of the microscope. [Figure 7] This figure schematically shows a further embodiment of a microscope that includes a reagent chamber located outside the sample chamber of the microscope. [Modes for carrying out the invention]

[0029] Figure 1 schematically shows one embodiment of a microscope according to the concept of the present invention. The microscope 100 for examining a sample 120 comprises a microscope housing 102, which surrounds an illumination optical system 118, a microscope stage 116, and an imaging optical system 124. An integrated sample chamber 106 is located within the microscope housing 102. The configuration shown in Figure 1 corresponds to an inverted transmission microscope in which the sample 120 is illuminated by the illumination optical system 118, and the light transmitted through the sample 120 is detected by the imaging optical system 124. In a preferred embodiment, the sample chamber 106 is formed by an isolated housing section 104 within the microscope housing 102. Typically, the sample chamber 106 is kept warm to maintain a desired warming atmosphere and temperature during the examination of the sample 120.

[0030] In the embodiment shown in Figure 1, the reagent chamber 150 is located outside the sample chamber 106 and includes a connecting tube 170 that connects the reagent chamber 150 and the sample chamber 106. The reagent chamber 150 is located inside the microscope housing 102 and is configured to supply reagents to the sample 120. The reagent chamber can be formed by another housing section of the microscope housing 102. The reagent chamber 150 includes a door 152 that provides direct access to the reagent chamber 150 by the user of the microscope 100. The connecting tube 170 includes a plug 172 at the end of the connecting tube 170 facing the sample chamber 106. The plug 172 partially closes the corresponding tube to prevent the insulating atmosphere from entering the reagent chamber 150. Particularly in the configuration shown in Figure 1, it is preferable that the reagent chamber 150, including the connecting tube 170, is thermally insulated and light-shielded. This allows for the preservation of reagents placed inside the reagent chamber and for maintaining the inside of the reagent chamber 150 at a desired temperature.

[0031] Figure 2 schematically shows another embodiment of a microscope according to the concept of the present invention. As seen in Figure 2, the reagent chamber 150 is incorporated into the sample chamber 106 within the microscope housing 102. In this embodiment, the sample chamber 106 is formed by an isolated housing section 104 of the microscope housing 102. Within this housing section 104 are the illumination optical system 118 and a portion of the microscope stage 116 for supporting the sample 120. Below the housing section 104, in another housing section, is the imaging optical system 124. Similar to the embodiment in Figure 1, the microscope in Figure 2 is also an inverted transmission microscope. However, it should be noted that the concept of the present invention is not limited to this type of microscope.

[0032] In the embodiment shown in Figure 2, the sample 120 is placed in the upper stage chamber 210, which is positioned on the microscope stage 116. Of course, depending on the type of test, the sample 120 may be placed directly on the microscope stage 116 without such an upper stage chamber 210. When using the upper stage chamber 210, it is convenient to keep only a small volume of the upper stage chamber 210 warm, rather than the entire sample chamber 106.

[0033] In the embodiment shown in Figure 2, the reagent chamber 150 is formed as a thermally insulated chamber including a closable door (not shown) for direct access to the reagent chamber 150. Various possibilities for internal fittings of the reagent chamber 150 in the embodiments of Figures 1 and 2 will be further described in relation to the following figures. The reagent chamber 150 includes an opening 252 which is partially closed by a plug 272 for insulation. A supply line 254 for supplying reagents into the upper stage chamber 210 extends from a reagent container (not shown) through the plug 272 into the upper stage chamber 210.

[0034] In the embodiment shown in Figure 2, the reagent chamber 150 includes another opening 112 that functions as a cable duct, which can be used for, for example, a power cord, a trigger cable, and / or a connection to an external temperature control unit (further described below in relation to the following figures). This opening 112 may also be partially closed by another plug, which includes a through-hole for the corresponding cable.

[0035] Figures 1 and 2 illustrate the basic principles of two possible configurations for an integrated reagent chamber within the microscope housing, but it should be noted that the following figures focus on possible internal accessories and temperature control of the reagent chamber 150 itself. It should be noted that the following embodiments can be combined with the alternative embodiments shown in Figure 1 or Figure 2.

[0036] Figure 3 shows another embodiment of the microscope according to the concept of the present invention in the option corresponding to Figure 1 described above. Therefore, for simplicity, only the differences from Figure 1 will be described.

[0037] The reagent chamber 150 in Figure 3 includes a reagent container 380 in the form of a Falcon tube, a pump system 374 in the form of a perfuser or syringe pump, and a reagent supply line 254. A cooling pack 360 is positioned at the rear of the reagent chamber 150 as a cooling unit. This allows the temperature inside the insulated reagent chamber 150 to be kept lower than the ambient temperature. The cooled reagent can be transported by the perfuser 374 from the Falcon tube 380 through the supply line 254 to the sample 120.

[0038] Figure 4 shows a slightly different embodiment from the embodiment in Figure 3. Instead of the cooling pack 350, a Peltier element 458 for cooling or heating the inside of the reagent chamber 150 is installed adjacent to or inside the outer wall of the reagent chamber 150. Such a Peltier element 458 can be easily used to control the temperature inside the reagent chamber 150 to a desired temperature using a corresponding temperature sensor (not shown) inside the reagent chamber.

[0039] Figure 5 shows another embodiment of the reagent chamber 150 connected to the temperature control unit 562. Separately, the embodiment in Figure 5 corresponds to the embodiments in Figures 3 and 4. Instead of having a cooling pack 360 or a Peltier element 458, the temperature inside the reagent chamber 150 is controlled by connecting the temperature control unit 562 to the reagent chamber 150. The temperature control unit 562 includes a fan 564, a Peltier element 458, and a heating unit 566. A heat exchange unit can be used instead of the heating unit 566. It should be noted that, depending on the type of test, it may be sufficient to use only the Peltier element 458, only the heater 566, or only the heat exchange unit. In this case as well, the Peltier element 458 is used to heat or cool the temperature inside the temperature control unit 562. Air is circulated by the fan 564 to ensure a uniform temperature inside the temperature control unit 562 and the reagent chamber 150. The heater 566 can assist the Peltier element 458 to achieve a higher temperature more rapidly, if necessary.

[0040] The embodiment in Figure 6 substantially corresponds to the embodiment in Figure 5, except that a Schott flask 680 is used as the reagent container instead of a Falcon tube 380. Instead of using a perfusion pump, a peristaltic pump 676 is used. Peristaltic pumps are typically used in tests such as biomedical chip tests, where reagents must be continuously supplied to the sample while simultaneously being removed from the sample. As shown in Figure 6, the reagent is transported from the Schott flask 680 to the sample 120 via the connecting tube 170 and plug 172 by the peristaltic pump 676 through the reagent supply line 254. At the same time, the reagent is removed from the sample 120 by the reagent removal line 656. The removed reagent is transported back to the same or a second (not shown) Schott flask 680 or another container.

[0041] It should be noted that this type of embodiment can also be implemented in the embodiment shown in Figure 3 or Figure 4, which does not have an external temperature control unit. Any of the embodiments described in relation to Figures 3 to 7 can also be implemented in a reagent chamber 150 as shown in Figure 2, i.e., a reagent chamber 150 incorporated into the sample chamber 106.

[0042] Figure 7 is substantially equivalent to Figure 3 and is intended solely to show the cooling pack 360 located at the rear of the reagent chamber 150. For further details, please refer to Figure 3. [Explanation of symbols]

[0043] 100 Microscopes 102 Microscope housing 104 Enclosure 106 Sample Room 112 Cable duct 116 Microscope Stages 118 Illumination optical system 120 samples 124 Imaging Optical System 150 Reagent Room 152 doors 170 connecting tube 172 plug 210 Upper Stage Room 252 Opening 254 Reagent supply line 272 plug 360 Cooler Pack 380 reagent containers, Falcon tubes 458 Peltier element 562 Temperature control unit 564 fans 566 Heating unit, heater 656 Reagent removal line 676 Peristaltic pump 680 Reagent containers, Schott flasks

Claims

1. A microscope (100) for examining a sample (120) placed in a multiwell plate or petri dish, wherein the microscope (100) A microscope housing (102) surrounds the illumination optical system (118), the microscope stage (116), and the imaging optical system (124), An integrated sample chamber (106) is located inside the microscope housing (102), An integrated reagent chamber (150) is located within the microscope housing (102) and configured to supply reagents to the sample (120), Equipped with, The reagent room (150) is located within the sample room (106), and the reagent room (150) includes a door (152) that provides direct access to the reagent room (150). Microscope (100).

2. The reagent chamber (150) includes an opening (252) which is at least partially closed by a plug (272). The microscope (100) according to claim 1.

3. The reagent chamber (150) is either thermally insulated or light-shielded, A microscope (100) according to claim 1 or 2.

4. The reagent chamber (150) is configured to be temperature-controlled. A microscope (100) according to any one of claims 1 to 3.

5. The reagent chamber (150) includes at least one of a cooling unit, a heating unit, and a heat exchange unit. The microscope (100) according to claim 4.

6. The reagent chamber (150) includes a Peltier element (458), The microscope (100) according to claim 4 or 5.

7. The reagent chamber (150) includes a cooling pack (360), A microscope (100) according to any one of claims 4 to 6.

8. The reagent chamber (150) is connected to a temperature control unit (562) configured to control the temperature inside the reagent chamber (150). A microscope (100) according to any one of claims 1 to 7.

9. The temperature control unit (562) includes at least one of a fan (564), a cooling unit, a heating unit (566), and a heat exchange unit. The microscope (100) according to claim 8.

10. The reagent chamber (150) is configured to allow at least one reagent container (380, 680) to be placed inside the reagent chamber (150). A microscope (100) according to any one of claims 1 to 9.

11. The reagent chamber (150) is configured such that at least one of the reagent supply line (254) and the reagent removal line (656) can be connected to one of the at least one reagent containers (380, 680). The microscope (100) according to claim 10.

12. The plug (272) is configured to guide at least one of the reagent supply line (254) and the reagent removal line (656) through the plug (272). A microscope (100) according to claim 11, relating to claim 2.

13. The reagent chamber (150) includes a pump system (374, 676) for transporting reagents through the reagent supply line (254) and / or the reagent removal line (656). The microscope (100) according to claim 11 or 12.

14. Light from the illumination optical system (118) is incident on the sample (120) from directly above the sample (120). A microscope (100) according to any one of claims 1 to 13.

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