JP7583664B2 - Sample Observation Module - Google Patents

Description

This disclosure relates to a sample observation module.

In recent years, research into optogenetics has made great strides worldwide in the medical field, including the field of neurology. One example of this type of research is the use of photoactivated adenylyl cyclase (PAC) (see, for example, non-patent documents 1 and 2). When PAC is stimulated with blue light, it produces the neurotransmitter cAMP depending on the light intensity. This makes it possible to induce a wide variety of physiological responses, such as glycogenolysis, lipolysis, muscle contraction, branching and elongation of nerve axons, vasodilation, inhibition of thrombosis, and activation of gene expression.

Conventionally, physiological responses such as those described above have been expressed by adding or injecting hormones or other substances into samples, but this requires complicated processes and has the problem of making it difficult to control the physiological response. In contrast, technology using PACs makes it possible to control physiological responses in time and space through optical manipulation. This is expected to lead to a variety of applications, such as the creation of disease model cells whose functions can be controlled through PACs and optical manipulation, and the restoration of lost biological functions through optical manipulation or cell medicine.

ZhouZ et al., “Photoactivated adenylyl cyclase (PAC) reveals novel mechanismsunderlying cAMP-dependent axonal morphogenesis” Scientific Reports 2016 Jan22: Volume 5, Article No.19679. doi:10.1038/srep19679 OhkiM et al., “Structural insight into photoactivation of an adenylate cyclase from aphotosynthetic cyanobacterium” Proceedings of National Academy of Sciences of the United States of America 2016 Jun 14; 113 (24): 6659-64.doi:10.1073/pnas.1517520113.

To activate the PAC in a sample and observe dynamic changes in the physiological response of the sample over time in response to light manipulation, a sample observation module that combines a light manipulation system and an observation system is required. Since it is expected that the sample observation module will be used in combination with a microscope system, for example, it is also required to have a simple configuration.

The present disclosure has been made to solve the above problems, and aims to provide a highly convenient sample observation module that allows for time-dependent observation of changes in a sample that accompany optical manipulation.

A sample observation module according to one aspect of the present disclosure is a sample observation module for observing changes in a sample over time due to optical manipulation, and includes a housing formed of a light-blocking material and having a mounting section therein on which the sample is placed, the housing is provided with a first window section for introducing stimulating light used for optical manipulation of the sample into the housing, and a second window section for introducing illuminating light used for observing the sample into the housing, and a multiplexing element is provided within the housing for multiplexing the stimulating light introduced from the first window section and the illuminating light introduced from the second window section and guiding the light to the mounting section.

In this sample observation module, a multiplexing element is provided in a light-tight housing that multiplexes the stimulating light introduced from the first window and the illuminating light introduced from the second window and guides the light to the mounting section. This makes it possible to simultaneously apply an optical stimulus to the sample and irradiate it with illuminating light, allowing the change in the sample accompanying the optical manipulation to be observed over time. Due to the simple configuration with the multiplexing element disposed in the housing, the sample observation module is also highly convenient when used in combination with a microscope system, etc.

A holder that holds the multiplexing element and is slidable between a first position facing the mounting section and a second position removed from the mounting section may be provided within the housing. In this case, by sliding the holder from the first position to the second position, it is possible to improve the ease of mounting the sample on the mounting section while saving space.

One or more attenuation filters that attenuate the intensity of the stimulating light may be removably provided inside the first window. In this case, the intensity of the stimulating light can be adjusted with a simple configuration. Therefore, appropriate light manipulation can be performed according to the type of sample.

The second window may be provided with a wavelength selection element that transmits the illumination light while cutting off light of the wavelength of the stimulus light. This prevents light other than the stimulus light introduced from the first window from contributing to the optical manipulation of the sample. This allows for more precise control of the optical manipulation of the sample.

The housing may be provided with a third window portion through which an external lens used for observing the sample is placed close to the mounting portion. In this case, observation of the sample using the external lens can be easily performed.

The housing may be provided with a temperature control section that adjusts the temperature of the sample placed on the mounting section. This allows the sample environment to be maintained in a good condition, making it possible to continuously observe changes in the sample that accompany optical manipulation.

The housing may be provided with a gas inlet for introducing a gas into the housing to maintain the environment of the sample. This allows the environment of the sample to be maintained well, making it possible to continuously observe changes in the sample due to optical manipulation.

The housing may be provided with a fan to circulate the gas within the housing. This allows the sample environment to be maintained in a good condition, making it possible to continuously observe changes in the sample that occur as a result of optical manipulation.

The housing may be provided with a humidity control section that adjusts the humidity inside the housing. This allows the sample environment to be maintained in a good condition, making it possible to continuously observe changes in the sample that accompany optical manipulation.

The present disclosure provides a highly convenient sample observation module that allows for the observation of changes in a sample over time that accompany optical manipulation.

FIG. 2 is a perspective view showing an embodiment of a sample observation module. 2 is a plan view showing the sample observation module shown in FIG. 1 with a lid of a housing removed. FIG. 2 is a cross-sectional view of the sample observation module shown in FIG. 1 . 13 is a cross-sectional view of the sample observation module in a state in which the holding part is slid. FIG. 1 is a cross-sectional view showing an example of use of the sample observation module.

Hereinafter, a preferred embodiment of a sample observation module according to one aspect of the present disclosure will be described in detail with reference to the drawings.
[Configuration of sample observation module]

Figure 1 is a perspective view showing one embodiment of a sample observation module. Figure 2 is a plan view of Figure 1 with the lid of the housing removed, and Figure 3 is a cross-sectional view of Figure 1. The sample observation module 1 shown in Figure 1 is a module for observing changes in a sample S (see Figure 5) caused by optical manipulation over time, and is used by being installed, for example, on the stage of a microscope system.

The sample S to be observed is, for example, a cell or a biological tissue. The sample S may be a microorganism. In this embodiment, the sample S is a cardiomyocyte into which a photoactivated adenylate cyclase (hereinafter referred to as "PAC") gene has been introduced (PAC-expressing cardiomyocyte). The animal from which the cardiomyocyte is derived is not particularly limited, and may be derived from a human, monkey, pig, cow, sheep, rabbit, rat, reptile, amphibian, fish, bird, or mouse. The cardiomyocyte may be an iPS cell-derived or ES cell-derived cardiomyocyte. For example, the cardiomyocyte may be a human iPS cell-derived cardiomyocyte.

Irradiating PAC-expressing cardiomyocytes with light (stimulating light L1: see Figure 5) causes PAC to produce cAMP in the cardiomyocytes. An increase in the intracellular cAMP concentration enhances the influx of calcium ions (Ca2 ⁺ ) into the cells. Cardiomyocyte contraction, i.e., cardiomyocyte pulsation, occurs due to an increase in calcium ion concentration. Therefore, by irradiating PAC-expressing cardiomyocytes with stimulating light L1, the pulsation speed of the PAC-expressing cardiomyocytes can be controlled.

As shown in Figs. 1 to 3, the sample observation module 1 includes a housing 2. The housing 2 is made of a strong material such as black hard plastic and has a light-shielding property against external light. The housing 2 has a rectangular parallelepiped shape that is slightly flattened in the height direction and includes a bottomed box-shaped main body 3 that is open at the top and a lid 4 that covers the top side of the main body 3. The main body 3 includes a bottom 5, a pair of side portions 6 and 7 that face each other in the longitudinal direction of the housing 2, and a pair of side portions 8 and 9 that face each other in the width direction of the housing 2. The lid 4 is detachably connected to the main body 3 by, for example, screws. An anti-slip member 10 (see Fig. 3) is provided on the outer surface side of the bottom 5 when the sample observation module 1 is placed on the stage of a microscope system.

A mounting section 11 on which a sample S is placed is provided within the housing 2. In this embodiment, as shown in Figs. 2 and 3, a recess 12 with a circular cross section is provided in the center of the bottom surface 5 of the main body 3, and the mounting section 11 is configured by the recess 12. For example, a circular receiving tray D (see Fig. 5) on which the sample S is placed can be placed on the mounting section 11.

The housing 2 is provided with a first window 13 for introducing the stimulating light L1 used for optical manipulation of the sample S into the housing 2, a second window 14 for introducing the illumination light L2 used for observing the sample S into the housing 2, and a third window 15 for arranging an external lens (not shown) used for observing the sample S adjacent to the mounting part 11. The first window 13 is formed, for example, by hollowing out the center part of the side part 7 of the main body part 3 in a circular shape. When optical manipulation of the sample S is performed, a light source 16 that outputs the stimulating light L1 is attached to the outside of the first window 13 (see FIG. 2). The light source 16 may be attached to the first window 13 in advance, or an external light source may be used. The light source 16 may be an LD, an LED, a laser light source, or the like.

When the sample S is a PAC-expressing cardiomyocyte, the stimulating light L1 is not particularly limited as long as it is light capable of activating PAC. PAC is activated by blue light. Therefore, light having a wavelength in the range of 350 nm to 500 nm is used as the stimulating light L1. The peak wavelength of the stimulating light L1 is preferably 400 nm to 500 nm, and more preferably 430 nm to 460 nm. The irradiation time and number of irradiations of the stimulating light L1 are adjusted according to the type of PAC gene, the expression level of the PAC gene, etc. The stimulating light L1 may be output continuously or may be output intermittently at a predetermined interval.

One or more attenuation filters 17 for attenuating the intensity of the stimulation light L1 are detachably provided inside the first window 13. By changing the number of attenuation filters 17 arranged, the intensity of the stimulation light L1 directed toward the sample S can be adjusted in stages. The intensity of the stimulation light L1 is adjusted depending on the type of PAC gene, the expression level of the PAC gene, etc. When the sample S is a PAC-expressing cardiomyocyte, the stimulation light L1 may have a photon flux density of, for example, 0.001 μmol/m ² /s to 100,000 μmol/m ² /s.

As shown in Figs. 1 and 3, the second window 14 is formed by hollowing out the center of the cover 4 in a circular shape. The second window 14 is provided in a position directly opposite the placement part 11 when the housing 2 is viewed from the height direction. The second window 14 is provided with a wavelength selection element 18 that transmits the illumination light L2 while cutting the light of the wavelength of the stimulus light L1. Examples of a light source (not shown) that outputs the illumination light L2 include a white light source and an LED that outputs visible light. In this embodiment, since blue light is used as the stimulus light L1, a red glass 19 as the wavelength selection element 18 is fitted into the second window 14. This red glass 19 cuts out short wavelength components, for example, wavelengths of 520 nm or less, of the illumination light L2 introduced into the housing 2 from the second window 14.

As shown in FIG. 3, the third window 15 is formed by hollowing out a circle in the central portion of the bottom surface of the mounting portion 11. The diameter of the third window 15 is smaller than the inner diameter of the mounting portion 11. As a result, the edge of the receiving dish D placed on the mounting portion 11 is supported by the bottom surface of the mounting portion 11, and the central portion of the receiving dish D is exposed from the bottom portion 5 of the housing 2 through the third window 15. An objective lens (not shown) of the microscope system in which the sample observation module 1 is installed is fitted into the third window 15. This allows the objective lens to be positioned close to the sample S placed on the mounting portion 11.

A multiplexing element 21 is provided in the housing 2, which multiplexes the stimulus light L1 introduced from the first window 13 and the illumination light L2 introduced from the second window 14 and guides the light to the mounting section 11. In this embodiment, the multiplexing element 21 is configured with a dichroic mirror 22. The dichroic mirror 22 reflects the stimulus light L1 introduced from the first window 13 and guides it to the mounting section 11, and transmits the illumination light L2 introduced from the second window 14 and guides it to the mounting section 11 coaxially with the stimulus light L1.

In this embodiment, the dichroic mirror 22 is disposed in the housing 2 while being held by the holder 23. The holder 23 is formed in a block shape, for example, from the same hard plastic as the housing 2. As shown in Figs. 2 and 3, the holder 23 has a fixing portion 24 to which the dichroic mirror 22 is fixed, an opening 25 that passes the illumination light L2 introduced from the second window portion 14, and a sliding portion 26 that slidably attaches the holder 23 to the inner wall of the housing 2.

As shown in FIG. 3, the fixing part 24 has an inclined surface 27 to which the edge of the dichroic mirror 22 is fixed. The dichroic mirror 22 fixed to the inclined surface 27 is disposed at an angle of 45° with respect to the optical axis of the stimulus light L1 introduced from the first window part 13 and the illumination light L2 introduced from the second window part 14. As shown in FIG. 2, the opening part 25 is provided on the upper surface side of the holding part 23 in correspondence with the second window part 14.

The dichroic mirror 22 can be seen through the opening 25 from the top side of the holding section 23. The shape of the opening 25 is not particularly limited and can be various shapes such as a circle, an ellipse, an oval, or a rectangle. In this embodiment, the opening 25 has rounded corners. In the holding section 23, below the dichroic mirror 22 is a space that allows the stimulating light L1 and the illumination light L2 combined by the dichroic mirror 22 to pass toward the mounting section 11 (see FIG. 3).

As shown in FIG. 2, the sliding portion 26 is provided on each of the end faces 23a, 23a of the housing 2 in the width direction of the holding portion 23. In this embodiment, the sliding portion 26 is configured by a pair of screws 28, 28. The screws 28, 28 are arranged on each of the end faces 23a, 23a at a predetermined interval in the longitudinal direction of the housing 2. The screws 28, 28 are screwed into screw holes (not shown) provided on the end faces 23a, 23a, and the amount of protrusion from the end faces 23a, 23a can be adjusted by adjusting the amount of screwing.

By adjusting the amount of protrusion of the screws 28, 28 and abutting the tips of the screws 28, 28 on the end faces 23a, 23a against the inner wall surfaces of the widthwise side portions 8, 9 of the housing 2, respectively, the holding portion 23 (dichroic mirror 22) can be slid in the longitudinal direction within the housing 2 while maintaining the orientation of the holding portion 23 within the housing 2. In this embodiment, the holding portion 23 can be slid in the longitudinal direction of the housing 2 between a first position P1 (see FIG. 3) facing the mounting portion 11 and a second position P2 (see FIG. 4) removed from the mounting portion 11.

The first position P1 is set in an area connecting the second window 14 and the mounting portion 11. The second position P2 is set on the opposite side of the first window 13 with respect to the first position P1. The second position P2 is set at a position where the holding portion 23 does not hit the fan 33 described later and the holding portion 23 does not overlap with the mounting portion 11 in a plan view of the housing 2. When the holding portion 23 is positioned at the first position P1, as shown in FIG. 3, the dichroic mirror 22 is disposed at a portion where the optical axis of the stimulus light L1 introduced from the first window 13 and the optical axis of the illumination light L2 introduced from the second window 14 intersect, and the stimulus light L1 and the illumination light L2 can be multiplexed. When the holding portion 23 is positioned at the second position P2, as shown in FIG. 4, the mounting portion 11 is exposed from the holding portion 23 in a plan view. Therefore, when the lid 4 is removed, the tray D can be placed on and removed from the placement section 11.

In this embodiment, the holding part 23 can be slid not only in the longitudinal direction of the housing 2, but also in the height direction of the housing 2. Therefore, when the lid part 4 is removed from the main body part 3, the holding part 23 can be removed from inside the housing 2 by sliding the holding part 23 in the height direction of the housing 2. This improves the ease of operation when replacing the holding part 23 with one that holds a dichroic mirror 22 with different optical characteristics.

The housing 2 is also provided with a temperature control section 31, a gas inlet section 32, and a fan 33 as components for maintaining the environment of the sample S placed on the mounting section 11. The temperature control section 31 is a section that adjusts the temperature of the sample S placed on the mounting section 11 using a temperature control liquid such as water. In this embodiment, as shown in Figures 2 and 3, a supply port 34 and a discharge port 35 for the temperature control liquid are provided at the bottom of the longitudinal side section 6 of the housing 2.

When constructing the temperature adjustment unit 31, the bottom surface 5 of the housing 2 has a double structure made up of an upper base portion 36 and a lower base portion 37. A predetermined pattern of cutouts is provided on the bottom surface side of the upper base portion 36 and the top surface side of the lower base portion 37, and a flow pipe 38 for the temperature adjustment liquid is formed by overlapping the upper base portion 36 and the lower base portion 37.

As shown in FIG. 2, the flow pipe 38 extends in the longitudinal direction of the housing 2 from the supply port 34 toward the mounting portion 11, makes a half turn around the outer periphery of the mounting portion 11, and then extends in the longitudinal direction of the housing 2 from the mounting portion 11 toward the discharge port 35. The temperature of the sample S can be maintained at a desired temperature by continuously supplying water (hot water) of a predetermined temperature to the flow pipe 38 as necessary.

The gas inlet 32 is a portion that introduces gas for adjusting the atmosphere of the sample S. The fan 33 is a device that diffuses the gas introduced from the gas inlet 32 within the housing 2. In this embodiment, as shown in Figs. 2 and 3, a gas inlet 39 is provided at approximately the center of the side portion 6 in the longitudinal direction of the housing 2. The fan 33 is fixed to the bottom portion 5 in the region between the side portion 6 where the inlet 39 is provided and the mounting portion 11 so as to face the inlet 39. Examples of the gas introduced from the gas inlet 32 include carbon dioxide, nitrogen, carbon monoxide, and nitric oxide. In this embodiment, the sample S is a PAC-expressing cardiomyocyte, and the gas introduced from the gas inlet 32 is, for example, carbon dioxide.

Furthermore, the housing 2 is provided with a humidity adjustment unit 40 that adjusts the humidity inside the housing. In this embodiment, as shown in Fig. 2, a water inlet 41 is provided near the gas inlet 32 on the longitudinal side surface 6 of the housing 2. For example, a tray in which a water-replenishment member such as a sponge is placed is placed inside the housing 2, and water is supplied to the water-replenishment member by a peristaltic pump or the like through a pipe inserted into the inlet 41, thereby adjusting the humidity inside the housing 2.
[Example of using the sample observation module]

Figure 5 is a cross-sectional view showing an example of using the sample observation module. Figure 5 illustrates an example in which a light source 16 is attached to the first window 13 in advance. When using the sample observation module 1, the holder 23 is positioned at the second position P2 with the cover 4 removed from the main body 3, and the tray D on which the sample S is placed is placed on the placement section 11. After placing the tray D, the holder 23 is positioned at the first position P1, and the dichroic mirror 22 is placed at the intersection of the optical axis of the stimulus light L1 introduced from the first window 13 and the optical axis of the illumination light L2 introduced from the second window 14. In addition, one or more attenuation filters 17 are placed inside the first window 13 as necessary based on the type of sample S, etc.

After the dichroic mirror 22 and the attenuation filter 17 are placed, the cover 4 is attached to the main body 3 to shield the inside of the housing 2 from light. Then, the sample observation module 1 is placed on the stage of the microscope system, and the objective lens K of the microscope system is fitted into the third window 15. After the sample observation module 1 is placed, the temperature control unit 31 continuously supplies water (hot water) at a predetermined temperature to the flow pipe 38 to maintain the temperature of the sample S at a desired temperature. In addition, the gas introduction unit 32 introduces gas into the housing 2 from the introduction port 39, and the fan 33 circulates the gas within the housing 2. Furthermore, if necessary, a tray in which a water-replenishment member such as a sponge is placed is placed within the housing 2. The humidity within the housing 2 is adjusted by supplying water to the water-replenishment member through a tube inserted into the water introduction port 41 (see FIG. 2) of the humidity control unit 40.

Next, the light source 16 is driven to introduce the stimulus light L1 from the first window 13. Also, the illumination light L2 is introduced from the second window 14. The stimulus light L1 introduced from the first window 13 is attenuated in intensity by the attenuation filter 17, reflected by the dichroic mirror 22, and incident on the sample S on the tray D placed on the mounting section 11. This allows optical manipulation of the sample S by the stimulus light L1 to be performed over time. The illumination light L2 introduced from the second window 14 passes through the wavelength selection element 18 of the second window 14, and the component corresponding to the wavelength of the stimulus light L1 is cut.

The illumination light L2 passes through the dichroic mirror 22 and is combined with the stimulus light L1. The illumination light L2 combined with the stimulus light L1 is incident on the sample S on the tray D placed on the placement unit 11 coaxially with the stimulus light L1. The illumination light L2 then passes through the tray D and is incident on the objective lens K fitted in the third window 15. As a result, on the microscope system side, bright-field observation of changes in the sample S accompanying optical manipulation is performed over time.
[Action and Effect]

As described above, in the sample observation module 1, a multiplexing element 21 that multiplexes the stimulus light L1 introduced from the first window 13 and the illumination light L2 introduced from the second window 14 and guides the light to the mounting part 11 is provided in the light-shielding housing 2. This makes it possible to simultaneously apply an optical stimulus to the sample S and irradiate it with the illumination light L2, allowing changes in the sample S associated with optical manipulation to be observed over time. Due to the simple configuration in which the multiplexing element 21 is arranged in the housing 2, the sample observation module 1 is also highly convenient when used in combination with a microscope system, etc.

In the sample observation module 1, a holder 23 is provided in the housing 2, which holds the multiplexing element 21 and is slidable between a first position P1 facing the mounting section 11 and a second position P2 removed from the mounting section 11. By sliding the holder 23 from the first position P1 to the second position P2, the workability of placing the sample S on the mounting section 11 can be improved while saving space.

In the sample observation module 1, one or more attenuation filters 17 that attenuate the intensity of the stimulus light L1 are detachably provided inside the first window portion 13. This allows the intensity of the stimulus light L1 to be adjusted with a simple configuration. Therefore, appropriate light operations can be performed according to the type of sample S.

In the sample observation module 1, a wavelength selection element 18 is provided in the second window 14, which transmits the illumination light L2 while cutting out light of the wavelength of the stimulus light L1. This makes it possible to prevent light other than the stimulus light L1 introduced from the first window 13 from contributing to the optical manipulation of the sample S. Therefore, the optical manipulation of the sample S can be controlled with greater precision.

In the sample observation module 1, a third window portion 15 is provided in the housing 2, which positions the objective lens K used to observe the sample S close to the mounting portion 11. This third window portion 15 makes it easy to observe the sample S using the objective lens K.

The sample observation module 1 is provided with a temperature adjustment unit 31, a gas introduction unit 32, and a fan 33 as components for maintaining the environment of the sample S. These components make it possible to maintain a good environment for the sample S, and to more continuously observe changes in the sample S accompanying light manipulation. The sample observation module 1 is also provided with a humidity adjustment unit 40 that adjusts the humidity inside the housing 2.
The humidity adjustment unit 40 also contributes to maintaining a good environment for the sample S, and can prevent the moisture in the sample S from evaporating due to the driving of the fan 33, for example.
[Modification]

The present disclosure is not limited to the above embodiment. For example, in the above embodiment, the intensity of the stimulus light L1 is adjusted using one or more attenuation filters 17, but the intensity of the stimulus light L1 may be adjusted by controlling the output of the light source 16 itself. In addition, in the above embodiment, bright field observation of the sample S is performed using the third window portion 15, but a fluorescent observation of the sample S through the third window portion 15 may be performed. In the above embodiment, water is supplied to the housing 2 by the humidity adjustment unit 40, but a gas-containing water such as carbonated water may be supplied. In this case, humidity adjustment and gas introduction can be performed simultaneously, so that the introduction of gas by the gas introduction unit 32 can be omitted.

1...sample observation module, 2...housing, 11...mounting section, 13...first window section, 14...second window section, 15...third window section, 17...attenuation filter, 18...wavelength selection element, 21...wave combining element, 31...temperature control section, 32...gas introduction section, 33...fan, 40...humidity control section, K...objective lens (external lens), L1...stimulation light, L2...illumination light, S...sample.

Claims (9)

A sample observation module for observing changes in a sample over time due to optical manipulation,
a housing formed of a light-shielding material and having a mounting portion therein on which the sample is placed;
the housing is provided with a first window portion for introducing a stimulating light used for optical manipulation of the sample into the housing, and a second window portion for introducing an illumination light used for observation of the sample into the housing;
A sample observation module in which a combining element is provided within the housing to combine the stimulus light introduced through the first window portion and the illumination light introduced through the second window portion and guide the combined light to the placement portion. The sample observation module according to claim 1, further comprising a holder in the housing that holds the multiplexing element and is slidable between a first position facing the mounting portion and a second position removed from the mounting portion. A sample observation module according to claim 1 or 2, wherein one or more attenuation filters that attenuate the intensity of the stimulating light are detachably provided inside the first window portion. The sample observation module according to any one of claims 1 to 3, wherein the second window portion is provided with a wavelength selection element that transmits the illumination light while cutting off light of the wavelength of the stimulation light. The sample observation module according to any one of claims 1 to 4, wherein the housing is provided with a third window portion for placing an external lens used for observing the sample adjacent to the placement portion. The sample observation module according to any one of claims 1 to 5, wherein the housing is provided with a temperature adjustment unit that adjusts the temperature of the sample placed on the placement unit. The sample observation module according to any one of claims 1 to 6, wherein the housing is provided with a gas inlet for introducing a gas into the housing to maintain the environment of the sample. The sample observation module according to claim 7, further comprising a fan provided within the housing for circulating the gas within the housing. The sample observation module according to any one of claims 1 to 8, wherein the housing is provided with a humidity adjustment unit that adjusts the humidity inside the housing.

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