JP2006115760A - Cell culture apparatus and optical observation device - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a cell culture apparatus and an optical observation device free from deterioration of image by cloudiness and stain and capable of preventing proliferation of bacteria and fungi. <P>SOLUTION: In the cell culture apparatus 3 equipped with a transmission window 23 applied to the optical observation device having at least either one of observation optical system and illumination optical system and used for transmitting light of the observation optical system or the illumination optical system, the transmission window 23 is coated with a photocatalyst. <P>COPYRIGHT: (C)2006,JPO&NCIPI

Description

  The present invention relates to a technique related to antifogging, antifouling, and antibacterial of a cell culture device and an optical observation device for observing living cells and living bodies.

First, observation of cultured cells using an inverted microscope equipped with a cell culture device will be described as a first conventional example.
When observing live cultured cells optically, as shown in the example of the inverted microscope of FIG. 9, the cell culture apparatus 3 for keeping the cells alive is loaded on the stage 14 and observed.

  Hereinafter, a detailed description thereof will be given with reference to FIGS. 9 and 10. FIG. 9 is an overall schematic view of an inverted microscope, and FIG. 10 is an enlarged view of the periphery of a specimen 20 (hereinafter referred to as “cultured cells”, “tissue cells”, or simply “cells”).

The configuration and operation of the inverted microscope 16 will be described in the order of transmission illumination observation and fluorescence observation.
In the case of transmission illumination observation, the illumination light emitted from the light source 4 illuminates the cultured cell 20 as a specimen via the transmission illumination optical system 5. Observation light from the illuminated cultured cell 20 is imaged by the objective lens 1 and a relay observation optical system 57 (shown in FIG. 9 shows an optical element arranged on the optical axis of the observation optical system). The eyepiece 17 can be visually observed. In the case of phase difference observation, a ring slit disposed in the transmission illumination optical system 5 is projected onto the phase film in the objective lens 1, or in the case of DIC observation, the illumination optical system 5 and the observation optical system 57 include polarizing plates. It is necessary to arrange a DIC prism.

  In the case of fluorescence observation, the illumination light from the light source 7 is condensed by the epi-illumination optical system 8 (in FIG. 9, an optical element disposed on the optical axis of the epi-illumination optical system is shown). At the same time, the wavelength is selected by the excitation filter 9 so as to obtain an optimum excitation wavelength for the fluorescent dye stained on the cultured cells 20, reflected by the dichroic mirror 10, and irradiated to the cultured cells 20. The fluorescence emitted from the cultured cells 20 passes through the dichroic mirror 10 and is selected by the absorption filter 11 only for the fluorescence wavelength necessary for observation. Thereafter, the eyepiece 17 can be visually observed as in the case of the above-mentioned transmitted illumination observation.

  The excitation filter 9, the dichroic mirror 10, and the absorption filter 11 are arranged in a turret-shaped mirror unit cassette 12, and the mirror unit cassette 12 is rotatable about a rotation shaft 13. As a result, the three types of filters 9, 10, 11 can be integrally inserted into and removed from the optical axis 2 of the objective lens.

  In the above, the optical operation in the case of performing the transmission illumination observation and the fluorescence observation has been described. Actually, the stage 14 loaded with the cultured cells 20 is operated by operating the stage handle (not shown) to observe the place where the cell is to be observed. It can be observed by bringing it onto the optical axis 2 and further operating the semi-focus handle 15 to move the objective lens 1 up and down to focus on the cell 20.

Next, the periphery of the cultured cell 20 will be described.
The cultured cells 20 are loaded on a cover glass 19 attached to a holed portion of the bottom of the glass bottom dish 21. The outer periphery of the glass bottom dish 21 is covered with a heat insulating box 22 and a transmission window 23 that can transmit the transmitted illumination light. A heater 32 is disposed on the bottom surface of the heat insulating box 22, and the cultured cells 20 can be maintained at 37 ° C. via the glass bottom dish 21.

On the other hand, a bath is provided around the glass bottom dish 21 in the heat insulating box 22, and water 24 is contained in the bath. By appropriately evaporating the water, the humidity in the heat insulation box 22 is kept at 100%, and the evaporation of the culture solution 58 is suppressed. Further, a tube 27 is provided on the side wall of the heat insulating box 22, and the cultured cell Ph is maintained at about 7 by supplying CO ₂ from the CO ₂ cylinder 26 through the tube 27.

  In addition, a hole 25 is provided in the bottom surface of the heat insulating box 22 so that the cultured cell is focused even on the objective lens 1 having a high NA and a short WD, and the objective lens can be inserted therein. The hole 25 is slightly larger so that the inner wall of the hole 25 does not interfere with the objective lens 1 even when the stage 14 is moved.

  In the configuration as described above, the inside of the heat insulation box 22 is a temperature of 37 ° C. and a humidity of 100%, while the objective lens 1 is in a room temperature environment of about 20 ° C., so that the objective lens 1 is cultured for focusing. When approaching the cell 20, the low-temperature objective lens is suddenly exposed to a high-temperature environment, and the temperature difference causes the surface of the front lens 18 of the objective lens 1 to condense and become cloudy. Since the objective lens of the microscope has a large NA, even a slight cloudiness leads to deterioration of the microscope image, and satisfactory observation cannot be performed. In particular, in the case of an inverted microscope, as described above, when an objective lens having a high NA and a short WD is used, the heat insulation box 22 must be moved together with the cultured cells 20, so that a hole 25 is formed on the bottom surface of the heat insulation box 22. The objective lens must be provided, and the objective lens is exposed to the high-humidity air leaking from the inside of the heat insulating box 22, and the front lens 18 of the objective lens 1 is likely to be cloudy. Further, since the WD is short, the front lens 18 of the objective lens 1 approaches the cultured cell 20 having a high temperature, which is also a factor that tends to be cloudy.

  Further, the transmission window 23 on the upper surface of the heat insulating box 22 is also in the same situation as the objective lens, and the lower side (the inner surface of the heat insulating box) tends to be cloudy. The transmission window 23 is a part through which the transmission illumination light is transmitted. In particular, in the case of phase difference observation, the ring slit in the transmission illumination optical system 5 is projected onto the billion phase film in the objective lens. If it is cloudy, the ring slit is not properly projected on the phase film, and the contrast of the image is deteriorated. In the case of DIC observation, the observation is performed with a contrast by interfering with the illumination light separated into two on the specimen. However, if the transmission window 23 is cloudy, appropriate interference does not occur, and the contrast of the image is also reduced. It gets worse.

  Although there is an example in which fogging is prevented by warming the transmission window with a heater, a heater must be purposely provided, which is not economical. In addition, although not shown, the culture solution 58 in the heat insulation box 22 and various chemicals added for observing the physiological reaction of the cells volatilize, and the lower surface of the transmission window 23 and the tip of the objective lens 1 are volatilized. The ball 18 may be soiled. This also deteriorates the microscopic image as in the above-described cloudiness.

  Furthermore, since the inside of the heat insulation box 22 is not completely sealed with respect to the outside as described above, bacteria or the like may enter from the outside. In some cases, the cultured cells are infected with bacteria and the cells become weak or die.

  Next, as a second conventional example, the prior art when observing a living body such as a tissue cell or a rat with an upright microscope is referred to in second and third embodiments described later with reference to FIGS. Will be described with reference to FIG.

  FIG. 4 is a diagram showing an overall schematic configuration, and FIGS. 5 and 6 are detailed views around the specimen. 4 to 6, the same parts as those in FIGS. 9 and 10 are denoted by the same reference numerals.

  The overall configuration and operation of the upright microscope will be described with reference to FIG. The basic configuration is the same as that of the first conventional example, the objective lens 1 for observing the specimen, the stage 14 for moving the specimen, and the semi-focus handle 15 for moving the objective lens 1 up and down to focus. , A light source 7 for epi-illumination, an epi-illumination optical system 8, an excitation filter 9 for fluorescence observation, a dichroic mirror 10, an absorption filter 11, and the three types of filters 9.10 and 11 are placed on the optical axis 2 of the objective lens. The rotating shaft 13 for insertion / removal, the light source 4 for transmitted illumination, the transmitted illumination optical system 5 and the condenser lens 33 are included, and can be visually observed with the eyepiece 17 by the same operation as in the first conventional example. It has become. The difference from the first conventional example is that the objective lens 1 is located above the specimen and is observed from above.

  The periphery of the tissue cell 20 will be described with reference to FIG. Since this case is also almost the same as the first conventional example, detailed description thereof is omitted, and only different portions will be described.

  The objective lens 1 is disposed on the specimen (tissue cell) 20 and is a water immersion objective lens for immersing the objective lens 1 in the culture solution 58 for observation. In addition, the upper surface of the heat insulation box 22 does not transmit transmitted illumination light as in the first conventional example, and is therefore made of an opaque material integrated with the side wall. Instead, the heat insulation box is provided so that the objective lens 1 can be inserted. The hole 59 is vacant on the upper surface of 22. Since the tissue cell 20 is moved by the stage 14 together with the heat insulating box 22, the hole 59 is slightly larger. In addition, the cell culture space in the heat insulation box 22 is fitted to the outer periphery of the objective lens 1 so that the cell culture space in the heat insulation box 22 can be sealed from the outside even when the tissue cells 20 move, and the flange portion of the cell 20 A ring 222b is provided so as to be in close contact with each other. However, it is not completely sealed because it is in close contact with its own weight. Unlike the first conventional example, a condenser lens 33 for condensing the transmitted illumination light is arranged under the heat insulation box 22 instead of the objective lens 1, but the WD is longer than the objective lens. In addition, it is not necessary to make a hole in the bottom surface of the heat insulation box 22, and instead, a transmission window 22 a made of a glass member that transmits transmitted illumination light is formed on the bottom surface of the heat insulation box 22 to improve the sealing performance.

  Thus, when observing tissue cells, unlike the first conventional example, the water immersion objective lens 1 is used, so there is no concern about the tip 18 of the objective lens 1 becoming cloudy, but the culture solution 58 is always present. Therefore, the front lens 18 of the objective lens 1 is easily contaminated, and this contamination causes deterioration of the image. Further, the transmission window 22a for transmitting the transmitted illumination light is likely to be clouded as in the first conventional example, and the image is deteriorated particularly by a phase difference mirror or a DIC microscope. Further, as in the first conventional example, the inner surface of the heat insulating box 22 becomes dirty due to volatilization of the culture solution or the like, or is contaminated by bacteria due to insufficient sealing, which also causes the same problem as in the first conventional example.

  With reference to FIG. 6, the case where the specimen is a rat 60, which is an original, will be described. The difference from the case of tissue cell observation is only the shape of the objective lens 1 as an apparatus configuration. In order to observe the internal organs of the rat, a part of the body surface is incised, a hole is made, and the objective lens 1 is inserted for observation. The objective lens 1 has a smaller outer diameter than that for tissue cell observation. , Φ10 mm or less. In this case, the illumination light hardly transmits, so that the epi-illumination is mainly used, and the condenser lens is unnecessary.

Thus, when observing a rat, since the objective lens 1 in a state close to room temperature of about 20 ° C. is inserted into the body of the rat at 37 ° C., the tip 18 is very easily clouded in the case of a dry objective lens. When the front lens 18 of the objective lens 1 is actually fogged, the objective lens 1 is pulled out and warmed with hot water or the like and then inserted again. However, this operation is complicated. In addition, since a living rat is being observed, it is necessary to finish the observation in as short a time as possible and sew and close the hole opened by surgery. Otherwise, the rat will be debilitated or in the worst case dead. This makes it impossible to continue the experiment next time, which is very wasteful. Of course, if the objective lens 1 suddenly becomes cloudy, the data that was about to be obtained in the experiment may be re-acquired, but such a case is also wasteful.
As described above, when observing a live rat, the outer periphery of the objective lens 1 and the like are easily soiled, causing bacteria and mold to propagate, and there is a risk of infecting the rat during the next experiment.

  In addition, photocatalysts that are excited by light and exhibit antifogging, antifouling, and antibacterial effects are generally known (see Patent Document 1, Patent Document 2, or Patent Document 3, etc.). However, there is no example applied to an objective lens of a microscope or a window of a cell culture device.

  In addition, the photocatalyst can not sustain its anti-fogging, antifouling and antibacterial effects unless it is irradiated with light, so it excites the photocatalyst in the case of optical observation devices and cell culture devices that are used indoors where sunlight does not shine. A separate light source is required, but it occupies a lot of space and is not economical.

Further, since the photocatalyst is reflected at the interface with the air, there is a problem in that the contrast of the image is lowered by flare when it is coated in the observation optical system of the optical observation apparatus.
Japanese Patent Laid-Open No. 5-4816 JP-A-6-65012 JP-A-6-298532

  It is an object of the present invention to provide a cell culture device and an optical observation device that can prevent the growth of bacteria and fungi without causing image deterioration due to cloudiness or dirt.

  The invention according to the aspect of the present invention is applied to an optical observation apparatus having at least one of an observation optical system and an illumination optical system, and includes a transmission window for transmitting light beams of the observation optical system or the illumination optical system. In the cell culture apparatus, the permeation window is coated with a photocatalyst. This technique can also be applied to an optical observation apparatus equipped with such a cell culture apparatus and an optical observation apparatus that directly observes a living body with an objective lens coated with a photocatalyst.

  According to the present invention, it is possible to provide a cell culture device and an optical observation device that can prevent the growth of bacteria and fungi without causing image deterioration due to cloudiness or dirt.

  Embodiments of the present invention will be described with reference to the drawings.

(First embodiment)
A first embodiment of the present invention will be described with reference to FIGS.
FIG. 1 is a diagram showing an overall schematic configuration of the optical observation apparatus according to the first embodiment, FIG. 2 is a detailed view around a specimen (cultured cell) 20 in FIG. 1, and FIG. 3 shows an epi-illumination portion from FIG. It is a figure for extracting and explaining another modification. The configuration of the present embodiment is almost the same as that of the first conventional example, and only different portions will be described.

First, the overall configuration of FIG. 1 will be described.
In the optical path of the transmission illumination optical system 5, an excitation filter 30 that selectively transmits only the 360-nm UV light necessary for exciting the photocatalyst in the illumination light from the light source 4 is detachable on the illumination optical path. Has been placed. Note that the switching means such as a known slider or turret may be used as the means for inserting / removing the excitation filter 30.

  In the state shown in FIG. 1, the filter in the mirror unit cassette 12 is provided with fluorescence observation filter sets 9, 10, 11 for excitation at 490 nm in the optical path, and filter sets 28, 29 for photocatalyst excitation are arranged on the opposite side. Has been. The excitation filter 28 selectively transmits only the 360 nm UV light that efficiently excites the photocatalyst among the illumination light from the light source 7, and the mirror 29 is a mirror coated with aluminum or silver. 90% or more from the visible region to the visible region.

  The cultured cells 20 are stained with a fluorescent dye FITC excited at, for example, 499 nm.

Next, the periphery of the specimen (cultured cell) 20 will be described with reference to FIG.
In the present embodiment, a photocatalyst made of titanium oxide is coated on a portion indicated by a thick solid line in the drawing, that is, on the outer surface of the front lens 18 of the objective lens 1, both surfaces of the transmission window 23, and the inner surface of the heat insulating box 22. It is. Further, a sterilizing lamp 31 for killing bacteria is provided in the heat insulating box 22, and the sterilizing lamp 31 also includes 360 nm UV light that can excite the photocatalyst.

The operation for exerting the effect of the photocatalyst with the above configuration will be described below.
In the state of FIG. 1, as described in the prior art, the filter set in the mirror unit cassette is in a state in which fluorescence observation filter sets 9, 10, 11 are arranged in the optical path and fluorescence observation is possible. In this state, the UV light for photocatalyst excitation from the light source 7 is cut by the excitation filter 9, so there is no concern that the UV light for photocatalyst excitation becomes a flare to the observation light. Of course, the epi-illumination light source 7 is turned on, and the transmitted illumination light source 4 and the germicidal lamp 31 are turned off. In this state, the fluorescent image is observed with the eyepiece 17. When the fluorescence observation is finished, the epi-illumination light source 7 is turned off.

Next, a procedure for exciting the photocatalyst after fluorescence observation will be described.
First, the filter sets in the mirror unit cassette are switched, and the filter sets 28 and 29 for photocatalyst excitation are inserted into the optical path. Next, when the epi-illumination light source 7 is turned on, the illumination light from the epi-illumination light source 7 is selected by the excitation filter 28 as UV light that can excite the photocatalyst, and then reflected by the mirror 29 toward the objective lens 1. The photocatalyst coated on the front lens 18 of the objective lens 1 and the transmission window 23 on the upper surface of the heat insulating box 22 is excited. Further, if the germicidal lamp 31 in the heat insulation box 22 is turned on as necessary, the incident light source 7 can illuminate the corners of the heat insulation box 22 that are difficult to irradiate. Can be excited.

  In order to excite the photocatalyst further, an excitation filter 30 may be inserted into the transmission illumination optical system 5 to turn on the transmission illumination light source 4. Thereby, the photocatalyst of the site | part which is hard to irradiate illumination light, such as the back side of the germicidal lamp 31, can also be excited.

  When the photocatalyst is sufficiently excited, each light source is turned off.

Next, a procedure for performing transmitted illumination observation will be described.
The transmitted illumination light source 4 is turned on. In this case, the epi-illumination light source 7 and the germicidal lamp 31 are turned off. Next, four types of filter sets in the mirror unit cassette can be switched. The filter sets in the mirror unit cassette are switched to form holes (not shown) and observed with the eyepiece 17.

  The above series of operations for fluorescence observation, photocatalyst excitation, and transmitted illumination observation may be automatically performed by motorization. Alternatively, the photocatalyst may be excited.

  In the present embodiment, the excitation filter 28 and the mirror 29 are used as a set as the photocatalyst excitation filter, but for example, a dichroic mirror that reflects 360 nm may be used instead of the excitation filter 28 and the mirror 29. In this case, since one dichroic mirror can be arranged at the position of the mirror 29 and the excitation filter 28 can be eliminated, the number of filters is reduced and the cost is reduced.

  Further, the excitation filter 28 and the dichroic mirror that reflects 360 nm may be used in combination. In this case, a filter set of an excitation filter, a dichroic mirror, and an absorption filter for observing a fluorescent dye such as 360 nm excitation DAPI can be used as it is, so there is no need to purchase a new one if you already have it. It is economical because it can be used for DAPI observation.

  Further, as shown in FIG. 3 in a state where the photocatalyst is excited, the dichroic mirror has a characteristic of reflecting 360 nm and 490 nm, and the absorption filter 11 has a characteristic for FITC. As shown in the figure, an excitation filter is arranged in the middle of the optical path of the epi-illumination optical system so that a plurality of types can be inserted and removed by a known switching function such as a slider and a turret. When performing FITC observation, it is possible to switch between photocatalytic excitation and single light observation by inserting the FITC excitation filter 9 into the optical path.

  If the excitation light for photocatalyst and the excitation light for fluorescence observation are sufficiently strong, a half mirror or glass may be used instead of the dichroic mirror. In this way, since the filter set space in the mirror unit cassette is not occupied exclusively for the photocatalyst, the four limited filter set mounting spaces can be used effectively for observation of other fluorescent dyes. it can.

  In this embodiment, the photocatalyst is UV-excited at 360 nm, and the fluorescent dye is FITC with 490 nm excitation. However, depending on the fluorescent dye to be observed, a photocatalyst having an excitation characteristic that does not excite the fluorescent dye is used. In addition, various filters as described above can be freely combined accordingly.

(Second Embodiment)
A second embodiment of the present invention will be described with reference to FIGS.
4 and 5 have been described in the second conventional example, detailed description thereof will be omitted. In the present embodiment, a method for preventing fogging and dirt in an upright microscope will be described. FIG. 4 is a diagram showing an overall schematic configuration, and FIG. 5 is a detailed view around the specimen.

  In the present embodiment, unlike the first embodiment, the objective lens 1 is disposed on the specimen (tissue cell) 20, and the immersion objective lens for immersing and observing the objective lens 1 in the culture solution 58. It has become. For this reason, in this embodiment, as shown in FIG. 5, as shown by a thick solid line, the inner surface of the heat insulating box 22, the inner exposed portion of the glass transmission window 22a, and the tip of the condenser lens 33 (thick broken line) ) Is coated with a photocatalyst. Thereby, the effect similar to 1st Embodiment is acquired.

(Third embodiment)
A third embodiment of the present invention will be described with reference to FIG. In the present embodiment, the overall configuration is substantially the same as in FIG. 4, and in the configuration according to FIG. 4, the cell culture device is a rat.

  In this case, in order to prevent fogging, contamination, and the like, a portion that may be in direct contact with the rat is coated with a photocatalyst. Specifically, the photocatalyst is coated on the outer surface of the main body of the objective lens 1, the tip portion, and the inner surface from the tip portion to the tip lens, and further on the exposed portion of the tip lens. In the present embodiment, a cover glass (not shown) may be provided at the tip of the objective lens 1 so that the front glass side is not soiled from the cover glass. In this case, the cover glass is coated with a photocatalyst. May be.

(Fourth embodiment)
A fourth embodiment of the present invention will be described with reference to FIG. In FIG. 7, the same parts as those in FIG.

  In FIG. 7, a light source unit 53, a detection unit including a detector 46, and a housing 36 including a scanning unit 35 are connected by a single mode fiber 47 and a multimode fiber 40, respectively. Laser light from light sources 50 to 52 of three different wavelengths arranged in the light source unit 53 is transmitted through the dichroic mirrors 49a and 49b and the mirror 49c, respectively, or combined light ("laser light" for convenience of explanation). Is referred to as the connection point of the single mode fiber 48. The laser beam guided to the detection unit via the connection end 45 of the single mode fiber 48 is reflected by the mirror 44 and the dichroic mirror 43 and guided to the multimode fiber 40 and guided to the housing 36.

  The laser light guided to the housing 36 is deflected by the scanning unit 35 having mirrors 35 a and 35 b and scanned on the rat 60 through the imaging lens 37, the relay lens 38 and the objective lens 1. The objective lens 1 is detachably held by the holding portion 1a of the drive mechanism 34, and can be moved in the optical axis direction by the drive mechanism 34. Light (including fluorescence and reflected light) from the rat 60 follows a path opposite to that of the laser light, and enters the multimode fiber 40 from the objective lens 1 through the scanning unit 35 and the coupling lens 39. Note that the core of the incident end face of the multimode fiber 40 has a pinhole function in the confocal microscope. The light emitted from the multimode fiber 40 is incident only on the detector 46 through the relay lens 41, the dichroic mirror 43, and the barrier filter 42 of the detection unit.

  In the above configuration, in the present embodiment, the photocatalyst is coated on the surface of the housing 36 and the exposed surface portion of the objective lens 1 indicated by a thick solid line in the drawing.

  In the present embodiment, unlike the third embodiment, the surface of the objective lens 1 is observed without contacting the objective lens 1 with the rat 60, but the portion close to the rat 60 is also at risk of contamination by bacteria, etc. Antibacterial effect by photocatalyst becomes effective.

(Fifth embodiment)
A fifth embodiment of the present invention will be described with reference to FIG. 8, the same parts as those in FIGS. 1 and 7 are denoted by the same reference numerals.

  In the present embodiment, the epi-illumination light source 7 is used as the light source. As in the third embodiment, a part of the body surface is opened and a hole is formed, and the objective lens 1 is inserted into the rat 60 and observed. ing. In the present embodiment, the detector (CCD) 15 and the portion including the light source 7 and the housing 36 are connected by the bundle fiber 54, and, as in the fourth embodiment, a thick solid line in the figure. The surface of the housing 36 and the exposed surface of the objective lens 1 are coated with a photocatalyst.

  In the present embodiment, unlike the fourth embodiment, the scanning unit 35 does not exist, and a two-dimensional image of the rat 60 is formed on the end surface of the bundle fiber 54 by the objective lens 1 and the coupling lens 37. The two-dimensional image is transmitted to the exit end face of the bundle fiber 54 as it is. Further, the transmitted image is picked up by a detector (CCD) 56 with a collimator lens 61 and a relay lens 55 and can be observed on a monitor (not shown). Here, the light source 7 is a white light source such as a mercury lamp, and the wavelength is selected to an excitation wavelength necessary for the fluorescent dye stained on the rat 60 by the excitation filter 62, and the end face of the bundle fiber 54 is formed by the collector lens 57 and the collimator lens 61. It is designed to be illuminated uniformly. Therefore, the rat 60 which is a sample surface optically conjugate with the end face of the bundle fiber 54 is uniformly irradiated. At this time, the dichroic mirror 43 reflects the excitation light, transmits the fluorescence emitted from the rat 60, is selected by the absorption filter 42 as an observable light receiving wavelength, and is guided to the detector 56.

  With the configuration according to each of the above embodiments, the following effects can be obtained.

  1. Since the photocatalyst is coated on the transmission window of the cell culture device for transmitting the light of the observation optical system or illumination optical system, the transmission window that has a large temperature difference and tends to become cloudy is less likely to cloud, and the observation image does not deteriorate and is appropriate. Lighting is possible. In addition, since the photocatalyst has an antibacterial effect, it is possible to prevent cells from being killed by bacteria when coated on the cell culture space side of the transmission window. Further, since the photocatalyst has a stain prevention effect, similarly to the fog prevention effect, the observation due to the stain and the adverse effect on the illumination are reduced, and a clear observation image can be obtained.

  2. Since the inner surface of the cell culture space is coated with a photocatalyst, an antibacterial effect is obtained. In other words, in a cell culture device equipped with an optical observation device, when an observation is made with a high NA, a small objective lens of the optical observation device is brought close to a specimen (cell), and an objective lens is used to change the observation position in that state. However, since the specimen (cell) must be moved relatively, it is difficult to completely block the observation space and the cell culture space, and bacteria from the observation space can easily enter the culture space. By coating the inner surface of the cell culture space with a photocatalyst, it is possible to prevent the cells from being killed by bacteria due to the antibacterial effect of the photocatalyst.

  3. In the optical observation apparatus, the photocatalyst coating is applied to a microscope in which the image is easily deteriorated due to the influence of cloudiness and dirt, so that the effect of preventing image deterioration is great, and a clearer microscope image can be observed.

  4). In many cases, the cell culture apparatus captures a germicidal lamp for sterilizing the cell culture space. If the germicidal lamp includes a wavelength capable of exciting the photocatalyst, the photocatalyst can be excited by the germicidal lamp. Therefore, it is not necessary to provide a dedicated light source for exciting the photocatalyst, which is space-saving and economical.

  5. When observing living organisms or living cells, the objective lens of an optical observation apparatus is often observed close to or inserted into the living organism or living cells. Usually, the objective lens is at a room temperature of about 20 ° C., and living cells as well as living cells are about 37 ° C. in a culture environment. By coating, in addition to the above effects, the lens is less likely to be clouded and a clear observation image is obtained.

  6). In an optical observation apparatus having a housing with a built-in scanner and an objective lens that is detachably mounted, the apparatus near the specimen can be made smaller, so that it may be used particularly for in vivo observation. Many. For this reason, it is easily affected by fogging, but by coating the objective lens tip with a photocatalyst, it has an antifogging effect and a clear observation image can be obtained. In addition, antibacterial and antifouling effects can be obtained similarly.

  7. In an optical observation device having a housing with a bundle fiber for transmitting a sample image and a detachably mounted objective lens, the device near the sample can be made smaller in the same manner as described above. Therefore, it is often used for living body observation. For this reason, it is easily affected by fogging, but by coating the objective lens tip with a photocatalyst, it has an antifogging effect and a clear observation image can be obtained. In addition, antibacterial and antifouling effects can be obtained similarly.

  8). When the optical observation apparatus is a microscope, the objective lens has a large NA, and therefore is easily affected by image deterioration due to fogging or dirt. In addition, since the WD is relatively small, even if the WD is not inserted into the living body, the tip is easily clouded due to the ripening of the living body, or blood or the like is easily applied to the lens. However, by coating the objective lens tip with a photocatalyst, the same antifogging and antifouling effects as described above can be obtained, and a clear observation image can be obtained.

  9. When the objective lens has an outer diameter of φ10 or less, it is often observed by inserting a hole in a living body or inserting it into a vacant hole such as an ear or a rod. The tip is easily cloudy and dirty, and at the same time, the fog cannot be removed unless the inserted objective lens is pulled out. It has antifouling and effects, and a clear observation image can be obtained.

10. In an optical observation apparatus having a housing with a built-in scanner and an objective lens detachably mounted, the apparatus near the specimen can be made smaller in the same manner as described above. Often used. For this reason, it is easy to be affected by fogging, but by coating the outer periphery of the housing or objective lens with a photocatalyst, it has an antifogging effect and a clear observation image is obtained. Furthermore, the antibacterial and antifouling effects can be obtained in the same manner, and the portions are kept clean, so that the generation of bacteria and mold can be suppressed, and careless infection to the living body and the examiner can be suppressed. Furthermore, the photocatalyst also has an effect of reducing friction with the living body, so that the objective lens can be smoothly inserted into the living body.
11. In an optical observation apparatus having a housing to which a bundle fiber for transmitting a specimen image is attached and an objective lens is detachably attached, the apparatus near the specimen can be further reduced in size. Since it is often used for observation, the outer periphery of the housing and objective lens is also placed near the living body, making it easy to get dirty. However, antifouling and antibacterial effects are demonstrated by coating the outer periphery of the housing and objective lens and a photocatalyst. Because it is kept clean, the generation of bacteria and mold is suppressed, and careless infection to the living body and the examiner can be suppressed. Furthermore, the photocatalyst also has an effect of reducing friction with the living body, so that the objective lens can be smoothly inserted into the living body.
12 With a dry objective lens, if the tip is coated with a photocatalyst with high reflectivity at the interface with air, the image deteriorates due to flare, etc. Since the reflectance at the boundary surface can be lowered, a clear image with no flare can be secured, and antifouling and stake fungus effects can be obtained.

  13. The optical observation device has illumination means for illuminating the sample coaxially with the optical axis of the objective lens, and the illumination means can excite the photocatalyst (that is, includes a wavelength that can be excited). Therefore, there is no need to provide a dedicated light source for exciting the photocatalyst, which is space-saving and economical.

  14 A light source for illuminating the specimen, and an optical path dividing means for reflecting the illumination light from the light source toward the specimen through the objective lens, guiding the light coaxially with the objective lens, and transmitting the light emitted from the specimen And a reflecting means that reflects only the wavelength that can excite the photocatalyst among the illumination light from the light source, and the optical path dividing means and the reflecting Since the means can be inserted into and removed from the optical axis of the objective lens, it is possible to observe by irradiating the specimen with only the illumination light necessary for observation by inserting the optical path dividing means into the optical path during observation. By inserting a reflection means that reflects only the wavelength that can efficiently excite the photocatalyst into the optical path, more photocatalyst can be excited and antifogging, antibacterial, and antifouling effects can be maintained for a long time.

  15. Because the photocatalyst is made of titanium oxide, the antifogging, antifouling and antifungal effects are great.

  16. Since the excitation wavelength of the photocatalyst is different from the excitation wavelength of the fluorescent dye dyed on the specimen, the fluorescent dye will not be excited if only the wavelength required for the photocatalyst is irradiated. There is no fear of fading, and bright fluorescence observation becomes possible. That is, for example, since the excitation wavelength of the photocatalyst is 360 nm UV light, unlike the excitation wavelength of 490 nm of the fluorescent dye FITC that is stained on the cultured cells 20, there is no concern that FITC will fade.

  17. Since the excitation filter for the photocatalyst is detachably disposed in the optical path of the transmitted illumination optical system, more powerful excitation of the photocatalyst is possible. Further, although the wavelength that is easily excited differs depending on the type of the photocatalyst, excitation of the photocatalyst by the transmitted illumination light source is effective even when the wavelength is weak at the wavelength of the epi-illumination light source or the germicidal lamp. Thus, economic effects and versatility are enhanced by using both the fluorescence filter and the photocatalyst filter, or by switching only the excitation filter.

  The following inventions can be extracted by the above embodiments. In addition, the following invention may be applied independently and may be applied in combination as appropriate.

The cell culture device according to the first aspect of the present invention is applied to an optical observation device having at least one of an observation optical system and an illumination optical system, and transmits light rays of the observation optical system or illumination optical system. A cell culture apparatus provided with a permeation window, wherein the permeation window is coated with a photocatalyst.
The cell culture device according to the second aspect of the present invention is applied to an optical observation device having at least one of an observation optical system and an illumination optical system, and transmits light rays of the observation optical system or illumination optical system. A cell culture device provided with a transparent window, characterized in that the inner surface of the cell culture space is coated with a photocatalyst. In the first and second aspects, the following embodiments are preferable.
(1) The optical observation device is a microscope.
(2) The cell culture device includes a germicidal lamp, and the germicidal lamp includes a wavelength capable of exciting the photocatalyst.
(3) The optical observation apparatus further includes an illuminating unit that illuminates the specimen, and light from the illuminating unit includes a wavelength capable of exciting the photocatalyst.
(4) A light source for illuminating the specimen, and the illumination light from the light source is reflected toward the specimen through the objective lens, and is guided coaxially with the objective lens and transmits the light emitted from the specimen. An optical path splitting means; and a reflecting means that can be arranged at the same position as the optical path splitting means and reflects only a wavelength capable of exciting a photocatalyst among illumination light from the light source, and the optical path splitting means; The reflecting means is configured to be detachable from the optical axis of the objective lens.
(5) The photocatalyst is titanium oxide.
(6) The excitation wavelength of the photocatalyst is different from the excitation wavelength of the fluorescent dye stained on the specimen.

An optical observation apparatus according to a third aspect of the present invention is an optical observation apparatus that includes an objective lens and a condenser lens that collects illumination light onto a specimen, and is capable of observing a living body or living cells, The tip of the objective lens or the condenser lens is coated with a photocatalyst. In the third aspect, the following embodiment is preferable.
(1) The periphery and the tip of the front lens of the objective lens or the condenser lens are further coated with a photocatalyst.
(2) In (1), the outer periphery of the objective lens or the condenser lens is further coated with a photocatalyst.
(3) A built-in scanner for scanning a light source image on a sample, a housing on which the objective lens is detachably mounted, and the outside of the housing is coated with a photocatalyst.
(4) A bundle fiber for transmitting the specimen image is mounted, the housing is provided with a detachable mounting, and the outside of the housing is coated with a photocatalyst.
(5) The optical observation device is a microscope.
(6) The objective lens has an outer diameter of φ10 or less that can be inserted into a living body.

  An optical observation apparatus according to a fourth aspect of the present invention is an optical observation apparatus having a housing with a built-in scanner for scanning a light source image on a specimen and having an objective lens detachably attached thereto. The outer periphery of each of the housing and the objective lens is coated with a photocatalyst.

An optical observation apparatus according to a fifth aspect of the present invention is an optical observation apparatus having a housing on which a bundle fiber for transmitting a specimen image is mounted and an objective lens is detachably mounted. And a photocatalyst is coated on the outer periphery of each of the objective lenses. In the third to fifth aspects, the following embodiments are preferable.
(1) The objective lens is immersion type.
(2) It further includes an illuminating means for illuminating the specimen, and the light from the illuminating means includes a wavelength capable of exciting the photocatalyst.
(3) In (2), the light source for illuminating the specimen and the illumination light from the light source are reflected toward the specimen through the objective lens, guided coaxially with the objective lens, and emitted from the specimen. An optical path splitting unit that transmits the light beam, and a reflecting unit that can be disposed at the same position as the optical path splitting unit, and reflects only the wavelength that can excite the photocatalyst in the illumination light from the light source, The optical path dividing means and the reflecting means are configured to be detachable from the optical axis of the objective lens.
(4) The photocatalyst is titanium oxide.
(5) The excitation wavelength of the photocatalyst is different from the excitation wavelength of the fluorescent dye stained on the specimen.

  An optical observation apparatus according to a sixth aspect of the present invention is an optical observation apparatus having at least one of an observation optical system and a proof optical system, and illumination light or observation light transmits or reflects the optical system. The photocatalyst is coated on the part. In the sixth aspect, the optical observation device is preferably a microscope.

  The present invention is not limited to the above-described embodiments, and various modifications can be made without departing from the scope of the invention at the stage of implementation. Further, the above embodiments include inventions at various stages, and various inventions can be extracted by appropriately combining a plurality of disclosed constituent elements.

  In addition, for example, even if some structural requirements are deleted from all the structural requirements shown in each embodiment, the problem described in the column of the problem to be solved by the invention can be solved, and the effect described in the effect of the invention Can be obtained as an invention.

The figure which shows schematic structure of the whole optical observation apparatus which concerns on 1st Embodiment. FIG. 2 is a detailed view around a specimen (cultured cell) in FIG. 1. The figure for extracting an epi-illumination part from FIG. 1 and demonstrating another modification. The figure which shows schematic structure of the whole optical observation apparatus which concerns on 2nd Embodiment. FIG. 3 is a detailed view around a specimen (cultured cell) in FIG. 2. The figure for demonstrating 3rd Embodiment. The figure which shows schematic structure of the whole optical observation apparatus which concerns on 4th Embodiment. The figure which shows schematic structure of the whole optical observation apparatus which concerns on 5th Embodiment. The figure which shows the schematic structure of the whole of the conventional optical observation apparatus. FIG. 10 is a detailed view around the specimen (cultured cell) in FIG. 9.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 ... Objective lens 1a ... Holding part 3 ... Cell culture apparatus 4 ... Transmission illumination light source 5 ... Transmission illumination optical system 7 ... Epi-illumination light source 8 ... Epi-illumination optical system 9 ... Excitation filter for FITC 10 ... Dichroic mirror 11 ... Absorption filter 12 ... Mirror unit cassette 13 ... Rotating shaft 14 ... Stage 14
DESCRIPTION OF SYMBOLS 15 ... Semi-focus handle 16 ... Inverted microscope 17 ... Eyepiece 18 ... Tip lens 19 ... Cover glass 20 ... Sample 21 ... Glass bottom dish 22 ... Insulation box 22a ... Transmission window 23 ... Transmission window 25 ... Hole 26 ... Cylinder 27 ... Tube 28 ... excitation filter 29 ... mirror 30 ... excitation filter 31 ... sterilization lamp 32 ... heater 33 ... condenser lens 34 ... drive mechanism 35 ... scanning unit 35a ... mirror 36 ... housing 37 ... relay lens 38 ... imaging lens 39 ... condenser lens DESCRIPTION OF SYMBOLS 40 ... Multimode fiber 41 ... Relay lens 43 ... Dichroic mirror 44 ... Mirror 45 ... Connection end 46 ... Detector 47 ... Single mode fiber 48 ... Single mode fiber 49a, 49b ... Dichroic mirror 49c ... Mirror 50-52 ... Light source 53 ... Light source 54 Multi-mode fiber 57 ... observation optical system 59 ... hole 60 ... rat

Claims (6)

In the cell culture apparatus, which is applied to an optical observation apparatus having at least one of an observation optical system and an illumination optical system, and has a transmission window for transmitting light beams of the observation optical system or the illumination optical system, the transmission window A cell culture device characterized in that is coated with a photocatalyst. A cell culture space, which is applied to an optical observation apparatus having at least one of an observation optical system and an illumination optical system, and includes a transmission window for transmitting light beams of the observation optical system or the illumination optical system. A cell culture device, wherein the inner surface of the substrate is coated with a photocatalyst. An optical observation apparatus having an objective lens and a condenser lens for condensing illumination light onto a specimen, wherein the objective lens or the front lens of the condenser lens is coated with a photocatalyst. An optical observation device. In an optical observation apparatus having a housing for scanning a light source image on a specimen and having a housing in which an objective lens is detachably mounted, a photocatalyst is coated on the outer periphery of each of the housing and the objective lens An optical observation device. In an optical observation apparatus having a housing in which a bundle fiber for transmitting a specimen image is mounted and an objective lens is detachably mounted, the outer periphery of each of the housing and the objective lens is coated with a photocatalyst. An optical observation device. An optical observation apparatus having at least one of an observation optical system and a proof optical system, wherein a photocatalyst is coated on a portion where illumination light or observation light transmits or reflects through the optical system.

Images