JP2006039171A - Environmental control device and environmental control type analyzer - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide an environment control device capable of preventing condensation on a cover of a well plate with a cover.
A temperature control device 1 includes a temperature adjustment plate 1a and a holding portion 1b for holding the temperature adjustment plate 1a. The temperature control plate 1a is placed in close contact with the upper surface of the well plate W with the cover C that accommodates the biological sample S, and a current is passed through the transparent conductive film 3 formed on the upper surface of the glass substrate 2 to thereby adjust the temperature control plate 1a. The surface temperature of the well plate W is adjusted by generating heat and monitoring with the temperature sensor 6. The temperature control plate 1a can be moved up and down by the electromagnetic solenoid 8 of the holding portion 1b.
[Selection] Figure 2

Description

  The present invention relates to an environment control apparatus that adjusts and controls the environment in a microscope observation container.

  In order to observe or analyze a biological sample during culture, a predetermined environment is required. For this purpose, there is known a microscope observation container for controlling a closed space in which a culture container for storing a biological sample is arranged in a predetermined environment (see, for example, Patent Document 1). Heaters are disposed above and below the closed space, and the heater disposed above is incorporated in a slide cover that forms the closed space, and prevents condensation of the slide cover.

JP 2003-107364 A (page 8, FIG. 6)

  When observing and analyzing a biological sample while culturing, a culture vessel for observation with a microscope is used to prevent foreign substances such as bacteria from entering the culture vessel. When such a culture container is used, the inside of the container is in a sealed state or close to a closed state, so that moisture or the like evaporates, and clouding or condensation tends to occur on the inner surface of the lid. In the technique of Patent Document 1, the heater is attached to the slide cover that covers the entire upper surface of the closed space, so that there is an effect of preventing condensation on the slide cover itself, but preventing condensation on the lid of the microscope observation container. There is a problem that can not be.

(1) In order to solve the above problem, the environmental control apparatus according to the first aspect of the present invention includes an air conditioner that adjusts the environment in the sample chamber, and the outer surface temperature of the lid of the culture vessel with a lid housed in the sample chamber. A temperature control means for adjusting the temperature, a control means for controlling the air conditioning means and the temperature control means, and a holding means for holding the temperature control means in close contact with or close to the lid of the lidded culture vessel.
(2) The environmental control apparatus according to the second aspect of the present invention is the environmental control apparatus according to the first aspect, wherein the holding means moves the temperature adjustment means so as to change the distance between the temperature adjustment means and the culture vessel. It is preferable to have.

(3) The environmental control device of the invention according to claim 3 is the environmental control device according to claim 1 or 2, wherein the temperature control means is in a plate shape facing the upper surface of the lid, and the sample in the culture vessel with the lid It is preferable that the light beam incident portion of the illumination light that illuminates is transparent. The temperature control means may include a transparent substrate and a transparent conductive film formed on the transparent substrate. The temperature control means also includes a transparent substrate and an annular shape that is formed on the transparent substrate and surrounds a light beam incident portion of illumination light. And an electric resistor film having the following pattern shape.
(4) An environmental control type analyzer according to a sixth aspect of the invention includes the environmental control device according to any one of the first to fifth aspects, and a microscope device or an optical analyzer.

  According to the present invention, since the temperature adjusting means is disposed in close contact with or close to the lid of the culture vessel, condensation on the lid of the culture vessel can be prevented.

Hereinafter, an environmental control apparatus according to an embodiment of the present invention will be described with reference to FIGS.
<First Embodiment>
FIG. 1 is an overall configuration diagram schematically showing the configuration of the environment control apparatus according to the first embodiment of the present invention. FIG. 2 is an enlarged cross-sectional view taken along the line II in FIG. 3 is an enlarged cross-sectional view taken along the line II-II in FIG. FIG. 4 is a block diagram showing the configuration of the control circuit of the environmental control apparatus according to the first embodiment of the present invention.

  In FIG. 1, a sample chamber 10 houses a well plate W with a lid (cover) and a temperature adjusting device 1 that adjusts the surface temperature of the well plate W. The entire sample chamber 10 is contained in a sealed container 20. It is stored in. The well plate W has a plurality of wells (small chambers) for storing the biological sample S together with the culture solution and the reagent, and has a cover C that seals the upper surface after the biological sample is stored.

  The sample chamber 10 is opened such that the ceiling portion is indicated by the open surface 10 </ b> A, and the opening 11 </ b> A is formed in the bottom plate 11. The well plate W with a cover is covered with a cover C in order to prevent contamination by germs and the like after the biological sample S and culture solution are placed in each well, and the bottom plate is closed so as to close the opening 11A of the sample chamber 10. 11 is mounted. The sample chamber 10 is placed on a two-dimensional moving stage 12 and can move two-dimensionally along a horizontal plane. In addition, an opening / closing window 13 is provided on the side surface of the sample chamber 10 for taking in and out the well plate W and for maintaining the inside of the sample chamber.

  The sealed container 20 houses the sample chamber 10, the two-dimensional moving stage 12, and the objective lens 22 of the microscope apparatus. The observation optical system 23, the imaging device 24, and the transmission illumination optical system 25 are disposed outside the sealed container 20. The open surface 10 </ b> A of the sample chamber 10 is close to and parallel to the lower surface of the upper plate 21 of the sealed container 20. Even when the sample chamber 10 is two-dimensionally moved along the horizontal plane, the open surface 10A of the sample chamber 10 and the lower surface of the upper plate 21 of the sealed container 20 are kept in close proximity to each other, so that the interior of the sample chamber 10 is sealed. Airtightness is maintained with respect to the inside of the container 20 and the outside.

The upper plate 21 of the sealed container 20 is provided with an opening 20A, and a shutter mechanism 26 that opens and closes the opening 20A and switches members. Thereby, biological sample S, culture solution, etc. can be inject | poured through the opening 20A from the exterior, or the illumination light from the transmission illumination optical system 25 can be permeate | transmitted. A temperature / humidity sensor 14 and a CO ₂ gas sensor 15 for detecting the temperature, humidity, and gas concentration in the sample chamber 10 are provided on the lower surface of the upper plate 21 of the sealed container 20. An open / close window 27 is provided on the side surface of the sealed container 20 to allow the well plate W to be taken in and out and to maintain the inside of the sample chamber 10 and the sealed container 20. A dehumidifier 28 is provided to protect the optical system from moisture.

The inside of the sample chamber 10 configured as described above is connected to the air conditioner 30 disposed outside by piping, and is maintained in a predetermined environment. The air conditioner 30 is a device that generates a gas having a desired temperature, humidity, and composition, and circulates the gas into the sample chamber 10. A CO ₂ gas cylinder 34, a temperature / humidity sensor 35, and a reserve tank 36.

Air conditioner 30 has a housing 30a for accommodating the heating / cooling device 31, the humidifier 32, in the housing 30a, CO ₂ gas is supplied from the CO ₂ gas cylinder 34. The air conditioner 30 is connected to the upper plate 21 of the sealed container 20 by a gas supply tube 37 and a gas discharge tube 38.

The heating / cooling device 31 can perform both heating and cooling using, for example, a Peltier element. The humidifier 32 can generate water vapor by ultrasonic vibration from the water 32a in the water tank using, for example, an ultrasonic atomizing element. Also, the humidifier 32 introduces the CO ₂ gas from the CO ₂ gas cylinder 34 through the electromagnetic valve 34a, supplied with water from the reserve tank 36, was pooled by dehumidification via the drain tube 32b from the dehumidifier 28 It is configured to collect water. As described above, the adjusted gas adjusted in the housing 30 a of the air conditioner 30 is supplied to the sample chamber 10 through the gas supply tube 37 by the blower 33, and the atmospheric gas in the sample chamber 10 is air-conditioned by the gas discharge tube 38. A circulation system is formed to be returned to the apparatus 30.

  In addition to the air conditioning device 30 described above, the environmental control device of the present embodiment further includes a temperature adjusting device 1 that adjusts the temperature of the well plate W.

  Referring to FIGS. 2 and 3, the temperature adjustment device 1 includes a temperature adjustment plate 1 a and a holding portion 1 b that holds the temperature adjustment plate 1 a. The temperature control plate 1a includes a glass substrate 2, a transparent conductive film 3 formed on the upper surface of the glass substrate 2, a transparent resin layer 4 that protects the transparent conductive film 3, an upper lid 5, and a temperature sensor 6. It is a structured. The area of the lower surface of the temperature control plate 1a is approximately equal to the area of the upper surface of the well plate W (the area of the cover C). The glass substrate 2 may be replaced with a transparent plastic substrate.

As shown in FIG. 3, the transparent conductive film 3 is formed in a pattern over almost the entire upper surface of the glass substrate 2, and is connected to a power source (not shown) by a conducting wire 3a, and generates heat when an electric current flows. For example, a known ITO (In ₂ O ₃ —SnO ₂ ) film is used as the transparent conductive film 3. The ITO film 3 is deposited on the glass substrate 2 to a thickness of 0.1 μm, and has an average transmittance of about 80% for light having a wavelength in the visible region. Therefore, the ITO film 3 has an arbitrary pattern shape regardless of the position of the well. Even if formed, there is no problem in the observation and analysis of the biological sample S. The temperature sensor 6 is fitted in a recess formed on the lower surface of the glass substrate 2, and is connected to a measurement circuit (not shown) by a conducting wire 6a.

  The holding part 1 b includes a frame 7 and an electromagnetic solenoid 8. The frame 7 is fixed to the axis 8 a of the electromagnetic solenoid 8, and when the temperature control plate 1 a is dropped into the frame 7, the frame 7 is engaged with the upper lid 5 to support the temperature control plate 1 a. If the chamfered portion 2a is formed on the lower surface of the glass substrate 2, the dropping operation is facilitated. The electromagnetic solenoid 8 is fixed to the bottom plate 11 of the sample chamber 10 and can move up and down the axis 8a by electromagnetic force. Therefore, the distance between the temperature control plate 1a and the cover C of the well plate W can be adjusted, and the temperature control plate 1a can be brought into close contact with or close to the cover C. Moreover, since the holding | maintenance part 1b can raise / lower the temperature control plate 1a with the electromagnetic solenoid 8, the replacement | exchange operation | work of the well plate W can also be performed easily. The well plate W can be replaced by moving the well plate W in the left-right direction by the actuator 29 in FIG.

  With reference to FIG. 1 again, the microscopic observation of the biological sample S in the well plate W will be described. In transmission image observation, illumination light from the transmission illumination device 25 passes through the shutter mechanism 26 and the opening 20A, passes through the temperature control plate 1a, and irradiates the biological sample S in the well. The light transmitted through the biological sample S enters the objective lens 22 through the bottom plate of the well plate W. The light incident on the objective lens 31 reaches the image pickup device 24 by the observation optical system 23 and forms an image on the image pickup element of the image pickup device 24. When observing the biological sample S injected into another well, the well plate W is moved along the horizontal plane by the two-dimensional moving stage 12.

  Hereinafter, the control of the air conditioning device 30 and the temperature control plate 1a of the temperature control device 1 will be described with reference to FIG. In FIG. 4, the sample chamber 10, the sealed container 20, the air conditioner 30, and the temperature control device 1 are the same as those shown in FIG. 1, but are simplified compared to FIG. 1.

The control device 40 includes a temperature control circuit 41, a humidity control circuit 42, a CO ₂ gas concentration control circuit 43, and an overall control microcomputer 44 that controls these three in an integrated manner. The overall control microcomputer 44 is electrically connected to the temperature control circuit 41, the humidity control circuit 42, and the CO ₂ gas concentration control circuit 43. The temperature control circuit 41 is connected to the heater / cooler 31 by the connection 41a, to the temperature / humidity sensor 35 by the connection 41b, to the temperature / humidity sensor 14 by the connection 41c, to the temperature control plate 1a by the connection 41d, and to the temperature control plate 1a by the connection 41e. 6 are connected to each other.

The humidity control circuit 42 is connected to the humidifier 32 by a connection 42a, to the temperature / humidity sensor 35 by a connection 42b, and to the temperature / humidity sensor 14 by a connection 42c. The CO ₂ gas concentration control circuit 43 is connected to the electromagnetic valve 34a by a connection 43a and to the CO ₂ gas sensor 15 by a connection 43b.

First, environmental adjustment in the sample chamber 10 by the air conditioner 30 will be described. The temperature adjustment is performed by feeding back temperature measurement values of the temperature / humidity sensor 35 in the air conditioner 30 and the temperature / humidity sensor 14 in the sample chamber 10 using the temperature control circuit 41, and the Peltier of the heating / cooling device 31. This is done by controlling the voltage applied to the element. The humidity adjustment circuit 42 uses the humidity control circuit 42 to feed back the measured humidity values of the temperature / humidity sensor 35 in the air conditioner 30 and the temperature / humidity sensor 14 in the sample chamber 10, so that the ultrasonic mist of the humidifier 32 is used. This is done by increasing or decreasing the drive voltage to the activating element. The CO ₂ gas concentration is adjusted by feeding back the measured value of the CO ₂ gas sensor 15 in the sample chamber 10 using the CO ₂ gas concentration control circuit 43 and opening the electromagnetic valve 34a (the number of rotations and the rotation angle of the valve). ) Is increased or decreased.

The gas thus adjusted is, for example, 37 ° C., 80% or higher RH, and 5% CO ₂ gas concentration in the sample chamber 10, and is supplied from the air conditioner 30 to the sample chamber 10 through the gas supply tube 37. Then, the sample chamber 10 returns to the air conditioner 30 through the gas discharge tube 38. By this circulation path, the inside of the sample chamber 10 is always maintained in a predetermined environment.

  Next, adjustment of the surface temperature of the well plate W by the temperature control plate 1a will be described. Adjustment of the surface temperature of the well plate W is performed by using the temperature control circuit 41 to feed back the temperature measurement value of the temperature sensor 6 of the temperature control plate 1a and increase or decrease the current flowing through the transparent conductive film 3. The surface temperature of the well plate W is set as follows.

  As shown in FIG. 2, the well plate W with the cover is covered with the cover C after the biological sample S and the culture medium M are put into each well, so the relative humidity in the well is 100%. Reach close. When the temperature of the outer surface or the outer surface of the well plate W becomes lower than the temperature in the well, water vapor or the like in the well reaches supersaturation, causing a phenomenon such as cloudiness or condensation. In order to prevent fogging or condensation, the temperature of the outer or outer surface of the well plate W may be set to be equal to or higher than the temperature in the well. In consideration of safety in terms of temperature control, the temperature of the outside or outer surface of the well plate W may be set slightly higher than the temperature in the well, for example, the temperature difference may be 0.2 ° C. That is, it is only necessary to input from the overall control microcomputer 44 so that the measured temperature of the temperature sensor 6 of the temperature control plate 1 a is set to be 0.2 ° C. higher than the measured temperature of the temperature / humidity sensor 14 in the sample chamber 10.

  As described above, in the present embodiment, the surface temperature of the well plate W is adjusted by bringing the temperature adjustment plate 1a into close contact with or close to the well plate W, so that the fogging and condensation of the well plate W can be efficiently prevented. Further, since the temperature control plate 1a can be moved up and down by the holding portion 1b, the temperature control plate 1a can always be set at a predetermined position even if the height of the well plate W changes, and the well plate W can be easily put into and out of the sample chamber 10. Can be done.

<Second Embodiment>
FIG. 5 is a partial cross-sectional view of the temperature control device of the environmental control device according to the second embodiment of the present invention. 6 is a cross-sectional view taken along the line III-III in FIG.
In the present embodiment, since only the temperature control device 101 is different from the temperature control device 1 according to the first embodiment, only the temperature control device 101 will be described, and the same components are denoted by the same reference numerals and description thereof will be omitted. To do. 5 is equivalent to the II cross section of FIG. 1, and the partial cross sectional view of FIG. 6 is equivalent to the II-II cross section of FIG.

  In FIG. 5, the well plate W with the cover is filled with the biological sample S and the culture medium M in each well, and the upper surface is covered with the cover C, and the opening 11A of the sample chamber 10 is closed as in FIG. It is placed on the bottom plate 11. The temperature control device 101 includes a temperature control plate 101a and a holding unit 1b that holds the temperature control plate 101a. The temperature control plate 101 a has a structure in which the Ni alloy resistor film 103, the heater holding sheet 104 that sandwiches the Ni alloy resistor film 103, the upper lid 5, and the temperature sensor 6 are integrated. The area of the lower surface of the temperature control plate 101a is substantially equal to the area of the upper surface of the well plate W (the area of the cover C).

  As shown in FIG. 6, the Ni alloy resistor film 103 is formed in a circular pattern corresponding to the position and size of each well of the well plate W, and is connected to a power source (not shown) by a conducting wire 3a. , Heat is generated by passing current. Since the Ni alloy resistor film 103 is not transparent to the illumination light, it is not formed in a region where the illumination light is transmitted. The heater holding sheet 104 is a sheet having six circular openings 102 as shown in FIG. 6 with the annular pattern Ni alloy resistor film 103 completely sandwiched therebetween. Therefore, when the illumination light passes through the temperature control plate 101a, there is no decrease in light intensity. The heater holding sheet 104 is made of, for example, silicone rubber. Silicone rubber easily adheres to the cover C of the well plate W, and can efficiently transmit the heat generation energy of the Ni alloy resistor film 103 to the well plate W. The heater holding sheet 104 does not need transparency but needs flexibility, and synthetic rubber or plastic other than silicone rubber can be used.

  The holding unit 1b that holds the temperature control plate 101a is the same as that of the first embodiment, and the temperature adjustment method and temperature setting method using the temperature control circuit 41 are the same as those of the first embodiment. The effect of.

  As described above, since the environmental control device of the present invention includes the air conditioner 30 and the temperature control devices 1 and 101, the temperature of the well plate W can be accurately set, and fogging or dew condensation of the well plate W can be set. Can be effectively prevented. The cover C can effectively prevent fogging and condensation on the inner surface of the cover C even when the inside of the well is kept in a hermetically sealed state or when the inside of the well is kept close to the hermetic state with a slight gap.

The present invention is not limited to the embodiments described above as long as the characteristics are not impaired.
In the first and second embodiments, a case has been described in which the environment control device of the present invention is combined with a microscope device, and the biological sample S in the well plate W whose environment is controlled is observed with the microscope device. The present invention is not limited to an apparatus, and can be used in combination with an apparatus that performs optical analysis and measurement. For example, it can be used in combination with a device that performs wavelength analysis of light emission by biochemical reaction of a biological sample or a device that measures fluorescence intensity. The temperature control plates 1a and 101a of the above embodiment correspond to temperature control means, and the holding unit 1b corresponds to holding means.

It is a whole block diagram which shows typically the structure of the environmental control apparatus which concerns on the 1st Embodiment of this invention. It is the arrow directional cross-sectional view which expanded the II cross section of FIG. It is arrow sectional drawing which expanded the II-II cross section of FIG. It is a block diagram which shows the structure of the control circuit of the environmental control apparatus by the 1st Embodiment of this invention. It is a fragmentary sectional view of the temperature control apparatus of the environmental control apparatus which concerns on the 2nd Embodiment of this invention. It is arrow sectional drawing by the III-III cross section of FIG.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1,101: Temperature control apparatus 1a, 101a: Temperature control plate 1b: Holding part 2: Glass substrate 3: Transparent conductive film 4: Transparent resin layer 5: Upper lid 6: Temperature sensor 7: Frame 8: Electromagnetic solenoid 10: Sample chamber 14, 35: Temperature / humidity sensor 15: CO ₂ gas sensor 20: Airtight container 30: Air conditioner 40: Control device 41: Temperature control circuit 42: Humidity control circuit 43: CO ₂ gas concentration control circuit 44: Overall control microcomputer 102 : Opening 103: Ni alloy resistor film 104: Heater holding sheet S: Biological sample W: Well plate

Claims (6)

Air-conditioning means for adjusting the environment in the sample chamber;
Temperature control means for adjusting the outer surface temperature of the lid of the lidded culture vessel housed in the sample chamber;
Control means for controlling the air conditioning means and temperature control means;
An environmental control apparatus comprising: holding means for holding the temperature adjusting means in close contact with or in proximity to the lid of the lidded culture vessel. The environment control device according to claim 1,
The environmental control apparatus, wherein the holding means includes a drive mechanism that moves the temperature adjustment means so as to change a distance between the temperature adjustment means and the culture vessel with a lid. The environment control device according to claim 1 or 2,
The temperature control means is in a plate shape facing the top surface of the lid, and the light incident portion of the illumination light for illuminating the sample in the culture vessel with lid is transparent. The environment control device according to claim 3,
The temperature control device includes a transparent substrate and a transparent conductive film formed on the transparent substrate. The environment control device according to claim 3,
The temperature control device includes an environment control device having a transparent substrate and an electric resistor film formed on the transparent substrate and having an annular pattern shape surrounding a light incident portion of the illumination light. The environmental control device according to any one of claims 1 to 5,
An environment control type analyzer comprising a microscope apparatus or an optical analyzer.

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