JP4357870B2 - Incubator for microscope stage - Google Patents

Description

[0001]
BACKGROUND OF THE INVENTION
In the present invention, when the cell culture process is continuously observed with a microscope for a long time, it is necessary to give a living cell in the culture medium an environment equivalent to normal body temperature and a hydrogen ion concentration (hereinafter referred to as pH) environment. The present invention relates to an incubator for a microscope stage that facilitates this.
[0002]
[Prior art]
Japanese Patent Application No. 2001-314700 discloses a temperature control device characterized by using a blowout port provided with a blower port on the top surface of an observation plate in addition to a ring duct for blowing air to the bottom surface of the observation plate.
[0003]
[Problems to be solved by the invention]
When living cell observation is performed continuously for a long time, it is necessary to keep the living cells in the culture solution constant at the normal living temperature and also to keep the pH of the culture solution constant.
In the box-type incubator that has been used for cell culture so far, when culturing mammalian cells, the temperature is generally set at 37 ° C and the atmospheric carbon dioxide concentration in the box is 5%. The concentration of 5% is a condition for maintaining the pH of the culture solution at the same pH 7.4 as the normal state of blood.
If the culture solution temperature can be maintained at 37 ° C. and the carbon dioxide concentration in the liquid surface atmosphere at 5% on the microscope stage , the cell culture process can be continuously observed.
0004
[Means for Solving the Problems]
According to claim 1, with supplying air containing a predetermined amount of carbon dioxide in the observation dish top can cover the culture surface without being diluted by the normal air supplied to the observation dish bottom, culture It becomes possible to maintain the pH of the liquid .
[0005]
According to claim 2 , air containing a predetermined amount of carbon dioxide supplied to the upper part of the observation dish can be heated to a temperature equivalent to normal air supplied to the bottom part of the observation dish for temperature control.
[0006]
According to claims 1 and 2, it is possible to apply the temperature control method shown in Japanese Patent Application No. 2001-314700 is prior art by preventing the occurrence of layer by vaporization heat the mineral oil culture solution .
[0007]
DETAILED DESCRIPTION OF THE INVENTION
FIG. 1 is a cross-sectional view of a double pipe outlet which is an embodiment of the apparatus of the present invention. The double pipe outlet is made of a synthetic resin such as nylon or epoxy, 11 is a ring duct surrounding the observation dish, and has a belt-like outlet on the inner wall. It is provided blowing port of the air containing a predetermined amount of carbon dioxide in its upper portion.
[0008]
Normal air controlled to a constant temperature supplied to the outlet nozzle, hereinafter referred to as hot air, is blown to the bottom of the observation dish through a ring duct and provided inside.
[0009]
The warm air blown to the bottom of the observation dish mainly heats the culture solution through a thin glass plate on the bottom of the observation dish.
[0010]
The skirt part indicated by 12 is provided at the bottom of the ring duct, and the position where the hot air intersects with the outside air is kept away from the observation plate and the observation plate and the objective lens are wrapped with the warm air, so that the observation plate proximity part of the objective lens Local heat dissipation to the objective lens is prevented by setting the same temperature.
[0011]
The upper cover shown in FIG. 13 is not fixed, and can be easily removed by retracting the light projecting portion, facilitating the operation of the observation dish. A donut-shaped protrusion is provided on the lower surface of the upper cover and is inserted into the upper end of the blowout barrel to form a part of a closed space for filling the air containing a predetermined amount of carbon dioxide supplied to the upper part of the observation dish. To do.
[0012]
As shown in Fig. 2, the ring duct lid shown in Fig. 14 is provided with a labyrinth seal structure at the inner peripheral edge that contacts the observation dish, so that the warm air supplied to the bottom of the observation dish can hardly blow up to the upper part of the observation dish. , air containing a predetermined amount of carbon dioxide supplied to the observation dish top it is possible to hold the concentration.
[0013]
FIG. 3 is a structural diagram of a double pipe hose which is an embodiment of the device of the present invention. The material of the hose is a resin such as vinyl chloride having heat retention and appropriate flexibility, and the outer tube is provided with a heat insulating material such as sponge rubber indicated by 33 on the outside of the outer tube. The air flowing through the pipe reaches the air outlet while maintaining almost the same temperature.
[0014]
Air in the inner tube containing a predetermined amount of carbon dioxide supplied to the observation dish top indicated by 31, warm air supplied to the observation dish bottom flows in the outer tube shown at 32.
[0015]
FIG. 4 is an explanatory view of a hot air supply box and a carbon dioxide supply box which are one embodiment of the apparatus of the present invention. The hot air supply box is a rectangular parallelepiped box having heat insulation performance mainly composed of foamed polystyrene, an external air intake port indicated by 41, a light bulb as a heat source indicated by 42, a circulation fan indicated by 43, an air supply fan indicated by 44, 45 the supply fan inlet thermometer shown in, PID control temperature controller indicated at 46, the copper tube coil for air heating comprising a predetermined amount of carbon dioxide indicated by 47, double hot air and carbon dioxide predetermined concentration air indicated by 48 Consists of a double tube nozzle for feeding into a tube hose.
0016
Air in the hot air supply box is agitated by a circulation fan to heat the air containing a predetermined amount of carbon dioxide flowing copper tube coil while being heated by the bulb is PID controlled to equal temperature.
[0017]
In this embodiment, the set temperature is room temperature +4 deg to 45 ° C. However, if an aluminum fin or the like to which a Peltier element is attached instead of a light bulb, not only heating but also cooling is possible.
[0018]
The control unit indicated by 49 reads the ring duct temperature indicated by 15 and the supply fan inlet temperature indicated by 45, and estimates and displays the observation dish temperature from the stored calculation formula.
[0019]
When the observation set temperature is set in the control unit, the control unit gives the set value to the PID control temperature controller indicated by 46 so that the observation dish temperature estimated value becomes the observation set temperature.
[0020]
The calculation formula stored in the control unit is as follows.
Observation dish temperature = A x ring duct temperature + B x supply fan inlet temperature + C
Here, A, B, and C are constants obtained by a performance test.
[0021]
The carbon dioxide supply box is a rectangular parallelepiped box made of steel plate, an outside air intake port shown by 51, a solenoid valve shown by 52, an air supply fan shown by 53, a carbon dioxide concentration meter shown by 54, and a PID control concentration controller shown by 55. And a hose for feeding air having a predetermined concentration of carbon dioxide into the copper tube coil of the hot air supply box.
[0022]
The air in the carbon dioxide supply box is divided into the amount supplied to the copper tube coil of the hot air supply box at the outlet of the air supply fan and the recirculation in the box. The recirculation in the box opens and closes the solenoid valve by PID control. The carbon dioxide supplied and the outside air flowing into the copper tube coil are agitated to make the detection response time of the carbon dioxide concentration meter appropriate.
[0023]
In this embodiment, the carbon dioxide concentration can be set to 0 to 10%. However, the oxygen concentration can be controlled by adding an oxygen concentration meter, a PID controller, a solenoid valve, a nitrogen cylinder or an oxygen cylinder. Become.
[0024]
【The invention's effect】
According to claim 1, with supplying air containing a predetermined amount of carbon dioxide in the observation dish top can cover the culture surface without being diluted by the ordinary air to be supplied to the observation dish bottom, culture It becomes possible to maintain the pH of the liquid .
[0025]
According to claim 2, it is possible to heat the carbon dioxide is supplied to the observation pan top to observe pan bottom same temperature and normal air supplied to for temperature control of air containing a predetermined amount.

[0026]
According to claims 1 and 2 , the following relational expression shown in Japanese Patent Application No. 2001-314700, which is a prior art relating to temperature control, was used by overlaying mineral oil on the culture solution to prevent the generation of heat of vaporization. Temperature control is possible.
Observation dish temperature = A x ring duct temperature + B x supply fan inlet temperature + C
(Where A, B and C are constants determined by performance tests)
[Brief description of the drawings]
FIG. 1 is a cross-sectional view showing a structure of a double pipe outlet according to an embodiment of the present invention.
2 is a partially enlarged view of FIG. 1 showing a labyrinth seal structure of an inner peripheral portion of a double pipe outlet according to an embodiment of the present invention.
FIG. 3 is a structural diagram of a double pipe hose that is one embodiment of the present invention.
FIG. 4 is an explanatory diagram illustrating configurations of a hot air supply box and a carbon dioxide supply box according to an embodiment of the present invention.
[Explanation of symbols]
11 Ring duct, 12 Skirt, 13 Cover
14 Ring duct lid, 15 Ring duct thermometer
31 Inner pipe (air flow path containing a certain amount of carbon dioxide, etc.)
32 Outer pipe (normal temperature controlled air flow path), 33 Insulation
41 Outside air intake, 42 bulbs, 43 circulation fans
44 Air supply fan, 45 Air supply fan inlet thermometer
46 PID controlled temperature controller, 47 copper tube coil, 48 double tube nozzle
49 Control unit,
51 Outside air intake, 52 Solenoid valve, 53 Air supply fan
54 CO2 concentration meter, 55 PID control concentration controller

Claims (2)

When observing viable cells by overlaying mineral oil on the microscope stage in order to prevent the generation of heat of vaporization in the culture medium in the observation dish, air is sent to the top of the observation dish in addition to the ring duct for blowing air to the bottom of the observation dish. Use a blowout outlet with a labyrinth seal structure to prevent air blown to the bottom of the observation dish at the part where the observation dish comes into contact with the ring duct. the observation dish top controlled normal air by supplying air containing a predetermined amount of carbon dioxide which is temperature controlled, and characterized in providing an incubation function necessary for cell culture observation dish on the microscope stage To incubator. A copper tube is coiled in a hot-air supply box that sends out temperature-controlled air for the bottom of the observation dish, and the observation dish is obtained by passing air containing a predetermined amount of carbon dioxide for the top of the observation dish through this copper pipe. 2. The incubator for a microscope stage according to claim 1, further comprising a hot air supply box that sends out the air for the bottom portion and the air for the upper portion at the same temperature.

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