JPS629305B2 - - Google Patents

Abstract

PURPOSE:To obtain the titled homogenizer having high disintegration efficiency, and operable easily, by placing a pestle in a tubular vessel, and jointing the rotary shaft of the pestle detachably to the driving shaft with a one-way clutch. CONSTITUTION:A pestle 13 is placed rotatably in a tubular vessel 9 interposing small gap between the inner wall of the vessel. The rotary shaft 11 of the pestle 13 is connected to the driving mechanism 15 in a vertically detachable manner with a one-way clutch 19 which makes the rotary shaft 11 rotatable in a nearly horizontal plane in one direction. The unemulsified tissue poured into the vessel through the inlet 5 is emulsified in the space between the outer circumference of the pestle and the inner wall of the tubular vessel, and is discharged continuously from the outlet port 7.

Description

Detailed Description of the Invention [Industrial Application Field] The present invention relates to a homogenizer used to disrupt cell membranes and separate intracellular organelles, etc. The present invention has a simple configuration, can be manufactured at low cost, has high disruption efficiency,
Moreover, the present invention relates to a continuous flow homogenizer that is convenient to handle.

[Description of the Prior Art] In biology, medicine, and other research fields, it is necessary to separate organelles such as Golgi bodies and mitochondria, oxygen, etc. in cells. To do this, the cells must be disrupted by some means, but conventional cell membrane disruption devices use ultrasonic waves or apply pressure using high-pressure nitrogen gas, etc., to rapidly remove this pressure. There are methods in which the cell membrane is crushed by using a hydraulic machine, and methods in which the cell membrane is crushed by applying pressure to the cell fluid and passing it through pores. There is a method in which cell membranes are broken by manually applying reciprocating motion in the vertical direction. Note that an existing rotating machine is also used to rotate the pestle inside the tubular container.

[Problems with the Prior Art] These conventional cell membrane disruption devices generally have poor disruption efficiency and are inconvenient to handle. For example, in the widely popular pot-type container, the tubular container must be moved up and down manually, which is inefficient. For example, other devices using ultrasonic waves or high-pressure nitrogen require large and expensive devices.

[Technical Means for Solving the Problems] The present invention solves the problems of the above-mentioned conventional technology, and is a device for cell membrane disruption that has a simple configuration, can be manufactured at low cost, has high disruption efficiency, and is convenient to handle. This homogenizer is characterized by a removably supported tubular container having an unemulsified tissue inlet at the upper end and an emulsified tissue outlet at the lower end; A pestle equipped with a rotating shaft is rotatably disposed inside the tube with an appropriate gap between the pestle and the inner surface of the tubular container, and a rotary drive section is provided at the upper end of the column, and the rotary drive section A one-way clutch is provided which allows the rotary shaft to rotate in one direction in a substantially horizontal plane with or without a suitable transmission shaft and also to be able to be attached and detached in the vertical direction.

[Function] When using the homogenizer for disrupting cell membranes having the above configuration, first, for example, animal sample material is minced, a buffer solution is added to obtain a cell solution, and the cell solution is placed inside the support column. It is poured into an unemulsified tissue inlet at the upper end of a tubular container equipped with a rotatable pestle, and then the pestle is rotated via a rotary drive shaft.

Then, the cell fluid is emulsified between the tubular container and the pestle, the cell membranes are crushed, and the cell fluid is sequentially discharged from the bottom outlet of the tubular container. or,
Since an unemulsified tissue introduction port is provided at the upper end of the tubular body container, unemulsified cell fluid can be continuously introduced through this introduction port, and continuous emulsification operations can be performed. can be performed extremely efficiently.

Furthermore, since a one-way clutch is provided that allows the pestle rotating shaft to rotate in one direction in a substantially horizontal plane and also to be detachable in the vertical direction, the pestle can be attached and detached by simply attaching and detaching it to the one-way clutch, which simplifies emulsification work. Good workability when starting, finishing, or changing samples.

[Description of Embodiment] Hereinafter, one embodiment of the present invention will be described in detail using the drawings.

Figure 1 is a perspective view of the entire continuous flow homogenizer device, Figure 2 is a sectional view of the tubular container equipped with a pestle, Figures 3 and 4 are illustrations of the one-way clutch, and Figure 5 is the rotating FIG. 3 is an explanatory diagram of an embodiment in which a coupling is provided on the shaft.

In Figure 1, a support 3 is erected on a base 1, and this support 3
The pestle is provided with a removably supported tubular body container 9 having an unemulsified tissue inlet 5 at the upper end and an emulsified tissue outlet 7 at the lower end, and a rotary shaft 11 provided inside the tubular body container 9. 13 to the tubular body container 9
The pillar 3 is rotatably disposed with an appropriate gap between it and the inner surface of the pillar 3, and a rotary drive section 15 is provided at the upper end of the support column 3, and the rotation drive section 15 is connected to the rotary drive section 15 with or without an appropriate transmission shaft. This shows a continuous flow type homogenizer characterized in that it is provided with a one-way clutch 19 that allows the shaft 11 to rotate freely in one direction within a substantially horizontal plane and also to be able to be attached and detached in the vertical direction.

For the base 1 and the support column 3, for example, a commercially available experimental stand may be used as is, but here, a rotation drive unit 15 is integrally provided above the support column 3. 21 is an outlet, and 23 is a voltage converter. Furthermore, mounting fittings 25 and 27 for gripping the tubular body container 9 are provided on the support column 3.

The mounting brackets 25 and 27 are semicircular arc-shaped fixing members 25a and 27a that open toward the tubular body container 9, and semicircular arc-shaped opening/closing members that are symmetrical to the fixing members rotatably equipped with thumbscrews 25b and 27b. Members 25c, 27c
One end of the fixing members 25a, 27a is provided with a tap hole into which the threaded portion of the thumb screw 25b, 27b is screwed, and the other end is provided with an opening/closing member 25c, 2.
A hinge (not shown) is provided to align the fixing members 7c with the fixing members 25b and 27b. Furthermore, cushions (not shown) made of rubber or the like are provided inside the semicircular arc portions of the fixing members 25a, 27a and the opening/closing members 25c, 27c, so that the tubular container can be fixed flexibly and reliably. I have to.

It is convenient if the cushion is colored white or gray so that the emulsified state can be clearly observed through the transparent tubular container 9.

The entire surface of the rotary drive unit 15 is provided with a power on/off switch 29, a start switch 31, a stop switch 32, a digital switch 33 for setting the rotation speed, an LED speed display device 35 for displaying the actual rotation speed, etc. . The rotating shaft 11 is the third
It is rotated in one direction (direction of arrow 17) at a speed determined by the digital switch 33 via a transmission mechanism detailed in FIGS.

As shown in FIG. 2, an unemulsified tissue (cell fluid) inlet 5 is formed at the upper end of the tubular body container 9 and is opened in the shape of an inverted cone so as to receive a large amount of tissue solution. An emulsified tissue outlet 7 is provided at the bottom of the tubular container 9. An example of the tubular body container 9 made of glass is shown, but the present invention is not limited to this.

FIG. 2 shows a state in which the pestle 13 is located at an intermediate position of the tubular body container 9. In this state, a first chamber 35 for storing emulsified tissue (cell fluid) is located above the tubular body container 9, and a first chamber 35 is located below the tubular body container 9. The liquid after emulsification is discharged from the outlet 7.
A second chamber 37 is formed for guiding. pestle 1
3 and the tubular body container 9 is the distance d necessary to separate the cell nucleus and other organelles.
(approximately 0.05 to 0.2 m), and this value is determined depending on the cell type.

In FIG. 2, a rotating shaft 11 provided on a pestle 13 rotates in the direction of an arrow 17. Here, the pestle 13 is made of Teflon, for example, and the unemulsified tissue in the first chamber 35 is placed on the upper outer peripheral surface of the pestle at the center of the pestle while gradually decreasing the distance from the inner peripheral surface of the tubular body container 9. Guide groove 4 that can be fed to 13a
1 and 31 are provided. The depth of the guide grooves 41, 41 is about 3 mm at the top, and gradually becomes shallower toward the center of the pestle until it reaches zero.

Since the pestle 13 is provided with the guide grooves 41, 41 as described above, as the pestle 13 rotates in the direction of the arrow 17, the emulsified tissue in the first chamber 35 is transferred to the lower part of the pestle 13 or to the tissue crushing part 13a. sent towards. The cells trapped between the surface of the tissue crushing part 13a and the inner surface of the tubular body container 9 are efficiently destroyed by causing a gentle shearing action within a small gap.

3 is a sectional view of the rotary drive unit 15, and FIG. 4 is an explanatory diagram showing the operation of the one-way clutch 19. FIG. 4 corresponds to a partially enlarged plan view of the one-way clutch described in FIG. 3. When the driving outer cylinder 43 is decelerated and rotationally driven in the direction of the arrow 17 via the motor 45 and the bevels 47 and 49, the plurality of shaped slopes 53 provided on the inner surface of the one-way clutch 19 shown in FIG. , 55, the rotating shaft 11 is driven in the direction of the arrow 17. In addition,
A roller 55 is disposed in the cage 51 to face the contract slope 53, and the roller 55 is disposed opposite to the contract slope 53.
3 is provided with a spring that biases it.

In FIG. 4, the contract slope 53 is
When rotated in the opposite direction, the rotating shaft 11 can be stopped.

When the motor 45 is stopped, as shown in FIG. 3, the position of the rotating shaft 11 can be adjusted in the vertical direction, and the rotating shaft 11 is also detachable. A fixing member 59 is attached to the. The fixing member 59 is preferably an O-ring or a one-touch rubber stopper so that it can be easily attached to and detached from the rotating shaft 11 by manual operation.

Hereinafter, the rotating shaft 11 can be vertically adjusted in position within the one-way clutch 19 and can be attached and detached by the structure of the drive section explained in FIGS. 3 and 4. Therefore, when starting work and arranging the pestle 13 as shown in FIG. It can be easily removed from the section. Note that the coupling 61 is shown in FIG. 3 by a two-dot chain line. This coupling is provided as appropriate and allows the rotation shaft 11 to be connected to the pestle 13.
It is divided into a side and a drive section 15 side, and an elastic member is interposed between the two sides, and has the effect of removing strain on both sides.

An example of the coupling 61 is shown in FIG. a
The figure is an assembled perspective view, and figure b is an exploded explanatory view thereof.
The coupling ring 61 connects the upper joint 63 and the elastic joint 6.
5 and a lower joint 67, the upper joint 63 is fixed to the upper rotating shaft 11, and the lower joint 67 is fixed to the lower rotating shaft 1 using screws 69, 69, respectively.

A small hole is provided on both sides of the upper joint 63, and the end of the clip 3, which has a holding part 71 that tilts downward and holds the lower end surface of the lower joint, is fitted into the small hole. It is supported so that it can rotate freely around the small hole. Also, detent gears 75, 75 with a height l are provided below the upper joint 63 and above the lower joint.

The elastic joint 65 is made of rubber (rubber hardness about 70 to 85 Hs),
Alternatively, it is made of an elastic material such as plastic, and has grooves 77 therein that fit with detent gears 75 provided in the upper joint 63 and the lower joint 67, respectively.
77 at both the upper and lower ends. The elastic joint 65 may be fixed to either the upper joint 63 or the lower joint 67.

Due to the configuration of the coupling ring 61, the lower rotating shaft 11 is at an angle θ (approximately 3
Since the pestle 13 is allowed to deviate within a range of 100 degrees, the attachment of the pestle 13 becomes easier, and unnecessary force is not applied to the entire device, so the durability of the working device can be improved.

A supplementary explanation of the emulsification work is provided based on Figure 1.

First, the apparatus is assembled in advance as shown in FIG. 1, and adjusted so that the pestle 13 is at a desired height within the tubular container 9. At this time, the height position can be adjusted by freely moving the rotary shaft 11 up and down within the one-way clutch 19. In addition, at this time, if a coupling 61 (see Fig. 5) is interposed in the middle of the rotating shaft 11, the lateral position within the tubular body container is allowed within the range of allowable angle θ (see Fig. 5). Therefore, it is not necessary to make very delicate adjustments.

Turn on the switch 29, set the digital switch 33 to the desired speed value, and then turn on the start switch 31.
When turned on, the rotating shaft 11 rotates at the desired speed,
The speed value is displayed on the speed indicator 35.

Unemulsified tissue (cell fluid) injected from inlet 5
enters the guide groove 41 from the upper end of the pestle 13 shown in FIG. and are sequentially and continuously discharged from the discharge port 7. Note that it is possible to temporarily stop the motor and change the pestle position during this work.

Once the cell fluid has been emulsified, the motor is stopped, the pestle 13 is raised upward, the thumbscrews 25b and 27b are loosened, and the tubular container 9 is removed. After that,
When cleaning the pestle 13, it is sufficient to lower the pestle downward again and remove its rotating shaft from the one-way clutch. Note that if the removable coupling shown in FIG. 5 is used, the rotating shaft can be completely removed.

In the above example, the speed of the rotating shaft was adjusted using the digital switch 33, but it is also possible to adjust the speed using a rotation knob (volume).

[Effects of the Invention] The present invention provides a tubular container having an unemulsified tissue inlet at the upper end and an emulsified tissue outlet at the lower end and is detachably supported on a support, and has a rotatable container inside the tubular container. A pestle with a shaft is rotatably disposed with an appropriate gap between the pestle and the inner surface of the tubular container, and a rotary drive section is provided at the upper end of the column, and an appropriate transmission shaft is connected to the rotary drive section. This continuous flow type homogenizer is characterized in that the rotary shaft is rotatable in one direction in a substantially horizontal plane with or without intervening, and is equipped with a one-way clutch that can be attached and detached in the vertical direction. Therefore, unlike conventional methods that use containers with limited capacity, work can be carried out continuously.

In addition, a one-way clutch is provided in the drive unit, so the position of the pestle can be adjusted freely, and if the internal side wall of the container is worn and the gap between the pestle and the external surface exceeds the allowable range, the rotating shaft can be moved. This has the advantage that the pestle can be moved up and down within the tubular container, and the useful life of the container can be significantly improved.

Note that the present invention is not limited to the above-described embodiments, and the present invention can be implemented in modes other than the above-described embodiments.

[Brief explanation of the drawing]

The drawings all show embodiments of the present invention; FIG. 1 is a perspective view of the entire device, FIG. 2 is an enlarged sectional view of the tubular container with a pestle attached, and FIG. 3 is an enlarged longitudinal sectional view of the drive unit. 4 is an explanatory plan view showing the operating state of the one-way clutch, FIG. 5 is an explanatory view of the coupling, FIG. 5a is a perspective view showing the assembled state, and FIG. 5b is an exploded perspective view thereof. It is. DESCRIPTION OF SYMBOLS 1... Base, 3... Support, 5... Unemulsified tissue introduction port, 7... Emulsified tissue introduction port, 9... Tubular body container, 11... Rotation shaft, 13... Pestle, 15... Rotation Drive unit, 19...One-way clutch, 61...
...Katsupring.

Claims (1)

[Claims] 1. A tubular container having an unemulsified tissue inlet at the upper end and an emulsified tissue outlet at the lower end is detachably supported on a support, and inside the tubular container,
A pestle equipped with a rotating shaft is rotatably disposed with an appropriate gap between the pestle and the inner surface of the tubular body container, and a rotary drive section is provided at the upper end of the pillar, and an appropriate power transmission is provided to the rotary drive section. A continuous flow homogenizer, characterized in that it is provided with a one-way clutch that allows the rotary shaft to rotate in one direction in a substantially horizontal plane with or without the shaft and to be able to be attached and detached in the vertical direction.