JPH01223348A - Particle immobilizing apparatus - Google Patents

Abstract

PURPOSE:To enable immobilizing of a particle such as cell simply and efficiently, by supporting a filter plate in which a plurality of tiny holes are arranged regularly with a min. diameter thereof smaller than the diameter of a particle to be immobilized. CONSTITUTION:A filter plate 3 in which tiny holes 4 are arranged regularly with a min. diameter smaller than a diameter of a particle (c) to be immobilized is supported into a culture liquid 9. A negative pressure generating means 6 is provided to put the side of one surface 3c thereof 3 at a negative pressure with respect to the other surface 3b. Therefore, the particle (c) is sucked into the tiny holes 4 by a negative pressure to be immobilized on the surface 3b of the filter plate 3. As the tiny holes 4 are arranged regularly, the particles (c) are also arranged regularly and immobilized. This facilitates the searching of the particles (c) immobilized to enable automation of injection of a chemical substance with a glass fine needle.

Description

DETAILED DESCRIPTION OF THE INVENTION (a) Field of Industrial Application The present invention relates to a particle fixing device capable of fixing particles such as cells and efficiently injecting chemical substances into the particles.

(b) Conventional technology Conventionally, in genetic engineering, when injecting genes into cells, cells C in the culture solution are observed using a microscope (not shown), as shown in Figure 4. The cells C were fixed by searching and suctioning with a dropper-like instrument 18. and,
A fine glass needle 19 was inserted into the nucleus n of cell C, and a chemical substance such as a gene was injected.

(c) Problems to be Solved by the Invention The above operations are performed one by one by humans while observing cells with a microscope, which requires skill, labor, and is inefficient. This has hindered the application of genetic engineering to produce useful substances cheaply and efficiently.

The present invention has been made in view of the above, and an object of the present invention is to provide a particle fixing device that can easily and efficiently fix particles such as cells.

(d) Means and action for solving the problem The structure of the particle fixing device of the present invention will be explained using FIG. 1 corresponding to the embodiment.The minimum diameter thereof is smaller than the diameter of the particle C to be fixed. A filter plate 3 in which small holes 4 are arranged regularly is connected to a liquid 9.
The filter plate 3 is characterized in that it includes a negative pressure generating means 6 which is supported inside the filter plate 3 and makes the one surface 3c side of the filter plate 3 a negative pressure with respect to the other surface 3b side.

Therefore, the particles C are attracted to the small holes 4 by the negative pressure and fixed to the surface 3b of the filter plate 3.

Since the small holes 4 are regularly arranged, the particles C are also regularly arranged and fixed. Therefore, it is easy to search for the fixed particles C, and it is also possible to automate the injection of chemical substances using a glass fine needle.

(E) Embodiment An embodiment of the present invention will be described below with reference to FIGS. 1 to 3.

FIG. 1 shows a longitudinal cross-sectional view of a fixing device fil for example particles. 2 is a container containing the culture solution 9. A filter plate 3 is supported within the container 2, and the interior of the container 2 is
divided into.

As shown in FIGS. 2(a) and 2(b), the filter plate 3 has a large number of small holes 4, arranged in a grid pattern on a substrate 3a.
..., 4 are provided. In this embodiment, the small hole 4 is a straight circular hole as shown in FIG. 2(b), but as shown in FIG. 2(C), the small hole 4 is formed into a funnel shape on the filter plate surface 3b side. It may be processed so that cells can be stably aspirated.

Moreover, instead of arranging the small holes 4 in a grid pattern, the small holes 4 are arranged in a grid pattern (see FIG.
It is possible to have a geometrical arrangement such as an equilateral triangle as shown in a) or a concentric circle as shown in FIG. 3(b).

The diameter d of this small pore 4 is made smaller than the diameter of the cell C, and is usually suitably 2 μm to 10 μm. The filter plate 3 is manufactured, for example, by drilling small holes 4 in a plastic plate using a laser. Alternatively, by applying semiconductor manufacturing technology,
Reactive ion etching is applied to the Si:+N, FJ film formed on the surface of the silicon substrate, and this Si*N4m
A small hole may be formed in the membrane (of course, the silicon substrate itself is also provided with a hole that communicates with the small hole).

The container 2 includes an inlet 2a that opens above the filter plate 3,
It has a discharge port 2b that opens at the bottom. The inlet 2a and the outlet 2b are connected by a channel 5, and a pump (means for generating negative pressure) 6 is provided in the channel 5, and the culture solution 9 sucked from the outlet 2b is pumped through the channel 5. It is returned into the container 2 through the inlet 2a.

A pressure sensor 7 is further provided at the bottom of the container 2 . The pressure sensor 7 detects the pressure in the lower half of the container 2, that is, below the filter plate 3. This pressure sensor 7
The pressure signal is data-processed by the pressure data processing device 8 and fed back to the pump 6, and the pump 6 is feedback-controlled so that a negative pressure suitable for suctioning and fixing the cells is obtained.

In this embodiment device 1, when the pump 6 operates, a negative pressure is generated in the lower half of the container 2, that is, on the filter plate surface 3c side. Due to this negative pressure, the cells C in the culture solution 9 are sucked into the small holes 4 and fixed to the filter plate surface 3b.

Since the small holes 4 are arranged in a grid pattern, the fixed cells C are also arranged in a grid pattern.

A microscope 11 for injecting a chemical substance 15 such as a nucleic acid into the fixed cells C is shown in FIG. The objective lens 12 of this microscope 11 is for water immersion, and a thin needle 13 passes through it on its central axis.

The proximal end of the silk needle 13 is supplied with a chemical substance 15 via a pump 14.
It is connected to the container 16 of.

To inject chemical substance 15 into the nucleus of cell C, use microscope 1.
While observing the cell C in step 1, a fine needle 13 is inserted into the cell C, and a chemical substance 15 is injected into the cell C using the pump 14.

When the injection into one cell C is completed, the fine needle 13 is pulled out, and if the cell C is suctioned and fixed in the nearest small hole 4, the small hole 4 is placed directly below the silk needle 13. The fixing device 1 is moved, the fine needle 13 is inserted into the cell C, and the chemical substance 15 is injected.

In this way, since the cells C are regularly arranged and fixed on the filter plate 3, there is no need to search for the next cell and fix it by suction as in the conventional method, and it is possible to efficiently apply chemicals to the cells. Can be injected.

Furthermore, since the cells C are regularly arranged and fixed on the filter plate surface 3b, the microscope 11 can automatically search for the cells C and the chemical substance 15 can be injected into the cells C. Chemical substances can be injected into cells, and useful substances can be efficiently produced using genetic engineering techniques.

(v) As described in detail, the particle fixing device of the present invention uses a filter plate having a plurality of regularly arranged small holes whose minimum diameter is smaller than the diameter of the particles to be fixed. To easily and efficiently fix a large number of particles by supporting the filter plate inside the filter plate and providing a means for generating negative pressure to make one surface side of the filter plate a negative pressure with respect to the other surface side. It has the advantage of being able to Furthermore, since the particles are regularly arranged and fixed, chemical substances can be automatically injected into cells, speeding up genetic engineering operations and efficiently producing useful substances.

[Brief explanation of the drawing]

FIG. 1 is a cross-sectional view of a cell fixation device according to an embodiment of the present invention, together with the main parts of a microscope applied to this cell fixation device, and FIG. An external plan view of the filter plate of the fixing device, FIG. 2(b) is a longitudinal cross-sectional view of the main part of the filter plate, and FIG. 2(C) is a longitudinal cross-sectional view of the main part showing a modification of the filter plate. Figure 3(a) and Figure 3(b)
) is a diagram showing a modification of the arrangement of small holes in the same filter plate, and FIG. 4 is a diagram illustrating a conventional particle fixing method. 1: Cell fixation device, 3: Filter plate, 4...
4: small hole, 6: pump, 9: culture solution,
C...C: Cell. Patent issuer: Tateishi Electric Co., Ltd. agent, patent attorney Shigeru Nakamura Figure 2 (a) 3: Phil 9th edition Figure 2 (b) Figure 2 (C) Figure 3 (a) Figure 3 (b) )

Claims (1)

[Claims] (1) A filter plate consisting of a plurality of regularly arranged small holes, the minimum diameter of which is smaller than the diameter of the particles to be fixed, is supported in a liquid, and one surface side of this filter plate is connected to the other surface. A particle fixing device comprising means for generating negative pressure against the side.