JPS63230072A - Cell fusion chamber - Google Patents

Abstract

PURPOSE:To obtain a cell fusion chamber, having a pair of electrodes in a specific direction and two special cell chambers for respectively containing heterologous cells and capable of fusing the heterologous cells with high efficiency by centrifugal force and voltage. CONSTITUTION:A pair of electrodes 5 and 6 opposite to the direction of centrifugal force are provided at the tip positions in the direction of centrifugal force in the interior of a cell fusion chamber 7. Two chambers 1 and 2 having different heights from the electrode 5 are provided in the interior thereof and gaps between the pair of electrodes are linked with capillaries 3 and 4. Heterologous cells are placed in the two chambers 1 and 2.

Description

DETAILED DESCRIPTION OF THE INVENTION (Industrial Application Field) The present invention relates to a chamber of a cell fusion device that fuses cells using high voltage pulse electrical stimulation.

(Prior Art) In commonly used cell fusion chambers, a pair of opposing electrodes is provided, and cells are transferred between the electrodes using i! A turbid cell suspension is placed in the cell suspension, and a high frequency voltage is first applied between the electrodes to cause the cells to electrophores and come into contact with each other, and then the voltage applied to the electrodes is switched to a DC high voltage pulse to bring them into contact. Fuse the cells together. When fusing different types of cells, the two types of cells are suspended in a cell suspension.

At this time, it is known that the better the contact, the higher the fusion rate. Therefore, some cell fusion chambers use centrifugal force to enhance contact.

(Problems to be Solved by the Invention) In conventional cell fusion chambers, even when two types of cells are fused to form a heterogeneous cell (hybrid cell), two types of cells (cells in one chamber) are mixed in the turbid solution. Cell fusion is performed with cells suspended.

Therefore, the fusion rate of heterozygous cells is low, and there is a limit to obtaining the desired heterozygous cells.

An object of the present invention is to provide a cell fusion chamber that can obtain heterogeneous cells with a higher probability by using two types of cells.

(Means for Solving the Problems) To explain with reference to FIG. Inside the chamber body (7), which has a pair of electrodes (5, 6) facing each other in the direction of the force, there are two chambers containing different cells, each having a different height from the electrode (5°6) in the direction of the centrifugal force. Cell chambers (1, 2) are provided, each cell chamber (1, 2) being connected between electrodes (5, 6) by a capillary tube (3, 4) extending in the direction of centrifugal force.

(Example) FIG. 1 represents one embodiment.

7 is a chamber body made of an insulator such as glass;
For example, it is formed in a cylindrical shape. A pair of electrodes 5 are provided at the bottom of the chamber body 7 and are opposed to each other in the depth direction of the chamber body 7.
, 6 are provided. The chamber body 7 can be housed in a centrifuge tube 10 of a centrifuge, and when the centrifuge tube 10 is attached to the centrifuge and rotated, the chamber body 7 has a depth direction extending from top to bottom in the figure. centrifugal force acts. Therefore, the depth direction of the chamber body 7 is the centrifugal force direction, and the electrodes 5 and 6 are provided facing each other in the centrifugal force direction on the tip side in the centrifugal force direction.

Two cell chambers 1 and 2 are provided within the chamber body 7 at different heights from the electrode 5 in the depth direction of the chamber body 7, that is, in the centrifugal force direction. Cell chamber I is installed at a lower position than cell chamber 2, that is, at a position closer to electrodes 5 and 6. Cell chamber 1
The cell chamber 2 passes through the electrodes 5 by a thin tube 3 extending in the direction of centrifugal force and is guided into the space between the electrodes 5.
, 6. Cell suspensions in which different cells are suspended are placed in cell chambers 1 and 2, respectively.

A lid 9 is attached to the upper opening of the chamber body 7, and by covering the chamber body 7 with the lid 9, contamination of cells within the chamber body 7 can be prevented.

A lead electrode 8 is taken out from the electrode 5 through the lid 9, and a portion of the electrode 6 protrudes outward from the bottom of the chamber body 7.

Next, the operation of this embodiment will be explained.

The lid 9 is removed and the cell suspension solvent 14 is poured into the chamber body 7 to a level above the upper surface of the cell chamber 2 at a high position. A cell suspension in which A cells 11 are suspended is then put into cell chamber 1 using a pipette, and a cell suspension in which B cells 12 are suspended is put into cell chamber 2 using a pipette.

The lid 9 is closed, the lead electrode 8 is attached, the chamber body 7 containing the cell suspension is placed in the centrifuge tube 10, and the centrifuge tube 10 is attached to a centrifuge.

Activate the centrifuge and rotate at a speed that does not destroy the cells 11 and 12. Since A cells 11 have a shorter centrifugal distance than B cells 12, they are first released between electrodes 5 and 6 and then collected on electrode 6.

Since the B cell 12 is located further from the electrode 5 than the A cell 11, the Al cell 1 is deposited after the A cell 11 precipitates on the electrode 6.
Aj41@11 and B cells 12 are collected in close contact with each other on electrode 6, as shown in FIG.

After the centrifugation operation is completed, the chamber body 7 is taken out from the centrifuge tube 10, and the electrode 6 and lead electrode 8 are removed as shown in FIG.
During this time, a high voltage pulse generator 13 is connected, and a high voltage pulse is applied to fuse the A cells 11 and the B cells 12.

In the example, after centrifugation is stopped, a high voltage pulse is applied between electrodes 5 and 6 to fuse cells 11 and 12; Alternatively, the cells 12 may be fused. This further strengthens the adhesion between the cells 11 and 12, increasing the probability of fusion.

Further, although the embodiment illustrates a case where the chamber body 7 is housed in the centrifuge tube 10, the inner body of the chamber body 7 may also serve as the centrifuge tube 10.

(Effects of the Invention) In the cell fusion chamber of the present invention, one of the cells is first precipitated on one of the opposing surfaces of a pair of electrodes using centrifugal force, and then one of the cells is precipitated on that surface using centrifugal force. Since the other cells were precipitated by using the high-voltage pulse, different cells were brought into contact with each other in the direction in which the high-voltage pulse was applied, increasing the probability that heterocells would be formed by cell fusion.

Since centrifugal force is used to bring cells into close contact with each other, the degree of adhesion is improved, the fusion rate is high, and there is no need to apply a high-frequency electric field to bring the cells into contact as in the past.

Even in the case of cells with a low cell density, cell fusion is possible because the cell layers can be stacked and brought into contact with each other in a layered manner on the electrode.

In addition, when the electrical resistance of the solvent is low, such as a cell suspension of seaweed cells that can only live in seawater, it is not possible to apply high voltage pulses with conventional cell fusion chambers, but the present invention Since a layer of cells can be formed to cover the surface of one electrode, the electrical resistance becomes high and high voltage pulses can be applied, making cell fusion possible.

[Brief explanation of drawings]

FIG. 1 is a cross-sectional view showing one embodiment, and FIG. 2 is a schematic diagram showing a state in which different cell layers are stacked on an electrode in the same embodiment.
FIG. 3 is a schematic diagram showing a state in which a high voltage pulse generator is connected to the same embodiment. ■, 2... Cell chamber, 3.4... Thin tube, 5.6... Electrode, 7... Chamber body, lO...・Centrifuge tube, 11.12...Cells,

Claims (1)

[Claims] (1) A chamber body that can be installed in a centrifuge and has a pair of electrodes facing each other in the direction of centrifugal force on the tip side in the direction of centrifugal force has a height from the electrodes in the direction of centrifugal force. A cell fusion chamber comprising two cell chambers each accommodating different cells, each cell chamber being connected between the pair of electrodes by a thin tube extending in the direction of centrifugal force.