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Method for electric-field-induced cell fusion based on composite pulse

A technology of cell fusion and electric field induction, which is applied in the direction of fusion cells, botany equipment and methods, biochemical equipment and methods, etc., can solve the problems of difficult fusion and low efficiency, and achieve the effect of improving the success rate

Active Publication Date: 2017-06-13
CHONGQING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The purpose of the present invention is to solve the problems of difficult fusion and low efficiency when cells of different sizes are fused under the action of microsecond pulses

Method used

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  • Method for electric-field-induced cell fusion based on composite pulse
  • Method for electric-field-induced cell fusion based on composite pulse
  • Method for electric-field-induced cell fusion based on composite pulse

Examples

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Effect test

Embodiment 1

[0050] The method for inducing cell fusion based on a composite pulsed electric field is characterized in that it comprises the following steps:

[0051] 1) Place cells and fusion solutions of different sizes in the cell culture tank;

[0052] 1.1) The cells of different sizes include large cells or small cells, the radius of the large cells ranges from 5 to 15 μm, and the radius of the large cells is 2 to 7 times the radius of the small cells;

[0053] The density of the large cells is 2 x 10 7 ~5×10 7 cells / L, the density of the small cells is five times that of the large cells;

[0054] 1.2) The cell is a plant cell or an animal cell; if the selected cell is a plant cell, the plant cell needs to be removed before fusion and then the fusion process is performed.

[0055] 1.3) The fusion solution includes PM buffer solution, enzyme solution and CPW salt solution with a volume ratio of (1-3):(1-3):(2-4);

[0056] The PM buffer solution consists of MgCl 2 , CaCl 2 , sorbi...

Embodiment 2

[0069] The method for inducing cell fusion based on a composite pulsed electric field is characterized in that it comprises the following steps:

[0070] 1) Place cells and fusion solutions of different sizes in the cell culture tank;

[0071] 1.1) Select cells

[0072] Large cells are: murine myeloma cells with a radius of 7.75±0.25μm;

[0073] Small cells are: mouse-derived B lymphocytes, with a radius of 3.85±0.35 μm;

[0074] 1.2) Select fusion medium (culture medium)

[0075] The main components of the fusion solution for electrofusion are glucose, calcium ions, and magnesium ions. The fusion solution in described step 1) includes PM buffer solution, enzyme solution and CPW salt solution with a volume ratio of 1:1:2;

[0076] The PM buffer solution consists of MgCl 2 , CaCl 2 , sorbitol and three distilled water; the MgCl 2 The concentration of 0.05mmol / L, CaCl 2 The concentration of sorbitol is 0.05mmol / L, and the concentration of sorbitol is 200mmol / L;

[0077]...

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Abstract

The invention discloses a method for electric-field-induced cell fusion based on composite pulse. The method comprises the following steps that firstly, cells different in size and a fusion liquid are placed in a cell culture tank; secondly, sine alternating voltage is applied to the two poles of the cell culture tank, and the cells are arrayed and tightly attached to each other; thirdly, the sine alternating voltage applied to the two poles of the cell culture tank is switched into nanosecond impulse voltage through a switch, the minute electroporation of the cells in the tank occurs; fourthly, the nanosecond impulse voltage continues to be switched into microsecond impulse voltage through the switch so as to act on the in-tank cells in the electroporation state, and the size of holes in the cells in the opening state is further increased, the outer membranes of the two cells tightly attached to each other are connected with each other, genetic materials in the two cells are mutually fused, and finally an intestinal fused cell is formed.

Description

technical field [0001] The invention relates to a method for cell fusion, in particular to a method for inducing cell fusion based on a compound pulsed electric field. Background technique [0002] Cell fusion can realize long-distance hybridization. . [0003] Cell fusion is a fundamental way of biological preparation. Whether cells can be effectively fused is the key to determining the feasibility of biological preparation, and even becomes a bottleneck restricting the development of biological preparation technology. Therefore, how to develop and improve existing cell fusion methods and improve the effect and efficiency of cell fusion has become a research hotspot in the field of biological preparation at home and abroad. [0004] According to the basic principle of electrofusion, the appearance of sufficient number and size of holes in the cell membrane (ie, cell electroporation) is a prerequisite for cell fusion. For cells to undergo electroporation, the transmembra...

Claims

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Application Information

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IPC IPC(8): C12N15/05C12N15/06C12N13/00
CPCC12N5/14C12N5/16C12N13/00
Inventor 李成祥柯强姚陈果米彦张媛媛董守龙吕彦鹏
Owner CHONGQING UNIV
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