Electroporator having an elongated hollow member

An electroporation and hollow technology, which is applied in the direction of electric/wave energy treatment of enzymes, electricity/wave energy treatment of microorganisms, microorganisms, etc., can solve problems such as increased surface resistance, harmful cells, and larger sample volumes

Active Publication Date: 2007-05-16
INVITROGEN SINGAPORE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Third, the generated ions (Al 3+ ) have adverse effects on cells
Fourth, due to the formation of oxide layer (Al 2 o 3 ), the surface resistance increases significantly
Fifth, the electric field is uneven
Sixth, the sample volume becomes larger, making it unsuitable for analyzing small numbers of cells
Eighth, high-throughput electroporation is not possible because tube handling is not easily integrated into automated systems
Ninth, the decomposition of water on the electrode surface causes severe pH changes, which are harmful to cells

Method used

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  • Electroporator having an elongated hollow member
  • Electroporator having an elongated hollow member
  • Electroporator having an elongated hollow member

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0081] Preferred Example 1: Electroporation experiment of human embryonic kidney HEK-293 cell line (cell line) using a pipette-type electroporation device

[0082] 1-1 Cell preparation

[0083] HEK-293 cell line (ATCC, CRL-1573) in 25 square centimeters (cm 2 ) culture flasks stored in medium supplemented with 10% fetal bovine serum (fetal bovine serum, FBS), in CO 2 Culture in an incubator and grow to 70% confluency. Next, the above medium was removed, and the cells were washed with phosphate buffered solution (PBS) and treated with trypsin. A medium supplemented with FBS was added and centrifuged, and then the cells were washed with a PBS buffer solution and suspended again in a medium supplemented with 10% FBS to prepare a cell sample.

[0084] 1-2 Electroporation

[0085] About 100 microliters (μl) of HEK-293 cell samples prepared in 1-1 were loaded into the reservoir at room temperature. 5 micrograms (µg) of plasmid DNA pEGFP (source: GenBank Accession: U55762; CLONT...

Embodiment 2

[0103] Preferred embodiment 2: use the electroporation device of channel structure to carry out electroporation experiment on SK-OV-3

[0104] 2-1 Manufacture of microchannel structure

[0105] In preferred embodiment 2, a biological experiment is performed using an electroporation device having a sample filler with a microchannel structure. An electroporation device is manufactured by a method such as moulding, having wells into which electrodes are inserted and channels for connecting the wells to hollow sample packs. Sample packings were fabricated with different channel structures, the channels had a height of 20 μm, a length of 2 cm, and a width of 100 to 500 μm. However, obviously, the channel pattern is formed by photolithography using a photomask. For example, first, a negative photoresist (SU-8, MicroChem, Massachusetts, USA) is spin-coated on a silicon wafer to form a 20 μm thick mold master. A soft bake was performed by a mask aligner (MA-6, Karl Suss GmbH, Germa...

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Abstract

An electroporation apparatus comprises an elongated hollo member in order to provide a uniform electric field during electroporation, wherein specifically, electroporation is carried out b applying electric pulses through a couple of electrodes from both end parts of the elongated hollow member, after the hollow member is charged with fluid specimen including cells and material which would be injected into the cells.

Description

technical field [0001] The present invention relates to an electroporation device having an elongated hollow member and an electroporation method whereby an electrical pulse or pulses are applied to a sample including cells to electroporate the cell membrane so that foreign substances can enter the cell. Background technique [0002] In general, electroporation is a technique in which macromolecules that cannot penetrate cell membranes enter cells by means of electrical pulses. Electroporation is a widely used and highly recommended method with direct application to cell assays and gene therapy. If a strong electric field is applied, the cell membrane temporarily becomes porous, exhibiting permeability to foreign substances. [0003] The electropermeabilization depends on various factors such as pulse width, pulse duration, number of pulses and other experimental conditions. In order to understand the mechanism of electroporation and enhance the effect of transfection, man...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N13/00
CPCC12M35/02
Inventor 张准根曹槿昌郑灿一申永植金贞我郑年哲
Owner INVITROGEN SINGAPORE
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