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Electroporation
What is electroporation?
Electroporation is a very popular and highly efficient method of transfection. During electroporation, an electrical pulse is used to create temporary pores in cell membranes through which substances like nucleic acids can pass. This versatile method can be used for all cell types and for transfection of DNA, RNA, mRNA, RNPs, or proteins.
Thermo Fisher Scientific offers the Neon NxT Electroporation System for convenient benchtop electroporation as well as the CTS Xenon Electroporation System for GMP-compliant cell therapy applications.
How electroporation works
The process of electroporation is fairly simplistic. Host cells and selected molecules are suspended in a conductive solution, and an electrical circuit is closed around the mixture. An electrical pulse at an optimized voltage and only lasting a few microseconds to a millisecond is discharged through the cell suspension. This disturbs the phospholipid bilayer of the membrane and results in the formation of temporary pores. The electric potential across the cell membrane simultaneously rises to allow charged molecules like DNA to be driven across the membrane through the pores in a manner similar to electrophoresis [1].
Figure 1. Overview of electroporation. Electroporation is based on a simple process in which an electrical pulse is used to create temporary pores in cell membranes through which payloads can pass. After electroporation, the cell membrane recovers, and expression of the transfected nucleic acid can occur.
To ensure successful transfection, optimization of electroporation parameters for your experimental conditions is extremely important. Factors such as strength of the electrical field, shape of the pulse, the number of pulses applied, and composition of the buffer can all impact transfection efficiency.
Basic steps for performing electroporation
Prepare cells by suspending in electroporation buffer.
Apply electrical pulse to cells in the presence of specialized buffer and nucleic acids.
Electrical pulse creates a potential difference across the cell membrane and induces temporary pores in the membrane for nucleic acid entry.
Return cells to growth conditions and allow them to recover.
Assay cells for gene expression or silencing.
Prepare cells by suspending in electroporation buffer.
Apply electrical pulse to cells in the presence of specialized buffer and nucleic acids.
Electrical pulse creates a potential difference across the cell membrane and induces temporary pores in the membrane for nucleic acid entry.
Return cells to growth conditions and allow them to recover.
Assay cells for gene expression or silencing.
Advantages and disadvantages of electroporation
Electroporation offers many advantages in comparison to other transfection methods, with the main benefits being its applicability for transient and stable transfection of all cell types and its ability to transfect a large number of cells in a short amount of time once optimum electroporation conditions are determined. The major drawback of electroporation is substantial cell death caused by high voltage pulses and only partially successful membrane repair, requiring the use of greater quantities of cells compared to chemical transfection methods.
Advantages
- Relatively simple to perform
- Can work well for difficult-to-transfect cell types
- Reproducible results
- No vector requirement
- Less dependent on cell type
- Rapid transfection of a large number of cells
Disadvantages
- Requires special instrument
- Parameters must be carefully optimized
- Potential for high toxicity as well as cell damage and mortality
More modern instrumentation, such the Invitrogen Neon NxT Electroporation System, overcome high cell mortality by distributing the electrical pulse equally among the cells and maintaining a stable pH throughout the electroporation chamber. However, optimization of pulse and field strength parameters is still required to balance electroporation efficiency and cell viability.
Neon NxT system: an electroporation system for physical transfection
The Neon NxT Electroporation System is an electroporation machine that uses an electronic pipette tip as an electroporation chamber. The design and performance of the electronic pipette transfection chamber result in increased cell viability and transfection efficiency compared to traditional cuvette-based electroporation systems.
The proprietary tip technology of Neon NxT Electroporation System enables scientists to reduce sample transfer loss while simplifying their workflow. The system was designed for electroporation of mammalian cells, but some customers have found it to be successful for other cell types such as insect cell cultures and parasites.
The advantages of this electroporation device are:
- Performance — Achieve >90% efficiency and >95% cell viability in over 150 mammalian cell lines
- Flexibility— Precisely optimize electroporation parameters to deliver DNA, RNA, or protein in as little as 2 x 10⁴ cells to as much as 6 x 10⁶ cells per reaction.
- Time-saving— Plug-n-play out of box, and transfect cells quickly with 3-step workflow, a single buffer kit and intuitive user interface
- Preserve sample — Minimize sample transfer loss and contamination risk of valuable cells with unique and compact electroporation instrument design
The design of the electroporation chamber distributes the current equally among the cells and maintains a stable pH throughout the chamber; these key benefits increase cell viability dramatically.
Xenon system: an electroporation device for cell therapy
The Gibco CTS Xenon Electroporation Instrument is part of a flexible closed electroporation system that helps enable rapid, efficient transfection with no sacrifice in cell viability or recovery. This large-volume electroporation device is designed for cell therapy development and manufacturing.
The CTS Xenon Electroporation Instrument offers reliably high transfection performance in volumes of 1–25 mL with exceptional cell viability and recovery. The intuitive programmable interface, process flexibility, sterile single-use consumables, and available software upgrade that helps enable 21 CFR Part 11 compliance allow the system to seamlessly scale with your cell therapy workflow from process development through clinical manufacturing.
The advantages of this system are:
- High speed, large volume—transfect up to 2.5 x 109 T cells in less than 25 minutes
- Proven performance and viability—up to 90% gene knockout and 80% viability
- Process flexibility—user programmable system helps enable you to create and optimize electroporation protocols for various cell types and payloads from process development through commercial manufacturing
- Efficient non-viral transfection—can be used to deliver DNA, RNA, and protein payloads
- Closed-system processing—MultiShot (MS) consumable enables sterile welding to PVC or C-Flex tubing
The 1 mL electroporation chamber enables efficient process development and scales directly to commercial manufacturing using the 5–25 mL cartridge. This larger-volume consumable enables aseptic processing in a system designed for cell therapy manufacturing.
Visit Transfection Basics to learn more about performing transfection in your lab.
Intended use of the products mentioned on this page varies. For specific intended use statements, please refer to the Instructions for Use (IFU). C-Flex is a trademark of Saint-Gobain Performance Plastics Corporation.
For Research Use Only. Not for use in diagnostic procedures.