Primary cells can prove much more difficult to transfect than other commonly used cell lines. While high transfection efficiencies are possible using commercially available transfection reagents in diverse, commonly used cell lines, primary cells but also diverse cell lines have proven much more difficult to transfect.
Despite these transfection difficulties, primary cells are often preferred over genetically modified or immortalized cell lines in cellular research since they more closely resemble the in vivo situation. Primary human umbilical endothelial cells (HUVECs) are, for example, a powerful tool in the study of physiological processes such as angiogenesis.
Unfortunately, many primary cells such as endothelial, epithelial or smooth muscle are notoriously hard to transfect and are susceptible to the toxic effects of certain transfection reagents. Monocytes and their progeny, primary hepatocytes, neuronal or stem cells also present a particular challenge for transfection protocols in comparison to cell lines or readily dividing primary cells.
Transfection success involves multiple factors
The success of transfection experiments is dependent on many factors and often involves a compromise between transfection rate and cell survival rate. General cell culture conditions such as cell density/confluency, and cell health shown by markers like viability and division rate impact the likelihood a transfection experiment will be successful. The cell’s specific ability to bind and internalize reagent-nucleic acid complexes via endocytosis also impacts transfection rate. Whether the delivered target gene is expressed is also strongly dependent on the host cell and its viability/metabolism, and release of the nucleic acids into the cytoplasm and nuclear uptake can be increased by the transfection reagent.
Keeping cells alive
Many well-known lipid-based transfection reagents have high cytotoxic effects – even when using robust cell lines – with up to 50% loss of viability. This means these reagents are not well suited for transfection of sensitive cell types such as primary cells. Electroporation to increase permeability of the cell membrane may increase transfection rates but also results in cell death, with up to 90% of cells killed. Primary cells are particularly sensitive to electroporation related death, and the electroporation process can be expensive.
Viral transduction is another option, but this can also be challenging in many highly differentiated and non-dividing end-stage cells such as macrophages, hepatocytes, dendritic or neuronal cells. Some vectors are unsuitable for non-proliferating cells while others induce potent anti-viral responses or may influence cellular functions. Viral strategies are often time-consuming and involve certain safety risks.
MATra for faster, easier transfection
Magnet Assisted Transfection (MATra) technology – now exclusively available at PromoCell – is an innovative approach for a highly efficient and gentle transfection of numerous hard-to-transfect cell lines and primary cells – including macrophages, hepatocytes, neurons or stem cells. It is extremely fast – cells are transfected within 15 minutes – which decreases cell stress. MATra is also convenient and simple, saving time and money. It is particularly well suited for transfection of non-adherent cells and can easily be adapted to high-throughput transfection assays using robotic stations and adapted protocols.
MATra is a mechanical transfection method that enables cells to internalize transfection complexes without physically damaging the cells like electroporation or gene gun. Exploiting a strong magnetic force, specifically coated magnetic nanoparticles that bind/complex the nucleic acids (e.g. plasmid DNA, siRNA) are rapidly drawn down and concentrated onto the cell surface, and eventually internalized by endocytosis.
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