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Sep 1, 2013 (Vol. 33, No. 15)

Transfection Methods Evolving

  • In Vivo Applications

    Click Image To Enlarge +
    Schematic illustrating the range of current transfection technologies. [Life Technologies]

    An exciting transfection milestone is development of RNAi transfection tools for use in research applications as well as for the delivery of new biologics as therapeutic modalities.

    “Small molecules, like RNAi, are less difficult to transfect than DNA, which needs to enter the nucleus, typically, when the cell is dividing. RNA just needs to get into the cytoplasm, eliminating a step in the transfection process,” said Xavier de Mollerat du Jeu, Ph.D., senior staff scientist, molecular biology, Life Technologies. “RNAi transfection, in general, works for more cells lines, primary cells and in vivo.”

    Previously, there were no tools to deliver RNAi in vivo. Invivofectamine 2.0, a lipid-based RNA transfection system that protects the nucleic acid from degradation in the bloodstream, allows researchers to deliver RNAi to small animal models, such as mice and rats, and to directly perform functional and biologic analysis. In vivo transfection technology opens the door to rapid design and production of knockout animals facilitating genomic and proteomic research.

    “Transfection technology development will continue. In addition to primary, stem cells and in vivo applications, efforts also need to focus on plant and algae cells. These systems have yet to be optimized,” added Dr. de Mollerat du Jeu.

  • Transfection Topics and Trends

    According to Louise Baskin, senior product manager, Thermo Fisher Scientific, the primary applications for transfection have not changed dramatically in the past few years. Gene and protein expression studies, RNAi, and protein production remain the key applications utilizing transfection.

    “Driving the market is the desire to work with primary and stem cells, which are often resistant to standard transfection reagents and technologies, yet hold greater biological and medical relevance in many experimental systems,” she said. “There are also trends toward higher throughput studies, for which the transfection technology must also be scalable.”

    Achieving a balance of efficient transfection with low toxicity is the universal challenge, maintains Baskin.

    “Researchers would love a ‘silver bullet,’ a single delivery technology that would transfect into any and all cell types with low cell death,” she continued.

    “However, the existing market would indicate that there is a necessary mix of broadly applicable technologies (which work well in easy-to-transfect cells) and niche products that are optimized for only a few cell types, but are more difficult to transfect with conventional means.”

    Baskin believes that transfection technologies to support stem cell research remains an under-served part of the life science research market. The low tolerance for cellular disruption and low overall transfection efficiency by most existing techniques remains a challenge.

    “There is also a strong desire for improved in vivo transfection methods, especially for RNA interference,” she added. “To advance as a therapeutic, RNAi reagents must be able to reliably be delivered to particular tissues or organs, or only certain cells within those areas, to achieve their therapeutic goal.”

    Derek Levison, Ph.D., managing director of emp Biotech, stressed that rapid, cost-effective, large-scale transient transfection for the production of huge amounts of affordable recombinant proteins is still an elusive goal.

    Currently, transient transfection requires special cell culture media and rather large amounts of both DNA and transfection reagent, he explained, adding that it is unavoidably necessary to exchange culture medium prior to transfection when transfecting with polymers. For batches of small to medium size, this procedure is easy to handle, but for larger scales it is next to impossible, according to Dr. Levison.

    “To achieve truly cutting-edge technology status, it is essential to have a novel cell culture media which would allow both cultivation/production and transfection to occur without the need for media exchange,” he said. “Furthermore, there remains the need to reduce the large amounts of DNA and transfection reagent required. And the use of transfection reagents for gene therapy needs to be established.”

    The most appealing applications of transfection will be gene therapy and personalized medicine, pointed out Dr. Levision.

    “Imagine repairing a defective gene or gene sequence by simple injection of a therapeutic oligonucleotide packed within an advanced transfection complex,” he said. “Transient gene expression, combined with the knowledge gained from the human genome project and other sequencing milestones, should eventually enable in vivo protein production and therefore offers a truly promising vision for the future of human health.”

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