Cell Transfection

Cell transfection is the deliberate artificial introduction of nucleic acid (DNA or RNA) into mammalian cells. Typically this process is utilized to study gene function and protein expression. Sometimes confused with term “transformation”, the key differential factor is transfection is nearly always used in eukaryotic cells, whilst transformation is commonly referred to in bacterial work. The terminology difference is largely historically based, however, and the distinction is minimal.
Among differentiating between the methods developed for transfection-based research, it is fundamental to understand both transient and stable process types considering their role in the end product. Specifically that transient transfection is typically applied for short term observation, whilst stable transfection is used for longer term gene study. Furthermore, their interaction with rAbs vary, and therefore need to be selected with care considering the desired effect.

The transfection method began as a research technique in the 1970’s when Vaheri and Pagano used DEAE-dextran, and Graham and Van der Eb used calcium phosphate for cell transfection experiments. These chemical-based methods were revolutionary at the time as they did not have to rely on viral based methods to introduce genetic material into cells. Once PCR was discovered, this accelerated the development of applications to introduce different types of genetic material into cells.

The modern era of cell transfection deals with introducing cargo such as plasmid DNA, mRNA, proteins, and small RNA (siRNA, microRNA, piRNA) into cultured cells in vitro and targeted tissue delivery in vivo. Experiments are designed and performed by testing the effect of these cargo molecules in cells. However, delivery efficiency depends on specific transfection reagent. Transfection reagents are specifically designed, developed, and optimized for each type of cancer cell, difficult-to-transfect cell lines, sensitive cells (such as primary and stem cells, as well as hormone-dependent neuronal cells, primary keratinocytes and aortic endothelial cells), adherent and suspension culture (such as primary T-cells).

The goal in any transfection experiment is high transfection efficiency, meaning good expression of the construct with minimal cytotoxicity. Achieving minimal stress to cells and reproducibility across experiments is influenced by the reagent used, and optimization of transfection protocol. When transfecting DNA or RNA, the quality and purity of nucleic acid also has an influence on transfection efficiency. Also, a low passage number of cell line allow cells to be more amenable to uptake the transfection complex with cargo nucleic acid. There is no single method which can be reproduced across cell types and experiments. Each individual transfection must be optimized. However, cell line specific transfection kits with optimized transfection protocols are available from Altogen Biosystems.

Primary cell cultures are used in biological and gene therapy studies and serve as important model systems that may more accurately represent the biology of normal cells. Many cultured cell lines, as well as the majority of primary cell cultures, are able to be transfected with exogenous nucleic acids when appropriate transfection approaches are employed. Since the majority of transfection methods causes significant toxicity in primary cell cultures, optimizing this procedure (specifically the protocol and reagents to be utilized) is essential for developing effective transfection strategies for a given cell type. Altogen kits for primary cells and sensitive cell lines have been designed to have significantly lower cytotoxicity than other alternatives.

In Vivo Transfection Reagents from Altogen Biosystems

RNAi has been used for in vivo target validation studies using animal models. The major challenge in performing RNAi studies in vivo is the effective, directed delivery of functional small RNA molecules into specific tissues. Altogen® In Vivo Transfection Reagents could be conjugated with siRNA (or microRNA) and administered intratumorally (i.t) or systemically via intravenous (i.v) tail vein injection in order to provide directed gene silencing in specific tissues, including liver, pancreas, kidney, and tumors. Selective knockdown could be seen as early as 24 hours after injection.

Featured articles:

siRNA Transfection || Transfection Methods || Stable Transfection || RNAi || siRNA Library Screening || RNAi Therapeutics || In Vivo Transfection Reagents || Cell Line Specific Transfection Reagents

Featured in vivo transfection products from Altogen Biosystems:

In Vivo Transfection Reagents  ||  In Vivo Nanoparticle Transfection Kit  ||  In Vivo PEG-Liposome Transfection Kit  ||  In Vivo Lipid Transfection Kit  ||  In Vivo Polymer Transfection Kit  ||  In Vivo Pancreas-targeted Transfection Kit

Altogen Custom Services provides cell transfection services and other specialized biotechnology and pharmaceutical services, including over 50 validated xenograft models, development of stable cell lines, RNA Interference (RNAi) services, assay development, ELISA and Western Blot services, siRNA library screening and transfection services. Generation of stably-expressing cell lines can be very expensive and time-consuming. Altogen Labs offers generation of stable cell line service completed in just 28 days (see service details).