ViaFect™ Reagent: Building Assays in Difficult Cells

The story of ViaFect begins with Promega Custom Assay Services (CAS), a group that uses Promega technologies to construct made-to-order assays, typically in a cell line. Many projects from the CAS group involve transfecting cells with expression vectors and reporter vectors. In some instances, customers contact CAS to have an assay constructed in a difficult cell line, after attempting and failing, or experiencing difficulty building the assay themselves.

CAS projects start with a proof-of-concept experiment using transient transfection before moving on to production of a clonal, stable cell line. For difficult cell lines, the CAS group previously turned to electroporation after exhausting lipid-based transfection options. Electroporation often worked, but success came with a price—cytotoxicity. The CAS group challenged R&D to find a better solution—better transfection with low toxicity for difficult-to-use cells. The result of that challenge is the ViaFect™ Transfection Reagent.

The figure demonstrates how ViaFect™ Transfection Reagent helped confirmed the concept, with a usable transient transfection prior to the selection of stable TF-1 suspension cells. On the other hand, as shown, electroporation did not result in significant transfection of the cells.

 Figure 1. ViaFect™ Transfection Reagent enables an assay for cytokine signaling in a hematopoietic cell model. TF-1 suspension cells were transiently transfected with pGL4.32[luc2P/NF-κB-RE/Hygro] Vector, an NF-κB response element luciferase reporter, using either ViaFect™ Transfection Reagent at a 2:1 reagent:DNA ratio or Amaxa Nucleofector® II (electroporation). The following day cells were stimulated with TNFα for 6 hours, and the response was measured with Bio-Glo™ Luciferase Reagent.


Figure 1. ViaFect™ Transfection Reagent enables an assay for cytokine signaling in a hematopoietic cell model. TF-1 suspension cells were transiently transfected with pGL4.32[luc2P/NF-κB-RE/Hygro] Vector, an NF-κB response element luciferase reporter, using either ViaFect™ Transfection Reagent at a 2:1 reagent:DNA ratio or Amaxa Nucleofector® II (electroporation). The following day cells were stimulated with TNFα for 6 hours, and the response was measured with Bio-Glo™ Luciferase Reagent.

Primary cells are often difficult to transfect due to their slow growth and high rate of death. ViaFect™ Reagent can be the perfect solution for such cells. These peer-reviewed articles demonstrate the use of ViaFect™ Reagent in the transfection of primary cells:

Cells Citation
Human Trophoblasts Straka, E. et al. (2016) Mercury toxicokinetics of the healthy human term placenta involve amino acid transporters and ABC transporters. Toxicology 340, 34–42.
Mouse Cortical Neurons Egusa, S.F. et al. (2016) Classic cadherin expressions balance postnatal neuronal positioning and dendrite dynamics to elaborate the specific cytoarchitecture of the mouse cortical area. Neurosci. Res. 105, 49–64.
Mouse Embryonic Fibroblasts Peng, Y. et al. (2016) AGE-RAGE signal generates a specific NF-κB RelA “barcode” that directs collagen I expression. Sci. Rep. 6, 18822.
Mouse Bone Marrow Macrophages Naujoks, J. et al. (2016) IFNs modify the proteome of Legionella-containing vacuoles and restrict infection via IRG1-derived itaconic acid. PLoS Pathogens 12, e1005408.
Mouse Alveolar Macrophages
Rat Aortic Smooth Muscle Cells Horita, H. et al. (2016) Nuclear PTEN functions as an essential regulator of SRF-dependent transcription to control smooth muscle differentiation. Nat. Comm. 7, 10830.
Human Renal Proximal Tubule Epithelial Cells Bethge, T. et al. (2015) Sp1 sites in the noncoding control region of BK polyomavirus are key regulators of bidirectional viral early and late gene expression. J. Virol. 89, 3396–411.
Mouse Male Germline Stem Cells Huang, Y. et al. (2015) Specific tandem 3’UTR patterns and gene expression profiles in mouse Thy1+ germline stem cells. PLoS One 10, e0145417.
Mouse Endothelial Fibroblasts
Human fibroblasts Takahasi, M. et al. (2015) Normalization of overexpressed α-synuclein causing Parkinson’s disease by a moderate gene silencing with RNA interference. Mol. Ther. Nucleic Acids 12, e241.
Human Umbilical Vein Endothelial Cells Yokota, Y. et al. (2015) Endothelial Ca2+ oscillations reflect VEGFR signaling-regulated angiogenic capacity in vivo. eLife 4, e08817.
Human Keratinocytes Nakmura, T. et al. (2014) Epiprofin orchestrates epidermal keratinocyte proliferation and differentiation. J. Cell Sci.  127, 5261–72.
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Kyle Hooper

A former Technical Services Scientist, Kyle has also worked with R&D for product development and now specializes in supporting Promega cellular analysis products in North America.

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