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Related Experiment Video

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Optimizing the process of nucleofection for professional antigen presenting cells.

Christina Susanne Mullins1,2, Tabea Wegner3, Ernst Klar4

  • 1Molecular Oncology and Immunotherapy, Department of General Surgery, University Hospital Rostock, Schillingallee 35, 18057, Rostock, Germany. christina.mullins@uni-rostock.de.

BMC Research Notes
|September 26, 2015
PubMed
Summary

Nucleofection effectively delivers antigens to both dendritic cells (DC) and B cells. While both cell types show similar nucleofection efficiency, plasmid DNA offers more potent and sustained antigen expression compared to in vitro-transcribed mRNA.

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Area of Science:

  • Immunology
  • Cell Biology
  • Molecular Biology

Background:

  • Antigen delivery is critical for immunological therapies.
  • Nucleofection, a form of electroporation, facilitates antigen delivery into cells, including the nucleus.
  • Dendritic cells (DC) and B cells are key antigen-presenting cells (APCs) for immune response induction.

Purpose of the Study:

  • To compare the nucleofection efficiency and antigen expression levels in B cells versus dendritic cells (DC).
  • To evaluate the suitability of plasmid DNA and in vitro-transcribed (IVT) mRNA for antigen delivery into APCs.

Main Methods:

  • Nucleofection optimization was performed using nearly 100 programs for both B cells and DC.
  • Cells were transfected with either green fluorescent protein (GFP) plasmid DNA or GFP mRNA.
  • Transfected cells were analyzed for GFP expression and viability using flow cytometry.

Main Results:

  • Nucleofection is feasible for both DC and B cells using plasmid DNA and IVT mRNA.
  • No significant differences in nucleofection efficiency were observed between DC and B cells.
  • A reverse correlation between nucleofection efficiency and cell viability was noted.

Conclusions:

  • Nucleofection is a viable method for antigen delivery into both dendritic cells and B cells.
  • Plasmid DNA constructs lead to more potent and longer-lasting antigen expression than IVT mRNA.
  • IVT mRNA offers the advantage of avoiding genomic integration, while plasmid DNA provides enhanced expression kinetics.