Xpress.4U- Transient Transfections

Axiogenesis exclusively offers a novel proprietary transfection technology for its human iPSC-derived cell port-folio. This opens up new opportunities for transient genetic modification of cells for drug development and disease modeling.

Xpress.4U technology overview

The novel liposomal formulation of Xpress. 4U facilitates single-step, rapid, and highly efficient transfection of iPSC-derived cells.

  • Transfection competent vesicles are formed around RNA cargo
  • Loaded vesicles fuse with plasma membrane of target cells
  • Cargo is directly released into cytosol, bypassing the endo- / lysosomal pathway
  • Instant bioavailability of cargo molecules in cytosol
  • Fluorescent tracer molecule in vesicles allows for verification of successful transfection or cell sorting in flow cytometry

 

 

Optimized transfection

Transfection has been optimized for efficiency and long-term stability using modified RNAs. 

Xpress.4U high efficiency transfectionHighly efficient transfection.  Overview images of Cor.4U® cells, transfected once for 10 minutes with ChR2-YFP mRNA using Xpress.4UTM. Cells were fixed 24 hours post-transfection and stained for nuclei with DAPI (blue).

   

 

Advantages

Protocols have been established for opto-genetic pacing of Cor.4U® iPSC-derived cardiomyocytes transfected with channel-rhodopsin-2 (ChR2; Figure 3) and for calcium transient analysis via transfection of Cor.4U® with GCaMP6f.

• Highly customizable
• siRNA-mediated knock-down
• Dominant-negative overexpression of diseased genes
• Genetically encoded sensors (e.g.,ChR2)
• Broad applicability

Application Examples

Cor.4U® can be paced up to 5 Hz after Xpress.4UTM-mediated transfection of ChR2 mRNA. Shown are representative MEA traces using Axion’s MaestroTM system with the LumosTM optical stimulation device. Optogenetic control of cardiomyocytes allows for investigation of beating rate-sensitive drug effects, avoids the need for frequency correction, and increases plate to plate reproducibility of drug effects.

 

Xpress.4U optical pacing of chr2 expressing Cor.4u

Optical pacing of ChR2-expressing Cor.4U®.  Light stimulation of control (upper panel) and transfected Cor.4U®

 

Physiological beat rates of ~70 bpm at 37 °C were obtained in measurements with the Hamamatsu FDSS 7000EX system. Using transfected cells circumvents the need for (often toxic) chemical fluorescent dyes, which may affect cardiomyocyte function. The use of encoded sensors also shortens experimental time, reduces manual work, and facilitates quality control of cells prior to the start of the experiment.

Xpress.4U calcium transients analysis using GCaMP6f transfected Cor.4UCalcium transients analysis using GCaMP6f-transfected Cor.4U®. 

 

Xpress.4U Benefits

• Non-toxic, highly efficient transfection (>80 %)
• Integration-free, transient transfection of RNA
• High stability through modified RNA
• Compatible with entire Axiogenesis iPSC-derived cell portfolio

 

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