Microbubble‐mediated sonoporation for highly efficient transfection of recalcitrant human B‐ cell lines

Abstract

Sonoporation has not been widely explored as a strategy for the transfection of heterologous genes into notoriously difficult-to-transfect mammalian cell lines such as B cells. This technology utilizes ultrasound to create transient pores in the cell membrane, thus allowing the uptake of extraneous DNA into eukaryotic and prokaryotic cells, which is further enhanced by cationic microbubbles. This study investigates the use of sonoporation to deliver a plasmid encoding green fluorescent protein (GFP) into three human B-cell lines (Ramos, Raji, Daudi). A higher transfection efficiency (TE) of >42% was achieved using sonoporation compared with <3% TE using the conventional lipofectamine method for Ramos cells. Upon further antibiotic selection of the transfected population for two weeks, we successfully enriched a stable population of GFP-positive Ramos cells (>70%). Using the same strategy, Raji and Daudi B cells were also successfully transfected and enriched to 67 and 99% GFP-positive cells, respectively. Here, we present sonoporation as a feasible non-viral strategy for stable and highly efficient heterologous transfection of recalcitrant B-cell lines. This is the first demonstration of a non-viral method yielding transfection efficiencies significantly higher (42%) than the best reported values of electroporation (30%) for Ramos B-cell lines.

A multiplexed microfluidic and microscopy study of vasodilation signaling pathways using microbubble and ultrasound therapy

“…Cell monolayers can give insight into cell viability, gene transfection, and drug uptake [107]. For example, Yong et al have shown an increase of 42% genre transfection rate in B-cell lines over conventional transfection using cell monolayers [109].…”

DNA vaccines for prostate cancer

“…Sonoporation, for example, involves the use of ultrasound to cause a temporary increase in cell membrane permeability through the creation of pores, allowing the intracellular diffusion of macromolecules such as DNA (Fan et al, 2014). Sonoporation has recently been described for high-efficiency transfection of B cells (Ling Yong et al, 2014) and has been employed to increase in vivo delivery of a DNA vaccines to APCs. Using ultrasound (US)-responsive and APC-selective lipolex capsules to carry plasmid DNA, this targeted delivery approach was found to generate potent and sustained effects against solid and metastatic melanomas (Un et al, 2011).…”

In vitro methods to study bubble-cell interactions: Fundamentals and therapeutic applications

“…20,38 Using the same monolayer model, enhanced gene transfection has been reported more recently for human B-cell lines using plasmid-coated microbubbles, which were activated by ultrasound. 41 Monolayers formed from endothelial cells can act as artificial blood vessels, and implementing them in a microchannel format allows studying flow-based delivery of microbubbles and recapitulating physiologically relevant cellular shear stress, as would occur in vivo. 42 Alternatively, cell population studies rely on the use of suspended cells.…”