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High Throughput siRNA Delivery In Vitro: From Cell Lines to Primary Cells

that the increased cytotoxicity at higher siRNA concentrations relates to the increased transfection efficiency of the method; we observe the same level of cytotoxicity when using standard transfection to deliver more than 100 nM siRNA.


96 Well Electroporation--For Primary Cells

While advances have been made in electroporation equipment and protocols [811], electroporation largely remains a process for single sample RNA and DNA delivery. We have created a device, the siPORTer-96 Electroporation Chamber, siPORT siRNA Electroporation Buffer, and associated protocols that enable the delivery of siRNAs to as many as 96 samples at once.

Efficiency and Cell Viability Associated with High Throughput Electroporation. We used the siPORTer-96 and siPORT siRNA Electroporation Buffer to deliver control siRNAs to several different cell types. The procedure consistently provided >70% reduction in target mRNA expression and >70% cell viability (Figure 4B). For many cell types, siRNA delivery efficiency and cell viability exceeded 90%. Time course studies using electroporation followed by real-time PCR revealed that target mRNA levels were reduced as soon as six hours after electroporation with maximal reduction at 2448 hours, depending on cell type (data not shown). Protein reduction lagged behind mRNA reduction with maximal protein reduction at 4872 hours post-transfection. Note that optimal electroporatio
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