( n conditionsnamely electrical pulse len...)High Throughput siRNA Delivery In Vitro: From Cell Lines to Primary Cells

Figure 4. Gene Silencing and Cell Viability After Electroporation with the siPORTer-96 Electroporation Chamber. Silencer GAPDH (Ambion) or Negative Control #1 siRNA (Ambion) (1.0 g) were electroporated using the siPORTer-96 (Ambion) and siPORT siRNA Electroporation Buffer (Ambion) and conditions listed in panel A into 8 cell types. Conditions listed were for electroporating eight identical samples at a time. (panel B): 24 h later, the cells were harvested and analyzed by real time RT-PCR for GAPDH mRNA levels. 18S rRNA levels were used to normalize GAPDH expression. Remaining gene expression was calculated as a percentage of GAPDH mRNA levels relative to levels obtained from cells electroporated with negative control siRNA.

Reproducibility of High Throughput Electroporation. To assess well-to-well reproducibility of the high throughput electroporation system, a GAPDH siRNA (48 samples) or negative control siRNA (48 samples) were simultaneously electroporated into NHDF-neo cells using the siPORTer-96 and siPORT siRNA Electroporation Buffer. Reduction in GAPDH levels was monitored 48 hours later by real-time PCR. As shown in Figure 5, GAPDH mRNA was reduced by 9296% (average reduction: 94.8%). In addition, cell viability remained above 85% across the plate. These results indicated that the siPORTer-96 provided highly reproducible results. Unlike other
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