TranslationBlocker Human ErbB2 / HER2 siRNA – DISCONTINUED

DISCONTINUED: Human ErbB2 / HER2 TranslationBlocker™ siRNA Duplex Product Attributes

1. 1. Starting with cultured cells in growth media seed 6 well plates with 3.5x10⁵ cells and incubate at 37°C.
   2. Prepare transfection reagent in serum-free growth media such as Opti-MEM-I. Allow to come to room temperature. The above data was generated with TransIT-TKO for delivery of ErbB2 / Her2 siRNA prepared in accordance with manufacturer’s instructions.
   3. Dilute siRNA to working concentration in serum-free growth media. For TransIT-TKO or similar transfections into 6 well plates, we recommend starting siRNA concentration optimization at around 75-100 pmol siRNA duplex in 200 ul of serum free media.Optimal dilution should be experimentally determined.
   4. Combine diluted siRNA and transfection reagent. Equivalent volumes of prepared TransIT-TKO or other transfection reagent and diluted siRNA should be combined and incubated at room temperature for 30 minutes with gentle mixing (optional).
   5. Add prepared siRNA solution to cells. For 6 well plates this will entail adding the solution to cells in approximately 2.5 ml serum-free media. Additionally, FITC labeled control siRNA (Product Number QC-1F) or positive control siRNA (Beta-Actin; Product Number QC-2) may be used to assess transfection efficiency.
   6. Incubate for several hours in serum free media. The above data was generated with a 18 hour initial incubation.
   7. Wash cells and add complete growth media
   8. Incubate and assay several hours after transfection. Optimal knockdown time should also be experimentally determined. Optionally, a second transfection can be performed 24 hours after the first transfection to enhance knockdown. ErbB2 / Her2 knockdown at the protein level was measured at 72 hours post transfection via western blot.