When performing siRNA-mediated knockdown experiments it is advisable to conduct a dose-response (concentration) analysis to determine the minimum siRNA concentration necessary for sufficient target knockdown on the mRNA, protein, or functional level. For Nucleofection®, the optimal siRNA concentration can range from lower than 2 nM up to 2 µM, depending on multiple factors such as the cell type, and the half-life of the mRNA and/or protein of the gene target. To determine the optimal concentration for your cell type and target, we suggest to perform an initial titration of the siRNA concentration within the range of 2nM-2µM (Nucleofector® 2b Device: 0.2 - 200 pmol in 100 µl; 4D-Nucleofector® X-Unit and 96-well Shuttle™ system: 0.04 - 40 pmol in 20 µl). Starting concentrations for a minimum titration would be 30 and 300 nM.