As the stability and half-life of various mRNAs and their protein products varies, it is important to empirically determine the best time points for assessing target knockdown.

For example, it has been documented that in mammalian cells, mRNA half-life can range from minutes to days (Ross J, 1995, Microbiol Rev 59:423-50) while the half-life of protein products can range from less than a few minutes to several days. Taking this into consideration, the experimental design should allow sufficient time for the siRNA to associate with RISC and deplete mRNA/protein concentrations to desired levels. In general, the recommended time course ranges are 12 to 72 hours to deplete target mRNA and 24 to 96 hours to adequately knockdown target proteins and assess phenotypic outcomes.