Cas9 nuclease may be used in vivo to create targeted genome modifications. There are several ways in which to introduce Cas9-guide RNA complexes into cells. Here we present a method for the transfection of Cas9 RNP’s into HEK293 FT cells using Thermo Fisher Lipofectamine® RNAiMAX. This is a ‘reverse transfection’ method that uses a final concentration of 10 nM RNP per transfection in a 96-well culture plate.