Protocol for simultaneous detection of dual subcellular localized dengue virus protease by co-transfection

It is difficult to track virus-coded proteins simultaneously if they localize to multiple subcellular organelles. Here, we present a protocol for the simultaneous detection of dual subcellular localized dengue virus protease by co-transfection. We describe steps for cell seeding, co-transfection with mitochondria targeted red fluorescent protein, cell fixation, permeabilization, and staining of transfected cells with Hoechst stain. Further, we describe how to generate fluorescent intensity profiles using the NIS-Elements software. We then detail procedures for subcellular fractionation followed by western blotting. For complete details on the use and execution of this protocol, please refer to Gandhi et al.¹.