Intracellular Detection of Viral Transcription and Replication Using RNA FISH.

Many hemorrhagic fever viruses require BSL-3 or BSL-4 laboratory containment for study. The necessary safety precautions associated with this work often contribute to longer assay times and lengthy decontamination procedures. Here we will discuss recent advances in RNA fluorescence in situ hybridization (FISH) that not only allow entirely new investigations into the replication of these viruses but also demonstrate how this method can be applied to any virus with a known sequence and how it can be rapidly performed to minimize time spent in high containment. We have adapted existing protocols for mRNA detection with appropriate changes for examining viruses in a variety of containment laboratories (Shaffer et al., PLoS One 8:e75120, 2013; Raj et al., Nat Methods 5:877-879, 2008).