Perturbations from environmental, genetic, and pharmacological sources can generate heterogeneous biological responses, even in genetically identical cells. Although these differences have important consequences on cell physiology and survival, they are often subsumed in measurements that average over the population. Here, we describe in detail how variability in adenoviral-mediated gene expression provides an effective means to map dose responses of signaling pathways. Cellcell variability is inherent in gene delivery methods used in cell biology, which makes this approach adaptable to many existing experimental systems. We also discuss strategies to quantify biologically relevant inputs and outputs. © 2011 Elsevier Inc. All rights reserved.