Notch signaling is probably the most widely used intercellular communication pathway. The Notch mutant in the fruit fly Drosophila melanogaster was isolated about 100 years ago at the dawn of genetics. Since then, research on Notch and its related genes in flies, worms, mice, and human has led to the establishment of an evolutionarily conserved signaling pathway, the Notch signaling pathway. In the past few decades, molecular cloning of the Notch signaling components as well as genetic, cell biological, biochemical, structural, and bioinformatic approaches have uncovered the basic molecular logic of the pathway. In addition, genetic screens and systems approaches have led to the expansion of the list of genes that interact and fine-tune the pathway in a context specific manner. Furthermore, recent human genetic and genomic studies have led to the discovery that Notch plays a role in numerous diseases such as congenital disorders, stroke, and especially cancer. Pharmacological studies are actively pursuing key components of the pathway as drug targets for potential therapy. In this chapter, we will provide a brief historical overview of Notch signaling research and discuss the basic principles of Notch signaling, focusing on the unique features of this pathway when compared to other signaling pathways. Further studies to understand and manipulate Notch signaling in vivo in model organisms and in clinical settings will require a combination of a number of different approaches that are discussed throughout this book.