Since the early analyses of gene expression in the Drosophila embryo (1), whole-mount in situ hybridization has become one of the most powerful and versatile tools in developmental biology. The ability to visualize a gene’s expression both in time and space is a necessary first step in investigating the roles of that gene in cell differentiation and morphogenesis in the developing embryo. Unlike conventional in situ hybridization to tissue sections, the whole-mount procedure provides a three-dimensional readout of the sites of gene expression. This, combined with sequence analysis, allows an initial prediction of gene function and provides the basis for further investigation. Unique patterns of gene expression have also been used to define regions of developing tissue within areas that otherwise appear anatomically uniform.