The extraordinary diversity and adaptive fit of organisms to their environment depends fundamentally on the availability of variation. While most population genetic frameworks assume that random mutations produce isotropic phenotypic variation, the distribution of variation available to natural selection is more restricted, as the distribution of phenotypic variation is affected by a range of factors in developmental systems. Here, we revisit the concept of developmental bias – the observation that the generation of phenotypic variation is biased due to the structure, character, composition, or dynamics of the developmental system – and argue that a more rigorous investigation into the role of developmental bias in the genotype-to-phenotype map will produce fundamental insights into evolutionary processes, with potentially important consequences on the relation between micro- and macro-evolution. We discuss the hierarchical relationships between different types of variational biases, including mutation bias and developmental bias, and their roles in shaping the realized phenotypic space. Furthermore, we highlight the challenges in studying variational bias and propose potential approaches to identify developmental bias using modern tools.