Comparative genomics has served as an essential guide in the identification of functional coding and non-coding sequences in vertebrate genomes. Human-mouse pair-wise comparisons have limited utility for identifying functional conserved non-coding sequences, owing to the large number of sequences shared between these species. In searching for more stringent filters to uncover non-coding elements more likely to be of functional importance in the human genome, human-fish sequence comparisons have emerged as an important strategy, leading to the efficient identification of enhancer elements. These sequences are unevenly distributed in the genome, tending to cluster around genes involved in key developmental processes, with recent studies suggesting that they represent genomic segments in which sequence variation can result in morphological changes and innovation. These elements, conserved over long evolutionary time, emerge as primary candidates that are likely to harbor sequence variation contributing to susceptibility of human disease phenotypes.