MicroRNAs (miRNAs) are small non-coding RNAs that repress the translation and reduce the stability of target mRNAs in animal cells. Post-transcriptional regulation mediated by miRNAs is a highly conserved mechanism utilized by organisms throughout phylogeny to fine tune gene expression. We document the approaches used to study the function of a single miRNA and miRNA regulation of biological pathways in the sea urchin embryo. The protocols that are described include selection of miRNA inhibitors, test of miRNA direct targets, and the use of target protector morpholinos to evaluate the impact of miRNA inhibition on its targets. Using the described techniques and strategies, the sea urchin researcher will be able to validate a miRNA's direct targets and evaluate how inhibition of the miRNA affects developmental processes. These results will contribute to our understanding of the regulatory roles of miRNAs in development.