The aberrant expression of microRNAs (miRs) has been linked to several human diseases. A promising approach for targeting these anomalies is the use of small‐molecule inhibitors of miR biogenesis. These inhibitors have the potential to (i) dissect miR mechanisms of action, (ii) discover new drug targets, and (iii) function as new therapeutic agents. Here, we designed Förster resonance energy transfer (FRET)‐labeled oligoribonucleotides of the precursor of the oncogenic miR‐21 (pre‐miR‐21) and used them together with a set of aminoglycosides to develop an interbase‐FRET assay to detect ligand binding to pre‐miRs. Our interbase‐FRET assay accurately reports structural changes of the RNA oligonucleotide induced by ligand binding. We demonstrate its application in a rapid, qualitativ ...