Isatin is an endogenous indole derivative that has been reported to possess various pharmacological and therapeutic activities of clinical importance. To date, isatin has been detected mainly by mass spectrometry. Here, we report for the first time the intense anodic electrochemiluminescence (ECL) of tris(2,2′‐bipyridyl)ruthenium(II) with isatin as an efficient co‐reactant. Interestingly, the ECL response of the Ru(bpy)32+/isatin system in borate buffer solution is about 12 times more intense than in phosphate buffer of the same pH; and approximately 114 times greater than the ECL intensity of the luminophore. Under the optimal conditions, the ECL of Ru(bpy)32+ increases linearly with the concentration of the isatin in the range of 1.0 μM to 1.0 mM (R2=0.998) with a limit of detection (LOD) of 0.28 μM (3σ/m). Moreover, the method has been successfully applied to the detection of isatin in human urine. The developed method has shown excellent recoveries in the range of 99.7–101.3 %. The anodic ECL assay reported in this work is superior in terms of linear range and selectivity than the previously reported methods and can be applied for the detection of isatin in real samples.