In the present work, electrogenerated chemiluminescence (ECL) behaviors of Ru(bpy)32+ were investigated at cucurbit[n]urils (CB[n]s, n = 5,6,7,8) modified electrodes in neutral condition, based on which a novel label-freestrategy for lysozyme (Lyz) detection was proposed. Strong anodic Ru(bpy)32+ ECL could be generated at themodified electrodes in the absence of coreactant. The ECL intensity increased in the following order: CB[5]bCB[6]bCB[7]bCB[8], and the strongest ECL signal was obtained at the CB[8] modified electrode. The ECL mechanismwas proposed based on the spectroscopic and the electrochemical results, and the excited state of Ru(bpy)32+ wasproven as the luminophor. Lyz exhibited apparent inhibiting effect on the ECL emission, and could be sensitivelydetected without any labeling process. Under the optimal conditions, the decreased ECL intensity varied linearlywith the logarithm of Lyz concentration in the range of 1.4 × 10−12-6.0 × 10−10 mg mL−1, and the detection limitwas calculated as 1.0 × 10−12 mg mL−1 (3σ). Moreover, the modified electrode exhibited high sensitivity andgood stability in the detection of Lyz. The results revealed a new role of cucurbit[n]urils in ECL sensing field,which will promote potential analytical application of macrocycle molecules.