The broad-spectrum expression of telomerase in most malignancies makes it a promising target for a cancer diagnostic and prognostic tool. Conventional polymerase chain reaction (PCR)-based telomerase activity assay is highly sensitive but susceptible to amplification-related errors. Here, we present a novel approach to telomerase activity detection. The detection of telomerase activity is accomplished by the hybridization of electrochemilumi-nescence (ECL) nanoprobes to telomerase reaction prod-ucts, subsequent capture by magnetic beads, and in situ measurement of the light signal from ECL nanoprobes. The ECL intensity directly reflects the quantity of telom-erasereactionproducts, thustelomeraseactivity. Thehigh sensitivity afforded by the current magnetic bead and nanoparticle based ECL detection platform allows mea-suring of telomerase activity from as little as 500 cultured cancer cells in crude cell extracts without the PCR amplification of telomerase reaction products. In addition, a comparative study of the ECL nanoprobe and linear telomere antisense ECL probe was executed. By the employment of the ECL nanoprobe, a gain of about 100-fold elevation of sensitivity was determined. The method described here is ideal for telomer aseactivity analysis due to its reliability and high sensitivity.