As crucial pieces in the puzzle of cancer and human aging, telomeres and telomerase are indispensable in modern biology. Here we describe a novel exponential Isothermal amplification of telomere repeat (EXPIATR) assay-a sensitive, simple, and reliable in vitro method for measuring telomerase activity in cell extracts. Through a strategically designed path of nucleic acid isothermal amplifications, EXPIATR abandons the expensive thermal cycling protocol and achieves ultrafast detection: telomerase activity equivalent to a single HeLa cancer cell can be detected in similar to 25 min.