Activation of telomere maintenance mechanisms (TMMs) is one of the major hallmarks of cancer. Cancer cells exploit two principal TMMs, telomerase (TEL) and Alternative Lengthening of Telomeres (ALT), to escape from normal proliferative constraints and achieve cellular immortality. Given that immortalisation is a characteristic of the great majority of human cancers, targeting TMMs has long been recognised as a promising therapeutic strategy for a broad range of cancer types. Yet direct inhibition of TMMs typically acts on a protracted timescale, because cancer cells must undergo multiple divisions before telomeres shorten sufficiently, allowing continued proliferation during treatment. Therefore, next-generation TMM-targeted therapeutics emphasise synthetic lethality (SL), targeting dependencies that arise upon activation of ALT or TEL. Therapeutics under development target either ALT or TEL, but there is evidence that some cancers utilise both TMMs. This raises the question whether these TMM-double positive (TMM-DP) tumours are heterogeneous (i.e., TMM-mixed) or contain cells with both TMMs active (TMM-dual) - scenarios with distinct implications for TMM-targeted therapy. To address this, we first surveyed 976 cancer cell lines and identified 17 that were classified as TMM-DP. We also developed methods for detecting telomerase activity by q-TRAP assay and the ALT marker, C-circles, in flow-sorted individual cells. We applied this methodology across human cancer cell lines, revealing the heterogeneity of telomerase activity and C-circle levels within single cells. Importantly, we demonstrated that TEL and ALT can co-exist within individual cells (TMM-dual) in vitro, and this dual-TMM state is maintained in vivo in xenografted tumours, confirming previous empirical evidence that these two TMMs can co-exist in cells. These results will have implications for the development of TMM-targeted cancer therapeutic strategies.