The invention relates to methods and assays for the detection of active Alternative Lengthening of Telomeres (ALT) activity in cells. The methods and assays involve detecting or assaying for partially double-stranded telomeric circles wherein the presence of said circles is specific for cells comprising an active ALT mechanism. In some embodiments the methods find application in, inter alia, determining the level of ALT activity in a cell, determining the ALT status of a cancer in a subject, diagnosing and/or treating disease, determining disease status, analysis of treatment efficacy, and the identification of novel therapeutic agents.