Circulating DNA can be released in the biological fluids by a physiological process and by different pathological conditions. The first reports detecting circulating DNA in human plasma date from the late 40s. Even when specific pathological conditions were analyzed, the clinical importance of circulating DNA remained unclear. After PCR introduction, genetic and epigenetic alterations in circulating DNA gained more prominence for understanding the mechanisms of cancer development and progression. Nowadays, the circulating DNA assays are highlighted for their clinical relevance for cancer screening in liquid biopsy. In this review, we described the landscape of studies on circulating DNA isolated from human plasma or serum and the molecular tools used to obtain these findings throughout the 20th century and the current application in cancer.