Liquid biopsies represent a valid alternative to conventional tissue biopsies, offering a real time molecular picture of tumors in a minimally invasive manner. Of the various circulating biomarkers available for liquid biopsy, circulating tumor cells (CTC) and circulating tumor DNA (ctDNA) are the most intensively studied to date. However, CTC and ctDNA represent different tumor components, therefore, complementary information from both sources might be beneficial. This protocol focuses on the description of a sample processing workflow that allowed for concurrent isolation of CTC and ctDNA from the same source sample. This single tube approach enables simultaneous analysis of multiple biomarkers to better monitor cancer drug resistance.