Blood samples and cell-free DNA (cfDNA) isolation

Blood was collected in Streck™ tubes during routine phlebotomy, and samples were shipped at room temperature overnight. 10 mL of blood was processed upon receipt to isolate plasma by centrifugation at 1,600 g for 10 minutes at 4°C. Plasma was immediately aliquoted and stored at −70°C. cfDNA was extracted from 1mL aliquots of plasma using the QIAamp circulating nucleic acid kit (Qiagen), concentrated using Agencourt Ampure XP beads (Beckman Coulter), and quantified by Qubit fluorometer (Life Technologies, Carlsbad, CA, USA). All cfDNA sequencing and analysis was performed at Guardant Health (Redwood City, Calif, USA).