Detection of circulating tumor DNA (ctDNA) by digital droplet polymerase chain reaction (dd-PCR) in liquid biopsies

Circulating tumor DNA (ctDNA) analyses are minimally invasive and accessible for risk stratification or treatment response monitoring of cancer patients. Compared to tumor biopsy analysis, they are not only less invasive, but also provide a more representative picture, allowing capturing both tumor heterogeneity and multiple tumor sites. Development of new technologies such as droplet-based digital PCR (ddPCR) has greatly improved the sensitivity, specificity and precision of the detection of rare ctDNA sequences in the pool of total circulating free DNA. In this chapter, we discuss the application of ddPCR in the analysis of ctDNA, and present the protocols for obtaining variant allele frequency or copy number variation analysis.