Table 1.

Comparison of PCR-based approaches and next-generation sequencing for their use in cfDNA analysis and applications.

Methods	PCR-based approaches	Next-generation sequencing
Features	Highly sensitive (MAF 0.001%)	Less sensitive (MAF 0.1–10%)
	Fast to run	Long to run
	Limited multiplexing	High levels of multiplexing
	Easy to interpret results	Requires bioinformatics and computational resources
	Low cost	High cost
	Absolute quantification	Approximate quantification
Applications	Identification of single or few mutations for molecular diagnostics and guiding therapy	Identification of multiple mutations for molecular diagnosis and guiding therapy
	Tracking mutations during therapy	Tracking mutations during therapy
	Limited to mutations in assay	Discovery of new mutations
	Measuring mutation load to predict prognosis	Measuring mutation load to predict prognosis
	Monitoring for minimal residual disease	Monitoring for minimal residual disease requires UMIs and sequencing to redundancy
	Copy number alteration detection in single or few genes	Broad copy number alteration detection across whole genome
	Studies of emerging resistance with well-defined mechanisms	Studies of all emerging resistance
	Tracking recurrence of known mutations	Tracking recurrence of known or unknown mutations

Abbreviations: PCR polymerase chain reaction, MAF mutant allele fraction, UMI unique molecular indexes, cfDNA cell-free DNA.