Table 3 Comparison of the advanced technologies deployed for circulating tumor DNA

Technology

PNA-based mutant enriched PCR and melting curve analysis

Digital PCR and flow cytometry

TaqMan-based mutant enriched PCR

Next generation sequencing

Droplet digital PCR

Sample

10 ng (plasma, 1–2 ml)

(plasma, 2 ml)

10 ng

10–250 ng

10 ng

Genotyping

YES

YES

YES

YES

YES

Multiplex

YES

YES

NO

YES

NO

Running time/Workflow

<3 h/Sample

10 days/complicated

<3 h/sample

2 days/complicated

2 days/complicated

Machine

Real time PCR

Droplet digital PCR, Flow cytometry

Real time PCR

Library machine/PCR/NGS sequencer

Droplet digital PCR/Droplet generator

Sensitivity

0.1–0.01 %

0.1–0.01 %

0.1 %

1–5 %

0.1–0.01 %

Advantages

Only a real-time PCR system is required; higher sensitivity, specificity, and reproducibility; multiplexing; and short run time.

Quantitative analysis; multiplexing

Requires only a real-time PCR system; and short run time.

Multiplexing (target gene panel); Barcoding samples; Quantitative analysis; Detects de novo mutations.

Quantitative analysis

Disadvantages

Cannot detect novel mutations

Cannot detect novel mutations; requires an expensive system; and a longer assay time.

Cannot detect novel mutations or perform multiplexing.

Requires an expensive system; and longer assay time.

Requires an expensive system; longer assay time; and cannot detect de novo mutations.