Overview - What is liquid biopsy?

DNA and RNA fragments are released into body fluids such as blood and urine via various physiological processes.  When released into the bloodstream, the so called cell-free DNA (cfDNA) and cell-free RNA (cfRNA) are collectively referred to as circulating nucleic acids in plasma or serum (CNAPS). The main source of cfDNA in the circulation is passive release from cells undergoing death mechanisms such as apoptosis and necrosis, as well as active secretion by living cells. The size of most cfDNA fragments ranges between 180 and 200 bp, but fragments as large as 300 bp also exist. The half-life of cell-free DNA is less than two hours before they are filtered out of the blood circulation by the spleen, liver and kidneys.

Similarly to healthy cells, primary and metastatic cancer cells release DNA fragments into the bloodstream. The resulting circulating tumour DNA (ctDNA) is characterised by genetic defects, such as point mutations, chromosomal rearrangements, abnormal epigenetic patterns and copy number aberrations. The genetic defects found in ctDNA are identical to those found in the tumor source and they are completely absent from normal circulating DNA. Since ctDNA is representative of the entire tumour genome, the analysis of tumour DNA from fluid samples is frequently referred to as whole body biopsy or “liquid biopsy”.

Graph: ctDNA release in blood circulation
Figure 1. Tumours can secrete fragments of ctDNA into the circulation where they are found in plasma. A liquid biopsy using blood samples obtained from cancer patients can be performed to detect ctDNA and to identify specific mutations that may have prognostic or therapeutic implications.

Other components of blood that could act as tumour surrogates include circulating tumour cells (CTC), circulating miRNA, exosomes and circulating vesicles.

Applications - What are the advantages of liquid biopsies?

The use of cell-free DNA holds great diagnostic potential for various clinical scenarios. Currently, the use of cfDNA as a non-invasive tumour tracker for cancer screening and diagnostics is captivating most of the attention of the life science community.

Cell-free DNA as a cancer biomarker

The ability to analyse circulating tumour DNA isolated from a simple blood draw provides rapid, affordable and non-invasive access to versatile biomarkers that accurately reflect changes in the tumour of origin. These characteristics of ctDNA make it attractive to a number of different areas in oncology.

Scheme workflow tumour diagnostic

The clinical utility of ctDNA analysis for cancer detection includes:

  • Assessment of molecular heterogeneity
  • Identification of specific genomic alterations to guide treatment selection
  • Monitoring of tumour burden
  • Early detection of therapy resistance
  • Detection of minimal residual disease
  • Evaluation of dissemination and recurrence risks
  • Understanding mechanisms of resistance

Liquid biopsy as a complement to tissue biopsy

Tissue biopsy is currently the standard of care for the molecular diagnosis of cancer. The conventional approach involves examination of tumour tissue by removing cells through a small needle or histological examination of a biopsy or surgical excision specimen. Molecular diagnosis of DNA mutations is then performed directly on fresh tissue samples or often on formalin-fixed paraffin embedded tissue (FFPE) material.

Despite the informative nature of solid tumour biopsies, tumor heterogeneity and clonal evolution present significant challenges in designing effective treatment strategies based on tissue biopsies alone. Molecularly targeted, personalised cancer therapies are dependent on serial monitoring of cancer genetics. Performing consecutive biopsies to capture a tumor’s spatial and temporal heterogeneity during tumour evolution is often difficult, risky, expensive and often times unachievable. Therefore, the need for novel approaches that could efficiently detect tumour heterogeneity in the course of cancer system treatments has arisen.  

Recent progress in genomic analysis of blood samples for ctDNA has made possible a real-time, affordable and non-invasive liquid biopsy approach for cancer detection and monitoring. Among other molecular insights, this novel diagnostics method provides important complementary information on therapeutic targets and drug resistance mechanisms in patients that are afflicted with cancer.

Other applications of cell-free DNA

Analysis of cfDNA analysis can be beneficial for other medical fields besides oncology. Analysis of cell-free fetal DNA is an established method for non-invasive prenatal testing (NIPT). Examination of fetal DNA can uncover point mutations and aneuploidy responsible for genetic disorders as early as seven weeks following conception. In Europe, fetal DNA found in the circulation of expecting mothers has been used for prenatal screening since 2012.

The state of cell-free DNA extracted from plasma can also serve as an indicator of pathogen infections, neurodegenerative, autoimmune and cardiovascular disease or as an early warning sign of organ rejection.

Workflow – How does liquid biopsy work?

Currently, blood-based cancer testing involves the assessment of biomarkers, most commonly cell-free DNA, circulating tumour cells (CTCs) or exosomes. Other prognostic and predictive markers of cancer that are at an earlier stage of development include cell-free RNA (cfRNA) fragments, methylated ctDNA and miRNA.

ctDNA is a particularly promising circulating biomarker for risk assessment in cancer patients due to the simplicity of obtaining plasma DNA, the cost-efficiency of the process and the high specificity and sensitivity of the resulting data.  As one of the most established techniques, liquid biopsies based on ctDNA analysis are closest to implementation in clinical settings. Several strategies exist for exploring tumour DNA found in plasma. Whole genome sequencing or whole exome sequencing approaches provide the most comprehensive view of tumour-associated mutations, but with lower sensitivity. Targeted approaches that are focused on specific genomic aberrations offer higher sensitivity, but no access to molecular information outside of the targeted regions of interest. Mutations that are most often assessed using targeted approaches for liquid biopsies include EGFR, PIK3CA, KRAS, BRAF, TP53, HER2, GNA11, KIT, MAP2K1, NRAS and others.

Next-generation sequencing of ctDNA isolated from blood follows a similar workflow regardless of whether specific targets or the whole genome is explored. The extraction of cfDNA from plasma is one of the most important steps and must be performed in optimal conditions to ensure downstream processing is successful. Following DNA extraction, library preparation protocols that are specialised for low-input DNA are followed. Next-generation sequencing is then performed on the appropriate platform and the resulting reads are subjected to thorough BioIT analysis. The final data is then delivered to a cancer researcher for a scientific interpretation or to a physician for a medical conclusion.

Workflow: sequencing of liquid biopsy samples
Figure 2. Example workflow for next-generation sequencing of ctDNA.

Scientific expertise: liquid biopsy

GATC Biotech is the first service provider to offer access to liquid biopsy-based assays with unprecedented flexibility for any stage or any project of blood-based tumour characterisation.

As a pioneer in the field of non-invasive diagnostics, GATC Biotech has already processed more than 40,000 blood and plasma samples under diagnostic conditions. Over the years, we’ve developed a proprietary protocol for highly efficient extraction of miniscule amounts of cell-free DNA from plasma. The DNA is sequenced in specialised NGS workflows. Additionally, proven approaches for target enrichment and variant analysis are included in the exome sequencing service.

All services offer sophisticated logistics, transparent LIMS-controlled sample and data processing, followed by convenient data delivery via our online platform myGATC. The GATCLIQUID services are processed in our own labs under ISO 17025 accreditation. ISO 13485 certification is available for special cases.


Find here, a list of selected research articles supported by GATC Biotech.

Wittenberger T. et al. (2014). DNA methylation markers for early detection of women’s cancer: promise and challenges. Epigenomics 6(3):311-327.

Related products to liquid biopsy

Are you ready to implement liquid biopsies in your cancer research studies? Find out how you can help advance personalised medicine through improved cancer detection and screening with our liquid biopsy service line, GATCLIQUID.

  • ONCOEXOME is the world’s only product for unbiased detection of all mutations in the protein coding regions of the cancer genome.
  • ONCOPANEL ALL-IN-ONE is a leading service for uniform and precise assessesment of more than 597 key cancer-associated genes.

Our GATCLIQUID services have complementary products which are fully compatible with tissue or FFPE samples (Figure 3.). The joint use of these services makes possible comparison studies of DNA from matched tumour tissue and plasma samples. These proof-of-concept publications are urgently needed for the clinical validation of liquid biopsies.

Cancer research related products
Figure 3. Overview of GATC Biotech products related to cancer research.

Further reading on liquid biopsies

Crowley E., Di Nicolantonio F., Loupakis F., Bardelli A. (2013). Liquid biopsy: monitoring cancer-genetics in the blood. Nat Rev Clin Oncol. 10(8): 472-484.

Diaz LA. Jr, Bardelli A. (2014). Liquid biopsies: genotyping circulating tumor DNA. J Clin Oncol. 32(6): 579-586.

Francis G, Stein S., 2015. Circulating cell-free tumour DNA in the management of cancer. Int J Mol Sci. 15: 14122-1442.

Gold B, Cankovic M, Furtado LV, Meier F, Gocke CD., 2015. Do circulating tumor cells, exosomes and circulating tumor nucleic acids have clinical utility? J Mol Diagn. 17(3): 210-224.

Heitzer E., Ulz P., Geigl JB. (2015). Circulating tumor DNA as a liquid biopsy for cancer. Clin Chem. 61(1): 112-23.

Qin Y, Ljubimov VA, Zhou C, Tong Y, Liang J., 2016. Cell-free circulating tumor DNA in cancer. Chin J Cancer 35: 36.