pathology and diagnostics Circulating tumour cells as liquid biopsy: technical aspects This article focuses on the current technologies used for the enrichment and detection of circulating tumour cells and the innovative technologies that have been developed to improve methods for detection Laure Cayrefourcq PhD Catherine Alix- Panabières PhD Laboratory of Rare Human Circulating Cells, University Medical Center of Montpellier, EA2415 University of Montpellier, France Cancer is one of the leading causes of death worldwide, and metastasis is responsible for more than 90% of the mortality of cancer patients. Metastasis occurs when tumour cells leave the primary tumour, travel through the bloodstream as circulating tumour cells (CTCs), and then colonise secondary tumour sites distant from the primary tumour. CTCs, which are cells that have become detached from primary tumours, are the most representative biomarkers of the biological functions of metastatic development and their analysis has the potential to reveal the key mechanisms of tumour progression. The analysis of CTCs in the blood of patients with cancer was termed ‘liquid biopsy’. 1,2 Blood samples can be obtained and analysed at the time of diagnosis and during treatment. The analysis of the liquid biopsy provides important information on the molecular properties of tumour lesions. This information contributes to the early detection of metastatic lesions and aids in the personalised treatment of cancer patients, such as prognostic evaluation, stratification of patients for targeted therapies, real-time monitoring of treatment efficacy, identification of therapeutic targets, and resistance mechanisms. Numerous clinical studies and meta-analyses, including large cohorts of patients, have shown that the number of CTCs is an important indicator of the risk of progression or death in patients with metastatic solid cancer (for example, breast, prostate, colon). 3–7 However, in-depth investigation of CTCs remains technically challenging. CTCs occur as very low concentrations of one tumour cell among millions of blood cells. Their identification and characterisation require extremely sensitive and specific analytic methods, which are usually a combination of enrichment and detection procedures. This article focuses on current technologies used for the enrichment and detection of CTCs. A number of innovative technologies to improve methods for CTC detection have been developed, including CTC microchips, filtration devices, quantitative reverse-transcription polymerase chain reaction (PCR) assays, automated microscopy systems and functional assays. Among the considerable number of promising CTC detection techniques that have been developed, the analytical specificity and clinical utility must be demonstrated for their introduction into clinical practice. 9 HHE 2018 | hospitalhealthcare.com Strategies for CTC enrichment CTC enrichment includes a large panel of technologies based on the different properties of CTCs that must differentiate them from the normal haematopoietic cells: (i) physical properties (for example, size, density, electric charges and deformability); and (ii), biological properties (for example, surface protein expression and invasion capacity; see Table 1). Biological properties Biological properties are mainly used in immunological procedures with antibodies against either tumour-associated antigens (positive selection) or common leucocyte antigen CD45 (negative selection). Positive enrichment typically attains high cell purity, which depends on antibody specificity. Negative enrichment technologies evade some of the pitfalls of positive enrichment; for example, CTCs are not tagged with a difficult-to-remove antibody, they are not activated or modified via an antibody–protein interaction and antibody selection does not bias the subpopulation of CTCs captured. However, these advantages come at the cost of purity, as negative enrichment strategies typically have a much lower purity than positive enrichment 8–10 and require a suitable CTC detection step.