Table 2: Summary of advantages and disadvantages of the methods of CTC enrichment. CTC, circulating tumor cell; CTC-Chip, circulating tumor cell chip; EPIPSPOT, epithelial immunospot; FACS, fluorescence-activated cell sorting = flow cytometric; FAST, fiber-optic array scanning technology; ICH, immunocytochemistry; ISET, isolation by size of epithelial tumor cells; MACS, magnetic cell sorting; MEMS, microelectromechanical system.

Methods of enrichmentAdvantagesDisadvantagesReferences

Nonspecific

Density
(OncoQuick, Ficoll, UNI-Set)
(1) Isolation of whole and living CTC,
(2) using another method of enrichment more specific (immunomagnetic beads),
(3) cytopathology, cytological staining, ICH, FISH, and so forth can be performed
(4) RNA, DNA extractions followed by RT-PCR or PCR, respectively, can be done.
(1) Nonspecific,
(2) rare CTC can be lost in the plasma fraction or trapped among erythrocytes and neutrophils,
(3) low and variable sensitivity,
(4) Depends on the type of CTC, temperature, and centrifuge,
(5) expensive.
[47, 65, 139, 140]

Size
(MEMS, ISET)
(1) Easy,
(2) precise counting of the cells per mL of blood, independently of the volume of the blood treated,
(3) Allowing cytopathology, cytological staining, ICH, FISH, and so forth,
(4) Allowing Microdissection followed by
(5) RNA, DNA extraction followed by RT-PCR or PCR respectively
(6) Avoiding multiple steps,
(7) Increasing sensitivity (1 single CTC can be detect from 1 mL of blood).
(1) Nonspecific,
(2) CTC can go through the filter,
(3) cells can be damaged,
(4) expensive, but less than the immunomagnetic beads.
[47, 68, 124, 141, 142]

Cytospin(1) Easy,
(2) Allowing cytopathology, cytological staining, ICH, FISH, and so forth,
(3) Allowing Microdissection followed by
(4) RNA, DNA extraction follow by RT-PCR or PCR, respectively.
(1) Increasing the mortality of the CTC.[65, 143146]

Lysis Buffer
(Qiagen)
(1) Easy,
(2) the cells harvested by this method can be reenriched or analysed by the methods already described,
(3) low cost.
(1) Increasing the mortality of the CTC,
(2) low sensitivity.
[147149]

Specific

ImmunoMagnetic Beads
(MACS system, CellSearch)
(1) Specific,
(2) morphological analysis of CTC, cytopathology, cytological staining, ICH, FISH, and so forth,
(3) multiple labelling of antigens on CTC,
(4) direct quantification of CTC.
(1) Subjective analyses for CTC identification,
(2) time-consuming screening of tumor cells,
(3) needing specific marker(s) and antibody available,
(4) expensive
[40, 150152]

EPISPOT(1) High sensitivity,
(2) detection of viable CTC,
(3) detection of secreted proteins
(1) Protein must be actively secreted, shed, or released,
(2) no identification and isolation of secreting cell possible.
[40, 150152]

FACS(1) High sensitivity,
(2) technique for counting, examining, and sorting microscopic particles (CTC) suspended,
(3) simultaneous multiparametric analyses of the physical and/or chemical characteristics.
(1) Need the apparatus,
(2) high cost.
[140, 153, 154]

FAST(1) High sensitivity,
(2) can detect rare events.
(1) Fluorescent dyeconjugated antibodies,
(2) specificity depend on the antibodies,
(3) very expensive.
[48, 89, 141, 155]

CTC-Chip(1) High sensitivity,
(2) detection of viable CTC,
(1) Detecting only cytokeratin ± CTC,
(2) needing to control precisely laminar flow conditions,
(3) expensive.
[40, 156]