Table 2.

CTC detection techniques.

CTC Detection Techniques
	Name	Commercially Available Formats or Providers	Mode of Detection	Antibodies	Material	Advantages	Disadvantages	References
Nucleic-acid-based detection	Reverse transcriptase polymerase chain reaction (RT-PCR)	Range of assays for selected diagnoses	Reverse transcription of CTC-specific mRNA to complementary DNA (cDNA) followed by PCR-amplification of cDNA	-	Blood, bone marrow, tissue, and other biological samples	High sensitivity	RNA instability, illegitimate expression, and false positivity; does not allow isolation of viable cells	[2,62,64]
Cytometry-based detection	Immunocytochemistry (ICC)	Range of assays for selected diagnoses	Antibody staining of tumor-specific antigens	Chosen based on proteins expressed in primary tumors	Blood, bone marrow, tissue, and other biological samples	Can be conjugated with automated imaging system	Limited number of cells evaluated, risks of cross-reactions with other epitopes, low sensitivity	[70,71]
	Flow cytometry (FC)	Range of assays for selected diagnoses	Quantification of surface and intracellular antigens using antibodies conjugated with fluorescent dye	Chosen based on proteins expressed in primary tumors	Blood, bone marrow, tissue, and other biological samples	Ability to measure multiple parameters of large numbers of cells relatively quickly. Cells can be isolated for further analysis	Low sensitivity, time-consuming	[72]
	Automated digital microscopy (ADM)	-	Fluorescence microscopy and robotic motion control system to automate imaging	Antibodies against tumor-specific biomarkers	Blood	Identification of very rare epithelial cells in whole blood samples	Enrichment step needed, long exposure time (800 cells/s)	[75,77]
	Fiber-optic array scanning technology (FAST)	FASTcell™ (SRI International, Menlo Park, CA, USA)	Image analysis of immunocytochemically labeled tumor cells	Antibodies to tumor-specific biomarkers	Blood	Does not require enrichment step	Special type of cytometer needed	[75,76,77]
Microscopy-based detection	Fluorescence microscopy	Range of assays for selected diagnoses, e.g., CytoTrack® (Cytotrack Aps, Lyngby, Denmark), RareCyte® (RareCyte, Inc., Seattle, WA, USA)	Optical microscopic examination of cells stained immunologically	Based on genes expressed in primary tumors, e.g., Anti-EpCAM and cytokeratins	Blood, bone marrow, tissue, other biological samples	Enrichment-free; automated microscopic imaging system; possibility of single-cell retrieval and further molecular characterization of CTCs; can be used for non-epithelial cells	Limited observation time, manual assessment necessary; long processing time; isolated cells are not viable	[11,25,79]