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Table 1 Summary of the CTC isolation and detection technologies

From: Detection of circulating tumor cells: opportunities and challenges

Name Basics Properties Limitations (Pre)clinical Application Recovery rate (%) Reference
Immunobead Assays
 CellSearch® system consists of ferrofluids coated with epithelial cell specific EpCAM antibodies; fluorescent labeling; immunohistochemical techniques omitting CTCs expressing low levels of EpCAM; low frequency and large blood volumes demand; cells captured appear to be more apoptotic FDA approved for advanced prostate, breast and colorectal cancers ≥85 [5, 7]
 Weissenstein et al a combination of anti-EpCAM and anti-cytokeratin magnetic cell separation cells captured appear to be apoptotic CTC levels were measured in MBC patients to assess prognostic value 78-90 [16]
 MagSweeper anti-EpCAM antibody-targeting immunomagnetic beads; characterize cells for multiple marker omitting CTCs expressing low levels of EpCAM; cells captured appear to be apoptotic capture live CTCs from BC patients for single cell analyses 55-69 [53]
 IsoFlux™ Rare Cell Access System a combination of flow control and immunomagnetic capture; Multiple kits for lab usage are available for cell enrichment and downstream analysis Maximum daily analysis is 12 samples biomarker heterogeneity of CTCs patients presenting ≥4 CTCs per blood draw were analyzed with prostate and colorectal cancers;
KRAS mutations rate in CRC were described
≥74 [90]
 AdnaTest® incubate blood samples with an antibody mixture (e.g. anti-EpCAM and anti-MUC1); a magnetic particle concentrator extracts the labeled cell detect CTCs via RT-PCR assay for tumor-associated transcripts biomarker heterogeneity of CTCs; magnetic beads may attach to the tube wall a combined analysis of CellSearch® and AdnaTest® leads to an improved detection of CTCs in mCRC patient;
prognosis prediction and efficacy evaluation in breast, prostate cancer
88 [43, 89, 109]
 CTC-μChip incubation with anti-EpCAM targeting immunomagnetic nanobeads; characterize gene expression using RT-ddPCR omitting CTCs expressing low levels of EpCAM CTC enumeration and genetic analysis in blood of patients with prostate cancer > 90 [91, 110]
 DynaBeads® bind to desired target and beads responding to magnetic field Only 3 types of DynaBeads are available for human tumor cell isolation enumerate CD4+ T lymphocytes in HIV-1-infected individuals;
not test in cancer patients
44 ± 23 [94]
 MACS CD45 leukocyte depletion method; utilize cytokeratin immunocytochemistry to analyze enriched cells CTC expressing CD45 maybe remove from the sample; erythrocyte lysis cause damage to CTCs CTC enumeration used in breast, lung, liver esophageal cancer patients;
morphologically intact tumor cells were not detected in the clinical application
70-88 [111, 112]
 EasySep anti-CD45 for removal of blood leukocytes CTC expressing CD45 maybe remove from the sample CTCs were detected in all of the BC patients (23/23) 24 ± 19 [94, 113]
 GILUPI CellCollector ex vivo Functionalized Structured Medical Wire is antibody coated and applied into peripheral arm vein
Isolation in vivo and overcomes sample blood volume limitations
Only used for extraction of CTCs directly from patient’s bloodstream in vivo isolation of CTCs in patients with different stages of prostate cancer;
distinguish between CTCs isolated from benign and malignant nodules
41 [106, 114, 115]
 3DPIC incubation with anti-EpCAM targeting immunomagnetic nanobeads
Utilize ATP luminescence assay for the detection of cancer cells in blood
extracellular ATP derived from non-CTCs may interfere with the measurement the ATP luminescence assay can detect as low as 10 cells in blood;
not test in cancer patients
80 [92]
Physical Property-Based Assay
 microfluidic ratchet mechanism distinguish CTCs based on cell deformability; deform cells in continuous flow without accumulating cells in the separation microstructure; the separated cells are available for downstream characterization cellular damage; throughput limitation detect CTC in a considerable proportion with clinically localized PC patients > 90 [67, 116]
 ISET® Utilizes a filter-based, size exclusion approach to isolate epithelial cells; high throughput morphology and size heterogeneity; damage or fragment CTCs on the result of multi-step cell processes ISET® has a relatively good detection rate for CTCs in BC and NSCLC patients;
fail to provide more information on pathological staging and molecular classification
75 [63, 64]
 Metacell Filtration® Size based separation technique driven by capillary-action; allow cytomorphological and immunocytochemical analysis of CTCs Filters have a larger pore size (8 μm) CTCs were detected in 66.7% evaluable PaC patients and the captured cancer cells displayed plasticity 66.7 [60, 117]
 ScreenCell size-based microfiltration; high CTC capture efficiency with processing 3 ml of blood per sample unable to capture CTCs smaller than WBCs; erythrocyte lysis may cause damage to CTCs the presence of CTCs does not influence prognosis in operated patients with NSCLC 89 [9, 118]
 Parsotrix™ size and compressibility-based platform for CTCs isolation; ability to capture CTC clusters; harvests CTCs with both epithelial and mesenchymal features CTC heterogeneity regarding size Parsortix-enriched and stained cells were successfully transferred with preservation of cell morphology;
not tested in clinical application
> 90 [65, 119]
 Dielectrophoresis (DEP) isolation based on polarizability and size; discriminate between cells of similar size having different morphological origins requires specific parameters such as cell type and electric field frequency; the extent to which DEP will be applicable of CTC isolation in different types of cancer is unclear concentrate MCF7 cancer cells from leukocytes;
not test in cancer patients
> 90 [51, 120]
 OncoQuick polypropylene tube is inserted above the separation medium which allows for elimination of unwanted blood cells;
High throughput, inexpensive
loss of sample while depleting mononuclear cells; detection depends upon only cytokeratine-20 biomarker detect epithelial cells by RT-PCR targeting CEA, CK20, and TEM-8 in colorectal carcinoma patients;
CTCs in breast cancer are correlated to bone marrow micrometastases
87 [55, 121]
 Ficoll density gradient centrifugation numerous cytospins had to be evaluated because of the low sensitivity; numerous “contaminating” MNCs in the enriched cell fraction lead to false-positive results detection of CTCs is of prognostic relevance in BCBM patients 84 [55, 122]
 AccuCyte® density-based cell separation; allows virtually complete harvesting of the red blood cells without cell lysis or wash steps cellular damage; viable cells recovery rate the median CTC count was 5 circulating prostate cancer cells/7.5 mL (range, 0-20) 90-91 [69, 123]
 RosetteSep unneeded cells are cross-linked with RBCs by specific antibodies to form a dense immune rose structure; unlabeled and highly purified target cells are left at the interface between plasma and density gradient centrifuge during density gradient centrifugation cause inherent cell loss and morphologic changes during the spinning and wash steps CTCs were detected in 54% (15/28) of MBC patients, 64% (16/25) of advanced stage HNC patients 36 ± 18 [9, 70, 95]
 SPPCN based on the surface charge of cancer cells serum protein-coated electrically charged nanoparticles can trap different cancer cells repeated magnetic separation and washing cause cells loss 2-8 CTCs has been isolated from 1 mL of blood;
only 0-1 CTC was detected from 10 healthy donors’ blood samples
50-89 [75]
 DEP-FFF Device DEP crossover frequencies of CTCs that are distinct from those of peripheral blood cell subpopulations and would permit them to be isolated from blood. throughput limitation;
Cannot be routinely applied in the biomedical and basic science labs
offer higher discrimination and throughput than earlier DEP trapping methods;
not test in cancer patients
92 [7]
 ApoStream® using dielectrophoretic technology in a microfluidic flow chamber; overcomes throughput limitations; high precision and linearity of recovery of viable; cancer cells may cause cellular damage be used to detect FRα(+) CTCs and may have clinical utility for assessing FRα levels in cancer patients;
detect EMT-CTCs among patients after neoadjuvant chemotherapy
75.4 ± 3.1;
71.2 ± 1.6
[72, 124, 125]
Functional Assays
 ELISPOT enriches cells via a depletion of the CD45+ hematopoietic cells and detects proteins shed/ secreted/ released from single epithelial cancer cells; a multi-parameter analysis revealing a CTC/DTC protein fingerprint requires efficient antigen binding and specific epitope presentation; high antigen levels demand; transition into in vitro cultures decrease cell viability and reduce detection rates measure the release of cytokeratin-19 (CK19) and mucin-1 (MUC1) in BC;
measure the release of PSA in prostate cancer;
[78]
 CAM assay based on CTC invasiveness compared to other cells; effective enrichment and identification based on CTC invasiveness; downstream analysis is possible. isolation step requires more than 12 hours; biomarker dependent capture invasive CTCs in mCRPC, mNSCLC and mPDAC 54 ± 9 [79]
 Nanoroughened Surfaces utilize the differential adhesion preference of cancer cells to nanorough surfaces adhesion strength of cancer cells might be affected by nanotopographic sensing; may cause cellular damage efficiently capture different kinds of cancer cells (MCF-7, MDA-MB-231, Hela, PC3, SUM-149);
not test in cancer patients
> 80 [79]
 TelomeScan Detects elevated telomerase activity via a telomerase-specific replication selective adenovirus May also detect hematopoietic stem cells for false-positive results The sensitivity of CTC detection was 69.1% in NSCLC patients;
Patients with positive EMT-CTCs at baseline had poor response to chemotherapy and decreased PFS
97 [81, 82, 126]
Microdevices
 eLoaD microfluidic platform Anti-EpCAM was immobilized on gold electrodes; quantifies CTCs by using label-free electrochemical impedance; CTCs expressing low levels of EpCAM are unlikely to be captured perform five different assays in parallel with linear dynamic range between 16,400 and (2.6 ± 0.0003) × 106 cancer cells/mL of blood;
not test in cancer patients
87 [98]
 NanoVelcro utilize an anti-EpCAM-coated SiNS to achieve significantly enhanced capture of CTCs Thermoresponsive NanoVelcro chips have demonstrated the capture and release of CTCs at 37 and 4 °C Only EpCAM-positive CTCs are detected clinical applications of each generation for various types of solid cancers (prostate cancer, pancreatic cancer, lung cancer, and melanoma) > 85 [99]
 iMECH deformation-based separation of CTCs from whole blood; enable label-free biomechanical profiling of individual cells for distinction; provide a low-cost yet high-throughput for single-cell level metastatic detection detect non-metastatic cells for false-positive results; may cause cellular damage MDA-MB-231 and MDA-MB-468 cells exhibit a loss of resistance;
not test in cancer patients
95 identified as metastatic [100]
 Size-Selective Microcavity Array separate cancer cells from the blood in accordance with differences in the size and deformability; approximately 98% of viable recovered cells; fast samples processing speed (200-1000 μL/min) clogging of cavities; size-heterogeneity detect approximately 97% of NCI-H358 cells in 1 mL whole blood spiked with 10-100 lung cancer cells;
not test in cancer patients
> 80 [127]
 PDMS microfiltration chip PDMS microfiltration membrane; size-based separation of CTCs from whole blood size-heterogeneity; balance the recovery rate and purity achieved great recovery from lung cancer cells spiked blood samples;
a high processing throughput of 10 mL/h;
not test in cancer patients
> 90 [53]