Table 2.
Culture protocols of CTC cell lines.
| Cell Line | Blood Volume | CTC Count/mL | CTC Enrichment and Isolation | Culture Type | Environment | Culturing Conditions | Culture Medium | References |
|---|---|---|---|---|---|---|---|---|
| CTC-1, CTC-2, CTC-3 | 20 to 45 mL | Undetectable by CellSearch | FACS (CD45, ALDH1, EpCAM) | 2D. single cell to colony formation | 5% CO2. Normoxic conditions | Adherent (1–8 days), medium change (8–21 days). The EpCAM(−)/ALDH1(+)/CD45(−) population was transferred to 24- or 6-well plates | Stem cell culture medium. Insulin, hydrocortisone, B-27, EGF, FGF-2, (1–8 days). EpiCult-C medium supplemented with 10% FCS (8–21 days). DMEM/F12 supplemented with 10% FCS (from day 22) | Zang et al. (2013) [9] |
| BRx33, BRx07, BRx68, BRx50, BRx42, BRx61 | 6 to 18 mL | 3–3000/6 mL | CTC iChip | 3D Spheroid | 4% O2 | Ultra-low attachment plate. Medium changes were performed under a microscope | Serum-free, EGF, FGF, B-27 | Yu et al. (2014) [10] |
| MSK-PCa5 | 8 mL | >100 count/8 mL | RosetteSep CD45-depleted Cocktail | 3D Organoid. Start to grow as spheroids after 10 days | N/A | Matrigel | DMEM/F12, EGF, R-spondin 1, Noggin, FGF10, FGF2, DHT, Nicotinamide Acros, A83-01, SB202190, Y-27632, B27, N-Acetyl-L-cysteine, Glutamax, HEPES, Primocin | Gao et al. (2014) [11] |
| CTC-MCC-41 | 10 mL | 302 count/7.5 mL | RosetteSep CD45-depleted Cocktail | 3D Spheroid | Initial environment: 2% O2. Maintenance: 5% CO2, Normoxic conditions | 24-well non-adherent plate. T25 flask (maintenance) | Initial medium: Stem cell culture medium, DMEM/HamF12 2% FCS, insulin, N2 component, EGF, L-Glutamine, FGF2. Second medium: RPMI1640 EGF, FGF2, insulin-transferrin-selenium supplement, L-Glutamine. Maintenance: N/A | Cayrefourcq et al. (2015) [12] |
| BHGc7, BHGc10 | N/A | N/A | Ficoll-Hypaque density gradient | 3D and 2D | Normoxic conditions | 12-well adherent plate. Normoxic conditions | Initial medium: Serum-free, RPMI-1640, insulin, IGF-1, transferrin, selenite, Maintenance: RPMI-1640, 10% FCS | Hamilton et al. (2015) [13] (2016) [14] |
| N/A (one line was named as CTC-TJH-01 in Que Z paper [20]) | 2 mL | 130 count/2 mL | Microfluidics-based immunomagnetic isolation. EpCAM coated and EGFR coated immunomagnetic microbeads | N/A | 3% O2, 5% CO2 (1–14 days). 5% CO2 Normoxic conditions | 96-well non-adherent plate. | Initial medium: RPMI1640, EGF, FGF, B27. Maintenance: RPMI-1640, 10% FCS | Wang et al. (2016) [15] |
| BHGc16, BHGc26, UHGc5 | N/A | N/A | N/A | 3D and 2D | Normoxic conditions | N/A | Initial medium: N/A. Maintenance: RPMI-1640, 10% FCS | Klameth et al. (2017) [16] |
| CTC44, CTC45 (both established from same patient) | 8–10 mL | N/A | RosetteSep CD45-depleted Cocktail | 3D | N/A | Ultra-low attachment 24-well plate | DMEM/F12, 2% FCS, L-Glutamine, N2 supplement, EGF, FGF2 | Grillet et al. (2017) [17] |
| BRx82, BRx142 | N/A | N/A | N/A | 3D | N/A | 6-well ultra-low adhesion plate | Initial medium: N/A, Maintenance: RPMI1640 EGF, FGF2, B-27 | Sandlin et al. (2017) [18] |
| CTC-MCC-41.4, 41.5A. 41.5B. 41.5C, 41.5D. 41.5E, 41.5F, 41.5G, (established from same patient) | 10 mL | 286/7.5 mL–3278/7.5 mL | RosetteSep CD45-depleted Cocktail | 3D | N/A | 24-well non-adherent plate (1st week). New 24-well non-adherent plate (2nd week). T25 flask (maintenance) | RPMI 1640, EGF, FGF-2, insulin-transferrin-selenium supplement, L-Glutamine | Soler et al. (2018) [19] |
| CTC-TJH-01 | 5 mL | 130 count | A mixture of EpCAM and EGFR coated immunomagnetic microbeads in microfluidic Herringbone-Chip | 2D | N/A | Non-adherent plate | Initial medium: RPMI1640, EGF, FGF, B27. Maintenance: RPMI-1640, 10% FCS | Que et al. (2019) [20] |
| CTC-3 | 6 mL | N/A | RosetteSep CD45-depleted Cocktail | 2D | Normoxic conditions, 5% CO2 | 6-well Matrigel-coated plate (2 weeks), normal 6-well plate (medium change every 2–3 days). Final culture in a T25 flask | DMEM/RPMI1640, 10% FCS, EGF, FGF, Nu-Serum, L-Glutamine | Zhao et al. (2019) [21] |
| CTC-ITB-01 | 7.5 mL | 1547/mL | RosetteSep CD45-depleted Cocktail | 3D and 2D | Normoxic conditions, 5% CO2 (1–14 days) | 96-well normal plate, then transferred to a 12-well culture dish | RPMI1640 10% FCS, L-Glutamine, insulin-transferrin selenium-A, FGF2, EGF, hydrocortisone, cholera toxin | Koch et al. (2020) [22] |
| UWG01CTC, UWG02CTC | 15 mL | 3/mL (UWG01CTC), 109/mL (UWG02CTC) | RosetteSep CD36-depleted Cocktail | 2D or 3D | Normoxic or hypoxic conditions | 24-well ultra-low attachment plate | DMEM/F12, EGF, FGF, N2 supplement (normoxic conditions) or DMEM/F12, 10% FCS (hypoxic conditions) | Brungs et al. (2020) [23] |
| Mel-167, PEM-22, Mel-182, PEM-78 | 10 mL | N/A | CTC iChip | 3D | Hypoxia, 5% CO2, 4% O2 | 24-well ultra-low attachment plate (4–8 weeks). Use of a 3D fibrin Matrigel culture. Finally, a switch to an anchorage-independent culture | RPMI 1640, EGF, FGF2, B-27, Heparin, Y-27632 | Hong et al. (2021) [24] |
| EMC-PCa-41 | 5 L | 5312/96 mL | Leukapheresis, RosetteSep CD45-depleted Cocktail | 3D Organoid | Normoxic conditions | 24-well plate Matrigel droplets. Organoids were collected and resuspended in a new plate | Initial medium: Prostate growth medium (PGM) or adjusted prostate cancer organoid medium (APCOM). Maintenance: AdMEM/F12 | Mout et al. (2021) [25] |
EGF: Epidermal growth factor, FGF: Fibroblast growth factor.