A498, Caki-1, and 786-O, human renal cell carcinoma cell lines, were purchased from ATCC. Human immortalized macrovascular endothelial cells (ECRF24) were obtained from VU Medical Center Amsterdam, The Netherlands and NHDFα cells were courtesy of Prof. M. Cuendet (University of Geneva, Geneva, Switzerland). BCL2-Jurkat (CRL-2899™) and THP-1 cells (TIB-202™) were purchased from ATCC. All cells (Table S1 ) were cultured in a humidified incubator with 5% CO2 at 37 °C. A498 and Caki-1 cells were maintained in DMEM medium (Thermofisher, Basel, Switzerland, Gibco, 31966021), 786-O in RPMI medium (Gibco, 61870010), and ECRF24 in a 50:50 mixture of DMEM and RPMI in a flask pre-coated with 0.2% gelatin (Sigma Aldrich, Buchs, Switzerland, G1393-100ML). All media were supplemented with 10% fetal bovine serum (Biowest, Nuaillé, France, S1810-500) and 1% penicillin/streptomycin (Bioconcept, Basel, Switzerland, 4-01F00-H). NHDFα cells were cultured in a specified culture medium for fibroblasts, including a supplement kit (Vitaris, Baar, Switzerland, C-23110-PRO).
Thp 1 tib 202 cells
Manufactured by American Type Culture Collection
Sourced in United States
THP-1 (TIB-202) cells are a human monocytic cell line derived from an acute monocytic leukemia patient. They are commonly used in immunological and cell biology research.
Automatically generated - may contain errors
Lab products found in correlation
Caki 1, by American Type Culture Collection (1 mentions)
Caki 1, by Thermo Fisher Scientific (1 mentions)
Penicillin streptomycin, by BioConcept (1 mentions)
Fetal bovine serum (fbs), by Biowest (1 mentions)
Rpmi 1640 medium glutamax supplement, by Thermo Fisher Scientific (1 mentions)
Rhgm csf, by Miltenyi Biotec (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
24 well plate, by Corning (1 mentions)
100 μm cell strainer, by BD (1 mentions)
Easysep cd11b positive selection kit, by STEMCELL (1 mentions)
Caco 2, by American Type Culture Collection (1 mentions)
Poly d lysine coated, by BD (1 mentions)
Countess automated cell counter, by Thermo Fisher Scientific (1 mentions)
Hek293t cells crl 3216, by American Type Culture Collection (1 mentions)
Universal mycoplasma detection kit, by American Type Culture Collection (1 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (1 mentions)
Phorbol myristate acetate (pma), by Merck Group (1 mentions)
Hek 293 crl 1573 cells, by American Type Culture Collection (1 mentions)
Sepmate tube, by STEMCELL (1 mentions)
Lymphoprep, by STEMCELL (1 mentions)
7 protocols using thp 1 tib 202 cells
1
Culturing Diverse Renal Cancer Cell Lines
2
THP-1 Macrophage Differentiation and Primary Monocyte Isolation
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The human monocytic cell line THP-1 cells (TIB-202) were purchased from ATCC. Cells were cultured in 24 well plates (Corning) and differentiated into THP-1 derived macrophages 24 hours prior to infection with 40 ng/mL phorbol 12-myristate 13-acetate (PMA) in RPMI 1640 Medium GlutaMAX™ Supplement (Gibco) at 37°C and 5% CO2. Experiments were realized within 15 passages and cell viability was measured before experiments by trypan blue exclusion and was greater than 97%. CD14+ primary human monocytes were extracted from peripheral blood of 5 healthy donors anonymously provided by the French Blood Establishment (EFS, Lyon). CD14+ monocytes were purified from whole blood using an autoMACS® Pro Separator, Whole Blood Column Kit and StraightFrom® Whole Blood CD14 MicroBeads (Mitenyi) according to the manufacturer’s instructions. Cell viability was measured before proceeding to macrophage differentiation, by trypan blue exclusion and was always greater than 80%. CD14+ monocytes were then plated in 24 well plates (Corning) at a density of 400 000 cells per well and differentiated into macrophages for 5 days prior to infection with 50 µg/ml Rh-GMCSF (Miltenyi) in RPMI 1640 Medium GlutaMAX™ Supplement (Gibco) with 10% FBS (Sigma-Aldrich) at 37°C, 5% CO2.
3
Murine Myeloid and Human Monocytic Cells
4
Evaluating Crystal-Induced Cytotoxicity in THP-1 Cells
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THP-1 cells (TIB202), a human monocyte derived cell line, were obtained from the American Type Culture Collection (Manassas, VA). THP-1 cells were cultured in RPMI medium 1640 supplemented with 10% fetal bovine serum and 2-mercaptoethanol (0.05 mM) in T-75 flasks. For all experiments, cultured THP-1 cells were treated with CaOx, Cystine or CaP crystals (50, 100, 200, 500, 1000 µg/ml) or NaOx (0.1, 0.5, 1, 1.5, and 2 mM) and incubated at 37 °C in 5% CO2 for 24 h. Following treatment, cell viability was determined by the Trypan Blue exclusion assay. In brief, cells were treated with 0.4% Trypan Blue (1:1 dilution) and counted using the Countess Automated Cell Counter (Thermo Fisher Scientific Inc., Waltham, MA).
5
Culturing HEK293T and THP-1 Cells
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HEK293T (CRL-3216) cells and THP-1 (TIB-202) cells were purchased from ATCC. HEK293T cells were grown in Dulbecco’s modified Eagle’s medium (DMEM) with L -glutamine, 10% fetal bovine serum (FBS), and PenStrep 100 U/ml. THP-1 cells were grown in Roswell Park Memorial Institute (RPMI) medium 1,640 with L -glutamine and 10% FBS. All cells were grown at 37°C in a 5% CO2 atmosphere incubator. Cell lines were regularly tested for mycoplasma using the Universal mycoplasma detection kit (ATCC, #30-1012K).
6
HEK 293 and THP-1 Cell Culture Protocol
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HEK 293 (CRL-1573™) cells were obtained from American Type Culture Collection (ATCC, Manassas, MDVA, USA) and cultured in Dulbecco’s modified Eagle’s medium (Gibco™, Waltham, MA, USA; 11965118) with 10% fetal bovine serum (Gibco™, 16000044) and 1% antibiotic-antimycotic (Gibco™, 15240062) in a humidified atmosphere in a 5% CO2 incubator at 37°C. THP-1 (TIB-202™) cells were obtained from ATCC and maintained in RPMI-1640 Medium (Gibco™, 21875034) containing 10% fetal bovine serum, 1% antibiotic-antimycotic and 0.05 mM 2-mercaptoethanol (Gibco™, 21985023). THP-1 cells were then stimulated with 50 nM phorbol myristate acetate (Sigma-Aldrich, St. Louis, MO, USA; P8139) for 24 h for induction of differentiation into macrophages.
7
Isolation and Treatment of PBMCs
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Peripheral blood mononuclear cells (PBMCs) were purified from patient blood samples using Lymphoprep (Stemcell Technologies) and SepMate tubes (Stemcell Technologies, Vancouver, Canada). Lymphoprep was added to the lower compartment of the SepMate tube. Blood was mixed with PBS + 2% FBS in a 1 to 1 ratio, then layered on top of Lymphoprep following the company protocol. Samples were centrifuged at 800g for 20 min at 18°C with the brake off. The upper plasma layer was discarded. The PBMCs layer was removed carefully then washed with PBS and centrifuged at 300g for 8’ at room temperature between each wash. The pellet was stored at -20°C or isolated PBMCs used immediately following the experiments. 10X106 fresh lymphocytes sample, ID W313715036917, was received from HemaCare Corporation (Van Nuys, CA, USA). THP-1(TIB-202) cells were purchased from ATCC (Manassas, VA, USA) and maintained following the manufacture protocol. PBMCs and THP-1 cells were treated with human recombinant proteins rhGCase or rhα-Gal A, rapamycin (RAP), or a combination of RAP and 3MA in phenol red-free RPMI media with 10% FBS.
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